Background

Human homologues of PIWI proteins identified are PIWIL1, PIWIL2, PIWIL3 and PIWIL4 (Sasaki et al. 2003). Aberrant expression of these proteins has been associated with hallmarks of cancer, and have also a potential prognostic and diagnostic markers for different cancers (Suzuki et al. 2012). However, their functional and clinical significance in pancreatic ductal adenocarcinoma (PDAC) remains unknown. The purpose of this study was to dissect the role of PIWI proteins and their prognostic relevance.

Methods

PIWI proteins expression were analyzed by western blot in human PDAC derived cell lines and one non-transformed pancreatic cell line. Functional experiments were performed with PIWIL3 and/or PIWIL4 downregulated PDAC derived cell lines and one non-transformed cell line. Immunohistochemistry was performed to evaluate expression of PIWI proteins in 124 PDAC samples from Fundacion Jimenez Diaz Hospital, and with 124 validation cohort from TGCA. Then, association between PIWI proteins expression and survival was assessed.

Results

Only PIWIL3 and PIWIL4 showed differential expression in PDAC cell lines. Both wound-healing and transwell assay showed a decrease in migration ratio after PIWIL3 and/or PIWIL4 downregulation (P < 0.05). Furthermore, both PIWIL3 and/or PIWIL4 are necessary for the maintenance of undifferentiated phenotype highlighted by a reduction in size and number of pancreatic spheres after PIWIL3 and/or PIWIL4 downregulation (P < 0.05). On the other hand, PIWIL1 associated with shorter survival (P = 0.056), and this finding was validated in the TGCA cohort (P = 0.021). PIWIL2 associated significantly to shorter survival (P = 0.046) in our training set, and validation set exhibited a high trend toward significance (P = 0.051). Although PIWIL3 and PIWIL4 did not show association with survival in our training set, validation set revealed a statistically significant association of PIWIL4 with shorter overall survival (P = 0.027).

Conclusions

The present study demonstrate that PIWIL3 and PIWIL4 regulates PDAC aggressiveness by inhibiting cell migration and regulate undifferentiated phenotype of cancer cells. Furthermore, PIWIL1, PIWIL2 and PIWIL4 are potential prognostic biomarkers in PDAC.

Legal entity responsible for the study

Fundación Instituto de Investigación Sanitaria - Fundación Jiménez Díaz (G-85874949).

Funding

Spanish Ministry of Economy and Competitiveness.

Disclosure

All authors have declared no conflicts of interest.

Background

Our understanding of the interaction between the host immune system and cancer has evolved rapidly over the previous 10 years due to the clinical success induced by checkpoint inhibition. Previous immunologic work can now be realized in combination strategies. Modified Vaccinia Ankara (MVA) offers significant opportunities due to its natural induction of innate and adaptive immunity, large payload, and excellent safety profile.

Methods

MVA vectors were administered subcutaneously (SC), intravenously (IV) or intratumorally (IT) into tumor-bearing mice. Cytokine secretion profile in serum was assessed by Luminex analysis. NK and T cell infiltration and activation in various organs and cytolytic activity against target cells were determined by flow cytometry-based assays. PD-1 immune checkpoint blockade (ICB), low dose cyclophosphamide, tumor-targeting Abs or 15 Gy radiotherapy were administered along with IV MVA.

Results

Recombinant MVA (rMVA) in which tumor antigens and costimulatory molecules are encoded can be customized for maximal effect by route of administration. rMVA administered intravenously (IV) causes superior induction of antigen specific T cells, cytokines and NK cells than previously seen with subcutaneous or intramuscular routes. Encoding CD40L in addition amplifies the effects and efficacy improves in relevant models. This is dependent on T cells and NK cells, indicating a potential solution to one tumor-resistance mechanism, MHC loss and/or mutation. Furthermore, the combination of rMVA with tumor targeting antibodies, checkpoint inhibition, radiotherapy or chemotherapy often showed additional synergistic therapeutic effects. Administration of MVA alone intratumorally (IT) causes innate immune activation through toll like receptors (TLRs) as well as the cGAS / STING pathway. Recombinant antigen encoding rMVA improves systemic and local antigen-specific T cell responses. These effects can be bolstered by encoding certain costimulatory molecules.

Conclusions

IV and IT recombinant MVA may offer off the shelf solutions to resolve many of the host – tumor immunity interactions that result in lack of efficacy of checkpoint inhibition alone in most patients. Bavarian Nordic plans to initiate clinical trials with existing agents in 2019 applied IV and IT and will create novel constructs to maximize clinical effect, planned to initiate in 2020 and beyond.

Legal entity responsible for the study

Bavarian Nordic, Inc.

Funding

Bavarian Nordic, Inc.

Disclosure

C. Heery: Employee and shareholder: Bavarian Nordic, Inc. C. Pico-Navarro, T. Adams, L. Bauman, J. Medina, M. Hinterberger, A. Heiseke, H. Lauterbach, H. Hochrein: Employee of Bavarian Nordic.

Background

To meet the rapidly growing I/O market, the demands for fast, relevant and cost effective mouse tumor model systems are also increasing. We developed a panel of straightforward humanized tumor models, designated as MiXeno platform.

Methods

CrownBio has established a sizable collection of MiXeno models where human PBMCs were reconstituted in the mouse system for the evaluation in vivo activity of immune checkpoint inhibitors or immune-modulators. These MiXeno models were characterized with tumor response to anti-PD-1 and anti-CTLA4 antibodies, and onset of possible graft versus host disease (GVHD) or graft versus tumor response (GVT) under different settings. To engage both host immune system and tumor antigens, we have developed some specific Mixeno tumor models by inoculating tumor cells over-expressing specific anti-tumor antigens (e.g. EGFR, CD47, Braf or PD-L1) into PBMC-humanized immunocompromised mice. Moreover, to improve capacity and consistency of MiXeno platform, we are conducting studies to characterize and validate commercialized frozen PBMCs for MiXeno model establishment.

Results

Models with over-expression of a variety of tumor antigens (e.g. EGFR, CD47, Braf, PD-L1...) were used to develop specific Mixeno tumor models. Meanwhile, in order to overcome the limitation of PBMC shortage, commercialized PBMC has been purchase and implanted into several xenograft models, and exhibit consistent tumor growth with fresh PBMC, as well as human immune component reconstitution. Up to date, a variety of test articles of different categories, including checkpoint inhibitors, T cell modulators and bispecific T cell engagers (e.g. EpCam-CD3, CD47/CD3, BCMA/CD3) have been evaluated using this platform. Merchandized I/O drugs, such as Pembrolizumab, are being tested in commercialized PBMC implanted immunocompromised mice.

Conclusions

MiXeno platform are valid model system for the human immuno-modulatory drugs including bispecific antibodies evaluation and will be optimized by introduction of specific Mixeno tumor models, commercial PBMC and B2M mice.

Legal entity responsible for the study

CrownBio.

Funding

CrownBio.

Disclosure

L. Bourre, L. Zhang, S. Qi, H. Wu, L. Zhao, X. An, M. Qiao, Q. Shi, W. Yang: Employee: CrownBio. All other authors have declared no conflicts of interest.

Background

Prostate cancer patients with distant metastasis have poor prognosis and develop resistance to all standard drugs at various time intervals. Therapeutic options which can alleviate symptoms and prolong survival are required for these patients. [¹⁷⁷Lu]prostate-specific membrane antigen ([¹⁷⁷Lu]PSMA) is a novel drug based on a theranostic concept. Here, we have presented the safety and efficacy profile of one cycle of [¹⁷⁷Lu]PSMA in metastatic castration-resistant prostate cancer (mCRPC) patients who have exhausted all standard therapeutic options.

Methods

Twenty two patients treated with at least first line anti-androgens and docetaxel were treated with one cycle of [¹⁷⁷Lu]PSMA therapy on a compassionate basis. Haemoglobin, total leukocyte counts, platelets and serum creatinine for toxicity profile while prostate specific antigen (PSA), Eastern Cooperative Oncology Group (ECOG) performance status, visual analogue scale (VAS) and analgesic quantification scale (AQS) for therapeutic efficacy were recorded pre and 8 weeks post therapy. Wilcoxon signed-rank and ANOVA tests were used for statistical analysis.

Results

Partial response (PR), stable disease (SD) and progressive disease (PD) for PSA were seen in 5 (22.7%), 13 (59.1%) and 4 (18.2%) patients, respectively, treated with mean 6.88GBq dose of [¹⁷⁷Lu]PSMA. 8/22 (36.4%) patients showed ≥ 30% drop in PSA. Grade 3 haemoglobin toxicity was seen in 5/22 (22.7%) patients. No patient developed grade 4 haemoglobin toxicity. No patients had grade 3 or 4 leukocytopenia or thrombocytopenia. Wilcoxon signed-rank test showed statistically significant (p < 0.05) difference in pre- and post-treatment ECOG, VAS, AQS scores while it was not significant for PSA (P > 0.05). ANOVA test showed a statistically significant difference in mean doses of [¹⁷⁷Lu]PSMA used in the three PSA response groups while the difference was non-significant for other variables.

Conclusions

We conclude that [¹⁷⁷Lu]PSMA therapy delivers adequate pain palliation in all end-stage mCRPC patients and it has a potential to become an effective therapeutic option in properly selected patients.

Legal entity responsible for the study

Manoj Gupta.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Immunotherapy with programmed death receptor-1 (PD-1) antibodies has changed the paradigm of advanced NSCLC treatment. These checkpoint inhibitors showed better outcomes compared with standard treatment but reliable predictive markers are still lacking. High pre-treatment NLR and PLR have been associated with poor prognosis in several tumor types and recent studies suggest a potential role also in NSCLC. We thus conducted this study to evaluate the prognostic significance of NLR and PLR in our patients.

Methods

All patients with locally advanced and metastatic NSCLC treated with nivolumab and pembrolizumab from February 2016 to October 2018 were enrolled. NLR and PLR were determined by the division of neutrophils and platelets by lymphocytes in peripheral blood. Kaplan Meier method and Cox proportional hazardous analysis were conducted to assess the impact of NLR, PLR and other clinical factors on overall survival (OS) and progression free survival (PFS).

Results

Thirty-two patients were treated, 20 with nivolumab and 12 with pembrolizumab. Median age was 61 (40-82); 63% were male; 91% had an ECOG PS ≤ 2; 37% received ≥ 2 prior systemic therapies and 78% had stage IV disease. Increased NLR or PLR values above mean were independent predictive factors for decreased PFS (11 vs. 6 months, HR 3.33 95%CI 0.97 - 11.3, p = 0.056 and 12 vs. 6 months, HR 3.9 95%CI 1.19 - 12.8, p = 0.025, respectively). NLR and PLR values higher than percentil 25 were predictive factors, when used in combination, for decreased OS (21 vs. 11 months, HR 12.363 95% CI 1.303 - 117.291, p = 0.028 and 13 vs. 11 months, HR 3.9 95%CI 1.19 - 12.8, p = 0.025, respectively). Other clinical factors (i.e. histology, tobacco use, age, gender, ECOG PS, metastatic sites) did not present any implication for OS and PFS, as determined by multivariate analyses.

Conclusions

Elevated pre-treatment NLR and PLR are associated with shorter OS and PFS in our cohort independently of other prognostic factors. Our results reinforce the potencial role of these markers as a predictive factor of poor prognosis for NSCLC patients. Prospective studies are warranted to validate these findings.

Legal entity responsible for the study

Susana Rocha Amaral.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Targeting the epigenome has demonstrated efficacy in hematological malignancies, and results of recent phase 1 (P1) trials have shown promising activity in solid tumors. The number of novel epidrugs is increasing exponentially, with several first-in-class, first-in-human selective compounds now evaluated in P1 trials. Accurate knowledge of their safety profile and toxicity management beyond cycle 1 is essential to appropriate P2 dose recommendation.

Methods

All patients (pts) with hematologic or solid tumors enrolled in at least one epidrug P1 trial at Gustave Roussy Drug Development Department were retrospectively analysed. Baseline pts characteristics, treatment-related adverse events (AEs) – type, grade, date of occurrence, duration, resolution - toxicity management (medication, dose modification) and outcome were collected.

Results

A total of 243 pts (43,6% hematologic, 23,1% non-Hodgkin lymphoma (NHL), 33,3% solid tumors excluding NHL) were included in 15 epidrug monotherapy P1 trials between Jan 2010 and March 2017; 62% were male; median age was 65 yo and median treatment duration was 119 days; 1980 treatment cycles and 335 AEs were analysed: 118 (35%) (64 G1-2; 54 G3-4) and 217 (65%) (114 G1-2; 103 G3-4) AEs occurred during and after cycle 1 (C1; DLT period), respectively; 58% of AEs were hematological toxicities. The risk of G3-4 toxicity for hematologic pts was 15% and 11% during and after C1 respectively, and was 12% and 18% for solid tumors excluding NHL, and was 29% and 24% for pts with NHL. DLT occurred in 10 pts (4%). Dose reduction occurred in 15% of pts, after a median duration of 21 treatment days. Temporary and definitive treatment interruption for toxicity occurred in 21% and 9% of pts, respectively; 87% of these occurred after C1.

Conclusions

In P1 trials of epidrugs, 65% of high-grade AEs occur after cycle 1 and 42% are non-haematological toxicities. More pts with NHL than pts with solid tumors (excluding NHL) or hematological malignancies present their first severe AE during C1. The dose recommendation process may require fine-tuning according to each pt population. Like molecularly targeted or immune therapies, epidrugs have distinct toxicity profile requiring specific attention in their development process.

Legal entity responsible for the study

Gustave Roussy Institut.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Many therapeutic methods existing for conventional cancer therapy have not been successful in achieving ideal outcomes or have noticeable side effects from off-targeting cytotoxicity. The photosensitizer-based cancer treatment approach has attracted great attention. Chemotherapy (CT), photodynamic therapy (PDT), and photothermal therapy (PTT), called combination treatments. In a single system could be potential solutions to address the above mentioned adverse effects in conventional therapy. Multimodal nanocomposites (NCs) are being used to achieve with innovative and noninvasive enhanced targeting synergistic anticancer phototherapy.

Methods

We prepared spherical-like titanium dioxide nanoparticles TiO₂NPs by the water in oil emulsification method, obtaining a novel theranostic nanocomplex FA-ICG-Qtn@PVPylated-TiO₂NPs. Nanocomplex (NCs) were characterized by various physicochemical techniques including UV via spectroscopy, TEM, DLS, FTIR, MTT, AO/EtBr, DAPI, cell cycle arrest, ROS, mitochondrial membrane potential loss, Western blot, RT-PCR, Histopathology, and immunohistochemistry. Studies were performed both in vitro/in vivo.

Results

The resulting TiO₂NPs achieved high drug loading in combination with low leakage at physiological pH, and minimal toxicity toward healthy cells. To assist drug delivery, we have prepared FA-ICG-Qtn@PVPylated-TiO₂NPs containing Qtn with high loading efficiency (35.2% w/w) as a novel drug delivery system. The NCs are taken up via FR endocytosis by MCF-7 cells and can generate intracellular reactive oxygen species (ROS) in order to increase mitochondrial membrane potential loss (MMPL) and enable release of cytochrome-c, followed by dysregulation of Bcl-xL into the cytosol and activation of caspase-7 to induce cancer cell apoptosis. These NCs can be utilized to improve cancer nanotherapy by induction of apoptosis in vitro. After intravenous in vivo direction of FA-ICG-Qtn@PVPylated-TiO₂NPs NCs could significantly accumulate in the tumour-bearing Balb/c mice, and effectively inhibit the tumor growth after 808 nm laser irradiation as confirmed by the cancer cell killing studies in vivo.

Conclusions

The present thermal/pH-coupling controlled and targeted drug delivery system paves the way for the next generation of nanotherapeutics working toward a potential proficient targeted anticancer treatment.

Legal entity responsible for the study

Department of Zoology, Bharathiar University, Coimbatore.

Funding

Has not received any funding.

Disclosure

C. Massard: Consultant/Advisory fees: Amgen, Astellas, Astra Zeneca, Bayer, BeiGene, BMS, Celgene, Debiopharm, Genentech, Ipsen, Janssen, Lilly, MedImmune, Novartis, Pfizer, Roche, Sanofi, Orion; Principal/sub-Investigator of Clinical Trials: Aduro Biotech, Agios Pharmaceuticals, Amgen, Argen-X Bvba, Arno Therapeutics, Astex Pharmaceuticals, Astra Zeneca, Aveo, Bayer Healthcare Ag, Bbb Technologies Bv, Beigene, Bioalliance Pharma, Biontech Ag, Blueprint Medicines, Boehringer Ingelheim, Bristol Myers Squibb, Ca, Celgene Corporation, Chugai Pharmaceutical Co., Clovis Oncology, Daiichi Sankyo, Debiopharm S.A., Eisai, Exelixis, Forma, Gamamabs, Genentech, Inc., Gilead Sciences, Inc, Glaxosmithkline, Glenmark Pharmaceuticals, H3 Biomedicine, Inc, Hoffmann La Roche Ag, Incyte Corporation, Innate Pharma, Iris Servier, Janssen, Kura Oncology, Kyowa Kirin Pharm, Lilly, Loxo Oncology, Lytix Biopharma As, Medimmune, Menarini Ricerche, Merck Sharp & Dohme Chibret, Merrimack Pharmaceuticals, Merus, Millennium Pharmaceuticals, Nanobiotix, Nektar Therapeutics, Novartis Pharma, Octimet Oncology Nv, Oncoethix, Oncomed, Oncopeptides, Onyx Therapeutics, Orion Pharma, Oryzon Genomics, Pfizer, Pharma Mar, Pierre Fabre, Rigontec Gmbh, Roche, Sanofi Aventis, Sierra Oncology, Taiho Pharma, Tesaro, Inc, Tioma Therapeutics, Inc., Xencor; Research Grants: Astrazeneca, BMS, Boehringer Ingelheim, Janssen Cilag, Merck, Novartis, Pfizer, Roche, Sanofi; Non-financial support (drug supplied): Astrazeneca, Bayer, BMS, Boringher Ingelheim, Johnson & Johnson, Lilly, Medimmune, Merck, NH TherAGuiX, Pfizer, Roche. All other authors have declared no conflicts of interest.

Background

HIF-1α and AP1 play important roles in hypoxia and activate anti-apoptotic pathways in tumor cells. No dual HIF-1α/AP1 inhibitors currently exist, so targeting these transcriptional factors is promising way to modulate hypoxia signaling in cancer cells. Aim of the study was obtaining and biological evaluation of hypoxia-selective 7-amino-6-halogen-substituted derivatives starting from 6,7-dihalogeno-3-phenylquinoxaline-2-carbonitrile 1,4-dioxides.

Methods

A series of 7-amino-6-halogeno-3-phenylquinoxaline-2-carbonitrile 1,4-dioxides was synthesized. Cancer cell lines were purchased from ATCC. The cytotoxic activity of compounds was evaluated in normoxia (21%O₂) and hypoxia (1%O₂). The cytotoxicity was assessed by MTT test (72 h growth with compounds). HIF-1α and AP1 activity was assessed by reporter analysis.

Results

Synthesis of 7-amino-6-halogen- substituted derivatives starting from 6,7-dihalogeno-3-phenylquinoxaline-2-carbonitrile 1,4-dioxides was carried out. A series was characterized by good solubility in water. Antiproliferative activity was evaluated in vitro on a panel of cancer cell lines including multidrug resistance variants. Novel synthesized compounds demonstrated higher hypoxia selectivity and cytotoxicity compared with tirapazamine. Some of the 7-amino-6-halogeno derivatives were 10-20-fold more potent than the reference drug. Selected 7-amino-6-halogeno derivatives LCTA-2425 and LCTA-2711 inhibited breast cancer cells growth in hypoxia at concentrations lower than 0.6 µM. Compounds LCTA-2711 and LCTA-2425 showed inhibitory effects on HIF-1α- and AP1-dependent luciferase activity, when tirapazamine revealed no potency to block these factors in MCF-7 breast cancer cells under hypoxic conditions.

Conclusions

A series of 7-amino-6-halogeno-3-phenylquinoxaline-2-carbonitrile 1,4-dioxides were more potent than reference drug tirapazamine in all tested cell lines and demonstrated high selectivity in hypoxia. Selected 7-amino-6-halogeno derivatives showed dual inhibitory activity against HIF-1α and AP1 factors, regulating anti-apoptotic pathways in hypoxia. RFBR grants 18-53-34005 (chemistry), 18-015-00422 (biology).

Legal entity responsible for the study

Alexander M. Scherbakov.

Funding

Russian Foundation for Basic Research, Grants 18-53-34005 (Chemistry) and 18-015-00422 (Biology).

Disclosure

All authors have declared no conflicts of interest.

Background

Anaplastic lymphoma kinase (ALK) is a tyrosine kinase receptor that is a clinical target of major interest in cancer. Mutations and rearrangements in ALK trigger the activation of the encoded receptor and its downstream signaling pathways. ALK mutations have been identified in both familial and sporadic neuroblastoma cases as well as in 30 to 40% of relapses, which makes ALK a bona fide target in neuroblastoma therapy. Tyrosine kinase inhibitors (TKIs) that target ALK are currently in clinical use for the treatment of patients with ALK-positive non–small cell lung cancer. However, monotherapy with the ALK inhibitor crizotinib has been less encouraging in neuroblastoma patients with ALK alterations, raising the question of whether combinatorial therapy would be more effective.

Methods

In this study, we established both phosphoproteomic and gene expression profiles of ALK activity in neuroblastoma cells exposed to first- and third-generation ALK TKIs, to identify the underlying molecular mechanisms and identify relevant biomarkers, signaling networks, and new therapeutic targets.

Results

The phosphoproteomic analysis identified several conserved oncogenic downstream signaling pathways of ALK, similar to those involved in insulin receptor (INSR)/tropomyosin receptor kinase (TRK) and fibroblast growth factor receptor (FGFR) signaling. In addition, signaling events involved in feedback and cross-talk were identified, including modulation of DUSP (dual-specificity phosphatase) family phosphatases. Furthermore, from analysis of the RNA-seq data, several transcription factors were predicted and validated as responsive to ALK signaling, including members of the FOXO (forkhead box O) and ETS (E26 transformation-specific or E-twenty-six) transcription factor families.

Conclusions

Although neuroblastoma is a complex heterogeneous disease, this in-depth investigation of downstream targets of ALK signaling offers future avenues to pursue to inhibit ALK-driven neuroblastoma.

Legal entity responsible for the study

Bengt Hallberg.

Funding

Barncancerfonden.

Disclosure

All authors have declared no conflicts of interest.

Background

Despite the major recent advances in cancer management, cure of cancer remains a serious challenge.

Methods

In the current study, we investigated the impact of butein, a biologically active flavonoid, on the human cancer cell survival and colony growth in vitro, and on tumor growth in vivo, alone and in combination with Frondoside-A, a triterpenoid glycoside isolated from an Atlantic sea cucumber, and camptothecin.

Results

We demonstrate that butein causes a concentration- and time-dependent decrease in the viability of the lung cancer cells (A549 and LNM35), the breast cancer cells (MDA-MB-231 and T47D), and a decrease in their related colonies growth in vitro. Similarly, treatment with butein significantly decreased the growth of A549 and MDA-MB-231 xenografts in the chick embryo model in vivo (P < 0.01) without significant toxicity. To determine whether the inhibition of cell viability and tumor growth by butein is due to growth arrest or to cell death, we assessed the induction of apoptosis by measuring caspase-3 activity and PARP cleavage. The treatment of lung and breast cancer cells with butein induce caspase-3 activation leading to PARP inactivation in a concentration and time-dependent manner. More than 50% of lung cancers and 40% of breast cancers show constitutive activation of STAT3. The role of this transcription factor in cancer progression is now well established and its targeting in cancer therapy is a very promising and challenging option. We observed that butein treatment induce a concentration and time-dependent inhibition of STAT3 phosphorylation without any impact on total STAT3. We finally demonstrate that butein at the IC25 and IC50 concentration enhances the anti-cancer effect of Frondoside-A as well as the effect of camptothecin on lung and breast cancer cells in vitro.

Conclusions

These findings increase our confidence of the potential benefit of using butein as an anticancer agent for the treatment of solid tumors either alone, and/or in combination with Frondoside-A and camptothecin.

Legal entity responsible for the study

Samir Attoub.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Most patients with advanced NSCLC don’t receive guideline recommended treatment as their poor performance status makes them ineligible for platin-based treatment protocols. While tumour-specific loss of ATM in early-resected NSCLC is associated with poor survival, these patients derive increased benefit from adjuvant chemotherapy, including cisplatin. We have previously reported that ATM-deficient cell lines in MCL and gastric cancer show increased sensitivity to PARP inhibition, and recent studies have indicated cisplatin sensitivity arising from DNA damage repair deficiencies. Here we present preclinical data suggesting that lower doses of platin in combination with PARP inhibition may be an option in patients whose tumours exhibit ATM loss.

Methods

We identified ATM-deficient NSCLC cell lines by protein expression and activation in response to ionizing radiation. Cells were treated with cisplatin and PARP inhibitor (olaparib) alone or in combinations to determine sensitivity by clonogenic survival assay. ATM was knocked out in an ATM-proficient cell line by Cas9-CRISPR gene editing, and cells were analyzed for the effect of cisplatin and olaparib on cell cycle progression.

Results

ATM-deficient NSCLC cell lines were sensitive to both cisplatin and olaparib alone, and this effect was amplified when drugs were given in combination, even at lower doses of both. ATM signalling was activated in response to these drugs in ATM-proficient cells, however when ATM is knocked out the cells become more sensitive to either drug alone or in combination. Interestingly, apoptosis does not appear to increase in ATM-KO cells, but rather cells accumulate in G2 – particularly cells treated with olaparib.

Conclusions

Our results suggest that ATM loss is sufficient to sensitize NSCLC cells to combinations of cisplatin and olaparib. PARP inhibition may arrest cells in G2, and while cisplatin does not appear to push cells into an apoptotic state, cell death might be triggered via another mechanism. This suggests that NSCLC patients with ATM loss in their tumour may benefit from lower doses of platin and PARP inhibitors, a combination that may allow increased uptake of palliative systemic treatment including the addition of immunotherapy.

Legal entity responsible for the study

D. Gwyn Bebb.

Funding

Glans-Look Lung Cancer Research.

Disclosure

All authors have declared no conflicts of interest.

Background

NSCLC patients undergoing tyrosine kinase inhibitor (TKI) therapy often develop resistance (9-12 months) resulting in low median survival (10-14 months) among stage IV patients. Studies showed that, in some cases, the resistance can be attributed to upregulation of pro-survival PI3K-Akt pathway. Indeed, EGFR-mutant patients with elevated levels of phospho-Akt have shown poor therapeutic response and the levels of p-Akt are increased upon prolonged EGFR TK treatment. In addition, several mechanistic studies attribute increased phospho-Akt (p-Akt) levels in tumor as one of the reasons for drug resistance. In an independent study, researchers have shown that suppression of phosphorylation of Akt can be achieved by inhibition of Phosphodiesterase- 4(PDE4) and in turn activate luminal apoptosis. Therefore, we hypothesized that PDE4 inhibition would sensitize drug resistant NSCLC to tyrosine kinase inhibitor therapy by abrogating the PI3K-Akt pathway.

Methods

TK resistant cell lines along with controls were used for the study. First generation TKIs, Dasatinib, Erlotinib and Gefitinib were used to study the efficacy of TKI. Roflumilast, a drug already used in chronic obstructive pulmonary disorder (COPD), was used as the PDE4 inhibitor.

Results

Inhibition of PDE4 followed by treatment with TKIs showed remarkable decrease in IC₅₀values, compared to either PDE4 inhibition or TK inhibition. Upon PDE4 inhibition, efficacy of TKIs is found to be increased several folds compared to individual inhibitors. Cells showed no signs toxicity to the PDE4 inhibitor at the dose levels used, thus making the inhibitor itself absolutely nontoxic at this treatment dose.

Conclusions

Our results suggest that PDE4 inhibition is a potential way to reverse TKI resistance and also to improve TKI therapy in patients with high levels of p-Akt.

Legal entity responsible for the study

University of Missouri - Columbia, Missouri, USA.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Prostate cancer (CaP) was the second most common cancer in men worldwide in 2012, and radiation therapy is one of the most common definitive treatment options for localized CaP. However, radioresistance is a major challenge for current radiotherapy, accumulating evidence suggests microRNAs (miRNAs), as an important regulator in cellular ionizing radiation (IR) responses, are closely correlated with radiosensitivity in many cancers.

Methods

We performed human miRNA probe hybridization chip analysis to identify the expression profile of miRNAs in CaP cells exposed to IR, and then we analysed the cell proliferation, cell viability, and cell cycle after transfection of miR‐16‐5p into the CaP cells. Analysis of the cyclin D1/E1–pRb–E2F1 pathway related proteins were performed by western blotting.

Results

microRNA- 16-5p (miR-16-5p) is significantly upregulated in CaP LNCaP cells following IR and can enhance radiosensitivity through modulating the cyclin D1/E1–pRb–E2F1 pathway. Overexpression of miR-16-5p suppressed cell proliferation, reduced cell viability, and induced cell cycle arrest at G0/G1 phase, resulting in enhanced radiosensitivity in LNCaP cells. Additionally, miR-16-5p specifically targeted the cyclin D1/E1-3′-UTR in LNCaP cells and affected the expression of cyclin D1/E1 at both mRNA and protein levels.

Conclusions

miR-16-5p enhanced radiosensitivity of CaP cells, the mechanism may be through modulating the cyclin D1/cyclin E1/pRb/E2F1 pathway to cause cell cycle arrest at G0/G1 phase. These findings provided new insight into the correlation between miR‐16‐5p, cell cycle arrest, and radiosensitivity in CaP, revealed a previously unrecognized function of miR‐16‐5p–cyclin D1/E1–pRb–E2F1 regulation in response to IR and may offer an alternative therapy to improve the efficiency of conventional radiotherapy.

Legal entity responsible for the study

Institute of Modern Physics, Chinese Academy of Sciences.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Prostate cancer is one of the most common types of cancer in men. Choline is an essential component of cell membrane phospholipids and is metabolized and internalized into cells by choline kinase, an enzyme that is overexpressed in certain tumors, such as prostate cancer. Two choline tracers are available for clinical use, which are labeled either with ¹¹C-choline or with ¹⁸F-choline. These choline PET tracers that target cell membrane metabolism have influenced prostate cancer imaging, particularly in biochemical relapse, and are therefore FDA approved for use in patients with recurrent disease. However, the uptake system for choline and the functional expression of choline transporters in prostate cancer are not totally understood.

Methods

We examined [3H]choline uptake in the prostate cancer cell line LNCaP, which depends on androgen. Cells were cultured in RPMI 1640 medium supplemented with 10 % fetal bovine serum and grown at 37 °C in 5% CO₂. The CellTiter-Glo Luminescent Cell Viability Assay is a homogeneous method of determining the number of viable cells. The Caspase-Glo 3/7 assay reagent was used for caspase detection in treated cells.

Results

[³H]Choline uptake is mediated by a single Na⁺-independent and intermediate-affinity transport system. Choline transporter-like protein 1 (CTL1) and CTL2 mRNA are highly expressed. CTL1 and CTL2 immunoreactivity were recognized in the plasma membrane and intracellular compartments, respectively. The anticancer drugs, flutamide, and bicalutamide inhibited cell viability and [³H]choline uptake in a concentration-dependent manner. The correlations between the effect of both anticancer drugs for cell viability and [³H]choline uptake were significant. The caspase-3/7 activity significantly increased by flutamide and bicalutamide. Furthermore, both flutamide and bicalutamide decreased the expression level of CTL1 in LNCaP cells.

Conclusions

These results suggest that CTL1 are functionally expressed in prostate cancer cells and are also involved in abnormal proliferation. Identification of this CTL1-mediated choline transport system provides a potential new target for prostate cancer therapy.

Legal entity responsible for the study

Tokyo Medical University.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Common mutations arise from MAPK and PI3K pathways are RAS, BRAF and PIK3CA. These mutations are focus for targeted inhibitors but also serve as integrators to drug resistance occurrence. Activated MAPK signaling will co-activate PI3K pathway which serves as by-pass mechanisms for drug resistance development. Many studies demonstrated that dual-inhibition of both pathways is a better strategy to overcome drug resistance. Our research investigates this rationale and tests if palbociclib (P; CDK4/6 inhibitor) can act synergistically with either gedatolisib (G; P13K/mTOR dual inhibitors) or PD0325901 (PD; MEK 1/2 selective inhibitor) to offer a highly potent antitumour efficacy in mutant CRC cell lines.

Methods

Five human CRC cells lines were profiled for mutations using Agena Massarray. The antiproliferative response to P, G and PD alone and in combination was assessed in each cell line. Finally, IC₅₀ and combination indexes at effective dose 50 (CI) were calculated using CalcuSyn.

Results

Table 19P:

Common mutations identified in each cell line and corresponding CI values for both drug combinations. The CI values <0.9 are synergistic, whereas CI values >1.0 are antagonistic. P+PD=palbociclib/PD0325901; P+G=palbociclib/gedatolisib; NR=not reached

Conclusions

Drug combination P+G has clear synergistic antiproliferative effect in CRC cell lines with common mutations arising from MAPK & P13K pathways except for BRAF_V600E. Combination P+G is also a potential option in CRC cells harbouring KRAS with(out) activating PIK3CA mutation, which associated with MEK inhibitor resistance. Our invitro data provide good rationale for further invivo evaluation and potential clinical drug development of P+G combination as future beneficial therapy in patients with treatment-resistant colorectal cancers.

Legal entity responsible for the study

Oncology Molecular Medicine, Royal College of Surgeons in Ireland (RCSI), Dublin.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Colorectal cancer is the fourth leading cause of cancer-related deaths worldwide. Current adjuvant chemotherapy regimens often lead to significant toxicities, so novel targets and targeted therapies are of great interest. Nischarin (NISCH, IR1, IRAS) is a non-adrenergic imidazoline-1 receptor protein that has been described as a tumor suppressor in breast and ovarian cancer. NISCH expression and its biological role in other types of cancer have not been investigated to date. Importantly, NISCH is a druggable target for which chemical agonists and antagonists have already been developed. The aim of this study was to examine the expression of nischarin in colon cancer and determine its potential role in disease progression and patient outcome.

Methods

The web resource for analyzing cancer transcriptome data from The Cancer Genome Atlas (TCGA) was used to analyze the NISCH mRNA expression levels in normal and colon cancer samples. Patients from the TCGA colon cancer database (n = 440) were divided into "high" and "low" groups based on mean NISCH expression and Kaplan-Meier survival analysis was performed. Gene Set Enrichment Analysis (GSEA) was performed on several publicly available datasets using the Hallmarks and KEGG gene sets to find gene expression signatures that correlate with NISCH expression.

Results

In the TCGA database there was no significant difference in expression of NISCH between normal and tumor tissue (p > 0.05). Survival analysis showed that patients with higher NISCH tumor levels had significantly shorter overall survival (p = 0.0141, Log-rank test). In the GSE41258 dataset, levels of NISCH were higher in primary tumors of patients with M1 status compared to tumors of patients without metastasis (p = 0.027, unpaired t test). GSEA revealed that gene expression sets of epithelial-mesenchymal transition, angiogenesis, focal adhesion assembly and extracellular matrix-receptor interaction, all of which are associated with metastasis, were significantly correlated with NISCH expression in primary tumors.

Conclusions

In colon cancer, unlike in breast and ovary, higher NISCH expression might be associated with poor prognosis. Given that nischarin is a druggable target, it is potentially a new therapeutic target for selective treatment of patients with advanced colon cancer.

Legal entity responsible for the study

Institute for Oncology and Radiology of Serbia.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Cancer, a heterogeneous disease is caused by genetic and epigenetic mutation. Irinotecan was indicated for treatment of colorectal cancer either alone or in combination with other chemotherapeutic drug. Due to associated toxicity, irinotecan was less explored in cancer therapy research. In recent years, magnetic nanoparticles (MNPs) were well explored for different biomedical applications including targeted drug delivery.

Methods

We hypothesized that the anticancer activity of irinotecan would improve when attached to the surface of MNPs. Magnetic nanoparticles were developed using in house developed method. Fourier transform infrared (FT-IR) spectra and scanning electron microscopy (SEM) were recorded to characterize the irinotecan MNPs. In Vitro and in vivo evaluations were perfomed to assess the feasibility of this novel formulation using colon cancer cell lines.

Results

Irinotecan MNPs were successfully developed with an average diameter of the magnetic core of 22 ± 5 nm and the thickness of the shell is around 7 ± 4 nm. Using colon cancer culture (HT-29 cells), we evaluated the effects of MNPs on cancer cell viability and apoptosis. Treatment of cancer cells with the irinotecan MNPs caused a greater decrease in cell viability and a higher rate of apoptosis compared with treatment using free irinotecan.

Conclusions

Although further clinical study is needed, the targeted uptake of irinotecan MNPs by colon cancer cells associated with their ability to eliminate and prevent further cancer cell growth indicates its potential for targeted and personalized cancer therapy with lower toxicity in patients with colon cancer.

Editorial acknowledgement

Not Applicable

Legal entity responsible for the study

Akhilesh Vikram Singh.

Funding

Has not received any funding.

Disclosure

The author has declared no conflicts of interest.

Background

Matricellular proteins are a subset of extracellular matrix (ECM) proteins which are dynamically expressed and serve many regulatory roles. Among matricellular proteins, the CCN family, a group of secreted proteins are known to regulate cell adhesion, migration, proliferation, differentiation, apoptosis, survival, senescence, and gene expression. Connective tissue growth factor (CTGF), also known as CCN2, is a member of the CCN family. CTGF is known to have many roles in biological processes such as cell proliferation, migration, adhesion, and angiogenesis.

Methods

To investigate the function of CTGF in triple-negative breast cancer cells (TNBC), we silenced CTGF in TNBC cell lines using short hairpin RNA. Knockdown of CTGF was verified by western blot, qPCR, and immunostaining. The effect of CTGF knockdown on TNBCs was examined by cell proliferation assay, adhesion assay, migration assays, metabolism assays, and cell cycle analysis.

Results

Knockdown of CTGF decreased cell proliferation, adhesion, migration, glucose uptake, ATP production and lactate production. Since CTGF is a secreted protein, we gave recombinant human CTGF (rhCTGF) to the cells and found that rhCTGF induced activation of the Src/FAK/MAPK pathway and led to expression of proteins related to cell cycle progression. Also, when CTGF-specific antibodies were given to the cells, they expressed cytotoxicity by neutralizing the extracellular CTGF and decreasing CTGF-mediated signalling.

Conclusions

We suggest that the secreted CTGF mediates tumor cell progression via modulation of cell proliferation, adhesion, migration and metabolism and could possess a potential for being a therapeutic target.

Legal entity responsible for the study

Hanyang University.

Funding

National Research Foundation of Korea.

Disclosure

All authors have declared no conflicts of interest.

Background

CCN3(NOV) is a matricellular protein belonging to the CCN family and is known to be involved in various cellular signaling processes including tumorigenesis. Previous studies have shown that CCN3 can promote cell migration in sarcoma, glioblastoma, prostate and renal cancer cells. In-silico analysis using the TCGA database shows CCN3 was highly amplified in breast cancer patients especially in TNBC (triple-negative breast cancer) patients. Accordingly, we confirmed that the expression of CCN3 is up-regulated in the TNBC cell lines. Based on these findings, it was thought that CCN3 would participate in TNBC tumorigenesis.

Methods

We generated CCN3 knockdown TNBC cell lines by using shRNA. Proliferation assays and wound healing assay were performed to check the phenotype changes in the established cell line, and western blotting and RT-qPCR were used to monitor changes in intracellular protein expression. In addition, in-silico analysis using the TCGA database confirmed gene sets affected by CCN3 expression.

Results

We identified that CCN3 knockdown cell lines exhibited decreased proliferation, migration ability and colony-forming ability compared with control cell lines. Then we performed immunoblotting to find altered signaling by CCN3 knockdown. Phosphorylation level of FAK and EGFR were decreased in CCN3 knockdown cell lines.

Conclusions

We have shown that CCN3 can be involved in proliferation and migration ability in TNBC cell lines. We thought that this regulation may involve EGFR, which is known to be overexpressed in TNBC, or phosphorylation of FAK through integrin signaling. This study shows that CCN3 expressed in TNBC affects tumorigenesis of TNBC and may be used as a possible molecular therapeutic candidate to target TNBC.

Legal entity responsible for the study

Hanyang University.

Funding

National Research Foundation Korea.

Disclosure

All authors have declared no conflicts of interest.

Background

Hypoxia plays an important role in ovarian cancer and understanding the role of reactive oxygen species and VEGF may identify patients who benefit the most from targeted therapies. The aims of the study were to determine the intensity of oxidative stress in ovarian cancer patients and to establish a connection between the presence of the reactive oxygen species (ROS) and VEGF.

Methods

Forty-one patients diagnosed with epithelial ovarian carcinoma stage II-IV between 2010 and 2017, who underwent multimodality treatment (surgery and chemotherapy) were included in the study. ROS measured in dynamic (four determinations between every cycle) were malondialdehyde to evaluate the lipid peroxidation, ceruloplasmin, SH- albumin thiols groups and total antioxidants.

Results

There was an increase in the value of ROS: malondialdehyde mean value was 8.5 μmol/100 ml (normal value 4 μmol/100 ml); ceruloplasmin mean value was 147.8U.I. (normal value 120U.I), both showing an active oxidative process in patients with ovarian cancer. A small decrease of the value of thiols (397 vs. 450 μmol/l) and a small increase of total antioxidants was noticed (1.45 vs. 1.4 μmol). All four compounds decreased between the first determination and the fourth one. There was a strong correlation between lipid peroxides levels and ceruloplasmin (Pearson correlation 0.315 p = 0.005) and between lipid peroxides and thiols groups (Pearson correlation 0.23 p = 0.039). There was a correlation between thiols and antioxidants (Pearson correlation 0.33 p = 0.003). Lipid peroxidation and ceruloplasmin were significantly higher in patients with residual disease (p = 0.039, p = 0.046) emphasizing that the tumor is a generator of oxidative stress. Mean VEGF levels were elevated 983.42 pg/ml, and VEGF levels statistically significant corelates with malondialdehyde (Pearson correlation 0.439, p = 0.036).

Conclusions

Tumours produce ROS in excess in patients with advanced ovarian adenocarcinoma. Those ROS are corelated between each other and with VEGF and act as signalling molecules.

Legal entity responsible for the study

National Institute of Oncology Al. Treistioreanu.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Recently the texture analysis has been applied to help predict the clinical outcome and treatment response of various tumor types. This study investigates the relationship between the contrast enhanced computed tomography (CECT) texture features of small cell lung cancer (SCLC) tumors tissue, and the survival of the patients.

Methods

Eighty-eight patients diagnosed with unresectable SCLC (Extended stage, 57; Limited stage, 31), and treated with platinum-based chemotherapy at West China hospital between January 2010 and 2015 were enrolled. All the patients are followed for at least 18 months or until death. The texture features of tumor tissues were extracted from CECT images taken prior to treatment. Based on the receiver operating characteristics curve analysis, the optimal cutoff value of each texture parameter was obtained for further calculation. The Kaplan-Meier analysis was performed to compare the eighteen-month overall survival (OS) rates and six-month event-free survival (EFS) rates of different patient subgroups. The multivariate Cox regression analysis was performed to determine the prognostic value of texture features on CECT images in patients with unresectable SCL C.

Results

Of all CT texture features analyzed, six (GLCM-Contrast, GLCM-Dissimilarity, HISTO-Energy, HISTO-Entropy, HISTO-Skewness and Std-value) were shown to be significantly related to 18-month OS and could be considered as independent predictors of patients’ survival. Two features (GLCM-Energy and GLCM-Entropy) were shown to be significantly correlated with the 6-month EFS but could not be taken as independent predictors according to the multivariate Cox regression analysis.

Conclusions

Texture features of contrast-enhanced computed tomography could potentially serve as prognostic predictors for SCLC patients taken platinum-based chemotherapy. In view of the present study and previous reports, the CECT-based teture analysis atlas can be recommended in the clinical practice of SCLC.

Legal entity responsible for the study

West China Hospital, Sichuan University.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

There is increasing interest in the use of circulating tumour DNA (ctDNA) to identify targetable genomic alterations for therapy selection. However, the feasibility of next generation sequencing on ctDNA in BC patients with varying disease burden merits further investigation.

Methods

The cohort of 35 BC patients included 30 metastatic cases with paired primary and metastatic specimens in addition to plasma taken prior to commencement of a new line of palliative systemic therapy (all subtypes), + 5 patients (3 stage III, 2 stage II) about to commence neoadjuvant systemic therapy. DNA from tumour, buffy coat and plasma was sequenced on a 77-gene capture panel customised for BC. Matched tumour/normal samples were processed to discover somatic alterations using a standard GATK pipeline. Plasma samples were processed using unique molecular identifiers to identify potential alterations at low frequency.

Results

Among the entire cohort, 74% (26/35) of patients had mutation(s) in at least 1 gene detected from ctDNA: 60% (3/5) for the neoadjuvant cases, 77% (23/30) for the metastatic cases. The most frequently mutated genes from ctDNA analysis were TP53 (17/35; 49%) and PIK3CA (8/35; 23%), with HER2, HER3, JAK2, NF1, MAGI3 and TSC2 mutations observed in 2 patients each (6%). There was no obvious correlation between detection of ctDNA mutation and disease burden, serum CA-15.3 tumour marker or circulating tumour cell levels using a microfluidic platform. Out of 35 patients, 21(60.0%) had ≥1 concordant mutation via both ctDNA and tumour genotyping. Concordance between the plasma and primary and/or metastatic lesions was observed for 59% (39/66) of the mutations detected in ctDNA. Among the metastatic cases, 14/37(38%) of the concordant mutations were shared between the plasma, primary and metastatic specimens, while 5(14%) were shared between the plasma and the primary only, and 18/37(49%) shared between the plasma and metastatic lesions only.

Conclusions

Detection of genomic alterations from ctDNA is feasible in BC patients, but concordance of mutations in ctDNA is better with the metastatic than the primary lesion. This is likely due to suboptimal quality of DNA from archived tissue, and spatial, temporal heterogeneity.

Legal entity responsible for the study

National Cancer Centre Singapore and Genome Institute of Singapore.

Funding

SingHealth Foundation.

Disclosure

Y.S. Yap: Personal financial interests, honoraria for consultancy and talks, travel support: AstraZeneca, Eisai, Lilly, Novartis, Pfizer, Roche. All other authors have declared no conflicts of interest.

Background

Recent studies have demonstrated of utility of quantifying circulating proteins for early disease detection, prediction of therapeutic response, and treatment monitoring. Although the utility is clear, standardization of measurement methods for a large number of proteins from plasma remains challenging. Here, plasma proteins are detected using hyper-reaction monitoring (HRM) mass spectrometry in samples from normal and pancreatic cancer subjects. The data is acquired and analyzed simultaneously with unbiased label-free quantification of all proteins and absolute quantification of proteins from a > 500-protein panel constructed from stable-isotope standard (SIS) peptides.

Methods

Plasma samples from subjects with Stage IV pancreatic cancer (PC, n = 6) and age matched healthy donors (n = 15) were prepared for mass spectrometry. Prior to analysis, a panel of SIS peptides, covering 582 plasma proteins, was spiked into each sample. All samples were acquired using nano-flow liquid chromatography with separation over a one-hour gradient coupled online to a Thermo Scientific Q Exactive HF mass spectrometer. Protein data was extracted using Spectronaut (Biognosys) and statistical analysis was conducted to identify disease associated biomarker candidates.

Results

Analysis of the PQ500 panel enabled absolute quantification of 282 proteins across all samples. Univariate statistical testing identified 29 candidate proteins (27 up- and 2 down-regulated; q-value > 0.05 and fold change > 1.5). Key dysregulated proteins include CRP, SAA1, and C9. With unbiased, label-free quantification, 414 proteins were quantified, with 87 significantly regulated. In addition to the acute phase protein candidates identified in the PQ500 panel, 12 adhesion related protein candidates were identified (e.g. TLN1, MYH9, TPM4), 9 of which were decreased in PC. Notably, membrane associated ICAM1 and VCAM1, both potential therapeutic targets, were increased in PC.

Conclusions

Combining PQ500 with unbiased label-free quantification enables comprehensive characterization of the plasma proteome, while at the same time providing absolute quantification for a large subset of well annotated, clinically relevant proteins.

Legal entity responsible for the study

Biognosys AG, Schlieren, Switzerland.

Funding

Biognosys AG, Schlieren, Switzerland.

Disclosure

H. Yu, J. Vowinckel, C. Escher, D. Heinzmann, N. Dupuis: Employee: Biognosys AG.

Background

Insufficient information is available on cancer types among patients enrolled in all-comer phase I oncology trials. We evaluated the global trends in cancer types of patients in these trials, including time-dependent changes and regional differences among North America, Europe, and Asia.

Methods

The PubMed database was searched to identify single-agent phase I trials published between 1991 and 2015, which enrolled patients with any type of solid tumor. The inclusion and exclusion criteria were predefined for article selection; trials recruiting from specific patient populations were excluded from the analysis.

Results

We identified 866 eligible clinical trials, which enrolled a total of 29,112 patients with advanced solid tumors. The selected trials were conducted mainly in North America (55.5%), Europe (27.8%), and Asia (10.5%). The distribution of cancer types in phase I trials was considerably different from cancer-related incidence and mortality. Colorectal cancers (n = 7,510, 25.8%) were the most prevalent in phase I trials, followed by lung cancer (n = 3,212, 11.0%), sarcoma (n = 1,756, 6.0%), breast cancer (n = 1,623, 5.6%), renal cancer (1589, 5.5%), and ovarian cancer (1473, 5.1%). The proportion of patients with either colorectal or lung cancer decreased with time. The proportion of trials, in which patients with either of these two cancers accounted for ≥50% of the total enrolled patients in each trial, had also decreased as follows: 31/67 trials (46.3%) from 1991 to 1995, 58/142 (40.8%) from 1996 to 2000, 59/223 (26.5%) from 2001 to 2005, 38/189 (20.1%) from 2006 to 2010, and 41/245 (16.7%) from 2011 to 2015. These trends were consistent across the three regions, with an increase in the proportion of various types of cancer enrolled in phase I trials.

Conclusions

The distribution of cancer types among patients enrolled in all-comer phase I trials has changed dramatically. Patients with common types of cancer, with poor general condition and vital organ dysfunction after multiple lines of therapy, are likely not to participate in phase I trials.

Legal entity responsible for the study

Noboru Yamamoto.

Funding

Has not received any funding.

Disclosure

T. Shimizu: Consulting: Takeda Oncology; Honoraria: ONO, ONO Pharma Taiwan, Boehringer Ingelheim, Taiho, Chugai; Research funding: Takeda Oncology, PharmaMar, BMS Japan, Daiichi Sankyo, SymBio, Five Prime Therapeutics, 3D Medicine. S. Kondo: Research funding: AstraZeneca, Eli Lilly, Pfizer. Y. Fujiwara: Consulting: ONO, BMS Japan; Participation in speakers’ bureau: ONO, BMS Japan, MSD, Taiho; Research funding: AstraZeneca, Chugai, Daiichi Sankyo, Eisai, Lilly Japan, Novartis, BMS Japan, MSD, Merck Serono, Abbvie, Incyte. N. Yamamoto: Consulting: Eisai, Takeda, Otsuka, OncoTherapy; Speakers’ bureau: BMS, Pfizer, AstraZeneca, Lilly, ONO, Chugai; Funding: Chugai, Taiho, Eisai, Quintiles, Astellas, Novartis, Daiichi Sankyo, Boehringer, Takeda, Kyowa Kirin, Bayer, Pfizer, BMS, ONO.

Background

Esophageal squamous cell carcinoma (ESCC) is the most important pathological type of esophageal cancer with high incidence and limited efficient therapies in China and worldwide. Understanding of ESCC genomic features is advantageous for exploring clinical therapeutic strategies.

Methods

Deep sequencing targeting 450 cancer genes was performed on FFPE and matched blood samples collected from 90 Chinese ESCC patients(pts) with a median age of 60 years old. Genomic alterations (GAs) including single nucleotide variations, short and long indel, gene rearrangements and copy number variations were analyzed. Tumor mutation burden (TMB) was assessed in all samples by standard NGS algorithms. The expression of PD-L1 in 44 samples was evaluated by IHC (28-8 Ab).

Results

The most commonly found GAs were mutations in TP53 (94.4%), FGF3 (43.3%), CCND1 (43.3%), FGF4 (42.2%), FGF19 (42.2 %), CDKN2A (36.7%), NOTCH1 (25.6%), PIK3CA (22.2%), and ERBB2 amp(2.2%).Compared with TCGA data, the frequency of TP53 and ERBB2 amp mutations was similar, while the frequency of NOTCH1 mutations was significantly higher (25.6% vs 11%) and the frequency of EGFR amp was lower (5.6% vs 19%) in Chinese cohort. In addition, 4 of 90 samples harbored germline mutations (SDHA, RAD51C, PALB2 and BCL2L11). Among these GAs, 73.3% mutations were in cell cycle (62.2%) and PI3K/AKT (42.2%) pathway. In Chinese ESCC pts, the median TMB was 7.0 muts/Mb, and some pts showed high TMB (33.3%pts≥10 muts/Mb,11.1%pts≥20 muts/Mb). 15.6% pts (TMB≥10 muts/Mb) harbored chromosome 11q13 (FGF3/4/19 and CCND1) amp, which may be related to immunotherapy hyper-progression. Based on the IHC analysis, 10 of 44 Chinese pts showed positive (TPS≥1%) PD-L1 expression, and 7 of them also showed high TMB (≥10 muts /Mb). However, no correlation was found between high TMB and PD-L1 expression.

Conclusions

In this study, we found that the frequencies of GAs, such as EGFR and NOTCH1 alterations, were different between Chinese and western ESCC pts. Compared to Western ESCC pts, the positivity of PD-L1 expression was similar, while TMB was higher in Chinese ESCC pts. These findings suggested potential therapeutic targets for targeted and immunotherapies of Chinese ESCC pts.

Legal entity responsible for the study

OrigiMed.

Funding

Has not received any funding.

Disclosure

M. Yao, A. Liu, Y. Zhou: Employee: OrigiMed. All other authors have declared no conflicts of interest.

Background

The clinical behavior of GISTs is highly variable. This study aims at detection of different histo-pathological tumor features and correlation with different clinical aspects after treatment with imatinib, based on C-KIT exon mutation status.

Methods

This is a retrospective study that included all cases diagnosed as GIST who presented to NCI from 2005 to 2017, The following data were collected from the patient’s files: age, gender, tumor site, size, mitotic rate, histological grade, capsular rupture, risk stratification by Joensuu criteria, treatment setting, date of start and end of treatment, dose and toxicity. KIT mutation was assessed on tumor tissues of all patients; clinical correlation between different clinical aspects after treatment with imatinib, based on C-KIT exon mutation status, was done.

Results

Eighty-nine cases of GIST presented to NCI in the period September 2005 to January 2017. Median age at diagnosis was 48 years old with a median follow up of 22 months. More than 75% of the patients had positive C-KIT mutation, while it was negative in 24.7 % of the patients. C-kit positive mutations were significantly associated with tumors more than 5 cm, high mitosis, and with high tumor risk stratification by Joensuu criteria in more than fifty percent of the patients. Exon 9 mutations had poor ORR (55.6 %) compared to those with exon 11(67.6%) (P = 0.33), with the latter having PFS of 55 months compared 120 months for exon 9 mutations, (P = 0.002). No statistically difference in OS was observed with exon 9 (70 months) compared to exon 11 mutations (77 months) (P = 0.55).

Conclusions

C-kIT-positive mutation per-se is an independent poor risk factor, associated with more aggressive tumor features whereas response to imatinib was affected by exon mutations with exon 11 having a tendency for better ORR, compared to exon 9 mutation, with the latter having longer PFS compared with exon 11, with no statistically difference in OS with exon 9 compared to exon 11 mutations.

Legal entity responsible for the study

NCI.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Metallothioneins (MTs) family is a intracellular and cysteine-rich proteins with a high affinity to metals. MT-3 could play important neuromodulatory and neuroprotective roles. MT-3 has been also found up-regulated in a number of cancers. Neuroblastoma (Nbl) is a cancer is the most common extra-cranial solid tumour of childhood. The main aim was to provide novel insights into the molecular mechanisms of hMT-3 up-regulation and to elucidate the effects beneath the MT-3 up-regulation in Nbl cells.

Methods

To increase the expression of MT-3 Nbl (UKF-NB-4) cells were transiently transfected with a plasmid containing MT-3 gene (pcDNA3.1-GFP-hMT-3-TOPO). Separations of tryptic digests were carried out on an Easy-nLC 1000 nano system. MS analysis was performed using a Q-Exactive MS. The mass spectrum *.raw file was searched against the human SwissProt 57.15 database using MASCOT search engine (version 2.3, Matrix Science).

Results

The efficiency of transfection analysed through a fluorescence of GFP tag expressed at the C-terminus of MT-3 showed more 70% transfection efficiency for both constructed plasmids (mock and MT-3). From the total of common proteins in dataset (hMT-3 vs. mock), 176, 20 and 1244 proteins were quantitatively identified with up-regulation, down-regulation, and no significant differences between hMT-3 and mock treatments. Noteworthy, the bioinformatical analyses revealed the exclusive expression (induced by MT-3) and up-regulation proteins of a number of proteins affecting biological pathways related to mitotic cell cycle, cellular responses to stress, positive regulation of proteolysis, negative regulation of cell cycle and programmed cell death.

Conclusions

Our proteomic data shed some light on the proteins involved in inducing senescence and apoptosis in tested Nbl cells with up-regulated MT-3. Organisms with renewable tissues had to evolve mechanisms to prevent the development of cancer. Cellular senescence and apoptosis are among those mechanisms. Further experiments will be performed to functionally verify these data to provide novel insights into the Nbl biology. These might be useful to develop novel therapeutic protocols utilizing MT-3 as prognostic biomarker or therapeutic target.

Legal entity responsible for the study

Department of Chemistry and Biochemistry, Mendel University in Brno, Brno, CZ.

Funding

European Research Council (ERC) under the European Uniońs Horizon 2020 Research and Innovation Programme (grant agreement No. 759585).

Disclosure

All authors have declared no conflicts of interest.

Background

PD-1/PD-L1 inhibitors are novel anti-cancer agents for various tumors. PD-1/PD-L1 inhibitors induced rash occurred in 20% to 30%. The therapy for rash includes anti-histamine and corticosteroid. Bilastine is a non-sedating second-generation H1-antihistamine. Bilastine showed the efficacy for urticaria, prurigo and cutaneous pruritus. However, its effectiveness for PD-1/PD-L1 inhibitors induced rash is unknown. The objective of this retrospective study was to evaluate the efficacy of bilastine for PD-1/PD-L1 inhibitors induced rash.

Methods

We identified 224 patients who received PD-1/PD-L1 inhibitors (nivolumab, pembrolizumab, atezolizumab) at the Kure Medical Center from September 2014 to October 2018. PD-1/PD-L1 inhibitors induced rashes were observed in 84 patients (37.5%). They were classified into 4 groups on the basis of the systemic antihistamine and topical corticosteroid therapy: the (1) bilastine and corticosteroid group (n = 18), (2) another anti-histamine and corticosteroid group (n = 22), (3) bilastine group (n = 20); and (4) another antihistamine group (n = 24). Adverse events were graded according to the Common Terminology Criteria for Adverse Events (version 4.0). This study was approved by the Kure Medical Center IRB.

Results

The bilastine and corticoegsteroid group had significantly shorter the median duration of topical corticosteroids and antihistamine than the another antihistamine and corticosteroid group (p<0.01). Bilastine group had significantly shorter the period of systemic medications than the another antihistamine group (p<0.01). The incidence of adverse events was observed as follows, somnolence in 3% (1/38), headache 3% (1/38) and dizziness in 3% (1/38) in the bilastine and corticosteroid group and bilastine group. There were no serious adverse events.

Conclusions

Bilastine treatment reduced the need for topical corticosteroids use and shortened the period of topical corticosteroids for PD-1/PD-L1 inhibitors induced rash with acceptable safety profiles. Bilastine may be more effective than another antihistamine for PD-1/PD-L1 inhibitors induced rash.

Legal entity responsible for the study

Taizo Hirata.

Funding

Has not received any funding.

Disclosure

The author has declared no conflicts of interest.

Background

The aim of the present study was to investigate the mechanisms underlying the radiation-sensitising effect from antennapedia proteins (ANTP)-SmacN7 on induction of apoptosis in lung cancer cells irradiated with high-LET ionizing irradiation (IR) from accelerated carbon and iron particles.

Methods

Two cultured human non-small lung cancer (NSCLC) cell lines, A549 and NCI-H460, were irradiated with low-LET X-irradiations or high-LET IR with or without treatment of ANTP-SmacN7. Change of cell survival, induction of apoptosis and cell cycle progression, and alterations in both death and survival signals for apoptosis, were studied by colony formation assay, flowcytometry, and Western blot analysis, respectively.

Results

Showed that at the LD₅₀ for clonogenic cell killing by high-LET iron particles, compared to the low-LET X-rays irradiations, high-LET IR was more efficient for clonogenic cell killing and induction of apoptosis, which was correlated with cell G₂/M phase progression. In addition, ANTP-SmacN7 markedly promoted apoptotic cell killing through inhibition of X-linked inhibitor of apoptosis protein (XIAP) and activation of caspase-3 and 9. Furthermore, both antiapoptotic and proapoptotic molecular response was correlated with the apoptotic cell killing and in accordance with the results of clonogenic cell killing.

Conclusions

These findings provide useful information to contribute to the improvement of high-LET clinical radiotherapy for NSCLC from the point of view of pharmaceutical radio-sensitization.

Legal entity responsible for the study

Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou, PRC.

Funding

Has not received any funding.

Disclosure

The author has declared no conflicts of interest.

Background

Triple negative breast cancer (TNBC) is the most aggressive breast cancer (BC) subtype. It has the worst prognosis, highest recurrence and metastatic rates. Due to the clinical and molecular heterogeneity of TNBC, there is an emerging need to identify new molecular therapeutic targets. Nitric oxide (NO) has a dual role in cancer depending on its levels. At low concentrations, NO promotes tumor growth, while at high concentrations, NO has an anti-neoplastic function. Natural compounds have emerged as signaling pathways’ regulators in various tumors. Cardenolides specifically have potent cytotoxic effects in different cancers as lung and liver cancer. Our group has isolated the cardenolides, Calotropin and 7,8-dehydrocalotropin from Calotropisgigantea (L.) Dryand (Apocyanaceae). Calotropin showed potent cytotoxicy against non-small cell lung cancer, glioblastoma and prostate cancer, but has never been investigated against BC. Our aim was to investigate the anticancer effects of the isolated compounds on MDA-MB-231 TNBC cells by functional characterization and unravel their role in regulating NO levels in BC.

Methods

MDA-MB-231 cells were treated with serial dilutions (1, 5, 10, 20, 60 and 100 μM) of calotropin and 7,8-dehydrocalotropin. Their cytotoxic activities were assessed using MTT for cellular viability and IC₅₀ values were obtained. Cellular migration and colony forming ability were measured using scratch and colony forming assays, respectively. NO production was measured using Greiss reagent.

Results

Both calotropin and 7,8-dehydrocalotropin were able to decrease cellular viability, migration and colony formation of MDA-MB-231 cells in a dose-dependent manner. Calotropin reduced NO levels in MDA-MB-231 cells. However 7,8-dehydrocalotropin did not have any significant effects in regulating NO.

Conclusions

Calotropin showed more potent cytotoxicity on MDA-MB-231 cells compared to 7,8-dehydrocalotropin. Calotropin acts as a negative regulator of NO production, whereas 7,8-dehydrocalotropin failed to regulate NO production. This could be attributed to the structural difference between both compounds. Thus calotropin can be developed as a targeted therapy against BC.

Legal entity responsible for the study

German University of Cairo.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Complimentary datasets and functionality that facilitate comparisons of genomic, molecular and pharmacological data within the NCI-60 cancerous cell lines, Cancer Cell Line Encyclopedia (CCLE), Genomics of Drug Sensitivity in Cancer (GDSC), Cancer Therapeutics Response Portal (CTRP), NCI/DTP small cell lung cancer (SCLC), and NCI Almanac cell line sets are provided by the CellMiner (http://discover.nci.nih.gov/cellminer) and CellMinerCDB (https://discover.nci.nih.gov/cellminercdb/) web-applications.

Methods

Pharmacogenomics analyses using CellMiner compare the 60 cancerous cell lines of the NCI-60 using five tools, and include 22 data sets. Pharmacogenomics analyses using CellMinerCDB compare the NCI-60, CCLE, GDSC, CTRP, NCI/DTP SCLC, and NCI Almanac cell line data six, using eight tools, and include 29 data sets. Both provide multiple ways to download or query that data, and are described in detail in their respective urls.

Results

Data for the NCI-60, providing the most extensive public set of cell line molecular and drug activity data (generated by the NCI Developmental Therapeutics Program https://dtp.cancer.gov), are made available through CellMiner. The substantially increased cell line numbers and tissue of origin types provided by the CCLE, GDSC, and CTRP are contained in CellMinerCDB. Separate but complimentary functionalities are provided by the two web-applications. Variable numbers and kinds of data types are available for the differing cell line sets. The composition and numbers of cell lines also varies within the different sets, with 60 for the NCI-60, 69 for the SCLC, and ∼1000 for the CCLE, GDSC, and CTRP. One may fill in data gaps by merging data from multiple sources, taking advantage of the partial cell line overlaps that exist between many of these cell line sets. Extended analysis and quality assessment are also made possible by comparisons of data from multiple institutions.

Conclusions

Exploration of the relationships between and among molecular alterations and pharmacological responses in cancer cell lines from the omic perspective is facilitated by this rich set of data and functionalities.

Legal entity responsible for the study

The National Cancer Institute, USA.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.