Background

ICBs are today the backbone of non-oncogene addicted NSCLC, both alone and in combination with chemotherapy (CT-ICB). To date, PD-L1 is the only predictive factor for ICBs benefit. Tumor burden is a new emerging additional biomarker to select pts who may derive a benefit from a CT combination compared to single agent ICBs. The biological correlates of tumor burden are largely unknown

Methods

In this multicentric study, 18F FDG PET scans were performed ≤ 42 days from 1st line initiation (CT, ICBs or CT-ICB). Total MTV was calculated with a threshold of 42% of SUV max. Progression Free Survival (PFS) and Overall Survival (OS) were analyzed with Kaplan Meyer method and log rank test. On a subset of pts with PET performed within 30 days from biopsy, transcriptomic data on fresh frozen tissue were correlated with MTV of the biopsied lesion.

Results

488 pts were enrolled, 162 treated with ICBs, 232 with CT-ICB and 94 with CT. The 3 groups were comparable in terms of MTV, sex, stage and PS, pts in ICBs are older (69y vs 64 for CT-ICB and 62 for CT).

MTV was negatively correlated with the outcome of ICI, with a median PFS of 31.8 months for 1^st quartile (1Q), 13.8 for 2Q, 4.7 for the 3Q and 1.8 for the 4Q (p < 0.001). A weaker correlation was found for CT-ICB, with a median PFS of 14.0m 1Q, 7.1 for 2Q, 7.0 for 3Q and 5.5 for 4Q (any PD-L1 expression), p 0.037, No correlation was found for CT ( p 0.696).

When comparing ICI to CT-ICB in patients with PD-L1 ≥ 50%, an advantage in PFS was seen for CT-ICB (10.7 vs 3.0m, p 0.039 ) for pts with MTV > median (100cc) as well as in 3 (73% vs 92%, p 0.043) and 6 months OS (66 vs 89%, p 0.033), but not in those with MTV < median.

Transcriptome analysis was performed on 48 pts and showed a negative correlation of MTV with GSEA immune signatures, as well as with RNAseq immune infiltrate quantification (Cybersort, p 0.016)

Conclusions

MTV identifies NSCLC pts with a worse prognosis and an increased rate of early progression and death on ICB. In the subgroups of PD-L1≥50%, pts with high MTV may do better on CT-ICB compared to ICB.

Transcriptomic analysis suggests that tumor microenvironment become increasingly immune suppressive with the increase in volume

Background

Adoptive cell therapy (ACT) using tumor-infiltrating lymphocytes (TILs) has proven efficacious in metastatic melanoma (Mel). Nevertheless, long-term clinical efficacy remains unsatisfactory for the majority of patients. Therefore, understanding TILs and the orchestrated tumor-microenvironment (TME) states associated with clinical response is of paramount importance.

Methods

We report the translational analysis of a single-center phase I study to assess feasibility and efficacy of TIL-ACT in Mel patients (NCT03475134). We performed comprehensive translational studies on patients` tumor longitudinal samples including multispectral immuno-fluorescence (mIF) imaging, bulk RNA-sequencing and single-cell RNA-sequencing before and after TILs infusion (30 days).

Results

Thirteen patients successfully completed TIL-ACT therapy and were included in the analysis. Best overall response was 46.1% (6/13) at 3 months, including two patients with ongoing complete response 3 years after infusion. Responders (Rs) exhibited immunogenic tumor cell states with higher inferred CNVs and overexpressed DNA sensing/IFN and class I antigen presentation-related genes. Multiplex IF imaging revealed that Rs had increased densities of polyfunctional intra-epithelial (ie)CD8⁺ T cells marked by higher PD-1 expression at baseline, and these cells persisted 30 days after TILs infusion. Single-cell transcriptomic analyses confirmed higher cytotoxic and exhaustion CD8 T cell states at baseline. PDCD1 (PD-1), CXCL13, TNFRSF9 (CD137), GZMB, HAVCR2 (TIM3) and PRF1 genes were overexpressed in CD8⁺ TILs and associated to clinical responses. Cell-cell interaction predictions corroborated by spatial neighborhood analyses revealed that tumors of Rs were highly enriched for ie-and stromal T-cell networks involving activated myeloid and costimulatory B cells. Successful TIL-ACT therapy reprogrammed the myeloid compartment and further increased CD8 TIL-myeloid cell networks.

Conclusions

Our systematic target discovery study reveals CD8 T-cell network-based biomarkers that could improve patient selection and guide the design of ACT clinical trials.

Clinical trial identification

NCT03475134

Background

The PRADO trial (n=99) tested different surgical and adjuvant therapy strategies based on the pathologic response after neoadjuvant IPI 1mg/kg and NIVO 3mg/kg in stage III melanoma patients (pts). The pathologic response rate (pRR: ≤50% viable tumor) was 72%, including 61% major pathologic responses (MPR: ≤10% viable tumor). After a median follow-up of 28.1 months, the 2-year (2y) event-free survival (EFS) rate was 80%. Here, we report the response and EFS data of PRADO according to the IFN-y signature and TMB.

Methods

TMB and the IFN-y gene expression signature (GES) were examined in baseline tumor biopsies by whole exome sequencing (n=76) and mRNA sequencing (n=80). Association with pRR, MPR or EFS was examined by logistic/Cox regression analyses. Cutoffs between high (H) and low (L) were calculated using ROC curves.

Results

Table 1 shows the association between IFN-y GES and TMB with pRR, MPR, and EFS. IFN-y GES and logTMB were not correlated (R = 0.065; p = 0.579). Pts with a high IFN-y GES had a higher pRR and MPR rate than pts with a low IFN-y GES (87% vs 50%, p=0.001 and 77% vs 35%, p<0.001, respectively), and also a higher 2y EFS rate (87% vs 68%, log-rank p=0.015). Pts with a high TMB were more likely to achieve a pathologic response (pRR 83% vs 58%, p=0.024) and MPR (80% vs 38%, p<0.001), but did not have a lower risk of relapse (2y EFS 77% vs 76%, log-rank p=0.531).

When combined, the pRR for pts with IFN-y H/TMB H was 90%, IFN-y H/TMB L 79%, IFN-y L/TMB H 67% and IFN-y L/TMB L 42%. For MPR rates this was 85%, 63%, 67% and 19%, respectively.

Conclusions

Similar to findings in our previous trials, baseline IFN-y GES and TMB were independent biomarkers for pathologic response and MPR, and the IFN-y GES was associated with EFS. However, TMB was not associated with risk of relapse, possibly owing to the different response-driven surgical and adjuvant therapy strategies in PRADO.

Clinical trial identification

NCT02977052

Background

Tumour-specific antigens are one of the principal targets in immune checkpoint inhibitor (CPI) treatment and fundamental to the development of personalised immunotherapy. The search for immunogenic neoantigens has primarily focused on mutations in protein-coding regions of genes. In this study, we investigate how novel open-reading frames (neoORFs) in the 5’ and 3’ untranslated region (UTR) of genes (generated by premature start-gain and stop-loss mutations respectively) contribute to the immune landscape of cancer.

Methods

We included two pan-cancer CPI-treated patient cohorts from the Hartwig Medical Foundation (HMF, n = 384), and Genomics England (GEL, n = 364). The frequency and length of UTR neoORFs for each patient tumour were predicted from whole genome sequencing data using PrimeCUTR, an R-based bioinformatics tool. Wilcoxon signed-rank test and survival analysis were used to assess associations with CPI treatment response and overall survival (OS).

Results

Within HMF, 75.5% of patients had one or more start-gain neoORFs while 30.5% had at least one stop-loss neoORF. Among those with start-gain neoORFs, 86.9% had neoORFs spanning 20 amino acids or more. These long neoORFs were most frequent in the grouping of lung and head and neck cancer. CPI treatment response was associated with a longer total length of start-gain (but not frameshift or stop-loss) neoORFs (P = 0.0047). Importantly, this association was also demonstrated in the subset of tumours with a low tumour mutational burden (TMB) (P = 0.014). Among GEL patients, survival benefit was specifically seen in patients with melanoma (n = 152), where those whose tumours had longer start-gain neoORFs (>29 amino acids, n = 63) had greater OS (HR 0.59, 95% CI 0.37 - 0.96, P = 0.03).

Conclusions

UTR neoORFs occur frequently in cancer, yielding a promising source of neoantigens. We designed a computational pipeline for identifying these neoORFs, which will be made available upon peer-review. We show that 5’ UTR start-gain neoORFs were associated with clinical response to CPI, even in the low TMB setting which is typically linked to reduced CPI response. These findings warrant further study to validate UTR neoORFs as a biomarker and target for personalised immunotherapy.

Background

ACT with TIL is personalized immunotherapy with promising results in MM. Recently, the positive outcome of a multicenter (National Center for Cancer Immune Therapy [Herlev, DK] and the Netherlands Cancer Institute [Amsterdam, NL]) phase 3 trial evaluating TIL in MM patients (NCT02278887) was recently announced. In this trial, the objective response (OR) rate for TIL-treated patients was 49% with 20% obtaining a complete response according to RECIST 1.1. However, there are currently no validated biomarkers that correlate with response to TIL. Identifying cellular subsets associated with response to treatment could in the future help guide clinicians in treating MM patients with TIL.

Methods

In this study, we analyzed the cryopreserved samples of the TIL infusion product from all patients with unresectable stage IIIC-IV MM treated with TIL in the phase 3 trial using multiparametric flow cytometry. The phenotypic characteristics of the TIL infusion product and OR to therapy according to RECIST 1.1 were investigated. The Mann-Whitney U test was used to test unpaired observations for statistical significance.

Results

In total, 80 patients received TIL, with 39 responders (complete + partial response) and 39 non-responders (stable + progressive disease). For 2 patients, the response was unevaluable. A higher number of TILs infused (Table 1) was significantly associated with response to therapy (p=0.013). Several lymphocyte subsets, including a higher number of CD8+ (p=0.004), CD8+CD28+ (p<0.001), CD8+BTLA+ (p=0.004), and CD8+CD39+ cells (p=0.007) favored response to therapy.

Median (95 % confidence interval)

Conclusions

In this study, we present an explorative immune characterization of the melanoma TIL infusion product and identify several cellular subsets that correlate with response to TIL therapy.

Background

ctDNA kinetics in the first 6 weeks and immune-related gene expression signatures can predict outcomes in patients (pts) treated with anti-PD-1 therapy. The earliest predictive timepoint in ctDNA kinetics for benefit to this therapy is unknown.

Methods

Pts with recurrent/metastatic head and neck squamous cell carcinoma treated with pembrolizumab or nivolumab monotherapy were prospectively enrolled. Plasma was collected at baseline (B), 2, 3, 8, 15, 22 and 29 days after first dose. Whole exome sequencing (WES) and RNA-seq were performed in archival tumor. ctDNA was analyzed using a bespoke 16 gene panel based on matched WES (Signatera^TM). Differential Gene Expression Analysis was performed with DESeq2. Gene Set Enrichment Analysis included different transcription factors (TF), molecular/immune pathways and PREDICT-IO signature (PMID 36055464).

Results

A total of 104 plasma samples from 15 pts were collected. ctDNA was detected in 90/104 (87%). Clinical Benefit (CB) rate, defined as complete (CR), partial response (PR) or stable disease (SD) > 6 months by RECIST 1.1, was 47% (1 CR, 3 PR, 3 SD). There were no differences in ctDNA quantity at B according to local/distant disease, primary site or PD-L1 status. A decrease in ctDNA at day 8 from B was associated with CB (100 vs 14%, p<0.01) and with longer progression-free survival (PFS) adjusted by Holm-correction for multiple comparisons (HR: 22.9 95%CI 2.5-211.3, p=0.02) compared to a stability/increase. This association was also seen at day 15, 22 and 29 but not at day 2 or 3. A steady decline in ctDNA beyond day 8 (↓ΔctDNA) was observed in 4 pts (3 PR, 1 SD). Immune-related pathways and TF were significantly enriched in CR/PR and ↓ΔctDNA. Oncogenic pathways and TF were enriched in pts with increase/stability beyond day 8 (↑ΔctDNA). PREDICT-IO High was associated with longer PFS (HR: 7.7 95%CI 1.5-38, p=0.005). All PREDICT-IO Low pts were ↑ΔctDNA (N=7). Within PREDICT-IO High pts, ↓ΔctDNA showed a trend to longer PFS than ↑ΔctDNA (HR: 6.9 95%CI 0.95-63, p=0.051).

Conclusions

A decrease in ctDNA by day 8 of anti-PD1 therapy is associated with CB and longer PFS. ctDNA kinetics can improve predictive value of PREDICT-IO High signature. Validations in larger pan-cancer cohorts are needed.

Clinical trial identification

NCT04606940

Background

Tertiary lymphoid structures (TLS) are ectopic lymphoid formations which are composed predominantly of B cells, T cells and dendritic cells. The presence in the tumor compartment of mature TLS - characterized by mature follicles containing germinal centers - has been strongly associated with improved survival upon cancer immunotherapies for patients with solid tumors. For this reason, TLS could be used as a predictive factor biomarker to identify the patients more likely to benefit from immune checkpoint inhibitors. However, the pathological assessment of the TLS status remains time-consuming and usually requires additional analysis including CD3/CD20 staining. This study aims to show that artificial intelligence techniques applied to digital pathology could offer a fast and cheap mass patient screening solution to assess TLS from Hematoxylin/Eosin (HE) digital pathology slides used in clinical workflow.

Methods

We analyzed a pan-cancer cohort of 289 HE-stained Whole Slide Images (WSI) from Institut Bergonié (43% NSCLC, 12% sarcoma, 45% other solid tumors) on which TLS were manually contoured by expert pathologists. A Deep Learning (DL) model was trained on WSI to predict TLS status at the patient level (presence or absence of TLS). Models were evaluated using five-fold cross validation. The best performing architecture included two main components: a predicted score of TLS presence on small areas (112x112 micrometers) of the WSI followed by an aggregation at the patient level. To assess the transferability of the model, a validation cohort (PEMBROSARC) of 236 sarcoma WSI was used.

Results

The best performing model achieved a ROC AUC score of 0.917 (std of 0.036) in five-fold cross validation on the pan-cancer cohort (specificity of 68% for a sensitivity of 90%). This model transferred very well on the PEMBROSARC study, the first clinical trial implementing TLS status as an inclusion criteria (Italiano et al Nature Medicine 2022) with a ROC AUC score of 0.89 (specificity of 64% for a sensitivity of 90%).

Conclusions

Our study demonstrates the predictive power of DL applied to predict the patient's TLS status from HE images. This model could be implemented in pathology labs as an efficient pre-screening tool.

Background

V-domain Ig suppressor of T cell activation, VISTA, is a type I transmembrane protein that functions as an immune checkpoint. VISTA can be expressed by a wide range of cell types. Malignant pleural mesothelioma (MPM) is a tumor type with the highest levels of VISTA expression. However, it remains unclear if VISTA could be a predictive or prognostic marker in MPM following PD-1 blockade.

Methods

The phase III PROMISE MESO study randomized 144 patients to either chemotherapy (vinorelbine or gemcitabine) or pembrolizumab. We analyzed 62 patients from the pembrolizumab arm and 57 patients from the chemotherapy arm with available tumour tissue. We performed multiplex IHC for the following markers/cell types: calretinin of WT1, CD8, CD14, CD66b, CD11c, VISTA, and DAPI.

Results

Most of the VISTA expression originated from the mesothelioma cells. Bioinformatic analysis of MPM RNAseq data showed enrichment for response to INFa and INFg with VISTA overexpression.

Responses to pembro or chemo were not influenced by global VISTA expression. However, VISTA expression on CD66b positive neutrophils negatively correlated with responses. The VISTA+/CD66b+ median value was 0.56 (IQR: 0.0-3.5), and the rounded median value of 1.0/mm² was used as threshold. Patients with VISTA+/CD66b+ high had an ORR of 5.6% whereas VISTA+/CD66b+ low tumors had 36.8%. Neutrophil infiltration alone did not correlate with responses. The difference in ORR also translated to prolonged PFS in patients with low VISTA+/CD66b+ cell counts (interaction p=0.0051), but no difference in overall survival was detected (interaction p=0.35). Neutrophils were not predictive of PFS or OS. In public single-cell expression data of lung cancer patients, VISTA expression indeed was higher in a subset of neutrophils, specifically in tN1 and tN3 subtypes.

Conclusions

Overall, our data show that VISTA expressing neutrophils correlate with outcomes. Our data underscore the complex biology of VISTA and suggest that specific targeting of VISTA on neutrophils might be a viable therapeutic option in a subset of MPM patients.

Clinical trial identification

ClinicalTrials.gov, number NCT02991482.

ETOP 9-15 PROMISE-meso trial

Background

DNA hypomethylating agents (DHA) combined with immune checkpoint inhibitors (ICI) are a promising strategy to improve the immunomodulatory and antitumor activity of ICI-based treatment. Providing initial support to this notion, guadecitabine plus ipilimumab in the phase Ib NIBIT-M4 trial, proved to be feasible, safe, and tolerable, with clinical and biological activity in metastatic melanoma patients (pts) (Di Giacomo AM, CCR 2019).

Methods

Pts enrolled in the NIBIT-M4 trial were assessed for long-term clinical outcomes. Molecular profiles of serial tumor biopsies performed at week 0, 4, and 12, including RNA, Whole Exome and Genome-wide methylation sequencing, were analyzed in an integrative manner and correlated with clinical data; pts were classified as Responder (R) or Non-responder (NR) based on disease control.

Results

Among the 19 pts enrolled (minimum follow-up 45 months), the 5-y OS rate was 29%; median duration of response was 20.6 months (95% CI,12.4-28.8). Analysis of longitudinal biopsies from 14 pts showed that primary/acquired resistance is underpinned by multi-level molecular mechanisms. Indeed, NR pts had higher mutation frequency in NRAS and harbored somatic alterations in genes involved in the Epithelial to Mesenchymal Transition (EMT) and chromatin organization pathways compared to R pts. Moreover, NR pts had a heterogeneous expression phenotype, characterized by activated patterns of proliferation/differentiation/EMT and by deactivation of INF-ɣ/HLA-II/immune-related genes. Conversely, R pts displayed dynamic patterns of activated immune signatures and favorable tumor-infiltrating microenvironment with T/NK cell clonal activation progressively increasing with therapy. We also developed a DNA-RNA immune stratification, simultaneously taking into account both the composition of the tumor microenvironment and the presence of reactive T-cells measured as the ratio between observed and predicted tumor neoantigens.

Conclusions

Our comprehensive longitudinal multi-omics analyses of tumors from the NIBIT-M4 study identify a landscape of biomarkers predictive of response that may guide patient stratification and selection in ICI plus DHA therapies.

Clinical trial identification

ClinicalTrials EudraCT, number 2015-001329-17,

ClinicalTrials.gov, number NCT02608437.

Background

Reactivation of tumor infiltrating T lymphocytes (TILs) with immune checkpoint inhibitors or co-stimulators has proven to be an effective anti-cancer strategy for a broad range of malignancies. However, epithelial ovarian cancer (EOC) remains largely refractory to current T cell-targeting immunotherapeutics. Therefore, identification of novel immune checkpoint targets and biomarkers with prognostic value for EOC is warranted. We here identified a TIM-3/CXCL13-positive tissue-resident memory (CD8/CD103-positive) T cell (Trm) population in EOC. Analysis of a cohort of ~175 patients with high-grade serous EOC revealed TIM-3-positive Trm were significantly associated with improved patient survival.

Methods

- Single cell mRNA sequencing data analysis.

- Immunohistochemical staining, image acquisition and analysis.

- Tumor Infiltrated Lymphocyte flow cytometric analysis.

Results

A cohort of EOC core samples was evaluated for the presence of tumor infiltrating CD8/CD103/TIM-3triple-positive T cells and subsequently correlated with patient survival. Interestingly, increased tumor infiltration of CD8/CD103/TIM-3triple-positive cells associated with improved patient survival in EOC, suggesting that CD8/CD103/TIM-3triple-positive TILs can serve as a prognostic marker for EOC. In line with this finding, a single-cell tumor immune transcriptomic dataset revealed upregulation of TIM-3, CXCL13 and CD103 within the terminally exhausted CD8-positive T cell fraction. Confirmatory flowcytometric evaluation of CXCL13 expression on isolated EOC TILs revealed that CXCL13 was predominantly found within the CD8/CD103/TIM-3triple-positive fraction compared to it’s single- and double-positive counterparts.

Conclusions

TIM-3 itself or as surrogate marker for prognostically favorable CXCL13-positive CD8-positive TILs may have prognostic value. As CXCL13-positive CD8-positive T cells have been strongly linked to patient response to anti-PD1 immune checkpoint blockade, combinatorial TIM-3 and PD-1 blockade therapy may be of interest for the (re)activation of anti-cancer immunity in EOC.

Background

Immunotherapy has become the standard of care for metastatic non-small cell lung cancer (mNSCLC) without a targetable driver alteration, yet we still lack insight into which patients (pts) will benefit from such treatments. To that end, we investigated characteristics of the immune infiltrate in the tumor microenvironment in relation to immunotherapy response. We report the results of an automated deep learning approach applied to digital H&E whole slide images (WSIs) of pre-treatment biopsies from the PEMBRO-RT clinical trial.

Methods

61 quality-checked H&E WSIs were processed with 3 deep learning algorithms. We extracted a tissue mask using an existing method (Bándi et al., 2019), and detected tumor and immune cells using HoVerNet (Graham et al., 2019). Tumor clusters were identified by combining the output of HoVerNet and tumor segmentation from an nnUnet (Isensee et al., 2021) model that we trained on external NSCLC images. From the output of this pipeline, we extracted immune infiltrate-based density metrics, calculated over all tissue (_allINF), stroma within 500um from the tumor border (_sINF), tumor region (_tINF), and the combination of stroma and tumor (_t+sINF). All metrics were used in ROC analysis after dichotomizing pts as responders and non-responders (response was defined as complete or partial response at any time point or stable disease for ≥12 weeks according to RECIST 1.1 measurement). Differences in metric distributions between the two groups were tested with a two-sided Welch t-test. Kaplan-Meier (KM) analysis was performed on progression-free survival (5-year follow-up).

Results

Our automated analysis reported denser immune infiltrates in responders, although not statistically significant (0.05<p≤0.2). All immune infiltrate metrics showed some predictive value with AUCs > 0.63, where _tINF reported an AUC of 0.70. KM analysis showed p=0.07 if pts were stratified based on the median _tINF, and p=0.02 if stratified based on the optimal operating point of its ROC curve.

Conclusions

Deep learning models that analyze the immune infiltrate density on H&E WSIs can identify mNSCLC responders to pembrolizumab.

Clinical trial identification

NCT02492568

Background

There is a lack of effective predictor for pCR of neoadjuvant immunotherapy in esophageal squamous cell carcinoma (ESCC) patients. High-throughput single-cell RNA sequencing (scRNAseq) can define the tumor microenvironment (TME), tumor heterogeneity and immune microenvironment. In this study, we sought to explore the immune pathways related to treatment response of immunotherapy by scRNAseq.

Methods

This was a single-arm phase II clinical trial, planned to enroll 20 locally advanced ESCC patients (cT2-4N0-1M0). Patients received neoadjuvant chemotherapy (paclitaxel-albumin + carboplatin) combined with tislelizumab and followed by minimaly invasive esophagectomy (MIE). We conducted a scRNAseq using fresh tumor tissues taken by endoscopy before neoadjuvant therapy. Histological outcome was divided into pCR (no residual tumor cells in tumor and lymph nodes), MPR (<10% residual tumor – excludes pCR) and non-MPR (>10% residual tumor).

Results

17 patients were enrolled since March,2022. 7 of 17 patients finished neoadjuvant therapy and MIE surgery. Among these 7 patients, 3 achieved pCR, 1 got MPR, and 3 were non-MPR, with a total of 42808 single cells sequenced. Compared to non-MPR, there were significantly increase in T lymphocytes (CD8+, T helper cell, Regulatory T cell) and mature dendritic cells, and significantly decrease in B cells and neutrophils in pCR patients. Morever, pCR patients showed greatly increase on the expression of PD-L1(CD274) in mature dendritic cells.

Conclusions

This study identified novel immunological predictor in baseline tissue for ESCC neoadjuvant immunotherapy. The enrichment of T lymphocytes (CD8+, T helper cell, Regulatory T cell) and mature dendritic cells together with the increase expression of PD-L1(CD274) in mature dendritic cells suggest a promising role of predicting completed pathological response (pCR) induced by neoadjuvant chemotherapy and immunotherapy in locally advanced ESCC patients.

Background

A number of coagulation-related studies found that fibrinogen was elevated in renal cell carcinoma (RCC) patients and correlated with more advanced disease. The impact of hyperfibrinogenemia on the clinical outcomes of immunotherapy remains uncertain.

Methods

In this prospective cohort study, fibrinogen levels were measured one week prior to the second-line nivolumab therapy (240 mg or 480 mg every 2 or 4 weeks) and then monthly until disease progression or unacceptable toxicity. High fibrinogen level was defined as = >5 g/L. Key eligibility criteria included metastatic clear cell RCC resistant to the first-line targeted therapy, <= 3 MSKCC risk factors, ECOG PS 0-2, platelet count <400×10(9)/L, no heart diseases, no anticoagulation therapy, and age 40-65 years. The primary endpoint was overall survival (OS) in cohorts with high (H) and normal (N) fibrinogen levels. Secondary endpoints included progression-free survival (PFS), objective response rate (ORR), and fibrinogen levels during therapy.

Results

Between November 2018 and January 2021, 82 patients were enrolled, of whom 41 and 41, respectively, were recruited in H and N cohorts. The groups were well balanced at baseline. The median follow-up time was 32.4 months. Compared to patients in N cohort, patients in H cohort had shorter OS (P=0.001), shorter PFS (P<0.01), and lower ORR (P=0.012). In H cohort, a below-median fibrinogen level was associated with longer observed OS (median, 23.7 vs. 19.4 months; P=0.03). In N cohort, 6 (15%) patients with stable disease had an increase in fibrinogen levels during nivolumab therapy. The table summarizes patients’ characteristics and clinical outcomes.

Conclusions

Detection of high baseline levels of fibrinogen in plasma derived from patients with metastatic RCC may be associated with poor clinical outcomes with nivolumab. These findings require large-scale prospective validation.

Clinical trial identification

KCRB20180412

Background

Accurately evaluating immuno-oncology therapeutics in pre-clinical models presents a significant technological challenge in translational oncology. Both human organoid models and humanized mouse models fail to recapitulate the complexity of the cancer tumor microenvironment found in the patient. Multiple stromal components such as immune cells, fibroblasts, blood vessels, and even bacteria have been shown to affect tumor response to treatment, suggesting the need for their inclusion and faithful reconstruction in pharmacological development.

Methods

To overcome these challenges, we have made use of a recently developed novel immune-competent ex vivo organ culture (EVOC) assay to evaluate the activity of CD3-based T-Cell Bispecific Antibodies (TCBs). In particular, we assessed the anti-tumor effects of a CEACAM5 TCB, which is designed to simultaneously bind CEA-expressing cancer cells and CD3-expressing T cells, activating the latter. Fresh tissues obtained during resection of NSCLC, CRC and PDAC tumors, were sliced and cultured ex vivo. The cancer tissue and microenvironment, including endothelial and immune cells, was preserved at high viability for 5 days. We then evaluated cytokine secretion in the assay after 24 and 48 hours.

Results

We found robust expression of T-Cell specific activation markers in response to treatment with CEACAM5 TCB antibodies. NSCLC and CRC tissues with abundant T-cells showed significant cytokine over-expression, while PDAC with low T-Cell presence showed none. Notably, comparing the histology of the fresh sample with the treated sample after 5 days, showed T-Cell infiltration from the tissue periphery into the cancer foci, demonstrating the efficacy of the bispecific antibody to recruit T-cells to tumor cells.

Conclusions

These results show the applicability of our ex vivo organ culture to evaluate immune activating compounds and suggests the ability to use EVOC, in the future, as a predictive biomarker for TCB therapy.

Background

Previous studies developed predictive biomarkers related to immune checkpoint inihibitors (ICI) based on clinical trials cohorts. Here, we leverage the ORIEN data conducted under the Total Cancer Care protocol across 18 collaborating cancer centers to evaluate the predictive value of published gene expression signatures in real-world data (RWD) of patients with cancer treated with ICI.

Methods

Overall Survival (OS) was the primary endpoint. RNA-seq performed on tumors was aligned to human reference genome (GRCh38) following RSEM pipeline. Gene expressions were quantified as TPM and log2(TPM+1) transformed. 28 published predictive gene expression signatures were collected and evaluated in our cohort (Table 1). Mann-Whitney U-test was used to compute the difference between groups, and Kaplan-Meier survival analysis was performed. Test with p<0.05 was considered statistically significant.

Results

659 patients treated with ICI for kidney cancer (n = 151), lung cancer (n = 138), melanoma (n = 123), head and neck cancer (n = 121), sarcoma (n = 78) and bladder cancer (n = 48) were included. We defined “good” and “poor” outcomes if OS was >24 or <24 months, respectively. Twelve immune active gene signatures were associated with ICI responses in melanoma (p < 0.05); Table 1. An angiogenesis signature (p = 0.0281) and a tertiary lymphoid structure signature (p= 0.0133) were associated with ICI responses in kidney cancer and head and neck cancer, respectively. None of the 28 evaluated gene signatures were associated with ICI responses in lung cancer, bladder cancer or sarcoma.

Conclusions

We validated the predictive value of immune related gene signatures in melanoma, kidney, head and neck cancers utilizing RWD. Ongoing analyses are taking on a discovery approach for predictive genes and related pathways to better understand the underlying mechanisms related to tumor immunogenicity across the different tumor types.

Clinical trial identification

ClinicalTrials.Gov NCT03977402

Background

SNPs of the cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) gene, an inhibitor of T cell priming, are associated with auto- and allo-immunity. Previously, studies implied a role for these SNPs as surrogate markers for outcome in melanoma patients treated with immune checkpoint inhibitors. However, no predictive SNPs are defined to date. The primary aim of this study was to analyze different CTLA-4 SNPs in a large real-world cohort of melanoma patients treated with ipilimumab and to correlate these SNPs with toxicity and survival.

Methods

Archival blood and/or tumor-tissue samples were collected from 317 advanced stage melanoma patients treated with ipilimumab (+/- nivolumab) in 5 Swiss and Dutch hospitals. DNA was extracted and used for MALDI-TOF MS based genotyping for 11 different SNPs. Associations between different allele genotypes and occurrence of grade ≥3 toxicity, treatment response and survival outcomes were tested using univariable logistic regressions or Cox proportional hazard models.

Results

DNA of 216/317 patients could be analyzed. The cohort was representative of a real-life population of metastatic melanoma patients receiving ipilimumab (+/- nivolumab): among the remaining 216/317 evaluable patients, 65.3% were male, median age at diagnosis was 59 years, 37% of patients had partial or complete response and 59% had ≥1 immune-related adverse events. A TT-genotype in the -1722 C/T SNP (rs733618, T=0.8345 (1000Genomes)), located in the promoter region of CTLA-4, was significantly associated with a decreased rate of ≥ grade 3 toxicity [odds ratio=0.40; CI₉₅=0.16 - 1.00; p=0.049]. The TT-genotype in the Jo27 T/C SNP (rs11571297, T=0.5575 (1000Genomes)) [hazard ratio (HR)=0.54; CI₉₅=0.28-1.02, p=0.056] and the GG-genotype in the Jo31 SNP (rs11571302, G=0.5713 (1000Genomes) [HR=0.54; CI₉₅ = 0.29-0.99, p=0.046], both located at the 3’ untranslated region, were associated with increased overall survival.

Conclusions

CTLA-4 SNPs might be predictive of toxicity and/or survival in melanoma patients receiving ipilimumab. Confirmatory studies are needed to exploit findings of this exploratory study and to investigate the exact role of CTLA-4 SNPs as possible biomarkers.

Background

Immunosenescence is a progressive remodeling of immune functions with a multifactorial etiology including aging and chronic antigenic stressors (inflammation, infections, cancer). We showed that a high pretreatment SIP (CD28^–CD57⁺KLRG1⁺CD8⁺ circulating T cells>39.5%, SIP+) was associated with resistance to ICB in patients with aNSCLC. Chronic viral infections may speed up immune aging, especially CMV which affects blood T cells phenotype (loss of CD28, overexpression of CD57). We aimed to assess antiviral serological profile and its association with SIP status.

Methods

Baseline SIP status was assessed by flow cytometry on fresh blood samples from ICB-treated and polychemotherapy-treated (PCT) aNSCLC patients. Sera from patients were analysed with VirScan (CDI Labs, USA), a high-throughput antiviral antibody (Ab) method (VirScan™) enabling simultaneous epitope-level antiviral antibody profiling of most viruses with human tropism (206 species, 1000 strains) via phage-display and immunoprecipitation sequencing (PhIP-Seq).

Results

132 aNSCLC patients were evaluable for SIP and VirScan™ assay. The antiviral serological profile seems similar between SIP+ and SIP- patients, except for CMV where the mean enrichment of anti-CMV antibodies was higher in SIP+. Of the 74 antiviral Ab associated with a higher SIP, 70 (94.6%) recognized CMV-peptides and CMV was the only virus globally associated with a higher SIP. SIP+ patients were predominantly CMV+ compared to SIP- (93% vs 57%, p<0.001), but 70.5% of CMV+ remained SIP-. In CMV+ patients, no difference was observed in the number of CMV epitodes targeted by antibodies (p=0.62) nor in protein immunisation profile between SIP+ and SIP- patients. Among all clinic-biological parameters studied only median age was higher in SIP+ (71 years vs 63 years, p=0.01) compared to SIP- in CMV+ patients.

Conclusions

Among 206 species, only anti-CMV Ab were associated with SIP+ status, which is associated with an older age in CMV+ patients. Further work investigating the predictive value of CMV in the response to ICB in older patients is ongoing.

Clinical trial identification

NCT03984318, NCT02105168

Background

Pre-existing immunity that describes the endogenous tumor-specific adaptive immunity, before treatment may represent a valuable novel predictive biomarker for ICI treatment. In this study we estimate the potential value of pre-existing cancer-antigen specific CD8⁺ T cells as circulating predictive biomarkers. Additionally, we evaluate the major differences of known immune cell phenotypes between Pre-existing positive (PreI+) and Pre-existing negative (PreI^-) NSCLC patients in circulation.

Methods

Blood was collected before initiation of immunotherapy from 24 NSCLC patients stage IIIb, PD-L1⁺ that receiving Durvalumab. PBMCs were isolated with Ficoll density gradient centrifugation including 15 healthy donors (HD). PreI was calculated by detecting endogenous IFNg expressing cells after in-vivo co-cultures of PBMCs with hTERT, MAGEA1, NY-ESO-1 and Survivin antigens. Immunophenotyping was performed by multi-color flow cytometry using antibodies against CD3,CD4,CD8,CD45RA,CD45RO,CCR7,PD-1,PD-L1 for CD4 and CD8 T-cells and CD3,CD4,FoxP3,CD25,CD127,CTLA-4,CD39 for Tregs.

Results

From 24 patients receiving Durvalumab 10/24 (41,6%) had peptide specific T-cells (PreI ⁺ patients). 60% had detectable peptide specific T cells for all (4/4) antigens tested. Survival analysis revealed better PFS in PreI⁺ than PreI ^– patients (Log-rank = 0.06, median 333,5 and 185 days ) and better OS (*Log-rank = 0.032, median undefined and 254 days). Patients with PreI⁺ and low CD25+CD127-CTLA-4+ Tregs (n=8) had better OS (*Log-rank 0,036, median undefined and 251 days) than those with PreI^- and high Tregs (n=9). The levels of CD3CD4PD-1 were higher in NSCLC patients compairing to HD (* 0,019) but there was not a difference between PreI⁺ and PreI^- patients. PreI⁺ patients had significantly higher numbers of CD3CD8PD-1 cells comparing to PreI^- (*p= 0,024) but not CD3CD8PD-L1. Prei⁺ patients had high numbers of Teff (CD3+CD8+CD45RA+CD45RO-CCR7-) compairing with both HD ( ** 0.0039) and PreI^- (*0.032). Both T cm and Tem do not differ between all tests.

Conclusions

Pre-existing tumor antigen specific T-cells in circulation before initiation of immune checkpoint inhibitors in NSCLC patients serve as a good prognostic factor of response.

Background

Although the use of immunocheckpoint inhibitors (ICIs) in cancer treatment has become widespread, the factors predicting the response to these treatments have not yet entered our clinical practice. Considering that the progression under ICIs is especially in the first 3-6 months, there is a need for biomarkers to help us determine which patients do not benefit from these treatments. It is important to detect an easy and inexpensive biomarker in order to be widespread in our clinical practice. In our study, we aimed to examine the relationship between mean platelet volume/lymphocyte ratio, which can be evaluated by hemogram test, and overall survival in cancer patients treated with nivolumab.

Methods

A total of 131 adult metastatic cancer patients treated with nivolumab for malignant melanoma (MM), renal cell carcinom (RCC), non-small cell lung cancer (NSCLC) and head and neck (HNC) cancer patients were included. Baseline demographics, ECOG performance status, type of tumors, and baseline blood count parameters were recorded. Possible predisposing factors were evaluated with univariate and multivariate analyses.

Results

The median age was 59.87 ± 11.97 years, and the median follow-up was 20.20 (IQR 12.80-27.60) months. Renal cell carcinoma (43.5%) and melanoma (25.9%) were most common diagnoses. Patients with ECOG 0-1 (mOS: 20.60 months (95% CI: 14.94-25.29) vs 5.24 months (95% CI: 0-16.42), p<0.001), LDH levels within the normal range (mOS: 24.54 months (95% CI: 14.13-34.96) vs 13.10 months (95% CI: 4.49-21.72), p=0.038) and low MPVLR (mOS: 33.70 months (95% CI: 25.99-41.42) vs 11.07 months (95% CI: 6.89-15.24), p<0.001) had a longer median overall survival. In multivariate Cox regression analysis, high MPV/Lymphocyte ratio, ECOG score 2-3 and high LDH level were associated with shorter overall survival time (p<0.001, p=0.001 and p=0.046 respectively).

Conclusions

In this study, we determined that MPVLR could be a new biomarker predicting response to nivolumab treatment.

Background

Novel immunotherapies targeting the adenosine pathway are in clinical trials, however, only modest monotherapy activity has been observed in non-prioritised patient groups. To optimise the chance of success with our A_2AR-selective antagonist, EXS-21546 (‘546; NCT04727138, discovered in collab. with Evotec), we work to identify an adenosine-induced immunosuppression biomarker signature for clinical trial patient selection.

Here we present initial transcriptional and functional data mapping the adenosine suppressed immune potential at the single cell level, and subsequent modulation through antagonism of A2AR with ‘546, along with first patient-selection modelling to prioritise patients for ‘546 therapy.

Methods

By leveraging disease-relevant primary human tissues, we model the patient specific anticancer immune potential, and begin to validate patient selection methods functionally with a translatable high content imaging platform amenable to primary human material. This platform is supported by end-to-end deep learning-driven image analysis; work is combined with orthogonal multi-omic characterisation of single cell effects induced by ‘546.

Results

We present preclinical mechanistic studies of A2AR antagonism on tumour infiltrating immune cells, leading to a foundational predictive adenosine suppression signature. The patient selection gene signature is correlated to functional profiling using a high content imaging platform with proven translational capabilities (Kornauth et al, Cancer Disc, 2022), demonstrating association of immune activity with antagonism of adenosine signalling by ‘546. Signatures and patient selection algorithms are cross-validated with publicly available data.

Conclusions

Combining deep learning of single cell functional and multi-omics profiling data of disease relevant primary model systems, we model the association of the immune response potential to A2AR antagonism in cancer to define a biomarker signature with the potential to identify patients likely to benefit from A2AR antagonism. This could be implemented in future studies of our clinical candidate ‘546 to deliver the right drug at the right time to the right patients.

Background

The introduction of immune-checkpoint inhibitors (ICIs) has revolutionized the treatment paradigm for advanced melanoma, leading to substantial benefit in overall survival (OS). However, several patients still do not benefit from ICIs and to date there are no predictive biomarkers for PD-1 inhibitors. PD-1 axis single nucleotide variants (SNVs) may affect receptor/ligand interactions in the tumor microenvironment and consequently the immune response and efficacy of ICIs. Based on this rationale, their predictive and prognostic role was investigated.

Methods

We analyzed, in metastatic melanoma patients treated with nivolumab or pembrolizumab, five PD-1 SNVs, namely PD1.3G>A (rs11568821), PD1.5 C>T (rs2227981), PD1.6 G>A (rs10204525), PD1.7 T>C(rs7421861), PD1.10 C>G (rs5582977) and three PD-L1 SNVs:+8293 C>A (rs2890658), PD-L1 C>T (rs2297136) and PD-L1 G>C (rs4143815) by pyrosequencing methods. Association of SNV genotypic frequencies with best overall response (BOR) to PD-1 inhibitors and development of immune-related adverse events (irAEs) were estimated through a modified Poisson regression. A Cox regression modelling approach was applied to evaluate the SNV association with OS. All regression analyses were adjusted for individual confounding factors.

Results

A total of 125 patients with advanced melanoma were included in the analysis. In patients carrying the PD-L1 C>T variant, a trend towards a lower relative risk (RR) of having progressive disease was observed in the C/T genotype (RR=0.60; 95%CI 0.25-1.41). In addition, a trend for a reduction in irAEs occurrence was observed in patients with PD1.7 C/C and PD-L1 +8293 C/A genotypes (RR = 0.35 [95%CI 0.09-1.31] and RR=0.45 [95%CI 0.22-0.93], respectively). Finally, a trend for a survival benefit was observed in PD1.7 C/C (HR=0.41 [95%CI 0.16-1.00]) and PD-L1 C/T (HR=0.53 [95%CI 0.24-1.18]) SNV genotypes.

Conclusions

Our study showed that SNV of PD-1 and PD-L1 may play a role as a predictive biomarker of response and development of irAEs to PD-1 inhibitor therapy. Some of the investigated SNVs were also associated with a reduction of the risk of death, although a larger group of patients is needed to confirm these results.

Background

HPD has been described in ≃14-25% of pretreated NSCLC pts upon single-agent (SA) ICI and has not been reported upon PCT-ICI. So far, no predictive biomarkers are available for HPD early detection.

Methods

NSCLC pts treated with 1^st line SA-ICI or PCT-ICI were assessed for HPD and circulating neutrophils. HPD was defined as delta tumor growth rate (TGR) >50% and/or TGR ratio ≥2. Circulating low density neutrophils (LDNs) were assessed by flow cytometry on peripheral blood mononuclear cells (PMBCs). LDNs were defined as CD66b⁺CD15⁺ cells among CD11b⁺ PBMCs and immature subtypes as CD10^- LDNs. The LDNs predictive role was assessed by penalized model-based tests. LDNs’ survival and sensitivity to cisplatin induced cell death were tested in-vitro.

Results

144 NSCLC pts were included: 75 treated with SA-ICI, 69 with PCT-ICI. In the SA-ICI cohort, HPD occurred in 8 (11%) pts. Baseline immature circulating CD10^- LDNs were significantly higher (p <0.01) in HPD [median (ME): 39.3, interquartile range (IQR): 28.7] vs pts with progression [ME: 7.4, IQR: 14.9] or response/stable disease [ME: 3.7, IQR: 12.6]. CD10^- LDNs were associated with HPD [odds ratio (OR): 1.17, 95% CI: 1.06; 1.29], with a good prediction capability [cross-validated AUC 0.97 (95%CI: 0.94;1.00)].

A 30.5% cut-off value was identified by Youden index to discriminate HPD from others. In the PCT-ICI cohort, 14 pts had CD10^- LDNs ≥30.5% being at high HPD risk. However, no HPD was observed in PCT-ICI cohort and dynamic LDNs evaluation in high HPD risk pts showed a 52.3% median reduction in CD10^- LDNs upon PCT-ICI, vs only an 8.9% reduction in HPD pts upon SA-ICI, suggesting that PCT prevents HPD by reducing immature LDNs. In vitro experiments showed that immature CD10^- LDNs had prolonged survival compared to mature CD10⁺ LDNs (alive cells after 5-days: 18.7% vs 0%), however, they were more sensitive to cisplatin induced necrotic cell death, confirming the role of PCT in killing preferentially immature LDNs.

Conclusions

Baseline immature CD10^- LDNs is a circulating easy to measure biomarker to early detect HPD upon 1^st line SA-ICI and could select NSCLC pts to be addressed to PCT-ICI.

Background

Tumor-derived genetic alterations can regulate tumor microenvironment (TME) and host immune responses via activation of oncogenic pathways. Therefore, detection of somatic mutations associated with TME and tumor immunity may be useful in identifying patients who could benefit from immunotherapy. Immune checkpoint inhibitors (ICIs), a type of immunotherapy, are considered to be the emerging standard of care for various cancers. However, biomarkers that can help in predicting response to ICIs remain unknown.

Methods

To identify genetic markers associated with sensitivity to ICIs, we established a transcriptome-based predictive model followed by integrative genomics analyses. The impact of in silico derived biomarkers on the TME and response to ICIs was then assessed using in vitro assays and an in vivo mouse model study.

Results

We generated a transcriptome signature associated with response to ICIs based on a small but well-defined immunotherapy study (GSE78220) and applied the signature to large melanoma cohort data from The Cancer Genome Atlas (TCGA) by estimating transcriptome similarity. Based on the analysis, TCGA melanoma cohort was classified into potential responders and non-responders, and subsequent genomic interrogation revealed that the distribution of splicing factor SF3B1 mutations was significantly higher in potential responders. Furthermore, comparative analyses based on patients genotypes and loss- and gain-of-function studies based on manipulation of SF3B1 mutations revealed that SF3B1 mutations promoted transcriptional reprogramming in response to ICIs. To substantiate these results in vivo, we established a syngeneic mouse model, wherein immune resistant B16F10 melanoma cells were implanted into mice. Strikingly, in comparison to mice bearing wild-type SF3B1 tumors, mice bearing mutant SF3B1 tumors showed improved TME and immune profiles suitable for immunotherapy, which were characterized by accumulation of CD4 and CD8 T cells, activated dendritic cells, and concomitant induction of immune effectors.

Conclusions

Our findings suggest that SF3B1 mutations serve as biomarkers that can used to preemptively predict response to ICIs and ensure rational use of immunotherapy in cancer patients.

Background

Melanoma, from its early stages, has a natural propensity to spread, that is regulated via constant immune counteraction. Unleashing adaptive immunity via ICIs has improved melanoma prognosis, but not all patients behave similarly under immunotherapy. Some relapse in the first few months of immunotherapy, while others experience durable disease control, even after discontinuation of ICIs.

Methods

In order to identify any cellular/molecular signature in the peripheral blood that could predict primary immune escape, we examined 100 consecutive patients with stage III or IV melanoma who were scheduled to start ICIs. A multiparameter flow cytometry was used to detect differences in immune subpopulations, gating them by standard conjugated antibodies and mRNA was extracted after sorting of CD3⁺CD4⁺T-cells. A gene ontology (GO) enrichment analysis was performed to recognize which specific genes and key pathways are upregulated in stage III and IV melanoma and between long responders (>36 months) and early progressors(<6 months) to ICIs.

Results

Although circulating immune cells did not differ numerically in patients with stage III and IV melanoma as well as in responders and non-responders to ICIs, transcriptomic analysis of CD3⁺CD4⁺T-cells from 24 selected patients showed that 189 genes were upregulated in stage IV vs. 92 in stage III; while 101 genes were upregulated in early progressors vs. 47 in long responders. These differentially expressed genes (e.g., HLA-DRB5, BCL3, TRIM37, NCAM1, FGFR1) were functionally implicated in distinct cellular pathways that were particularly activated in these two settings. In fact, biological GO pathways of adaptive immunity, negative regulation of DNA-binding transcription factor activity, and lipid metabolism were significantly more upregulated in stage IV compared to stage III melanoma; while phosphorylation of CD3 and TCR chains, PD-1 and IFN signaling were more activated in early progressors compared to long responders to ICIs.

Conclusions

Extrapolating the rules of intra-tumoral plasticity to the blood, specific epigenetically altered pathways in the peripheral CD3⁺CD4⁺T-cells may differentiate the melanoma evolution from stage III to IV and drive the resistance to ICIs.

Background

Immune checkpoint blockade (ICB) improves survival outcomes for patients with mesothelioma, however the mechanisms underpinning response remain elusive with only a minority of patients exhibiting response to ICB. Better understanding could facilitate therapeutic advances in both patient stratification and the rational treatment of relapse. The goal of this study was to gain greater insight into the cellular and molecular regulation of response to dual PDL1-VEGF inhibition (atezolizumab/bevacizumab) in patients with relapsed mesothelioma treated in the MiST4 single arm phase II trial (NCT03654833)

Methods

Diagnostic FFPE tumour blocks acquired from the MIST4 cohort, were subjected to RNA extraction and RNA sequencing(n=18) STAR and featureCounts were used for alignment and read counts Differential gene expression (DEseq2), gene set enrichment analysis (broadinstitute.org/gsea, Clusterprofile) were conducted pathway signatures scores for individual patient were calculated by single-sample GSEA. Immune microenvironment cells type deconvolution were deconvoluted using CIBERSORTx (https://cibersortx.stanford.edu/) and was compared with multiplex immunofluorescence as ground truth. Response was dichotomised into those patients exhibiting tumour growth (G) or reduction (R) measured by mRECIST1.1. Random forest machine learning, and non-parametric statistical testing was used to identify enriched transcriptional patterns in R (n=9) vs G (n=9) groups

Results

The G group exhibited significant enrichment of epithelial-mesenchymal transition (EMT) as estimated by GSEA compared with R. Conversely, inflammatory signatures were highly enriched in the R group versus G. Immune cell deconvolution revealed that NK cells are more abundant in R group (p value = 0.03). Chemokine analysis revealed a significant increase in CXCL12 (p = 0.039) and CXCL17 (p = 0.013) in R group.

Conclusions

EMT and inflammatory transcriptional signatures are exhibit reciprocal association with response to dual anti PDL1-VEGF therapy in relapsed mesothelioma. Further exploration is needed and ongoing to fully understand the genomic, immune microvironment and gut microbiome and their influence on these potentially predictive features.

Background

Recent trials have shown a clinical efficacy of checkpoint inhibitors combined with radiotherapy for a subset of patients with pancreatic ductal adenocarcinoma (PDAC). However, predictive biomarkers are needed. Multiple proteins are released into the bloodstream due to cancer and immune reactions, and several circulating proteins have been associated with response to immunotherapy in other cancers. We investigated 90 immuno-oncology (I-O) related serum proteins in 78 patients with PDAC treated with nivolumab with or without ipilimumab combined with radiotherapy in a randomized phase 2 study (CheckPAC, NCT02866383).

Methods

Proteins were measured in serum samples collected at baseline and after 2 months of treatment using Olink Target 96 I-O panel (Olink Proteomics, Uppsala, Sweden). Serum protein levels were correlated with efficacy data using Wilcoxon test or t-test. Survival was analyzed with univariate and multivariate Cox-regression adjusting for performance status, Glasgow prognostic score, bilirubin, study arm, and weight loss. An unadjusted p-value of 0.05 was considered significant.

Results

Patients with clinical benefit (partial response (n=7) or stable disease (n=15)) had significantly higher levels of fas ligand (FASLG) and galectin 1 (Gal-1), and decreased C-C motif chemokine (CCL) 4 compared to patients without clinical benefit. High Gal-1 and FASLG and low angiopoietin-2 and mucin-16 were associated with longer progression-free survival in univariate analysis. In multivariate analysis, the association remained significant for Gal-1 (Hazard ratio (HR) = 0.25, confidence interval (CI) 0.12-0.56, p<0.001), and borderline significant for FASLG (HR=0.52, CI: 0.26-1.04, p=0.06). For 14 proteins, an increase in protein level during treatment was associated with progressive disease, whereas a decreasing level of 3 proteins (including CCL4) was associated with clinical benefit. Changes in FASLG and Gal-1 were not associated with clinical benefit rate.

Conclusions

The study identified Gal-1 and possibly FALSG as potential novel predictive biomarkers of immunotherapy efficacy in patients with PDAC. The results need to be confirmed in future studies.

Clinical trial identification

NCT02866383

Background

Endemic nasopharyngeal carcinoma (NPC) is associated with persistent Epstein Barr Virus (EBV) infection. Radiotherapy (RT) is standard-of-care for NPC, where clearance of EBV DNA in plasma is a recognized predictor of response. The relationship between post-RT clearance of EBV and concomitant changes in circulating immune cell populations in blood is not known.

Methods

In this study, serial blood samples were prospectively collected from 24 NPC patients before, during and post-RT, and peripheral blood mononuclear cells (PBMCs) were analysed with multiplex flow cytometry to assess the frequency of T cell subsets according to >20 markers of differentiation, co-signalling and chemotaxis. Sera were analysed for the number of EBV DNA copies by PCR, and patients were classified either as having cleared or not cleared EBV post-RT.

Results

We observed that patients with EBV clearance demonstrated a lower frequency of PD1+ CD8 +T cells as well as CXCR3+ CD8+ T cells during and post-RT when compared to patients with no EBV clearance. Most notably, these patients showed a temporal increase in frequencies of CD8+ T cells expressing the chemo-attractant receptors CCR1, 4 and/or 5 upon RT, which was not observed in patients with no EBV clearance. The increase in frequency of CCR+ CD8+ T cells was accompanied by a drop in frequency of naïve CD8+ T cells and an increase in frequency of OX40+CD8+ T cells. Moreover, blood of non-recurrent NPC patients contained higher quantities of CCL14 and CCL15, constituting ligands for CCR1 and CCR5, prior to RT when compared to recurrent NPC patients.

Conclusions

our findings suggest that plasma clearance of EBV DNA post-RT is preceded by increased frequencies of CCR1, 4 and/or 5+ CD8+ T cells, potentially serving as a predictor for EBV clearance in NPC patients, which requires validation in larger cohorts.

Background

Treatment with immune checkpoint inhibitors (ICIs) is successful, but only in a subset of patients with advanced stage non-small cell lung cancer (NSCLC). Blood-based kinase profiling of peripheral blood mononuclear cells (PBMCs) has previously shown its predictive value for response to ICI treatment in a discovery cohort (Hurkmans et al., JITC 2020). The aim of the current prospective study was to perform a blind validation of the predictive value of kinase activity profiles in PBMCs for ICI response in patients with NSCLC in a standardized setting.

Methods

PBMCs from patients with advanced stage NSCLC included in this multicenter study were collected prior to ICI treatment. Tyrosine kinase activity of PBMCs were profiled using a micro-array with 144 different peptide-substrates (Pamchip). Classification analysis was based on grouping of patients with no progression vs. progression (RECIST v1.1) ≤24 weeks after start of treatment. Only patients treated with first-line pembrolizumab (+/- chemotherapy) and ≥24 weeks follow-up were included.

Results

The model was first established based on tyrosine kinase activity profiles, determined in a calibration cohort (N = 61) and then applied to an independent validation cohort (N = 63). The model had a predictive accuracy of 60% (CI₉₅ = 47-72%), which was improved when the model was combined with PD-L1 expression: for patients with a PD-L1 score <50%, the accuracy was 72% (CI₉₅ = 55-86%). For this subgroup of patients, the kinome analysis showed a significant lower progression-free survival for patients for whom progression was predicted vs. those for whom no progression was predicted (p <0.01, hazard ratio= 3.6, CI₉₅ = 1.6-8.1).

Conclusions

The tyrosine kinome of PBMCs before ICI treatment can be applied to predict progressive disease in patients with advanced stage NSCLC, in particular when combined with low PD-L1 expression. This may imply enhanced predictive value of kinome profiling for tumors with low presence of CD8 T cells. The latter needs confirmation, for which data from an ongoing extension study will be used. The final aim is to implement the kinome analysis for clinical use.

Background

Due to lack of reliable markers that determine metastatic potential and guide cancer treatment; metastasis remains a major cause of cancer-related deaths. Immune cells, especially myeloid cells, contribute significantly to metastasis and could function as potential prognostic markers. However, due to their tremendous heterogeneity and plasticity; no reliable and clinically translatable markers to define pro-metastatic immune cell subsets have been identified. With this study we aim to gain deeper insights into myeloid immune cell activities in metastatic tumors to reveal potential markers that define pro-metastatic immune cells. We expect such markers will improve assessment of prognosis and further help devise therapeutic strategies.

Methods

To identify pro-metastatic functions and markers, we performed RNA sequencing on immune cells of 67NR (non-metastatic) and 66cl4 (metastatic) primary tumors, derived from the syngeneic 4T1 mouse breast cancer model. After data analysis, we validated the candidate of interest using flow cytometry, immunoblotting, ELISA, qPCR among other techniques. The translational potential of the identified marker was studied by immunohistochemistry on 487 breast cancer patient biopsies.

Results

We identified several biological processes specifically upregulated in immune cells of metastatic tumors. Interestingly, a metabolic enzyme- arginase1 (ARG1), with known wound healing and immunosuppressive functions was associated with the top enriched processes. Therefore, we investigated it further and found that ARG1 was expressed in myeloid cells, specifically immature neutrophils and macrophages, infiltrating the metastatic tumors but not in non-metastatic ones. This indicated the potential of myeloid cells that express ARG1 to serve as prognostic markers. Further, we analyzed breast cancer patient biopsies and showed that a higher number of ARG1+ infiltrating cells were associated with breast cancer tumors that are more prone to metastasis (basal and HER2 type).

Conclusions

We identified ARG1-containing myeloid immune cells as pro-metastatic markers and indicated their translational potential to clinic.

Background

The circulating predictive factors for survival of advanced non-small-cell lung cancer (NSCLC) patients receiving immune checkpoint inhibitors (ICIs) remain elusive. We aimed to identify the predictive value of circulating cytokines and programmed cell death ligand-1 (PD-L1) expression for survival.

Methods

Serum samples from 102 advanced-stage NSCLC patients who underwent immunotherapy were collected at baseline. PD-L1 expression was also analyzed.

Results

Positive PD-L1 expression (≥1%) was detected in 49.0% of patients. Compared with PD-L1-negative patients, PD-L1-positive patients had a significantly higher ORR (70% vs. 28.8%, p<0.05) and an increased mPFS (25.35 vs. 4.64 months, p=0.003), and tended to have increased mOS (44.84 vs. 20.42 months, p=0.087). CXCL12 levels in the top 33% (1/3) was a poor prognostic factor for durable clinical benefit (DCB, 23.5% vs. 72.1%, p<0.001), PFS (3.76 vs. 14.4 months; p<0.001) and OS (12.2 vs. 44.84 months; p=0.008), and a signature comprising PD-L1<1% and top 33% CXCL12 was associated with lowest ORR (27.3% vs. 73.7%, p<0.001), DCB (27.3% vs. 73.7%, p<0.001) and the worst mPFS (2.44 vs. 25.35 months, p<0.001) and mOS (11.97 vs. 44.84 months, p=0.007). Area under the curve (AUC) analysis for PD-L1 expression, CXCL12 level or PD-L1 expression plus CXCL12 level to predict DCB or NDB showed AUC value as 0.680, 0.719 and 0.794 respectively.

Conclusions

Our findings suggest that the combination of circulating cytokine CXCL12 level and PD-L1 status can predict survival of advanced NSCLC patients treated with ICIs .

Background

Despite potential life-changing morbidity or even mortality, there are no validated biomarkers for identifying patients at risk of developing serious immune-related adverse events (irAE) from immune checkpoint inhibitor (ICI) therapy. Autoimmune conditions and irAE both involve loss of tolerance to endogenous antigens with similar clinical manifestation. We have previously shown that individuals with irAE have higher baseline IgG autoantibody (autoAb) levels with a greater increase in IgG and IgM autoAb after ICI administration, compared to those without irAE. Previous studies have identified associations and potential mechanistic relationships between the gut microbiome and development of irAE. We hypothesize that changes in gut microbiome, autoAb profiles, and peripheral immunophenotype with ICI therapy are associated with the development of irAE and are influenced by immunosuppressive treatment for irAE.

Trial Design

INSPECT-IO is a prospective Princess Margaret Cancer Centre initiative that aims to identify correlates of irAE in adults (>18 years) with advanced solid tumors [NCT04107311]. Patients receiving ICI-based combination immunotherapy are included given their higher risk of irAE compared to those on ICI monotherapy. Those with a history of autoimmune disease and a flare episode within one year are excluded. Whole blood and stool samples are collected from all patients at baseline (≤ 28 days prior to ICI), at an early timepoint (2 – 6 weeks after starting ICI) and end of treatment (≤ 28 days after completion of ICI). Additional blood and stool samples are collected within 96 hours of onset, and upon resolution of every significant irAE (≥ grade 2 by CTCAE v5.0 or requiring systemic immunosuppression). Shotgun sequencing will be used to determine gut microbial taxonomic and metagenomic composition; autoAb profiling will be performed by multiplex proteomic arrays and immunophenotyping will be done using flow cytometry. Archival tumor tissue and organ-specific tissue biopsies for histologic irAE diagnosis will be stored for additional molecular analyses. Study enrolment is ongoing (37 patients to date) with a target accrual of 120 patients.

Clinical trial identification

NCT04107311

Background

Metastatic Pancreatic Ductal Adenocarcinoma (PDAC) is a root cause of lethality in patients. The metastasis is promoted by abundant proteases in the PDAC microenvironment. Synthetic immuno-oncology opens new avenues towards overcoming limitations of standard immunotherapy agents, previously unsuccessful in PDAC. Anti-tumour cytokines such as IL-12 poorly infiltrate the stroma of solid tumours and trigger toxicity. Here, by exploiting the abundant proteases in PDACs, a targeted and tolerable masked Collagen-Binding Domain CBD-Il12 (mCBD-Il12) was produced by fusing the mask to the p35 subunit of Il-12 through a cleavable linker. This linker is cleaved by PDAC-specific proteases, leading to activation of mCBD-Il12 upon tumour infiltration.

Methods

Human PDAC samples were analysed for the expression of MMP2/9 and PLAU in established subtypes (classical and quasi-mesenchymal). mCBD-IL12 with a linker sensitive to such protease activity was therefore produced in HEK293F cells and purified using immobilised metal affinity and size exclusion chromatography. The ability of various murine tissues and their associated proteases to cleave the linker was verified through western blotting. Finally, the molecule was tested in a metastatic and mesenchymal pancreatic syngeneic mouse model.

Results

The quasi-mesenchymal subtype was found to express MMP2/9/PLAU to a higher degree than the other Collisson/Sadanandam PDAC subtypes. In addition, we found the linker to be readily cleaved by murine pancreatic tumour lysate in contrast with non-cancerous pancreatic tissue in vitro. Interestingly, mCBD-Il12 was found to considerably reduce the number of liver metastases and ascites volume compared to wild-type Il-12 or controls in a murine PDAC model. The ascites colour varied from white in wild-type Il-12 to light red in mCBD-Il12, and dark red in controls. Analysis of the blood-based cytokines is pending.

Conclusions

MCBD-Il12 is a novel bioengineered immunotherapy molecule that seems to suppress liver metastases in murine PDAC models. Further studies are required to investigate the changes in the tumour microenvironment and immune infiltrates involved.

Background

Recent ground-breaking results from a randomized phase III clinical trial showed tumor-infiltrating lymphocyte (TIL)-therapy to be superior to ipilimumab in anti-PD-1 refractory advanced melanoma patients, paving the way for TIL-therapy to become a standard treatment (Ref 1). Responses to TIL-therapy are thought to be primarily mediated by expanded CD8+ TILs (CD8+ REP-TILs) via secretion of cytotoxic molecules, including granzymes (Gzms) of which five distinct types (A, B, H, K, and M) have been identified in humans. To better understand the role of Granzymes in TIL-therapy, we characterized the granzyme profile of CD8+ REP-TILs during tumor cell recognition.

Methods

Matched CD8+ REP-TILs and autologous tumor cell line (TCL) pairs were generated using fresh metastatic melanoma samples. TIL/TCL pairs with known high tumor-reactivity were selected and employed in in vitro co-culture systems reproducing TIL:tumor recognition. Gzm mRNA upregulation was assessed via bulk mRNA (n=10) and single-cell RNA sequencing (n=6) of expanded TILs, +8 hours post co-culture. Gzm protein secretion was quantified by ELISA and flow cytometry post co-culture (+2, +8, and +24 hours, n=11).

Results

Bulk and single-cell RNAseq data analysis demonstrated distinct upregulation of only GzmB by CD8+ REP-TILs post TCL recognition. Protein levels were similarly dominated by GzmB, representing 61%, 73%, and 48% of total secreted Gzm at +2, +8 and +24 hours, respectively. In addition, GzmB showed by far the greatest post TCL recognition fold induction, up to 23.8-fold, whereas other Gzms were below 6.0-fold. Overall, GzmA was secreted at high levels, although the response was largely unspecific and only minimally induced by TCL recognition (up to 4.9-fold increase). All remaining granzymes were secreted and induced at minimal levels.

Conclusions

GzmB dominates the tumor-specific granzyme-related response of CD8+ REP-TILs, suggesting GzmB as the primary effector molecule in patients treated with TIL-therapy. GzmB secretion is therefore a potential candidate for a TIL-therapy potency biomarker. The potential of less inducible Gzms, such as GzmA, requires further clarification.

Ref 1: Haanen et al. 2022 ESMO Congress (LBA3, Presidential Symposium I).

Background

Adoptive transfer of tumour infiltrating lymphocytes (TIL therapy) has shown high efficacy in a phase III trial for melanoma patients (M14TIL), and in phase I trials for other solid cancers. However, not all patients respond to TIL therapy. Good prediction tools would help to select which patients may benefit most. The tumour microenvironment is infiltrated by different types of lymphoid and myeloid cells, which communicate with each other. We therefore hypothesized that the presence of specific immune cell types may explain the variation in TIL products.

Methods

To define tumour-specific alterations of immune infiltrates, we characterized the myeloid and lymphoid cell populations present in tumour lesions and healthy adjacent tissue from 26 early-stage NSCLC patients by flow cytometry. We generated TIL products according to the clinical expansion protocol and determined their tumour reactivity based on cytokine production upon co-culture with the autologous tumour digest. Polyfunctional T cells were defined as cells producing at least two cytokines. Data were analysed using Cytotree, a R/Bioconductor package to analyse flow data in an unbiased fashion. Spearman’s Rank Correlation was used to correlate immune infiltrates with expansion rate and percentage of polyfunctional T cell.

Results

The majority of immune cells in the tumour tissue consisted of lymphoid cells (T cells 67%; B cells 16%; NK(T) cells 3.5%), which were all increased compared to the healthy adjacent tissue (T cells 33.6%; B cells 2.6%; NK(T) cells 13%). This increase was at cost of monocyte and dendritic cell infiltrates, which were decreased in tumour tissue (9%) compared to the healthy adjacent tissue (18%). None of the immune infiltrates correlated with the expansion rate of TILs. Monocytes, dendritic cell and NK cell infiltrates showed a negative association with the percentage of polyfunctional T cells. Interestingly, the presence of B cells was positively associated with the percentage of polyfunctional T cells.

Conclusions

We found that specific immune infiltrates in the tumour tissue associate with the functionality of TIL products, which may help select for TIL products with highly responsive T cells against NSCLC tumours.

Background

Colorectal cancer (CRC) is a devastating disease that kills ~700,000 people worldwide annually, with immunotherapies struggling against the immunosuppressive CRC tumour microenvironment (TME). Tumour infiltrating γδ T cells confer a prognostic benefit to CRC patients and can kill cancer via multiple innate and adaptive mechanisms, including antibody-dependent cellular cytotoxicity (ADCC) against tumour antigens (e.g., B7-H3). We hypothesise that γδ T cells can be exploited as an anti-CRC biotherapeutic but their complex interactions within the CRC TME need to be elucidated.

Methods

We performed 3D TME cultures of CRC patient-derived organoids (PDOs) with CRC cancer-associated fibroblasts (CAFs) and human Vγ9Vδ2 T cells, either unmodified or engineered to secrete a modified IL15 cytokine. The addition of anti-B7-H3 IgG also allows the modelling of anti-PDO ADCC, with 3D CRISPR-Cas9 PDO models generated for demonstration of antigen specificity. Using 126-plex Thiol Organoid Barcoding in situ (TOBis) mass cytometry (MC), we measure over 60 parameters per cell across hundreds of culture conditions (Qin et al., Nature Methods, 2020) (Sufi and Qin et al., Nature Protocols, 2021). Cell-type specific signalling analysis across multiple γδ donors and CRC PDOs including post-translational modifications (PTMs), cell-state and immunological phenotype were analysed computationally.

Results

Engineered γδ T cells exhibit superior proliferation, purity, cytotoxicity, and viability, with γδ donor-specific responses to cytokine engineering (¹²⁷IdU⁺ and pSTAT5 [Y694]⁺) and PDO co-culture (GranzymeB⁺ and CD69⁺). Multiple engineered γδ T cells generate a substantial killing phenotype against various CRC PDOs (cCaspase3 [D175]⁺ and cPARP [D214]⁺). We also detect the transfer of Granzyme B from γδ T cells to PDOs, executing PDO apoptosis. ADCC varies between γδ donors and is associated with FcγR CD16 expression.

Conclusions

We have identified marker signatures for the selection of candidate γδ biotherapeutics. CAFs can protect PDOs from γδ ADCC and work is continuing to understand the mechanistic basis of biotherapeutic stromal protection.

Background

Several pivotal studies established the high prognostic value of tumor-infiltrating lymphocytes (TILs) in patients with solid tumors, including colorectal (CRC) and ovarian (OvCa) cancer. Albeit the association between clinicopathological outcomes and tumor infiltration supports the intervention with immunotherapeutic strategies, TIL-adoptive cell transfer (ACT) only led to clinical results in a subset of patients with non-melanoma tumors, raising the question of the abundance of tumor-reactive T cells in these cancers. In the present study, we interrogated CRC and OvCa patients for the presence and features of neoantigen (neoAg) and tumor-specific T cells in in vitro-expanded and also in situ TILs.

Methods

We collected tumors from 30 patients with either early and advanced CRC or advanced OvCa. Neoepitope- and tumor-specific CD8 T cells were identified and FACS-sorted from blood or in vitro-expanded TILs. In parallel, T-cell receptor sequencing (TCR-Seq), including single cell TCR-seq (scTCR-seq) was performed on tumor samples followed by TCR annotation based on a robust in-house pipeline for TCR cloning and screening.

Results

Tumor-specific in vitro-expanded TILs were identified in most CRC and OvCa patients and originated from clonotypes of a broad range of intratumoral frequencies. In CRC, the presence of TILs targeting NeoAg correlated with the density of activated and exhausted TILs in both tumor and stroma. Furthermore, higher structural affinity TCRs preferentially infiltrated tumors. By interrogating libraries of scRNA/TCRseq from tumor samples, several additional tumor-specific TCRs were identified and their transcriptomic profile led to a signature and a predictor of tumor specific TCRs.

Conclusions

Although tumor-specific TILs are consistently detected in CRC and OvCa patients, they are not clonally expanded in tumors and do not efficiently proliferate in conventional cell culture conditions. Their transcriptomic profiles represent a unique dataset that can be exploited to determine cell culture conditions to guide next generation immunotherapies as well as biomarkers discriminating patients eligible for TIL-ACT.

Background

Treatment with autologous tumor infiltrating lymphocytes (TILs) can induce remarkable clinical responses. The absolute numbers of CD8+ T cells in TIL products have been shown to correlate with clinical responses upon TIL therapy. With the current production process, the numbers of CD8+ T cells in the TIL products vary greatly between patients. By using a targeted cytokine that preferentially activates CD8+ T cells, we aimed to increase the cytotoxic potential of the TIL products.

Methods

A cis-targeted CD8-IL2 molecule (Asher Biotherapeutics) was used to promote CD8+ T cell outgrowth from tumor digests. In the rapid expansion protocol (REP), TILs were subjected to polyclonal stimulation using anti-CD3 antibodies (OKT-3) or anti-CD3/CD28 polymers (TransAct™) in the presence of CD8-IL2. The standard ‘young TIL’ production process using high dose IL-2 and OKT-3 was used as a standard comparison. TIL product composition, T cell differentiation phenotype and tumor-reactivity was assessed by flow cytometry and ELISA.

Results

7 tumors (5 melanoma, 1 cervical carcinoma and 1 endometrial carcinoma) were subjected to enzymatic digestion. At the end of the REP phase, TIL products cultured with CD8-IL2 contained 97.5% (range 92-98%) CD8+ T cells, compared to 77.2% (range 35-81%) for products cultured with conventional IL-2 (p = 0.036). Highest total CD8+ T cell numbers were obtained using CD8-IL2 in combination with TransAct in the REP. T cell differentiation phenotypes of the TIL products generated with CD8-IL2 were similar to the standard production process, showing that the improved expansion and CD8+ T cell enrichment did not come at the expense of a more exhausted phenotype. Upon restimulation with tumor digest clear anti-tumor reactivity of the TIL products could be demonstrated based on IFN-y production and tumor cell kill.

Conclusions

This study shows that CD8 cis-targeted IL-2 can be used to generate TIL products mainly comprising CD8+ T cells, thereby potentially improving cytotoxic potential and therapeutic efficacy. The use of CD3/28 TransAct, compared to anti-CD3 stimulation (OKT-3) improved the final yield of CD8+ T cells in the TIL products.

Background

LNP-mediated delivery of long coding RNA has been clinically validated for vaccines and gene editing. We have been developing a novel, synthetic, circular coding RNA platform (oRNA technology) which exhibits significant improvements in production, expression and formulation compared to mRNAs. Lacking the cap structure of mRNA, our oRNA technology uses a proprietary sequence-based IRES element to initiate protein translation in target cells. At the same time, ex vivo generated chimeric antigen receptor (CAR) T cell therapies have had tremendous success in treating hematologic malignancies, yet manufacturing, safety and efficacy challenges remain. At Orna Therapeutics, we are combining oRNA technology with novel immunotropic LNPs to address these challenges, by creating off-the-shelf “autologous” in situ CAR (isCAR™) therapies.

Methods

Orna’s immunotropic LNPs show preferential biodistribution to the spleen, with oRNA reporter expression detected in multiple immune cell subsets, including T cells, macrophages and NK cells. Delivery to immune cells is preserved across mice, rats and non-human primates. In vitro, expanding human T cells expressing an anti-human CD19 CAR oRNA show potent and sustained cytotoxicity and pro-inflammatory cytokine production compared to controls. To maximize protein expression, we developed FoRCE (Formulated oRNA Cell-based Evaluation)[AB1] [AW2] : a robust high-throughput platform that enables parallel arrayed synthesis, purification, lipid nanoparticle (LNP) formulation, and cell-based screening of oRNAs. We applied FoRCE to almost 3,000 unique oRNAs containing UTRs extracted from viral genomes and discovered hundreds of IRESs that drive translation from synthetic oRNA in primary human T cells, hepatocytes, and myotubes.

Results

Select IRESs from this screen drove high levels of CAR expression in primary human T cells. This elevated CAR expression translated to signficantly improved tumor regression in a human PBMC-engrafted NALM6 tumor-bearing mouse model. Tumor regression was dose-dependent, and the novel imunotropic LNP was well tolerated. oRNA-enabled isCAR therapies promise a re-dosable and scalable immune cell therapy

Conclusions

Background

Glioblastoma (GBM) is a very aggressive brain tumor, associated with poor prognosis and survival. So far, the efficiency of available therapies is limited. After proving their effectiveness in targeting hematological malignancies, chimeric antigen receptor (CAR) T cells might provide a promising therapeutic approach for GBM. Here, we present our switchable Reverse CAR technology (RevCAR). Unlike conventional CAR T cells, RevCAR T cells contain an epitope in their extracellular receptor domain, and can only be activated via bispecific target modules (RevTM) which recognize the RevCAR T cells on one side and tumor cells on the other side. Once these RevTMs are eliminated, RevCARs are switched off. In addition, we have developed dual-targeting RevCARs, allowing the control of T cells according to the AND gate logic of Boolean algebra.

Methods

Novel RevTMs specific for GBM were developed. Subsequently, they were expressed in eukaryotic cells and purified with affinity chromatography. The ability of these RevTMs to bind GBM cells and RevCAR T cells was analyzed using flow cytometry. Moreover, the capability of the mono-specific and the dual-targeting RevCAR T cells to kill GBM cells was analyzed using luminescence-based cytotoxicity assay in the absence or presence of a range of RevTM concentrations. Secreted pro-inflammatory cytokines were also evaluated by ELISA. In addition to the in vitro assays, a proof of concept co-injection experiment was performed in vivo.

Results

In this study, we show that GBM-specific RevTMs can bind both the RevCAR T cells and GBM cells. Importantly, the RevCAR T cells can be activated to secrete pro-inflammatory cytokines and to efficiently kill GBM cells via the RevTMs. Moreover, we were able to prove that dual-targeting RevCAR T cells can only be activated upon recognition of two different GBM targets, thereby allowing a highly specific and selective killing of GBM both in vitro and in vivo.

Conclusions

In conclusion, the switchable RevCAR platform is a novel therapeutic approach that provides improved safety, and allows combinatorial targeting of GBM.

Background

Background: Most solid tumors are of epithelial origin and express Mucin 1 (MUC1), a heterodimer of MUC1-N and the oncogenic subunit MUC1-C. Many drugs targeting MUC1 in clinical trials have been primarily directed against MUC1-N. Since MUC1-C is present broadly in tumor due to loss of cell polarity, exposure via hypoglycosylation and MUC1-N shedding, it may represent a more tumor-selective target than MUC1-N. P-MUC1C-ALLO1 is an allogeneic CAR-T targeting MUC1-C and is manufactured using transposon-based integration (piggyBac® DNA Delivery System) and the Cas-CLOVER™ Gene Editing System to knockout the TCR and MHC class I proteins resulting in an enriched T stem cell memory product. Thus, P-MUC1C-ALLO1 addresses multiple common solid tumor indications.

Methods

Methods: MUC1-C epitope expression was investigated by IHC using the scFv binder for P-MUC1C-ALLO1 CAR in epithelial tumor and normal frozen tissue arrays. Pre-clinical efficacy of P-MUC1C-ALLO1 was tested in xenograft models for triple-negative breast (TNBC) and ovarian cancers. Clinical safety has been evaluated in three patients in a phase I trial (NCT05239143).

Results

Results: MUC1-C epitope was positive in multiple tumor samples. While tumor expression was relatively nonpolarized, normal tissue expression was restricted to the apical surface. P-MUC1C-ALLO1 demonstrated robust infiltration and activity in TNBC and ovarian cancer xenografts, with >90% of tumor mass comprised of CAR-Ts at day 10, and 100% tumor elimination at 2 weeks. In the phase I trial, 4 pts (esophageal, colorectal, breast, and pancreatic carcinomas) have been infused either at 0.75x10⁶ (pts 1-3) or 2x10⁶ cells/kg (pt 4). No P-MUC1C-ALLO1 related toxicities were observed. Early efficacy was seen at the low dose with one partial response in pt 3 (HR+, Her2- Breast cancer).

Conclusions

Conclusions: MUC1-C epitope is highly expressed across common epithelial cancers and is apically restricted in normal tissues. Potent anti-tumor activity is seen in preclinical models. In early phase I experience, P-MUC1C-ALLO1 is safe and tolerable with an early signal of efficacy at a low starting dose. P-MUC1C-ALLO1 phase I trial enrollment is on-going.

Clinical trial identification

NCT05239143

Background

P-BCMA-ALLO1 is an allogeneic Chimeric Antigen Receptor T-cell (CAR-T) targeting B-cell Maturation Antigen (BCMA) being investigated in RRMM. P-BCMA-ALLO1 is manufactured using non-viral transposon-based integration (piggyBac®DNA Delivery System) that introduces a humanized anti-BCMA VH-based CAR producing a highly enriched T stem cell memory product. The Cas-CLOVER™ Site-Specific Gene Editing System eliminates endogenous T cell receptor (TCR) expression via knockout of the TCR beta chain 1 gene to prevent graft-vs-host disease, and the beta-2 microglobulin gene to reduce MHC class I expression to eliminate host-vs-graft responses. P-BCMA-ALLO1 demonstrated compelling activity in MM xenografts, providing rationale for this first-in-human phase I study.

Methods

The primary objective is to assess the safety and maximum tolerated dose based on dose limiting toxicity (DLT) in RRMM patients who have received a proteasome inhibitor, immunomodulatory drug and anti-CD38 monoclonal antibody. Secondary objectives will assess the anti-myeloma effect. The protocol utilizes standard 3+3 dose escalation to treat 40 patients. Patients receive lymphodepleting chemotherapy (LDC) with cyclophosphamide (300 mg/m²/day) / fludarabine (30 mg/m²/day) on days -5, -4 and -3 followed by a single dose of P-BCMA-ALLO1 on Day 0.

Results

As of 21SEP2022, 7 patients were treated with P-BCMA-ALLO1. Six patients received the cohort 1 dose of 0.75 X 10⁶ CAR-T cells/kg and 1 patient received the cohort 2 dose of 2 X 10⁶ cells/kg. To date, 4 cohort 1 patients have completed the DLT evaluation period and are evaluable for response. Most adverse events (AE) were grade 1 and 2. One patient had a serious AE of G3 febrile neutropenia. DLTs, cytokine release syndrome and neurotoxicity have not been observed. To date, 1 patient achieved very good partial response, 2 patients achieved partial response, and 1 patient had stable disease. Responses were seen starting at week 2, and overall response rate is 75%.

Conclusions

Early results demonstrate acceptable toxicity profile and promising efficacy for P-BCMA-ALLO1. Dose escalation is ongoing. Updated safety and efficacy results will be presented.

Clinical trial identification

NCT04960579

Background

Glioblastoma (GBM) is a poor prognosis malignant grade IV glioma. After surgical resection, standard therapy consists of concomitant radiotherapy (RT) and temozolomide (TMZ) followed by TMZ alone. Multiple phase I/II trials and at least 3 meta-analysis showed improved survival (OS) and progression free survival (PFS) with dendritic cell (DC) vaccination in high-grade gliomas (HGGs) patients (pts). In those developing antitumor immunity, DC vaccine increases the amount of intratumoral activated cytotoxic T lymphocytes and decreases the number of FoxP3 positive regulatory T cells. Based on these data we have developed a phase II study with DC vaccine concomitant to standard RT-CT in pts undergoing radical surgery for GBM.

Methods

This is a single-arm, monocentric, phase II trial evaluating progression free-survival (PFS) and safety of a DC vaccination integrated to standard therapy in resected GBM. All pts receive a DC vaccine loaded with autologous tumor homogenate for up to one year. The vaccine administration starts at the end of the RT-CT (Induction Phase) and then is alternated to TMZ (Maintenance Phase). Primary end points are PFS and safety, among secondary end points the in vitro (Elispot, Plasma Cytokines, Tumor tissue analysis) and in vivo (DTH skin test) immune response biomarkers are evaluated. A Simon's two-stage design has been used for the sample size calculation. In the first stage, 9 pts will be accrued and a total of 28 pts will be enrolled.

Results

The first 9 pts have been enrolled since October 2021, 4 females and 5 males with a median age of 58 years. Four pts had no MGMT methylation. Eight out of 9 pts concluded the induction phase and 1 is ongoing. To date 4 pts have progressed with a median PFS from the date of diagnosis of 7.5 months (range 5-11). DTH test became positive in 4 out of 7 evaluable pts. No gr3-4 vaccine related toxicities have been observed and gr1-2 toxicities were mostly due to local skin reactions.

Conclusions

This combination therapy seems very well tolerated. The 2 end points of the first step have been reached so the study will proceed to enrol the remaining 19 pts.

Clinical trial identification

Eudract number: 2020-003755-15

Background

Dendritic Cells (DCs) account for the best adaptive immune stimulators promoting their usage as advanced therapy medicinal products (ATMP). Despite the well tolerated profile and the ability to induce anti-tumor immunity, DC vaccine has nowadays failed to translate into prolonged long-term clinical benefit, requiring an update of its therapeutic application.

Methods

We conducted a retrospective study correlating clinical and multiple biologic data with the aim to define biomarkers of DC vaccine clinical activity and to exploit its capacity to expand peripheral autologous tumor-specific effector cells. Immunohistochemistry on pre and post FFPE samples, proteomic analysis on plasma and tumor homogenate, multiparametric flow cytometry on the starting apheresis and on DCs, RNAseq and scRNAseq on peripheral monocytes and manufactured DCs respectively, were applied.

Results

Analysis of post-vaccine biopsies highlighted the vaccine ability to induce intratumoral CD8 up-regulation (p=0.0195). The presence of multiple checkpoint molecules was a common denominator of our ATMP (PD-L1 90,1%-98,9%; PD-L2 94,9%-%99,5; VISTA 21,9%-%39,3; TIM-3 17%-59,4%; B7-H3 86,3%-98,8%). Intriguingly, high dimensional analysis of the DC final product revealed the coexistence of B cell clusters (CD19, MS4A1), higher (p=0.0286) in patients displaying clinical response. Of note, GMPc DCs induced a sustained expression of CD137 in co-cultured CD8+ T cells, and anti-PD1 addition enhanced the fraction of CD137+ T cells.

Conclusions

In vivo mechanisms of adaptive immune resistance (e.g. tumoral PDL1 upregulation p=0.0353) as well as the observed expression of checkpoint molecules by our ATMP support the clinical investigation of DC/ICI combination. The association of B cells with tumor response has to be deepened and the B cell frequency assessed also in pretreatment blood samples. Of note, the expansion of bona fide antigen-reactive CD137+ T cells resulting from the designed autologous DC ex vivo platform could translate into the manufacturing of a novel GMPc T cell product.

Background

Tumor-infiltrating lymphocytes (TILs) and natural killer (NK) adoptive cell therapies have shown promising results in advanced-stage melanoma and haematological malignancies, respectively. However, their limited persistence in vivo in absence of an exogenous source of IL-2, and migration to neoplastic sites have impaired their effectiveness in immunosuppressive tumor microenvironments, such as ovarian cancer. Here, we propose the use of an engineered oncolytic adenovirus encoding a vIL-2 cytokine, Ad5/3-E2F-d24-vIL2 (vIL-2 virus), to improve said cell therapies. Oncolytic adenoviruses are immunogenic agents that lyse infected cancer cells and recruit immune cells to the neoplastic site. Moreover, the vIL-2 virus continuously expresses a vIL-2 cytokine that preferentially stimulates effector lymphocyte proliferation over T regulatory cells.

Methods

Fragments of resected human ovarian cancer tumors were received and processed into single-cell suspensions. Autologous TILs were expanded from said sample fragments, while PBMC cells from healthy donors were used as the source of allogeneic NK cells. To evaluate cancer cell killing, ovarian cancer ex vivo tumor cultures were co-cultured either with autologous TILs or with allogeneic NK cells in the presence or absence of the vIL-2 virus. Additionally, in vivo combination therapies efficacy was assessed with a patient-derived xenograft (PDX) ovarian cancer model. Immune cell profiling was performed following patient sample co-cultures and PDX tumor treatments.

Results

The addition of vIL-2 virus to the ovarian cancer ex vivo tumor cultures improved both TIL and NK cell therapies efficacy in vitro. Similarly, significantly better tumor control was achieved when the vIL-2 virus was given in conjunction with cell therapies compared to their respective controls. Mechanistically, vIL-2 virus treatment enhanced cell cytotoxicity of adoptively transferred TILs and NK cells in PDX tumors, and in ovarian cancer tumor cultures.

Conclusions

Ad5/3-E2F-d24-vIL2 virus treatment seems to be a promising immunotherapeutic candidate to improve the response of adoptive cell therapies in human immunosuppressive solid tumors

Background

In recent years, cancer therapy has witnessed the successful development and clinical implementation of a wide array of tools to fight cancer, including, for example, monoclonal antibodies and adoptive cell therapies (ACT). Immunotherapies employing such tools to reinvigorate T-cells continue to change patient care by providing durable benefit in patients with advanced solid tumors. The latter is offset, however, by a large share of patients that present little response to immunotherapy, in part due to T-cell dysfunction. To overcome this issue, we constructed a chimeric oncolytic adenovirus (TILT-123; Ad5/3-E2F-D24-TNFa-IRES-IL2) capable of expressing tumor necrosis factor alpha (TNFa) and interleukin-2 (IL-2). The use of this agent in preclinical cancer models led to complete responses and favorable immunological effects, both as monotherapy or, in combination with immune checkpoint inhibitors and ACT using tumor-infiltrating lymphocytes (TIL) or chimeric antigen receptor T-cells.

Methods

Blood from patients enrolled in TILT-T115, a clinical study using TILT-123 as monotherapy in patients with injectable advanced solid tumors (NCT04695327), was collected at screening and during treatment, and peripheral blood mononuclear cells (PBMCs) and serum were extracted. Immunological effects were evaluated by detecting immune cell populations in PBMCs by flow cytometry, and by detecting cytokines in blood serum by a targeted multiplexed immuno-assay.

Results

Preliminary immunological data points at immune effects both in PBMCs and blood serum of patients undergoing treatment. Immunological data (e.g. flow cytometry and/or cytokine data) will be presented.

Conclusions

TILT-123 can induce systemic immunological effects supporting the continued development of the agent as cancer immunotherapy.

Clinical trial identification

NCT04695327 (first posted January 5, 2021)

Background

Bispecific T cell engager (BsTe) is a fusion recombinant protein comprised of two single–chain variable fragments with dual specificity for a tumor–associated antigen (TAA) and T cell receptor (usually CD3ε). Immunotherapy with BsTe has shown efficacy in patients with hematologic malignancies and uveal melanoma. However, the antitumor efficacy of BsTe in most solid tumors has been limited due to their short serum half-life and insufficient tumor concentration. We designed a novel serotype 5/3 oncolytic adenovirus encoding for a BsTe cross-linking Mucin1 (MUC1) to CD3, Ad5/3–E2F–d24–aMUC1aCD3. The BsTe (aMUC1aCD3) is designed for the treatment of human solid tumors, where the BsTe links CD3 molecules on the surface of T cells and MUC1 on the target cancer cells.

Methods

Cell viability assays were used to assess the oncolytic potential of the novel Ad5/3–E2F–d24–aMUC1aCD3 construct. The functionality of virus-derived aMUC1aCD3 was evaluated in vitro using T cell activation, proliferation and cancer cell killing assays. The efficacy of the aMUC1aCD3 coding virus was investigated in vivo using a humanized MUC1 expressing lung cancer xenograft mouse model.

Results

The addition of aMUC1aCD3 transgenes did not compromise virus replication capacity. When Ad5/3–E2F–d24–aMUC1aCD3 is combined with allogenic T cells, rapid tumor cell lysis upon infection was observed using different cancer cell lines. TILT-321 infected cells secreted functional aMUC1aCD3 engagers as evidenced by increased T cell activity and competitive binding to MUC1+ cells. Ad5/3–E2F–d24–aMUC1aCD3 treatment also led to effective anti–tumor efficacy in vivo in a human MUC1 expressing lung cancer xenograft model. This response was associated with increased intratumoral T cell activation mediated by the aMUC1aCD3-armed adenovirus.

Conclusions

This study provides proof–of–concept for an effective strategy to overcome some of the limitations of recombinant BsTe delivery in the treatment of solid tumors. The proposed technology could be beneficial for the treatment of solid tumors preferentially those expressing MUC1.

Background

One of the main reasons for the low efficiency of immunotherapy with immune checkpoint inhibitors in non-inflamed (cold) tumors is the lack of anti-tumoral T cell responses. Arenavirus-based cancer vaccines are ideally suited to induce tumor specific T cells but are hampered by the presence of immunosuppressive factors within the tumor microenvironment. Due to their capacity to promote activation, expansion, and effector function of activated T cells, we hypothesized that 4-1BB agonists could help to overcome intratumoral immune suppression by maintaining or even enhancing the functionality of vector-induced T cell responses.

Methods

To test this hypothesis, we initially combined replicating LCMV based vectors (artLCMV) encoding the tumor-associated antigens gp70 or Trp2 with agonistic 4-1BB monoclonal antibody (mAb).

Results

Single administration of vector and 4-1BB mAb prolonged tumor growth control and increased the number of complete responders in the cold B16.F10 tumor model. The combination induced a moderate increase of tumor-infiltrating CD8+ T cells, and positively affected expression of granzyme B, Ki67 and Bcl-XL in tumor-infiltrating antigen-specific T cells. Next, we generated artLCMV vectors co-expressing 4-1BBL together with gp70 or Trp2. In the immunogenic MC38 model, vector-encoded 4-1BBL did not further enhance the already high anti-tumor efficacy of artLCMV-gp70. In the cold B16.F10 model, however, 4-1BBL co-expression increased median survival time and number of complete responders, especially after intratumoral application. This contrasted with systemic application of vector and 4-1BB mAb, as described above. These data indicate that 4-1BB costimulation of T cells is most effective within the tumor, supporting the development of tumor-targeted 4-1BB agonists, which could be applied systemically, as intratumoral injections are not applicable to all cancer patients.

Conclusions

Overall, these experiments confirm the potential of combination therapies with arenavirus vectors and 4-1BB agonists, especially for the treatment of cold tumors, which lack functional T cells.

Background

The efficacy of checkpoint inhibitors are far from satisfied in lung cancer patients with liver metastases. Oncolytic virus can enhance the efficacy of PD-1/PD-L1 antibody in vitro/vivo. In this study, we evaluate the safety and efficacy of oncolytic virus H101 plus PD-1 antibody Toripalimab for liver metastasis-lung cancer advanced patients who have progressed and failed after EGFR-TKIs, chemotherapy or checkpoint inhibitors treatment.

Methods

The patients refractory to previous several lines of standard treatment were injected with recombinant human adenovirus 5 (10¹² vp or 2×10¹²vp) locally for liver metastases plus toripalimab (3mg/kg) systemic therapy every 2 weeks until progression or intolerable toxicity. The two primary end points were investigator-assessed safety and objective response rate (ORR) and the secondary end points included progression-free survival (PFS) and disease control rate (DCR). Efficacy was assessed every 2 months by investigators according to mRECIST v1.1 criteria. Blood samples were collected from patients prospectively.

Results

From January 26, 2021 to July 15, 2022, ten patients were enrolled and received at least 1 cycle of the combination regimen. The most common treatment-related adverse events (TRAEs) were grade 1-2 fever, which was occurred in 9 (90%) of 10 patients. No deaths and other SAE were judged to be treatment-related. Six among the 10 patients received at least 3 cycles of the combination regimen and 5 were evaluable for efficacy analyses as one patient was lost to follow-up. Two of five (40%) patients (one small cancer and one adenocarcinoma) acquired partial remission (PR) after 3 cycles of the combination therapy, including one patient remained remission for 11 months. Two patients (40%) (one small cancer and one adenocarcinoma) achieved stable disease (SD) and the other one adenocarcinoma patent (20%) had progressive diseases (PD).

Conclusions

Our results confirm that the combination of recombinant human adenovirus type 5 plus toripalimab has shown an acceptable safety profile and the preliminary efficacy in those who refractory to previous several lines of standard treatment, deserved further investigation in randomized trials.

Background

In the clinic, immune checkpoint immunotherapy (ICI) is used to re-activate immune reactions against tumor neoantigens, leading to striking remission in cancer patients’ tumors. However, complete or durable responses to ICI treatment only occur in a minority of patients. While the level of tumor mutational burden (TMB) can be used as a predictive marker for responsiveness, we questioned whether the subcellular localization of the neoantigens within the tumor cell additionally plays a role. Using 3 human datasets with 1722 patients treated with ICI, we previously highlighted that patients bearing a high proportion of tumor neoantigens at the membrane of cancer cells responded better to anti-PD1. To decipher underlying immunological mechanisms, we developed a melanoma mouse model that expresses membrane-bound or soluble antigens and analyzed local and systemic anti-tumor immune responses upon anti-PD1 immunotherapy.

Methods

We engineered B16F10 melanoma cells to express membrane-bound or soluble OVA and analyzed intratumoral immune cell infiltration upon implantation in C57BL6 mice. We then compared tumor growth upon anti-PD1 treatment in the wild-type mice or in mice depleted for specific immune cells population. In addition, we compared systemic responses upon tumor implantation via ex vivo antigen-specific restimulation of the splenocytes.

Results

We demonstrated that mice bearing tumors with membrane-bound OVA have increased local and systemic anti-tumor immune reactions compared to soluble OVA, which rendered these tumors highly susceptible to ICI, leading to complete tumor rejection in mice. We additionally observed that tumor rejection was dependent on the level of antigen expressed in the cancer cells, with an increased rejection rate observed for the high dose membrane-bound OVA tumors compared to low ones. We surprisingly found that tumor rejection was independent of immunoglobulin G (IgG), of NK cells and of BatF3⁺ cross-presenting dendritic cells, and mostly relied on CD8⁺ T cells cytotoxicity, and partially on CD4⁺ T cells.

Conclusions

In this study, we show that the subcellular localization of tumor neoantigens plays an important role in the immunogenicity of tumors and subsequent responsiveness to anti-PD1 immunotherapy.

Background

Treatment options for advanced melanoma have recently increased with the introduction of the anti-LAG3 and anti-PD-1 relatlimab/nivolumab combination. To date, ipilimumab/nivolumab is the benchmark of overall survival (OS), despite a high toxicity profile. Furthermore, in BRAF-mutant patients, BRAF/MEK inhibitors and the atezolizumab/vemurafenib/cobimetinib triplet are also available treatments, making the first-line therapy selection more complex. To address this issue, we conducted a systematic review and network meta-analysis of the available first-line treatment options in advanced melanoma.

Methods

Randomised clinical trials (RCTs) of previously untreated, advanced melanoma were included if at least one intervention arm contained a BRAF/MEK or an immune-checkpoint inhibitor (ICI). The aim was to indirectly compare the ICIs combinations ipilimumab/nivolumab and relatlimab/nivolumab, and these combinations with all the other first-line treatment options for advanced melanoma (irrespective of BRAF status) in terms of activity and safety. The co-primary endpoints were progression-free survival (PFS), overall response rate (ORR) and grade ≥3 treatment-related adverse events (TRAEs) rate, defined according to Common Terminology Criteria for Adverse Events (CTCAE).

Results

A total of 9070 patients treated in 18 first-line clinical trials of metastatic melanoma were included in the network meta-analysis. No difference in PFS nor ORR between ipilimumab/nivolumab and relatlimab/nivolumab was observed (HR=0.99 [95%CI 0.75 – 1.31] and RR=0.99 [95%CI 0.78 – 1.27], respectively). The PD-(L)1/BRAF/MEK inhibitors triplet was superior to ipilimumab/nivolumab in terms of PFS (HR=0.56 [95%CI 0.37 – 0.84]) and ORR (RR=3.07 [95%CI 1.61 – 5.85]). Relatlimab/nivolumab showed a trend to a lower risk of grade ≥3 TRAEs (RR=0.71 [95%CI 0.30 – 1.67]) compared to ipilimumab/nivolumab, which showed the highest probability of grade ≥3 TRAEs.

Conclusions

Relatlimab/nivolumab showed similar PFS and ORR compared to ipilimumab/nivolumab, with a trend for a better safety profile. The PD-(L)1/BRAF/MEK inhibitors triplet combinations showed the highest probability to achieve better PFS and ORR.

Clinical trial identification

PROSPERO registration number: CRD42022303279.

Background

Incidence of cardiac metastasis (mets) is rising among pts with cancer. Data on the safety and clinical outcomes of ICIs on pts with cardiac mets is limited.

Methods

In an international multi-center retrospective study, pts with solid tumors and mets to the heart who had received ICIs were included. Immune-related adverse events (irAEs) were graded according to CTCAE v5.0. Objective response rates (ORR) were evaluated by RECIST when available. Overall survival (OS) and progression-free survival (PFS) were calculated from time of ICI initiation using the Kaplan-Meier Method.

Results

62 pts across 16 institutions were identified with a median follow-up of 38.4 months (mo). Melanoma (33.9%, n=21) and non-small cell lung cancer (NSCLC, 22.6%, n=14) were the most common cancers. Median age at ICI start was 65 (23-89) years and 29.0% (n=18) received combination therapy with anti-PD1/PD-L1 and anti-CTLA4, and 58.1% (n=36) received ICIs as 1^st line. Most common locations of cardiac mets were the ventricles (38.7%, n=24) and atria (37.1%, n=23). At ICI initiation, 20.1% (n=13) had a cardiac thrombus. Median diameter of cardiac mass was 4 (0.5-9.0) cm. Cardiology referrals and cardiac MRIs were done on 37/62 (59.7%) and 28/62 (45.2%) pts, respectively. 33.9% had cardiac complications from cancer mets, including arterial/venous emboli (8.1%, n=5), tamponade and arrhythmias (each 6.5%, n=4). For NSCLC, ORR, PFS, and OS were 28.7% (11.7-54.6), 4.3 mo (1.5-11.2), and 9.9 mo (2.0-14.4), respectively. For melanoma, ORR, PFS, and OS were 38.1% (20.8-59.1), 7.5 mo (2.2-19.3), and 35.0 mo (5.4-62.9), respectively. Cardiac mass ORRs for NSCLC and melanoma were 21.4% (7.6-47.6) and 42.9% (24.5-63.5), respectively. iRAEs are shown in table 1.

Conclusions

Cardiac mets are frequently associated with cardiac complications. ICIs have activity in cardiac mets and are safe.

Background

Atezolizumab plus bevacizumab and lenvatinib have not been compared in a randomized controlled trial. We conducted a retrospective multi-center study to compare the clinical efficacy and safety of lenvatinib and atezolizumab with bevacizumab as a first-line treatment for patients with unresectable HCC in the real-world scenario.

Methods

Clinical features of lenvatinib and atezolizumab plus bevacizumab patients were balanced through inverse probability of treatment weighting (IPTW) methodology, which weights patients' characteristics and measured outcomes of each patient in both treatment arms. Overall survival was the primary endpoint.

Results

The analysis included 1,341 patients who received lenvatinib, and 864 patients who received atezolizumab plus bevacizumab. After IPTW adjustment, atezolizumab plus bevacizumab did not show a survival advantage over lenvatinib HR 0.97 (p=0.739). OS was prolonged by atezolizumab plus bevacizumab over lenvatinib in viral patients (HR: 0.76; p=0.024). Conversely, OS was prolonged by lenvatinib in patients with NASH/NAFLD (HR: 1.88; p=0.014)

In the IPTW-adjusted population, atezolizumab plus bevacizumab provided better safety profile for most of the recorded adverse events.

Conclusions

Our study did not identify any meaningful difference in overall survival between atezolizumab plus bevacizumab and lenvatinib. Although some hints are provided suggesting that patients with NASH/NAFLD might benefit more from lenvatinib therapy and patients with viral etiology more from atezolizumab plus bevacizumab.

Background

The addition of immunotherapy (either atezolizumab or durvalumab) to platinum-etoposide for patients with extensive stage SCLC (ES-SCLC) has been recently established as standard first-line treatment based on IMPOWER133 and CASPIAN trials. Yet, the efficacy and safety in a real-world setting remains unclear.

Methods

We retrospectively evaluated patients with ES-SCLC treated with either atezolizumab or durvalumab plus platinum-etoposide between October 2018 and October 2021 in ten centres in the UK and ten centres in Switzerland. Responses were assessed using RECIST v1.1 criteria. Median PFS and OS were analyzed by the Kaplan-Meier method.

Results

A total of 436 patients were included. Median age was 67 years, 228 (52.3%) were males, 209 patients (47.9%) were current and 203 (46.6%) former smokers. 63 patients (14.4%) had an ECOG performance status (PS) ≥2. 385 patients (88.3%) were initially diagnosed with ES-SCLC. Liver and brain metastases were diagnosed in 170 (39.0%) and 87 (20.0%) of patients, respectively. At the time of analysis, 284 patients (65.1%) had died. Most of the patients received atezolizumab (n=427, 97.9%) in combination with chemotherapy, 2.1% received durvalumab. Most patients (n=422, 96.8%) were treated with carboplatin/etoposide. Overall response rate was 71.8% (95%CI 67.3-76.0%) with a median duration of response of 3.5 months (95%CI 3.3-4.0). Median PFS and OS were 5.5 (95%CI 5.3-5.7) and 9.3 months (95%CI 8.4-10.4), respectively. Immune related adverse events (AEs) were seen in 21.8% of patients. 5 patients (1.1%) developed a grade 5 AE. 174 patients (39.9%) received a subsequent systemic therapy. Among patients with brain metastases at diagnosis, mPFS and mOS were 4.8 months (95%CI 4.4-5.5) and 8.6 (95%CI 6.9-10.8), respectively.

Conclusions

Data from our large series show shorter OS than what reported in the trials. Known poor prognostic factors (mainly ECOG PS ≥2 and brain metastases) were more common in our cohort of patients at baseline and may have determined a shorter OS. This is the largest real-world evidence dataset evaluating patients with ES-SCLC treated with first-line chemo-immunotherapy, and could help to optimise clinical management of these patients.

Background

Atezolizumab plus bevacizumab has been recently approved as new first-line standard of care for patients with unresectable hepatocellular carcinoma (HCC). We perform a real word study to evaluate the impact in term of outcome of the inclusion criteria from the IMbrave150 trial on safety and efficacy of treatment.

Methods

We enrolled patients treated with atezolizumab plus bevacizumab for unresectable HCC from 4 different countries. No specific inclusion and exclusion criteria were applied, except for the absence of previous systemic therapies for HCC. The whole population was split in two groups according to the concordance with the inclusion criteria as reported in the IMbrave150 trial in “IMbrave150-in” and “IMbrave150-out” patients, and survival outcomes in the two groups of patients have been evaluated.

Results

766 patients were enrolled in the study: 561/766 (73%) were included in the “IMbrave150-in” group, and 205/766 (27%) were included in the “IMbrave150-out” group. mOS and mPFS were 16.3 Vs 14.3 months (HR 0.48, 0.35–0.65; p < 0.0001] and 8.3 Vs 6.0 months (HR 0.79, 0.63–0.99; p = 0.0431) in “IMbrave150-in” and “IMbrave150-out” patients, respectively. Multivariate analysis confirmed that patients included in the “IMbrave150-in” group had significantly longer OS compared to patients included in the “IMbrave150-out” group (HR 0.76, 0.47-0.97; p=0.0195). In “IMbrave150-in” patients the ALBI grade was not significantly associated to OS, whereas in “IMbrave150-out” patients, those with an ALBI grade 1 reported a significant benefit in terms of OS compared to those with an ALBI grade 2 (16.7 Vs 5.9 months HR 4.40,2.40-8.08, p>0.0001). No statistically differences were reported in “IMbrave150-in” and “IMbrave150-out” groups in terms of safety profile.

Conclusions

The accordance with the inclusion criteria of the IMbrave150 trial positively impact prognosis of patients receiving Atezolizumab plus bevacizumab. Between patients who do not meet the inclusion criteria, those reporting an ALBI grade of 1 could benefit from treatment with the combination.

Background

Conventional time to event endpoints, such as overall survival and progression-free interval fail to reflect quality of life when characterizing outcomes with immune checkpoint inhibitors (ICI). Treatment-free survival (TFS) represents an alternative approach, to more accurately characterize the time free of systemic therapy, providing a more patient-centric view of ICI therapy. There remains a paucity of studies evaluating TFS outcomes in advanced melanoma patients receiving immunotherapy, especially in the non-clinical trial context. Here, we present the first real-world observational analysis of TFS outcomes for patients with advanced melanoma receiving first line ICI therapy.

Methods

Demographic, clinical and survival characteristics of patients with advanced melanoma receiving first-line ICI therapy (n=316) was collected retrospectively from a multi-center observational cohort study in Alberta, Canada. Treatment-free survival was defined as the difference in the 36-month restricted mean survival time between 2 survival endpoints, time to ICI cessation or censored at last follow up, and time to subsequent systemic anti-cancer therapy, death or censored at last follow up.

Results

The restricted mean TFS was longer for nivolumab plus ipilimumab (mean 12.38 months, 95% CI 8.79-16.00) when compared to nivolumab (mean 8.93 months 95% CI 4.38-13.47) and pembrolizumab (mean 11.14 months, 95% CI 8.47, 13.80). During the 36 month follow up, interval patients treated with Nivolumab plus ipilimumab spent 34.4% of their time off systemic anticancer treatments, compared to the 30.9% and 24.8% of their time for the pembrolizumab and nivolumab treatment groups respectively.

Conclusions

TFS represents a patient-centric, informative endpoint for advanced melanoma patients treated with first-line ICI therapy. Of note, patients treated with combination nivolumab plus ipilimumab spent more time alive and free from systemic anti-cancer therapy than those treated with anti-PD-1 monotherapy alone. This information may inform therapeutic decision making, given the number of treatment choices available to clinicians and patients in this context.

Background

Immune checkpoint inhibitors (ICIs) combined with chemotherapy have become standard first-line treatments for advanced esophageal cancer. However, effective therapeutic strategies are limited after failure of immunotherapy. The combination of ICIs and anti-angiogenic agents may produce synergistic effects. Anlotinib, a multi-targeted anti-angiogenic agent, has been approved with good tolerance and efficacy in advanced ESCC. Therefore, we aimed to retrospectively analyze the efficacy and safety of anlotinib plus PD-1 inhibitors in advanced ESCC previously treated with ICIs in the real world (NCT 04984096).

Methods

We retrospectively analyzed the effects of anlotinib plus PD-1 inhibitors with unresectable locally advanced or recurrent/metastatic ESCC and must have received anti–PD-1 or anti–PD-L1 therapy previously. Efficacy was assessed using the Response Evaluation Criteria in Solid Tumors version 1.1. Baseline characteristics and adverse events (AEs) were recorded throughout the entire study.

Results

Between July 2020 to March 2022, 29 eligible patients were included in the final analysis. At data cutoff (July 3, 2022), the ORR was 31.0 % and the DCR was 86.2%, including 9 patients with partial response (PR), 16 patients with stable disease (SD) and 4 patients with progression disease (PD). 8 (27.6%) of 29 patients had received first-line treatment, 20 patients (69.0%) had received second-line treatment and 1 (3%) patient received three lines system chemotherapy. The median PFS was 5.33 months (95% CI 4.28-6.38) and the median OS was 10.38 months (95% CI 6.26-14.48). Any treatment-related adverse events (TRAEs) occurred in all patients, anemia and lymphopenia were the most two common TRAEs, only 2 patients (6.9%) with grade 3-4 TRAEs of lymphocytopenia. All AEs could be relieved after symptomatic treatment. No treatment-related deaths occurred.

Conclusions

Anlotinib plus PD-1 inhibitors showed encouraging anti-tumor activity and manageable safety in advanced ESCC previously treated with ICIs, providing a feasible and well-tolerated treatment option for this patient population. The data further need to be validated in prospective study.

Clinical trial identification

NCT 04984096

Background

Sint has been used to treat patients (pts) with advanced NSCLC. This observational study aims to describe the real-world use of Sint and to explore its real-world effectiveness in the first-line (1L) treatment of advanced NSCLC.

Methods

Pts with advanced NSCLC initiating 1L therapy with Sint between Aug 1^st 2018 and Aug 1^st 2020 in 4 tertiary hospitals in mainland China were included. Data in demographic and clinical characteristics, treatment patterns and outcomes were retrospectively collected through medical records. Descriptive analysis was used to describe the baseline characteristics and treatment patterns. Kaplan-Meier method was used to calculate the median progression-free survival (mPFS).

Results

102 pts (85.3% men) were included with mean age of 63.9±9.5 years (median follow-up time: 4.8 months (mo) (range 0.0-32.0)). The most commonly-used treatment regimen was Sint + platinum (Pt) + taxanes (56.0%) for squamous NSCLC (sqNSCLC) and Sint + Pt + pemetrexed (38.5%) for non-squamous NSCLC (nsqNSCLC) (Table). The mPFS was 15.8 mo (95% CI: 11.5-NA) and 12.0 mo (95% CI: 6.3-NA) respectively for sqNSCLC and nsqNSCLC pts. The mPFS was 17.0 mo (95% CI 11.5-NA) in sqNSCLC pts using Sint + Pt + taxanes as 1L therapy and 7.2 mo (95% CI: 0.7-NA) in nsqNSCLC pts. The ORR was 57.1% and 53.6% respectively for sqNSCLC and nsqNSCLC pts.

Table: Baseline characteristics and treatment pattern of NSCLC

Conclusions

For sqNSCLC pts with Sint as 1L therapy, taxanes and gemcitabine are more preferred, while for nsqNSCLC pts, pemetrexed is more often combined. The real-world effectiveness of 1L treatment of Sint for advanced NSCLC pts is promising from the study.

Background

NSCLC has to date a poor prognosis with life expectancy at five year of 26%. The development of precision medicine is going to improve the prognosis, but most of alterations are still lacking of clinical significance. Somatic BRCA alterations, is reported to be quite frequent in NSCLC, with a prevalence of 5.3-6.6%. However, no specific study to date specifically evaluated its prognostic and predictive role in this population of patients.

Methods

We evaluated BRCA status of aNSCLC patients undergone liquid biopsy at Istituto Nazionale dei Tumori in Milan and Luigi Vanvitelli Hospital in Naples. Guardant360CDx^® was used and mutations pathogenicity were interpreted based on the COSMIC database. Only mutations interpreted as pathogenic were considered. Chi square and Fisher tests were used to match clinical characteristics with BRCA status and survival outcomes were analysed with Cox model.

Results

307 patients had BRCA status available on liquid biopsy, of which 33 (10.7%) had a pathogenic mutation in BRCA1/2 genes. BRCAm patients were older, more frequently male and smoker, had a higher PS and higher PDL1 expression on tissue samples; however, none of these features was significantly associated except for PS. Among patients treated with first-line platinum-based CT, a trend for better PFS was observed among BRCAm patients vs wt, without reaching statistical significance (HR 0.76, CI 95% 0.43-1.35, p=0.354). An increased platinum ORR was observed in BRCA mut vs wt patients (47.6% vs 34.0%, p=0.012). Conversely, among patients treated with any-line IOT, a worse PFS was observed among BRCAm patients vs wt (mPFS 3.5 vs 7.2 m; HR 2.14, CI 95% 1.35-3.40; p=0.001) and numerically reduced ORR was also observed (13.0% vs 26.8%, p=0.192). Finally, BRCA mutations seem to do not have impact on OS (BRCAm vs wt: 28.2 vs 28.8 m; HR 1.17, CI 95% 0.76-1.79; p=0.482).

Conclusions

In our retrospective cohort of aNSCLC patients, presence of a pathogenic mutation in BRCA1/2 genes resulted in reduced benefit to IOT, without affecting overall survival, probably due of the counterbalance effect of the increased sensitivity to platinum-CT. These data could be used to guide therapeutic choices and therefore deserve to be confirmed in a prospective cohort.

Background

To evaluate the therapeutic efficacy and safety of neoadjuvant PD-1 inhibitor camrelizumab combined with TPF induction chemotherapy versus camrelizumab alone in patients with locally advanced resectable oral squamous cell carcinoma (OSCC). This study also aims to build predictive models for treatment decision based on a multivariable logistic regression analysis of multi-dimensional data.

Methods

A phase II study (NCT04649476) was conducted. Untreated stage III or IVA (UICC 8th edition) OSCC patients received neoadjuvant therapy with three 2-week cycles of camrelizumab (200 mg, d1, q2w) (arm A, n=34) or three 2-week cycles of camrelizumab (200 mg, d1, q2w) combined with two 3-week cycles of TPF induction chemotherapy [docetaxel (T) 75 mg/m2, cisplatin (P) 75 mg/m2 and 750 mg/m2 5-fluorouracil (F)] (arm B, n=34), followed by surgery and adjuvant radiotherapy. The primary end point was pathological response rate.

Results

Of the 68 patients enrolled into this trial, 60 completed the full treatment protocol. The pathological response rate in arm B was 83.4%, which was much higher than arm A (16.7%). The pathological complete response (pCR) rate was 30.0% in arm B, while no patient with pCR was observed in arm A. The median follow-up duration was 9.7 months, and the 1-year event-free survival (EFS) of arm A and arm B were 66.7% and 96.7%, respectively. Grade 3-5 neoadjuvant therapy-related adverse events were occurred in 7 patients from arm B and one patient from arm A. A decision tree, mainly based on lymph node metastasis, tumor site, and infiltrated immunocytes in biopsy tissue, was developed to screen patients with high responsivity and low adverse effects.

Conclusions

Neoadjuvant camrelizumab plus TPF induction chemotherapy for locally advanced resectable OSCC showed high pathological response rate, with acceptable adverse effects. Newly developed predictive models may benefit precise treatment decision for OSCC patients in clinical practice.

Clinical trial identification

ClinicalTrials.gov identifier: NCT04649476

Background

This is a multi-cohort, phase II trial to evaluate the safety and preliminary antitumor activity of 70 mg (cohort A) or 100 mg (cohort B) sitravatinib QD plus tislelizumab in pts with locally recurrent or metastatic TNBC, and their combination with nab-paclitaxel in untreated locally recurrent inoperable or metastatic TNBC pts (cohort C). The preliminary result of cohort A has been reported with ORR of 38.1% (Lei Fan. JCO 2022 40:16_suppl). This analysis aims to report the updated analysis in cohort A and the interim results in cohort B.

Methods

Pts with locally recurrent or metastatic TNBC were included and received 70 mg (cohort A) or 100 mg (cohort B) sitravatinib QD PO and 200 mg tislelizumab IV Q3W until disease progression or intolerable toxicity. The primary endpoints included ORR (cohort A and B) and rate of grade ≥3 treatment-related adverse events (TRAEs) (cohort B). Secondary endpoints included DCR, PFS, and safety/tolerability. Updated analysis was provided for cohort A (Simon's 2 stage design). A Bayesian optimal phase II design with one interim analysis (on the first 20 pts out of the total 40 efficacy evaluable pts) was employed for cohort B to monitor both efficacy and safety primary endpoints simultaneously. The stopping boundary of interim analysis was the number of pts with ORR ≤1 or with grade ≥3 TRAEs ≥13.

Results

The data cutoff date of this analysis is 29 July 2022. In cohort A, with the median follow up of 10.7 months, the median PFS was 9.7 (95% CI: 2.8, 12.5) months among 21 efficacy evaluable pts. The rate of grade ≥3 TRAEs was 33.3% (7/21). In cohort B, among the first 20 efficacy evaluable pts with the median follow up of 4.1 months, 9 (45.0% [95% CI: 23.8%-67.9%]) pts were with confirmed ORR and 4 (20%) pts experienced grade ≥3 TRAEs, which met the interim goal. DCR was 80.0% (95% CI: 56.3%-94.27%). At the data cutoff, cohort B achieved the predefined enrollment target.

Conclusions

Sitravatinib combined with tislelizumab demonstrated clinically meaningful anti-tumor activity and had a manageable safety profile in the targeted pts population.

Clinical trial identification

NCT04734262

Background

Atezolizumab (atezo; anti–PD-L1) is used to treat pts with locally advanced/metastatic UC after plt chemo or who are cisplatin-ineligible and whose tumours have PD-L1 expression ≥5%. IMreal (NCT03782207) is a non-interventional, global, multi-centre, multi-cohort prospective study evaluating the short- and long-term outcomes and safety with atezo in the real-world setting. We report the second interim effectiveness and safety data for Cohort 1.

Methods

Eligible pts were adults with UC receiving atezo either as 2L+ treatment following plt chemo or after disease progression within 12 mo of adjuvant plt chemo. Enrolment occurred from 7 Feb 2019 to 15 Sept 2020. Clinical outcomes and safety data before, during and after treatment with atezo were collected. Pt history data were retrospectively collected. The primary endpoint is overall survival (OS) rate. Key secondary endpoints include progression-free survival (PFS), overall response rate (ORR), pt characteristics and safety.

Results

Results are based on 116 pts with a data cutoff of 1 Jun 2021. See table for key endpoints and pt characteristics. Overall, 105 pts (91%) had ≥1 AE. Treatment-related (TR) AEs occurred in 38 pts (33%), with Grade 3/4 TRAEs in 6 pts (5%). AEs of special interest occurred in 27 pts (23%); they were considered related in 14 pts (12%). Three pts (3%) had a Grade 5 AE, including 2 (2%) due to complication of device insertion/device obstruction and 1 (1%) due to a bacterial infection; neither was considered related.

Conclusions

The OS rate with and safety profile of atezo remain consistent with those observed in pivotal studies, with no new or unexpected safety signals identified.

^a Per various tests.

^b Categories are not mutually exclusive.

Clinical trial identification

NCT03782207

Editorial acknowledgement

Medical writing support was provided by Marcia Gamboa, PhD of Health Interactions, funded by F. Hoffmann-La Roche.

Background

Pembro has well characterized, manageable safety in the advanced setting. Clinical data suggest pembro has a similar profile in the adjuvant setting; further characterization is important given the extended life expectancy of pts with resectable disease.

Methods

Safety data from pts with resected stage IIIA-C (AJCC 7th ed) melanoma (KEYNOTE-054), resected stage IB-IIIA (AJCC 7th ed) NSCLC (PEARLS/KEYNOTE-091), RCC post-nephrectomy/metastasectomy (KEYNOTE-564), and resected stage IIB-C (AJCC 8th ed) melanoma (KEYNOTE-716) were pooled. Pts received adjuvant pembro 200 mg (2 mg/kg pediatric pts) Q3W or placebo for up to ~1 yr. Safety was monitored for ≤30 days after treatment end (90 days for serious AEs). AEs were graded using NCI CTCAE v4.0.

Results

Data from 4125 pts were pooled (pembro, n=2060; placebo, n=2065). Median age was 61.0 yrs; 1351 (65.6%) pts treated with pembro and 1343 (65.0%) with placebo were male. Any-grade treatment-related AEs (TRAEs) occurred in 1620 (78.6%) pts treated with pembro and 1212 (58.7%) with placebo. Grade 3-5 TRAEs occurred in 336 (16.3%) pts treated with pembro and 72 (3.5%) with placebo; most common were diarrhea (n=23; 1.1%), ALT increased (n=20; 1.0%), and colitis (n=20; 1.0%) with pembro and lipase increased (n=13; 0.6%) and fatigue (n=6; 0.3%) with placebo. 4 pts in the pembro group died due to a TRAE. TRAEs led to discontinuation in 326 (15.8%) pts treated with pembro and 44 (2.1%) with placebo. Immune-mediated AEs (imAEs) and infusion reactions occurred in 761 (36.9%) pts treated with pembro and 193 (9.3%) with placebo (grade 3-5: n=181 [8.8%] vs n=23 [1.1%]). Most common imAEs were hypothyroidism (n=382; 18.5%), hyperthyroidism (n=227; 11.0%), and pneumonitis (n=82; 4.0%) with pembro and hypothyroidism (n=76; 3.7%), pneumonitis (n=29; 1.4%), and hyperthyroidism (n=27; 1.3%) with placebo. Median time to onset of first imAE or infusion reaction was 2.6 mo with pembro and 4.1 mo with placebo.

Conclusions

Data from this pooled analysis of >4000 patients across several tumor types show that adjuvant pembro has a manageable safety profile consistent in severity and management with the established profile of pembro in advanced disease.

Clinical trial identification

NCT02362594 (first posted to clinicaltrials.gov: Feb 13, 2015); NCT02504372 (first posted to clinicaltrials.gov: July 21, 2015); NCT03142334 (first posted to clinicaltrials.gov: May 5, 2017); NCT03553836 (first posted to clinicaltrials.gov: Jun 12, 2018)

Editorial acknowledgement

Medical writing and/or editorial assistance was provided by Jemimah Walker, PhD, and Holly C. Cappelli, PhD, CMPP, of ApotheCom (Yardley, PA, USA). This assistance was funded by Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

Background

T-cell immunoreceptor with Ig and ITIM domains (TIGIT) plays an important role in tumor immunosurveillance in addition to well-established immunosuppressive checkpoint receptors such as PD-1 and CTLA-4. We report the preliminary safety and anti-tumor activity of IBI939 (anti-TIGIT mAb) in combination with sintilimab (anti-PD-1 mAb) in patients (pts) with previously untreated locally advanced unresectable or metastatic PD-L1-selected NSCLC.

Methods

Eligible pts with systemic treatment-naïve, advanced or metastatic NSCLC, PD-L1 TPS ≥50%, and driver gene negative were enrolled and randomized 2:1 to IBI939 20 mg/kg plus sintilimab 200 mg IV Q3W (arm A) or sintilimab 200 mg monotherapy IV Q3W (arm B). The primary objective was to evaluate the ORR per RECIST v1.1. The secondary objectives include evaluation of PFS per RECIST v1.1, OS, and safety.

Results

Of 42 pts enrolled, 28 pts (median age: 65; adenocarcinoma: n=19; brain metastasis: n=7) and 14 pts (median age: 58; adenocarcinoma: n=6; brain metastasis: n=1) were in arm A and arm B, respectively. As of June 30^th, 2022, among 40 response-evaluable NSCLC pts ( 27 in arm A vs 13 in arm B), the confirmed ORR was 66.7% (95% CI, 46.0-83.5) vs 61.5% (95% CI, 31.6-86.1)(arm A vs B). Nine pts with the event (progressive disease or death) had occurred in both arms. The median PFS was not reached (95% CI, 6.8-NA) in arm A vs 6.0 months (95% CI, 1.4-NA) in arm B (HR: 0.43; 95% CI, 0.17-1.10). The incidence of TRAEs was 85.7% vs 71.4% (2 and 5 pts experienced ≥ grade 3 events in each arm), respectively. The most common TRAEs were hypothyroidism (35.7%), aspartate aminotransferase increased (21.4%), blood urea increased (17.9%), hyperthyroidism (17.9%) in arm A. Immune-related AEs (determined by the investigator) were reported in 64.3% of pts in arm A and 50.0% of pts in arm B.

Conclusions

IBI939 plus sintilimab demonstrated improved PFS benefit and manageable safety profile in PD-L1 TPS ≥ 50% NSCLC patients with no prior systemic treatment.

Clinical trial identification

NCT04672369

Background

The aim of this multicenter study is to compare response obtained by AB o L and identify any factors that may predict the emergence of this response to one of two treatments in a real-world setting.

Methods

Study population derived from retrospective analysis of prospectively collected HCC patients (P) treated with AB or L as first-line treatment. ORR with AB vs L was the primary endpoint. The secondary endpoint compared OS with L vs AB in terms of best response.

Results

1312 P were treated with L, and 823 P were treated with AB. ORR was 38.6% for P receiving L, and 27.3% for P receiving AB [p < 0.01; odds ratio (OR) 0.60)]. In responding P (CR + PR), OS was 22.3 months (m) for P receiving L, and 22.5 m for P treated with AB (p 0.20; HR 0.81, reference L). In non-responding P (SD + PD), OS was 10.8 m for P receiving L, and 11.5 m for P receiving AB (p 0.36; HR 0.85, reference L). For P who achieved CR, OS was not reached in both arms, but the result from univariate Cox regression model showed 62% reduction of death risk for P treated with AB (p 0.05). In all multivariate analyses, treatment arm was not found to be an independent factor conditioning OS. Comparing ORR achieved in the two arms, L was shown to be statistically significant in all subgroup P except for Child Pugh B, presence of portal vein thrombosis (PT), αFP ≥ 400 ng/mL, presence of extrahepatic disease (EHD), ALBI 2, and no previous locoregional procedures. P who achieved CR were compared to the rest of the population in the same way. The only statistically significant difference was in favor of L in P with NLR > 3 (p 0.03; OR 0.39). Finally, we evaluated separately which P had more response in AB arm and then in L arm. Only in L arm, three factors favoring response were highlighted: absence of EHD (p < 0.01; OR 1.88), BCLC B (p < 0.01; OR 0.54), absence of PT (p 0.05; OR 1.34), and NLR ≤ 3 (p < 0.01; OR 0.70).

Conclusions

L achieve higher ORR in all P subgroups. P who achieve CR with AB can achieve OS so far never recorded in HCC patients. This study didn’t highlight any factors that could identify HCC P subgroups capable of obtaining CR.

Background

Zimberelimab is a novel, fully human anti-PD-1 monoclonal antibody with high affinity and selectivity for PD-1.The objective of this study (ClinicalTrials.gov identifier: NCT03972722) was to evaluate zimberelimab, a novel,anti-programmed cell death protein 1 monoclonal antibody, in patients with programmed death ligand-1-positive recurrent or metastatic cervical cancer that had progressed after first- or subsequent-lineplatinumcontaining standard chemotherapy.

Methods

In this single-arm, phase II study, eligible patients in 27 Chinese sites were assigned to receive intravenouszimberelimab 240 mg as monotherapy every 2 weeks until confirmed disease progression, death, intolerableadverse effects, or withdrawal from the study. The primary endpoint was the objective response rate (ORR)assessed per Response Evaluation Criteria in Solid Tumors (version 1.1) by an independent review committee.Secondary endpoints included duration of response (DoR), disease control rate (DCR), progression-free survival(PFS), overall survival (OS), and safety.

Results

Ninety participants were included in the full analysis set, with a median follow-up of 11.5 months. Completeand partial responses were achieved by 4 and 21 patients, respectively, corresponding to an ORR of 27.8%(95% confidence interval [CI], 18.85 to 38.22; P < .0001 vs historical controls). Median OS and DoR were notreached during the study: 12-month OS rates were 54% (95% CI, 41 to 66) and 6-month DoR rates were 84% (95% CI, 58 to 95). Median PFS was 3.7 months and the 12-month PFS rate was 15% (95% CI, 2 to 42). Treatment-related adverse events (TRAEs) occurred in 78.1% of participants, with hypothyroidism (25.7%) and anemia (19.0%) being the most frequently reported. Grade ≥ 3 TRAEs occurred in 22.9% of participants.

Conclusions

Zimberelimab monotherapy demonstrated durable antitumor activity and an acceptable safety profile in patients with recurrent or metastatic cervical cancer that had progressed after first- or subsequent-line platinum-containing standard chemotherapy. Further investigation of zimberelimab in patients with cervical cancer is warranted.

Clinical trial identification

NCT03972722

Background

The combination of immune checkpoint inhibitors and tyrosine kinase inhibitors (TKIs) manifested high efficacy in uHCC. Anlotinib was a novel oral multi-targeted TKI selective for VEGF receptors 1/2/3, FGF receptors 1-4, PDGF receptors α and β, and c-kit. Penpulimab, a humanized anti-PD-1 IgG1 monoclonal antibody, was engineered to eliminate FcγR binding, consequently eliminating ADCC, ADCP and ADCR effects. This AK105-203 trial (NCT04172571) aimed to explore the efficacy and safety of penpulimab plus anlotinib in patients (pts) with histologically or cytologically confirmed uHCC.

Methods

In this single-arm, multicenter phase Ib/II trial, pts with uHCC and no prior systemic treatment were eligible if they were 18-75 years, and classified as BCLC stage B (not amenable for locoregional therapy) or C, Child-Pugh score ≤7 and ECOG PS of 0-1. Pts received anlotinib (8 mg, p.o., qd, d1-14, q3w) plus penpulimab (200mg, iv, d1, q3w). The primary endpoint was ORR (RECIST v1.1). Secondary endpoints were safety, DCR, DoR, TTP, PFS and OS.

Results

31 pts (median age 56 years [23-74], ECOG 0/1 [64%/36%], BCLC B/C [23%/77%], HBV/HCV [61%/7%]) received combined therapy. As of August 5, 2022, median follow-up time was 23.0 months (range 3.7-31.9). The ORR was 31.0% (95% CI, 15.3-50.8%), and DCR was 82.8% (95% CI, 64.2-94.2%). The median PFS and TTP for 31 patients were 8.8 months (95% CI, 4.0-12.3) and 8.8 months (95% CI, 4.0-14) respectively. OS events were observed in 16 patients (51.6%), and the median OS was 23.0 months with 12-months OS rate was 67.9%. Treatment-related adverse events (TRAEs) occurred in 90.3% of pts (≥G3 in 25.8% [8/31]). No G5 AE occurred. Most common TRAEs (≥25%) were increased AST (41.9%) and ALT (35.5%), general disorders and administration site conditions (35.5%), skin and subcutaneous tissue disorders (32.3%), platelet count decreased (25.8%), asthenia (25.8%).

Conclusions

Anlotinib combined with penpulimab showed encouraging efficacy and acceptable safety in pts with uHCC. The further randomized, phase 3 study of penpulimab plus anlotinib at a higher dose (10 mg) in this setting is ongoing (NCT04344158).

Clinical trial identification

The trial protocol number was NCT04172571 and release date was November 21, 2019.

Background

Platinum-based chemotherapy is still the standard of care for Epidermal growth factor receptor (EGFR) mutated non-small cell lung cancer (NSCLC) patients after developing EGFR-TKI resistance. However, no study focusing on the role of PD-1 inhibitor based treatments for EGFR mutated NSCLC patients who carried PD-L1 TPS ≥ 50% progressed after EGFR-TKI therapy. Thus, we aimed to investigate the outcomes of PD-1 inhibitor based treatments for these patients and to explore the population that may benefited from PD-1 inhibitor based therapies.

Methods

We retrospectively collected data of EGFR mutated advanced NSCLC patients with PD-L1 TPS≥50% who have failed prior EGFR-TKI therapies without T790M mutation at Shanghai Chest Hospital between January 2018 and June 2021. Progression-free survival (PFS) and overall survival (OS) were utilized to evaluate the outcomes.

Results

A total of 146 patients were included. The median follow-up was 36.7 months (IQR, 12.5-44.2 months). Among the population, 66 patients (45.2%) received chemotherapy, the remaning (54.8%) received PD-1 inhibitor based therapies, including 56 patients(70.0%) received PD-1 inhibitor combined with chemotherapy (PC) and 24 patients (30.0%) received PD-1 inhibitor monotherapy (PM). Survival analysis shown that patients who received PD-1 inhibitor based therapies had better PFS and OS compared with those treated with other therapy (median PFS, 4.0 vs. 10.0 months, P＜0.001; median OS, 39.5 vs. 24.2 months, P＜0.001). What’s more, patients who treated with PC treatment had a superior survival time than those received PM treatment (median PFS, 10.3 vs. 7.0 months, P＜0.001; median OS, 32.4 vs. 41.6 months, P＜0.001). Subgroup analysis found that the PFS and OS benefit of PC was evident in all subgroups.

Conclusions

For advanced NSCLC patients with EGFR mutations and PD-L1 TPS≥50% who have failed prior EGFR-TKI therapies without T790M mutation, PD-1 inhibitor based treatment could provide a more favorable survival than classical chemotherapy. What's more, compared with PD-1 inhibitor monotherapy, PD-1 inhibitor combined with chemotherapy seems to be the preferred treatment.