Nuclear pCHK1 as a potential biomarker of increased sensitivity to ATR inhibition.

Excessive genomic instability coupled with abnormalities in DNA repair pathways induces high levels of 'replication stress' when cancer cells propagate. Rather than hampering cancer cell proliferation, novel treatment strategies are turning their attention towards targeting cell cycle checkpoint kinases (such as ATR, CHK1, WEE1, and others) along the DNA damage response and replicative stress response pathways, thereby allowing unrepaired DNA damage to be carried forward towards mitotic catastrophe and apoptosis. The selective ATR kinase inhibitor elimusertib (BAY 1895344) has demonstrated preclinical and clinical monotherapy activity; however, reliable predictive biomarkers of treatment benefit are still lacking. In this study, using gene expression profiling of 24 cell lines from different cancer types and in a panel of ovarian cancer cell lines, we found that nuclear-specific enrichment of checkpoint kinase 1 (CHK1) correlated with increased sensitivity to elimusertib. Using an advanced multispectral imaging system in subsequent cell line-derived xenograft specimens, we showed a trend between nuclear phosphorylated CHK1 (pCHK1) staining and increased sensitivity to the ATR inhibitor elimusertib, indicating the potential value of pCHK1 expression as a predictive biomarker of ATR inhibitor sensitivity. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Decoding transcriptomic intra-tumour heterogeneity to guide personalised medicine in ovarian cancer.

The evaluation of intra-tumour heterogeneity (ITH) from a transcriptomic point of view is limited. Single-cell cancer studies reveal significant genomic and transcriptomic ITH within a tumour and it is no longer adequate to employ single-subtype assignment as this does not acknowledge the ITH that exists. Molecular assessment of subtype heterogeneity (MASH) was developed to comprehensively report on the composition of all transcriptomic subtypes within a tumour lesion. Using MASH on 3431 ovarian cancer samples, correlation and association analyses with survival, metastasis and clinical outcomes were performed to assess the impact of subtype composition as a surrogate for ITH. The association was validated on two independent cohorts. We identified that 30% of ovarian tumours consist of two or more subtypes. When biological features of the subtype constituents were examined, we identified significant impact on clinical outcomes with the presence of poor prognostic subtypes (Mes or Stem-A). Poorer outcomes correlated with having higher degrees of poor prognostic subtype populations within the tumour. Subtype prediction in several independent datasets reflected a similar prognostic trend. In addition, paired analysis of primary and recurrent/metastatic tumours demonstrated Mes and/or Stem-A subtypes predominated in recurrent and metastatic tumours regardless of the original primary subtype. Given the biological and prognostic value in delineating individual subtypes within a tumour, a clinically applicable MASH assay using NanoString® technology was developed as a classification tool to comprehensively describe constituents of molecular subtypes. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

The effect of inhibition of receptor tyrosine kinase AXL on DNA damage response in ovarian cancer.

AXL is a receptor tyrosine kinase that is often overexpressed in cancers. It contributes to pathophysiology in cancer progression and therapeutic resistance, making it an emerging therapeutic target. The first-in-class AXL inhibitor bemcentinib (R428/BGB324) has been granted fast track designation by the U.S. Food and Drug Administration (FDA) in STK11-mutated advanced metastatic non-small cell lung cancer and was also reported to show selective sensitivity towards ovarian cancers (OC) with a Mesenchymal molecular subtype. In this study, we further explored AXL's role in mediating DNA damage responses by using OC as a disease model. AXL inhibition using R428 resulted in the increase of DNA damage with the concurrent upregulation of DNA damage response signalling molecules. Furthermore, AXL inhibition rendered cells more sensitive to the inhibition of ATR, a crucial mediator for replication stress. Combinatory use of AXL and ATR inhibitors showed additive effects in OC. Through SILAC co-immunoprecipitation mass spectrometry, we identified a novel binding partner of AXL, SAM68, whose loss in OC cells harboured phenotypes in DNA damage responses similar to AXL inhibition. In addition, AXL- and SAM68-deficiency or R428 treatment induced elevated levels of cholesterol and upregulated genes in the cholesterol biosynthesis pathway. There might be a protective role of cholesterol in shielding cancer cells against DNA damage induced by AXL inhibition or SMA68 deficiency.

GAGE mediates radio resistance in cervical cancers via the regulation of chromatin accessibility.

Radiotherapy (RT) resistance is a major cause of treatment failure in cancers that use definitive RT as their primary treatment modality. This study identifies the cancer/testis (CT) antigen G antigen (GAGE) as a mediator of radio resistance in cervical cancers. Elevated GAGE expression positively associates with de novo RT resistance in clinical samples. GAGE, specifically the GAGE12 protein variant, confers RT resistance through synemin-dependent chromatin localization, promoting the association of histone deacetylase 1/2 (HDAC1/2) to its inhibitor actin. This cumulates to elevated histone 3 lysine 56 acetylation (H3K56Ac) levels, increased chromatin accessibility, and improved DNA repair efficiency. Molecular or pharmacological disruption of the GAGE-associated complex restores radiosensitivity. Molecularly, this study demonstrates the role of GAGE in the regulation of chromatin dynamics. Clinically, this study puts forward the utility of GAGE as a pre-screening biomarker to identify poor responders at initial diagnosis and the therapeutic potential of agents that target GAGE and its associated complex in combination with radiotherapy to improve outcomes.

Recommendations to improve the clinical adoption of NGS-based cancer diagnostics in Singapore.

Next-generation sequencing (NGS)-based diagnostics have demonstrated clinical utility in predicting improved survival benefits with targeted treatment in certain cancer types, and positive cost-benefit in several healthcare systems. However, clinical adoption in Singapore remains low despite commercial availability of these diagnostics. This expert opinion review examines the key challenges to the clinical adoption of NGS-based diagnostics in Singapore, provides recommendations on impactful initiatives to improve adoption, and also offers practical guidance on specific cancer types in which NGS-based diagnostics are appropriate for use in Singapore. Limited patient affordability is one major challenge to clinical adoption of NGS-based diagnostics, which could be improved by enabling patient access to more funds for specific cancer types with clear benefits. Expert opinion based on current evidence and clinical experience supports the upfront use of hotspot panels in advanced non-small cell lung cancer (NSCLC), metastatic colorectal cancer, advanced and recurrent ovarian cancer, and acute myeloid leukemia. Comprehensive genomic profiling could be considered for upfront use in select patients with NSCLC and ovarian cancer, or in refractory patients with the four cancer types. Wider adoption of NGS-based diagnostics will improve the delivery of cancer care in Singapore and Asia-Pacific, and thus lead to better patient outcomes.

Analysis of gene expression signatures identifies prognostic and functionally distinct ovarian clear cell carcinoma subtypes.

BACKGROUND: Ovarian clear cell carcinoma (OCCC) is a histological subtype of epithelial ovarian cancer (EOC) with distinct pathological, biological, and molecular features. OCCCs are more resistant to conventional treatment regimen of EOC and have the worst stage-adjusted prognosis amongst EOC subtypes. As the OCCC incidence rate in Asian populations has significantly increased in recent decades, it is critical to elucidate its molecular features that could lead to OCCC-tailored therapeutic strategies. METHODS: Gene expression profiles of 222 OCCC were analyzed by hierarchical clustering and statistical analyses. FINDINGS: We identified two OCCC gene expression subtypes: EpiCC-epithelial-like, which is associated with early-stage disease, with a relatively higher rate of gene mutations in the SWI/SNF complex; and MesCC-mesenchymal-like, associated with late-stage and higher enrichment of immune-related pathway activity. Genetic, copy number and transcriptomic analyses showed that both EpiCC and MesCC carried OCCC-associated aberrations. The EpiCC/MesCC classification was reproducible in validation cohorts and OCCC cell lines. MesCC tumors had a poorer progression-free survival (PFS) than EpiCC tumors (HR: 3·0, p = 0·0006). Functional assays in cell lines showed that the MesCC subtype was more proliferative and more anoikis-resistant than the EpiCC. By applying the EpiCC/MesCC classification to the TCGA renal clear cell carcinoma cohort, our results indicated interoperability of the subtyping scheme, and revealed preferential drug response of MesCC to bevacizumab. INTERPRETATION: The EpiCC/MesCC classification shows promise for prognostic and therapeutic stratification in OCCC patients and warrants further investigation in the context of OCCC gene expression subtype-tailored treatment strategies.

Update on immune checkpoint inhibitors in gynecological cancers.

In recent years, progress in our understanding of immune-modulatory signaling pathways in immune cells and the tumor microenvironment (TME) has led to rejuvenated interest in cancer immunotherapy. In particular, immunotherapy targeting the immune checkpoint receptors such as cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4), programmed cell-death 1 (PD-1), and programmed cell-death ligand 1 (PD-L1) have demonstrated clinical activity in a wide variety of tumors, including gynecological cancers. This review will focus on the emerging clinical data on the therapeutic role of immune checkpoint inhibitors, and potential strategies to enhance the efficacy of this class of compounds, in the context of gynecological cancers. It is anticipated that future biomarker-directed clinical trials will provide further insights into the mechanisms underlying response and resistance to immunotherapy, and help guide our approach to designing therapeutic combinations that have the potential to enhance the benefit of immunotherapy in patients with gynecologic cancers.

Targeting the AXL signaling pathway in ovarian cancer.

In a recent publication in Science Signaling, we showed that a Mes molecular subtype of epithelial ovarian cancer (EOC) harboring epithelial-mesenchymal transition (EMT) features has a unique signaling network downstream of the GAS6/AXL pathway. Our finding leads to a potential strategy for treating the Mes subtype of EOC by targeting AXL.

Limb Hypothermia for Preventing Paclitaxel-Induced Peripheral Neuropathy in Breast Cancer Patients: A Pilot Study.

BACKGROUND: Peripheral neuropathy (PN) due to paclitaxel is a common dose-limiting toxicity with no effective prevention or treatment. We hypothesize that continuous-flow limb hypothermia can reduce paclitaxel-induced PN. PATIENTS AND METHODS: An internally controlled pilot trial was conducted to investigate the neuroprotective effect of continuous-flow limb hypothermia in breast cancer patients receiving weekly paclitaxel. Patients underwent limb hypothermia of one limb for a duration of 3 h with every paclitaxel infusion, with the contralateral limb used as control. PN was primarily assessed using nerve conduction studies (NCSs) before the start of chemotherapy, and after 1, 3, and 6 months. Skin temperature and tolerability to hypothermia were monitored using validated scores. RESULTS: Twenty patients underwent a total of 218 cycles of continuous-flow limb hypothermia at a coolant temperature of 22°C. Continuous-flow limb hypothermia achieved mean skin temperature reduction of 1.5 ± 0.7°C and was well tolerated, with no premature termination of cooling due to intolerance. Grade 3 PN occurred in 2 patients (10%), grade 2 in 2 (10%), and grade 1 in 12 (60%). Significant correlation was observed between amount of skin cooling and motor nerve amplitude preservation at 6 months (p < 0.0005). Sensory velocity and amplitude in the cooled limbs were less preserved than in the control limbs, but the difference did not attain statistical significance. One patient with a history of diabetes mellitus had significant preservation of compound muscle action potential in the cooled limb on NCS analysis. CONCLUSION: This study suggests that continuous limb hypothermia accompanying paclitaxel infusion may reduce paclitaxel-induced PN and have therapeutic potential in select patients and warrants further investigation. The method is safe and well tolerated.

Low Levels of NDRG1 in Nerve Tissue Are Predictive of Severe Paclitaxel-Induced Neuropathy.

INTRODUCTION: Sensory peripheral neuropathy caused by paclitaxel is a common and dose limiting toxicity, for which there are currently no validated predictive biomarkers. We investigated the relationship between the Charcot-Marie-Tooth protein NDRG1 and paclitaxel-induced neuropathy. METHODS/MATERIALS: Archived mammary tissue specimen blocks of breast cancer patients who received weekly paclitaxel in a single centre were retrieved and NDRG1 immunohistochemistry was performed on normal nerve tissue found within the sample. The mean nerve NDRG1 score was defined by an algorithm based on intensity of staining and percentage of stained nerve bundles. NDRG1 scores were correlated with paclitaxel induced neuropathy. RESULTS: 111 patients were studied. 17 of 111 (15%) developed severe paclitaxel-induced neuropathy. The mean nerve NDRG1 expression score was 5.4 in patients with severe neuropathy versus 7.7 in those without severe neuropathy (p = 0.0019). A Receiver operating characteristic (ROC) curve analysis of the mean nerve NDRG1 score revealed an area under the curve of 0.74 (p = 0.0013) for the identification of severe neuropathy, with a score of 7 being most discriminative. 13/54 (24%) subjects with an NDRG1 score < = 7 developed severe neuropathy, compared to only 4/57 (7%) in those with a score >7 (p = 0.017). CONCLUSION: Low NDRG1 expression in nerve tissue present within samples of surgical resection may identify subjects at risk for severe paclitaxel-induced neuropathy. Since nerve biopsies are not routinely feasible for patients undergoing chemotherapy for early breast cancer, this promising biomarker strategy is compatible with current clinical workflow.

CSIOVDB: a microarray gene expression database of epithelial ovarian cancer subtype.

Databases pertaining to various diseases provide valuable resources on particular genes of interest but lack the molecular subtype and epithelial-mesenchymal transition status. CSIOVDB is a transcriptomic microarray database of 3,431 human ovarian cancers, including carcinoma of the ovary, fallopian tube, and peritoneum, and metastasis to the ovary. The database also comprises stroma and ovarian surface epithelium from normal ovary tissue, as well as over 400 early-stage ovarian cancers. This unique database presents the molecular subtype and epithelial-mesenchymal transition status for each ovarian cancer sample, with major ovarian cancer histologies (clear cell, endometrioid, mucinous, low-grade serous, serous) represented. Clinico-pathological parameters available include tumor grade, surgical debulking status, clinical response and age. The database has 1,868 and 1,516 samples with information pertaining to overall and disease-free survival rates, respectively. The database also provides integration with the copy number, DNA methylation and mutation data from TCGA. CSIOVDB seeks to provide a resource for biomarker and therapeutic target exploration for ovarian cancer research.