Background

Chemotherapy is the main mode of treatment for the ER negative (ER-ve) subtype of breast cancer. However, despite an initial good response there is a high risk of recurrence in this subtype. microRNAs play an important role in mediating this drug resistance by regulation of the tumor-stromal interactions. Here we have explored the role played by miR-18a in immune modulation and chemo-responsiveness in ER-ve breast cancer.

Methods

Chemosensitivity to Paclitaxel (20-200 μM) was evaluated using MTT assay in an ER-ve cell line; MDA-MB-468 after inhibition of miR-18a using synthetic inhibitors. Expression of integrin β3 was evaluated by immunohistochemistry and that of miR-18a by q-PCR in surgically excised breast tumors with residual disease from partial and non-responders to neo-adjuvant chemotherapy (NACT) (n=57). The expression of miR-18a was further analysed in ER-ve breast tumors from the TCGA (n=116) and the METABRIC cohort (n=107). Differentially expressed genes (DEGs) and deregulated pathways were analysed between miR-18a high and low groups using G:profiler. Immune cell identification was done using CIBERSORT analysis.

Results

miR-18a inhibition in MDA-MB-468 led to increased chemoresistance at 200 μM paclitaxel (p=0.005). In the residual tumors of non-responders to NACT, 78% of the ER-positive and 60% of the ER-ve tumors had a lower expression of miR-18a. miR-18a low tumors also expressed high levels of integrin β3, a marker for chemo-unresponsiveness (p=0.01); the same association was observed in residual tumors of ER-ve subtype (p=0.1). Functional enrichment of DEGs in miR-18a low tumors of the TCGA and METABRIC cohorts demonstrated activation of pathways involving integrin cell surface interactions and drug transporter genes like ABCC11, ABCG1, & ABCG2. (p<0.05). Immune cell stratification using CIBERSORT algorithm revealed a higher proportion of M2 macrophages (p<0.008) and a lower proportion of M1 macrophages in the low miR-18a expressing tumors (p=0.01).

Conclusions

Our results support the existence of a miR-18a mediated chemoresistance within ER-ve tumors. Immunomodulation brought about by the less expression of miR-18a may contribute to immune suppression and chemoresistance in these tumors.

Legal entity responsible for the study

St. John's Research Institute.

Funding

Department of Health Research and ICMR-India Welcome Trust-DBT.

Disclosure

All authors have declared no conflicts of interest.

Background

Breast cancer is the commonest cancer among women requires multimodality treatment including adjuvant radiotherapy for better loco regional control. Long term toxicity to heart and lung following chemotherapy especially anthracyclines and Trastuzumab.Deep inspiratory breath-hold technique is a state of art technique for reducing heart doses without compromising target coverage or other organ-at-risk. The technique is based upon the observation that during inspiration, the flattening of the diaphragm and expansion of the lungs pulls the heart away from the chest wall. Surface guided radiotherapy (SGRT) is a magnificent devise helps us to do deliver and monitor radiotherapy with DIBH technique easily. This study explores the benefit of using DIBH with SGRT in left sided breast cancer radiotherapy treatment.

Methods

Eighty four breast cancer patients (both BCS and MRM), received adjuvant radiotherapy, between December 2019 and February 2022 were studied in this study. Patients were immobilized using breast boards with both arms positioned above the head. All patients had 2 sets of planning CT images (one in FB and another in DIBH) with a HD CT scanner in the same positional setup.Doses to the target and OARs and patient’s positional shits with three point laser and Align RT (VisionRT) surface guided RT were compared. The standard fractionation regimen of 50Gy in 25 fractions over a period of 5 weeks was used for all patients in this study.

Results

Among Eighty-four enrolled patients, average Dmean, V5%, V10%, V30% to heart with DIBH is 6.27 Gy, 33.04 GY, 19.49 Gy, 3.85Gy respectively. Average Dmean, V5%, V10%, V30% to heart with free breathing is 8.87Gy, 42.2 Gy, 29.9 Gy, 4.9Gy. which shows a statistically significant reduction in dose delivered to heart by using DIBH technique. Average dose to lung in terms of V30 Gy, V20Gy, V12Gy, V5Gy with DIBH is observed to be 24.38%, 28.9%, 36.51%, 48.98% respectively. While using FB technique lung doses in terms of V30 Gy, V20Gy, V12Gy, V5Gy with DIBH is observed to be 27.86%, 32.74, 37.58, 49.45 respectively.

Conclusions

Surface guided radiotherapy helps in precise positioning and patient monitoring which aims at more accurate treatment delivery.

Legal entity responsible for the study

Institutional Review Board.

Funding

Has not received any funding.

Disclosure

The author has declared no conflicts of interest.

Background

Biosimilars have the potential for cost savings and thus help increase access to treatment. The study aims to understand the physicians’ perception and usage of biosimilars in HER2+ breast cancer (BC) and changes in trends from pre-biosimilar (only innovator trastuzumab was available) to current biosimilar era in India.

Methods

This virtual qualitative survey was conducted at private and government hospital settings in India between April and June 2022. Medical oncologists (MO) with experience ≥10 years, and with BC patient load ≥10 per month were included.

Results

Thirty MOs (24 private and 6 government) across 10 cities participated in the survey. The survey reported the following: All BC patients undergo HER2+ screening of which 29% test positive; 29% of patients are diagnosed at metastatic stage (MBC); 80% of patients diagnosed in non-metastatic (NMBC) stage undergoes surgery, of which 70% receive neo-adjuvant therapy. In the pre-biosimilar era, up to 30% of the prescribed patients were able to access the trastuzumab treatment. While in the current biosimilar era, access to trastuzumab in NMBC itself has increased to ≥75%, with biosimilars accounting for 84% of the total usage of trastuzumab. Currently, trastuzumab is usually taken for an average of 12 cycles in NMBC and 6 cycles in MBC as 3 weekly regimens. In patients with financial constraints, a majority (70%) of MOs offer a modified treatment plan for trastuzumab, reducing the number of cycles to 6-8 vs standard 17. MOs generally do not prefer to switch patients between different trastuzumab brands, however, the average switch from an innovator to a biosimilar is 14%. Only 16% of patients receive pertuzumab with the major limitation being a financial constraint, due to the non-availability of a biosimilar.

Conclusions

The access to trastuzumab therapy in HER2+ BC patients has increased many folds in the current biosimilar era in India, leading to reduced financial burden and improved overall healthcare.

Legal entity responsible for the study

Viatris and IPSOS Research Pvt Ltd.

Funding

Viatris.

Disclosure

N. Ramanjinappa, J. Agarwal, K.H. Upveja: Financial Interests, Personal and Institutional, Full or part-time Employment: Viatris.

Background

There are sparse data on outcomes with combined chemotherapy (CT) and hormonal therapy (HT) in MBC patients, including those with multiple prior treatments.

Methods

This was a retrospective analysis of HR-positive, HER2-negative MBC patients who were treated with a combination of CT (capecitabine or oral cyclophosphamide) and HT (tamoxifen or aromatase inhibitor or fulvestrant or megestrol acetate) between Jan 2015 and Dec 2020, with data cutoff in May 2022. Outcomes were progression-free survival (PFS) and overall survival (OS) from the start of CHT, and toxicity.

Results

A total of 224 patients with median age of 53 (26-91) years and median 3 (0-12) prior treatment lines were included. At median follow-up of 21.2 (1.7-87.0) months, 195 (87.1%) patients had experienced progression and 154 (68.8%) had died, with median PFS 8.8 (95% CI 7.0-10.6) months and median OS 16.7 (95% CI 13.5-19.9) months. In univariable analyses, ECOG PS [(</=1, n=82, 36.6%) vs (>/=2, n=142, 63.4%)] was significantly associated with PFS (14.9m vs 6.0m, HR 0.32, 95% CI 0.23-0.44, p<0.001) and OS (36.2m vs 11.0m, HR 0.29, 95% CI 0.20-0.42, p<0.001), duration of most recent HT [(>12 m, n=74, 33.0%) vs (</=12 m, n=124, 55.4%)] was significantly associated with PFS (11.7m vs 6.9m, HR 0.32, 95% CI 0.23-0.44, p=0.023) and OS (21.9m vs 15.0m, HR 0.70, 95% CI 0.49-0.99, p=0.047), prior treatment lines [(</=3, n=143, 63.8%) vs (>/=4, n=81, 36.2%)] was not significantly associated with PFS and OS, visceral metastasis [(no, n=57, 25.4%) vs (yes, n=167, 74.6%)] was not significantly associated with PFS and OS, and CHT type [(capecitabine, n=196, 87.5%) vs (cyclophosphamide, n=26, 11.6%)] was not significantly associated with PFS and OS. In multivariable Cox analyses, better PS and fewer prior lines were significantly associated with higher PFS while better ECOG PS and capecitabine-based CHT were significantly associated with higher OS. CHT was well tolerated with 21 (9.4%) patients experiencing any grade>/=3 toxicity.

Conclusions

Chemo-hormonal therapy is an effective treatment in heavily pre-treated MBC patients, including those with visceral metastases.

Legal entity responsible for the study

The authors.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Lapatinib at an approved dose of 1250 to 1500 mg/day in Her2 positive metastatic breast cancer contributes to a high pill burden and financial toxicity. Hence the concept of “value meal” has been suggested, i.e. increasing the bioavailability by giving lower doses of lapatinib with meal, as suggested by phase I studies.

Methods

In our institution, for patients with Her2 positive metastatic breast cancer who cannot afford the approved dose of lapatinib or other Her2 agents, low dose or metronomic lapatinib, 500mg daily with meal, is offered. We conducted a retrospective review of the safety and efficacy of this regimen, including data from January 1, 2014 to December 31, 2020.

Results

A total of 47 females were enrolled. The majority had de novo metastatic disease (n=27,57.4%) and multiple visceral metastases (n=23,48.9%). The median number of lines of treatment before lapatinib was one. Lapatinib was given in combination, but 4 patients received it as a single agent. The disease control rate with lapatinib was 61.7%, out of whom 4.3% (n=2) experienced complete remission. The median progression free survival was 7 months (95% CI: 5.6 - 8.4 months), with a median overall survival of 12.5 months (95% CI: 9.7 - 15.3 months). The median duration of response was 4.5 months, ranging from 1.3 months to 45.8 months. The low-dose lapatinib was well-tolerated. Only eleven patients (23.4%) experienced some toxicity, with grade 3 in only one (2.1%) and no grade 4 toxicities. The most common toxicities were skin rashes and hand-foot syndrome. Dose interruptions were necessary in 3 (6.4%) patients, while drug was discontinued in one (2.1%).

Conclusions

The disease control rate and the progression free survival with low dose lapatinib is equivalent to the standard dose in published literature. The overall survival is short here as many did not receive any further anti-Her2 agents in view of financial toxicity. As a well tolerated regimen with less financial burden, this is an approach that needs further validation in larger randomised controlled trials.

Legal entity responsible for the study

Malabar Cancer Centre.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Breast cancer is the second most common cancer worldwide. Some studies have begun investigation of Absolute Lymphocyte Count (ALC) to predict prognosis of breast metastasis. This review aims to analyze use of ALC to predict overall survival (OS) in breast metastasis patients.

Methods

A total of 73,233 literatures were found through PubMed, PMC, Science Direct, and Google Scholar using a combination of keywords including ALC, breast metastasis, and OS. Publications included are limited to English manuscripts published within the last ten years. We excluded publications with insufficient datas and non-breast metastatic studies. Studies were evaluated by all four authors using the Newcastle-Ottawa Scale (NOS), JADAD scale, funnel plot to assess publication bias, and Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) to assess quality of this review.

Results

A total of 11 studies consisting 2,433 patients were included in this review. Nine retrospective studies were of good quality using NOS, two clinical trial studies were of low quality using JADAD scale. GRADE assessment revealed this review to be moderate in quality and publication biases are minimal. Most studies revealed an association between low ALC or lymphopenia and worse prognosis, while 1 study showed otherwise. A pooled analysis of hazard ratio (HR) from log-rank test (HR = 0.57; 95% CI, 0.46-0.71; P = 0.52), univariate cox regression analysis (HR = 0.69; 95%CI, 0.61-0.78; P = 0.01) and multivariate cox regression analysis (HR = 0.70; 95%CI, 0.60-0.81; P = 0.30) showed minimal correlation.

Conclusions

While most studies used in this review showed strong correlation between low ALC and OS with statistically significant results, pooled analysis from this review revealed only the univariate cox regression analysis of HR showed statistically significant results. We conclude that lymphopenia may affect OS in patients with metastatic breast cancer. Further studies are required, especially high quality clinical trials.

Legal entity responsible for the study

The authors.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.

Background

Imaging-based assessment for tumor progression in bone metastasis is difficult because of the nature of fixed bony defects and their complexity. The aime of this study was to investigate the circulating osteocalcin-positivecells (cOC) in the peripheral blood as a biomarker for monitoring the therapeutic effects on bone metastasis.

Methods

An animal model of bone metastasis was established by intratibial injection of MDA-MB-231-luc cells into nude mice and treated with zolendronate (ZA) for 4 weeks. cOC was characterized as CD15^-CD45^-Osteocalcin⁺ cell fractions from the peripheral blood mononuclear cells using a flow cytometry. Breast cancer patients treated with ZA for bone metastasis were recruited, and cOC was measured.

Results

In animal model, ZA significantly reduced % tumor area and TRAP+ osteoclasts in histological analysis and tumor growth in bioluminescence images from 2-3 weeks after treatment. Serum TRAP5b or C-terminal telopeptide (CTx) levels were significantly decreased from 2 weeks in ZA-treated group. Earlier than these, cOC in peripheral blood was significantly decreased from 1 weeks after treatment. In patients with bone metastasis, the decrease in cOC was significantly frequent in the ZA-responsive group than in the ZA-nonresponsive group, whereas the serum CTx levels did not differ between the groups.

Conclusions

cOC can predict therapeutic response in bone metastasis of breast cancer.

Legal entity responsible for the study

Sun Wook Cho.

Funding

Cellus, Inc.

Disclosure

M.J. Kim: Financial Interests, Personal, Full or part-time Employment: Cellus, Inc. S.W. Cho: Financial Interests, Personal, Officer: Cellus, Inc. All other authors have declared no conflicts of interest.

Background

The objective of this study was to assess the effect of an educational activity on the knowledge of oncologists regarding the rationale for using next-generation antibody drug conjugates (ADCs) for the treatment of breast cancer and the latest data from clinical trials.

Methods

This CME activity consists of a 2000-word article presenting expert insights in an interview style. Effectiveness of this education was assessed with a repeated-pairs pre-/post-assessment study design using 3 knowledge questions and 1 confidence question, in which each individual served as their own control. A paired sample t-test assessed mean differences in average number of correct responses pre- to post-assessment, and a McNemar’s test assessed significant improvement at question level. P values <.05 are statistically significant. The activity launched on January 27, 2022, with data collected through to April 1, 2022.

Results

By June 22, 2022 this activity had reached 1,019 physicians of whom 375 were oncology specialists (41% from Europe, 22% from Asia, 21% from Middle East and North Africa, and 13% from Latin America). Sixty-seven oncologists who completed all assessment questions were included in the study analysis. The average knowledge of oncologists increased significantly from 45% pre to 77% post-education (P < .001) as assessed by correct responses to the knowledge questions. There was a significant improvement in the knowledge of the following learning objectives as shown in the table below. Table: 27P

There was a measurable improvement in confidence in around 55% of oncologists regarding understanding of the treatment implications of the latest clinical trial data, and a 71% average confidence shift for those whose confidence had improved.

Conclusions

This study showed a positive educational impact of a 2000-word article presenting expert insights in an interview style, on the latest developments regarding emerging ADCs for the treatment of breast cancer. Further education is needed to update physicians on new data and guideline recommendations as these evolve in the future

Legal entity responsible for the study

Medscape.

Funding

The educational activity was supported by an independent educational grant from Daiichi Sankyo.

Disclosure

N. Dorkhom, D. Grainger, Y. Ren: Financial interests, Personal, Full or part-time Employment: Medscape.

Background

A target-specific mAb for the HER2 receptor is trastuzumab, which is used for treatment of breast cancer. This mAb coupled with radionuclides β or α particle emitters are used for radioimmunotherapy. However, radioimmunotherapy with the whole mAb is still limited due to slow pharmacokinetics. This leads to the slow elimination of trastuzumab from the blood and low tumor to blood and tumor to background ratios. To limit this, a small molecule Fab was introduced but the role in HER2 detection by immuno-PET imaging is unknown.

Methods

Fab has been generated by papain digestion and conjugation with NOTA. The characterization of Fab and NOTA Fab were performed by the SDS-PAGE and mass spectrometry technique MALDI-TOF. In vitro stability and target specificity for HER2/neu receptor were performed in plasma and receptor binding with bio-layer interferometry technique. After standardization of radiolabelling with 68GaCl3 the in vivo target efficiency was performed in HER2.positive breast cancer patients, which compared with 18F-FDG and corelated with immunohistochemistry.

Results

Fab generation was standardized at molar ratio 140:1 wt/wt of trastuzumab and papain, temp 37°C, stirrer at 850 rpm, time 20-24h.The purity were found >99% on SDS-PAGE (band 43 and 29 kDa) and MALDI-TOF (45 kDa). The conjugation was done at molar ratio of 1:25 and 1:30. The radiolabelling yield 48-70% were standardized at parameters noted pH 4.5-5, temp 37-40°C, incubation time 15 min for Rxn Vol 2.5mL. The trastuzumab affinity for HER2 were found in the picomolar range (Kon and Koff 3.79 E+05M-1s-1 and <1.0E-7s-1), Fab 1.9nM (Kon and Koff 3.9 E+05M-1s-1 and 4.77E-4s-1) and NOTA Fab 20 nM (Kon Koff 7.4 E+04M-1s-1 and 1.6E-3s-1). The stability of the radiopharmaceutical was found to be more than 4h and sterile up to 14 days. The endotoxin limit was found 5-10 EU/mL. In vivo targeting potential was checked in HER2-positive patients with uptake at primary lesion (SUV)max 3-5.8 and at lymph node 2.0-3.1, while liver and kidney also showed high uptake.

Conclusions

The 68Ga trastuzumab Fab can be formulated with high radiochemical purity and demonstrates high receptor affinity for HER2/neu receptor. The 68Ga trastuzumab Fab can be used as a diagnostic tool in immunoPET imaging for the HER2/neu receptor.

Legal entity responsible for the study

The authors.

Funding

Board of Research in Nuclear Sciences (BRNS) and University Grants Commission (UGC).

Disclosure

All authors have declared no conflicts of interest.

Background

Although CXC chemokine receptor 4 (CXCR4) blockades have facilitated antitumor in many cancer types, the mechanisms of CXCR4 blockades underlying antitumor activity, especially in immunity, remain to be determined.

Methods

Single-cell transcriptome data of multi-tissues and T cells of multi-cancers were analyzed to identify the expression patterns of CXCR4 in exhausted CD8⁺ T cells. In addition, in vitro and in vivo assays were used to explore the effects of CXCR4 blockades in promoting anti-PD-1 immunotherapy via JAK2/STAT3/TOX axis of exhausted CD8⁺ T cells.

Results

Single-cell transcriptome analysis of multi-tissues and multi-cancers demonstrated that CXCR4 was a specific marker of T cells, and it was highly expressed in PD-1⁺ exhausted CD8⁺ T cells. By targeting CXCR4, the expressions of TOX and PD-1 was down-regulated, with the upregulation of p-JAK2/p-STAT3 in T cells. Importantly, TOX-mediated CD8⁺ T cell activation of CXCR4 blockades was reversed via the p-JAK2/p-STAT3 inhibitor. In vivo, single-cell transcriptome analysis revealed CXCR4 blockades promotes anti-PD-1 immunotherapy of cervical cancer, breast cancer, and ovarian cancer via activating exhausted CD8⁺ T cells.

Conclusions

Overall, we found that targeting CXCR4 promotes antitumor immunity through TOX-mediated CD8⁺ T cell activation, which paves the way toward immunotherapy in antitumor activity.

Legal entity responsible for the study

The authors.

Funding

National Key R&D Program of China (2021YFC2701201), National Natural Science Foundation of China (82072895 and 82141106), Shenzhen Science and Technology Innovation Committee (JCYJ20210324105808022, RCBS20210706092345027), China Postdoctoral Science Foundation (2021M702223).

Disclosure

All authors have declared no conflicts of interest.

Background

While the currently available KRAS^G12C-selective inhibitors including Sotorasib have shown promising efficacy, acquired resistance eventually occurred in most cancer patients following treatment. Blocking SHP2, one of critical upstream nodes, represents a rationalized strategy to overcome resistance. We have previously developed JAB-21822, a selective covalent KRAS^G12C inhibitor and JAB-3312, a selective allosteric SHP2 inhibitor, both of which have entered multiple clinical trials.

Methods

Cell growth inhibitory effect of JAB-21822 was evaluated in various human cancer cell lines, as well as Ba/F3 cells expressing different KRAS^G12C mutants with or without secondary point mutations that contribute to KRAS^G12C inhibitor resistance. Tumor growth inhibitory effect of JAB-21822 was also evaluated in cell line- and patient-derived xenografts. Using similar approaches, the effect of JAB-21822 in combination with JAB-3312 was evaluated in KRAS^G12C inhibitor-resistant tumor cells and xenograft models. Furthermore, RNA-seq was performed to identify genes with altered expression in KRAS^12C inhibitor-resistant NCI-H358 cells compared to their parental counterparts. Expression of selected candidate genes were further confirmed by real-time PCR.

Results

As a single agent, JAB-21822 showed early potent antitumor effect both in vitro and in vivo. RNA-seq further identified potential genes involved in KRAS^G12C inhibitor resistance. Significantly, combination of JAB-21822 with JAB-3312 exerted synergistic effect in the KRAS^G12C inhibitor-resistant tumor cells and xenograft models tested.

Conclusions

JAB-21822 is a promising KRAS^G12C-targeting drug and, when combined with JAB-3312, can overcome adaptive resistance to KRAS^G12C inhibition. These preclinical data have provided rationale for our clinical trial featuring the two drugs combination in treating KRAS^G12C inhibitor-resistant cancer patients.

Legal entity responsible for the study

Jacobio Pharmaceuticals Co., Ltd.

Funding

AbbVie.

Disclosure

P. Wang, Q. Zheng, D. Kang, X. Sun, S. Zhu, Y. Wang, W. Long, Y. Lin : Financial interests, Personal, Full or part-time Employment: Jacobio Pharmaceuticals Co.

Background

Pancreatic cancer is the fourth most dominant disease leading the high mortality with a survival rate of 10%. Conventional therapeutics cause adverse effects as well as reoccurrence of pancreatic malignancy. Therefore, combination therapeutics including chemotherapy, photothermal, photodynamic, and immunotherapeutics has received great attention. The nanotechnological approaches are applied to develop the multifunctional nanocarrier to efficiently target the cancer cells without causing noticeable side effects. Therefore, herein we developed the multi-stimuli (pH, photothermal, and cancer cell receptor) responsive drug delivery system (DDS) for synergetic photothermal ablation of pancreatic cancer cells.

Methods

We synthesized the Au/Pd nanoparticles and conjugated them with 5-Fu using the CTAB. The 5fu-Au/Pd were loaded in CS through TPP ionic gelation. Then, apt was functionalized with 5fu-Au/Pd CS NPs by NHS and EDS covalent bonding method. The DDS (Apt-5fu-Au/Pd-CSNPs) were characterized for morphology, size, drug loading, entrapment, stimuli-responsive drug release, and photothermal conversion efficiency. Further, anticancer activity was evaluated in various molecular level cytotoxicity assays and animal tumor model experiments.

Results

DDS exhibited high drug loading and entrapment efficiency, and its size were exhibited <200 nm with a PDI of 0.23. The higher release of 5-Fu and Au/Pd were observed at the pH 5.4. This DDS exhibited the photothermal conversion efficiency of 28 %. The Apt-5fu-Au/Pd-CSNPs result in higher anticancer activity evidenced by the in-vitro cytotoxicity and animal experiments. The in vitro studies revealed that the treatment of Apt-5fu-Au/Pd-CSNPs triggers the cytotoxicity in the PANC-1 cells by regulating the MMP, ROS, apoptotic, and pro-apoptotic related proteins. The in-vivo studies reveal that the treatment of these DDS inhibited tumor proliferation and increase the mice's survival rate. And histopathological H & E staining studies confirmed the non-toxicity.

Conclusions

The present work delivered the novel nano DDS (Apt-5fu-Au/Pd-CSNPs) as an alternative to the conventional method of pancreatic cancer therapy.

Legal entity responsible for the study

The authors.

Funding

Has not received any funding.

Disclosure

All authors have declared no conflicts of interest.