Evaluation of ABT-751, a novel anti-mitotic agent able to overcome multi-drug resistance, in melanoma cells.

PURPOSE: Drug efflux transporter associated multi-drug resistance (MDR) is a potential limitation in the use of taxane chemotherapies for the treatment of metastatic melanoma. ABT-751 is an orally bioavailable microtubule-binding agent capable of overcoming MDR and proposed as an alternative to taxane-based therapies. METHODS: This study compares ABT-751 to taxanes in vitro, utilizing seven melanoma cell line models, publicly available gene expression and drug sensitivity databases, a lung cancer cell line model of MDR drug efflux transporter overexpression (DLKP-A), and drug efflux transporter ATPase assays. RESULTS: Melanoma cell lines exhibit a low but variable protein and RNA expression of drug efflux transporters P-gp, BCRP, and MDR3. Expression of P-gp and MDR3 correlates with sensitivity to taxanes, but not to ABT-751. The anti-proliferative IC50 profile of ABT-751 was higher than the taxanes docetaxel and paclitaxel in the melanoma cell line panel, but fell within clinically achievable parameters. ABT-751 IC50 was not impacted by P-gp-overexpression in DKLP-A cells, which display strong resistance to the P-gp substrate taxanes compared to DLKP parental controls. The addition of ABT-751 to paclitaxel treatment significantly decreased cell proliferation, suggesting some reversal of MDR. ATPase activity assays suggest that ABT-751 is a potential BCRP substrate, with the ability to inhibit P-gp ATPase activity. CONCLUSION: Our study confirms that ABT-751 is active against melanoma cell lines and models of MDR at physiologically relevant concentrations, it inhibits P-gp ATPase activity, and it may be a BCRP and/or MDR3 substrate. ABT-751 warrants further investigation alone or in tandem with other drug efflux transporter inhibitors for hard-to-treat MDR melanoma.

The prognostic value of the derived neutrophil-to-lymphocyte ratio (dNLR) in patients treated with immune checkpoint inhibitors.

BACKGROUND: The (derived) neutrophil-to-lymphocyte ratio (dNLR) is a potential predictive biomarker in the era of checkpoint inhibitors (CPI). An elevated dNLR is associated with worse outcomes across several malignancies. However, there is no clearly defined cut-off in the clinical setting. AIM: To compare outcomes in patients prescribed CPI with a baseline dNLR0 > 3 and dNLR0 ≤ 3. The dNLR6 was measured 6 weeks later to determine its impact on patient overall survival (OS). METHODS: Prospectively maintained pharmacy databases in a regional cancer centre were interrogated for patients who were prescribed CPI in the advanced setting between January 2017 and May 2020. RESULTS: There were 121 patients with advanced cancer and a median age of 68 (range 30 to 88) years. Forty-four percent (n = 53) received prior systemic therapy. Patients with an initial dNLR0 > 3 when compared with a dNLR0 ≤ 3 had significantly shorter median progression-free survival (PFS), 3 vs. 14 months (p = 0.001) and median OS, 6.4 vs. 30.2 months (p = 0.001). Patients with an initial dNLR0 > 3 and increased dNLR at 6 weeks (dNLR6) had significantly reduced median PFS (3.5 vs. 14.7 months, p = 0.03) and OS (5.7 vs. 16.3, p = 0.03) when compared with those whose dNLR decreased. In the dNLR0 ≤ 3 cohort, any increased dNLR when compared with decreased dNLR after 6 weeks of CPI had significantly reduced PFS (8.4 months vs. NR, p = 0.01) and OS (24.2 months vs. NR, p = 0.02). CONCLUSIONS: Lower pre-CPI treatment dNLR is associated with improved OS. A decrease in dNLR during treatment confers improved OS.

Clinical and Genomic Characterization of Bladder Carcinomas With Glandular Phenotype.

PURPOSE: To compare oncologic outcomes and genomic alteration profiles in patients with bladder and urachal adenocarcinoma, urothelial carcinoma (UC) with glandular differentiation, and UC, not otherwise specified (NOS) undergoing surgical resection, with emphasis on response to systemic therapy. METHODS: We identified patients with bladder cancer with glandular variants who underwent surgical resection at Memorial Sloan Kettering from 1995 to 2018 (surgical cohort) and/or patients who had tumor sequencing using a targeted next-generation sequencing platform (genomics cohort). Pathologic complete and partial response rates to neoadjuvant chemotherapy (NAC) and recurrence-free and cancer-specific survival were measured. Alteration frequencies between histologic subtypes were compared. RESULTS: Thirty-seven patients with bladder adenocarcinoma, 46 with urachal adenocarcinoma, 84 with UC with glandular differentiation, and 1,049 with UC, NOS comprised the surgical cohort. Despite more advanced disease in patients with bladder and urachal adenocarcinoma, no significant differences in recurrence or cancer-specific survival by histology were observed after adjusting for stage. In patients with UC with glandular differentiation, NAC resulted in partial (≤ pT1N0) and complete (pT0N0) responses in 28% and 17%, respectively. Bladder and urachal adenocarcinoma genomic profiles resembled colorectal adenocarcinoma with frequent TP53, KRAS, and PIK3CA alterations while the genomic profile of UC with glandular differentiation more closely resembled UC, NOS. Limitations include retrospective nature of analysis and small numbers of nonurothelial histology specimens. CONCLUSION: The genomic profile of bladder adenocarcinomas resembled colorectal adenocarcinomas, whereas UC with glandular differentiation more closely resembled UC, NOS. Differences in outcomes among patients with glandular bladder cancer variants undergoing surgical resection were largely driven by differences in stage. Cisplatin-based NAC demonstrated activity in UC with glandular differentiation, suggesting NAC should be considered for this histologic variant.

Pregnancy-associated breast cancer: evaluating maternal and foetal outcomes. A national study.

PURPOSE: Pregnancy-associated breast cancer (PABC) is defined as breast cancer diagnosed during the gestational period (gp-PABC) or in the first postpartum year (pp-PABC). Despite its infrequent occurrence, the incidence of PABC appears to be rising due to the increasing propensity for women to delay childbirth. We have established the first retrospective registry study of PABC in Ireland to examine specific clinicopathological characteristics, treatments, and maternal and foetal outcomes. METHODS: This was a national, multi-site, retrospective observational study, including PABC patients treated in 12 oncology institutions from August 2001 to January 2020. Data extracted included information on patient demographics, tumour biology, staging, treatments, and maternal/foetal outcomes. Survival data for an age-matched breast cancer population over a similar time period was obtained from the National Cancer Registry of Ireland (NCRI). Standard biostatistical methods were used for analyses. RESULTS: We identified 155 patients-71 (46%) were gp-PABC and 84 (54%) were pp-PABC. The median age was 36 years. Forty-four patients (28%) presented with Stage III disease and 25 (16%) had metastatic disease at diagnosis. High rates of triple-negative (25%) and HER2+ (30%) breast cancer were observed. We observed an inferior 5-year overall survival (OS) rate in our PABC cohort compared to an age-matched breast cancer population in both Stage I-III (77.6% vs 90.9%) and Stage IV disease (18% vs 38.3%). There was a low rate (3%) of foetal complications. CONCLUSION: PABC patients may have poorer survival outcomes. Further prospective data are needed to optimise management of these patients.

Frequency and outcomes of brain metastases in patients with HER2-mutant lung cancers.

BACKGROUND: Mutations in human epidermal growth factor receptor 2 (HER2; also known as ERBB2) are found in approximately 2% of lung adenocarcinomas. The frequency and clinical course of brain metastases in this oncogenic subset are ill defined. METHODS: Baseline and subsequent development of brain metastases was evaluated in consecutive patients with HER2-mutant (n = 98), epidermal growth factor receptor (EGFR)-mutant (n = 200), and KRAS-mutant lung cancers (n = 200). RESULTS: At metastatic diagnosis, the odds ratio (ORs) for brain metastases was similar for patients whose tumors harbored HER2 mutations (19%) in comparison with patients with KRAS mutations (24%; OR for HER2 vs KRAS, 0.7; P = .33) but lower compared to patients with EGFR mutations (31%; OR for HER2 vs EGFR, 0.5; P = .03). Patients with lung cancer and HER2 mutations developed more brain metastases on treatment than patients with KRAS mutations (28% vs 8%; hazard ratio [HR], 5.2; P < .001) and trended more than patients with EGFR mutations (28% vs 16%; HR, 1.7; P = .06). Patients with HER2 YVMA mutations also developed more brain metastases on treatment than patients with KRAS mutations (HR, 5.9; P < .001). The median overall survival (OS) was shorter for patients with HER2-mutant (1.6 years; P < .001) or KRAS-mutant lung cancers (1.1 years; P < .001) than patients with EGFR-mutant lung cancers (3.0 years). Brain metastases occurred in 47% of patients with HER2-mutant lung cancers, which imparted shorter OS (HR, 2.7; P < .001). CONCLUSIONS: These data provide a framework for brain imaging surveillance in patients with HER2-mutant lung cancers and underpin the need to develop HER2-targeted agents with central nervous system activity.

Tumor mutational load predicts survival after immunotherapy across multiple cancer types.

Immune checkpoint inhibitor (ICI) treatments benefit some patients with metastatic cancers, but predictive biomarkers are needed. Findings in selected cancer types suggest that tumor mutational burden (TMB) may predict clinical response to ICI. To examine this association more broadly, we analyzed the clinical and genomic data of 1,662 advanced cancer patients treated with ICI, and 5,371 non-ICI-treated patients, whose tumors underwent targeted next-generation sequencing (MSK-IMPACT). Among all patients, higher somatic TMB (highest 20% in each histology) was associated with better overall survival. For most cancer histologies, an association between higher TMB and improved survival was observed. The TMB cutpoints associated with improved survival varied markedly between cancer types. These data indicate that TMB is associated with improved survival in patients receiving ICI across a wide variety of cancer types, but that there may not be one universal definition of high TMB.

ERCC2 Helicase Domain Mutations Confer Nucleotide Excision Repair Deficiency and Drive Cisplatin Sensitivity in Muscle-Invasive Bladder Cancer.

PURPOSE: DNA-damaging agents comprise the backbone of systemic treatment for many tumor types; however, few reliable predictive biomarkers are available to guide use of these agents. In muscle-invasive bladder cancer (MIBC), cisplatin-based chemotherapy improves survival, yet response varies widely among patients. Here, we sought to define the role of the nucleotide excision repair (NER) gene ERCC2 as a biomarker predictive of response to cisplatin in MIBC. EXPERIMENTAL DESIGN: Somatic missense mutations in ERCC2 are associated with improved response to cisplatin-based chemotherapy; however, clinically identified ERCC2 mutations are distributed throughout the gene, and the impact of individual ERCC2 variants on NER capacity and cisplatin sensitivity is unknown. We developed a microscopy-based NER assay to profile ERCC2 mutations observed retrospectively in prior studies and prospectively within the context of an institution-wide tumor profiling initiative. In addition, we created the first ERCC2-deficient bladder cancer preclinical model for studying the impact of ERCC2 loss of function. RESULTS: We used our functional assay to test the NER capacity of clinically observed ERCC2 mutations and found that most ERCC2 helicase domain mutations cannot support NER. Furthermore, we show that introducing an ERCC2 mutation into a bladder cancer cell line abrogates NER activity and is sufficient to drive cisplatin sensitivity in an orthotopic xenograft model. CONCLUSIONS: Our data support a direct role for ERCC2 mutations in driving cisplatin response, define the functional landscape of ERCC2 mutations in bladder cancer, and provide an opportunity to apply combined genomic and functional approaches to prospectively guide therapy decisions in bladder cancer.See related commentary by Grivas, p. 907.

YES1 amplification is a mechanism of acquired resistance to EGFR inhibitors identified by transposon mutagenesis and clinical genomics.

In ∼30% of patients with EGFR-mutant lung adenocarcinomas whose disease progresses on EGFR inhibitors, the basis for acquired resistance remains unclear. We have integrated transposon mutagenesis screening in an EGFR-mutant cell line and clinical genomic sequencing in cases of acquired resistance to identify mechanisms of resistance to EGFR inhibitors. The most prominent candidate genes identified by insertions in or near the genes during the screen were MET, a gene whose amplification is known to mediate resistance to EGFR inhibitors, and the gene encoding the Src family kinase YES1. Cell clones with transposon insertions that activated expression of YES1 exhibited resistance to all three generations of EGFR inhibitors and sensitivity to pharmacologic and siRNA-mediated inhibition of YES1 Analysis of clinical genomic sequencing data from cases of acquired resistance to EGFR inhibitors revealed amplification of YES1 in five cases, four of which lacked any other known mechanisms of resistance. Preinhibitor samples, available for two of the five patients, lacked YES1 amplification. None of 136 postinhibitor samples had detectable amplification of other Src family kinases (SRC and FYN). YES1 amplification was also found in 2 of 17 samples from ALK fusion-positive lung cancer patients who had progressed on ALK TKIs. Taken together, our findings identify acquired amplification of YES1 as a recurrent and targetable mechanism of resistance to EGFR inhibition in EGFR-mutant lung cancers and demonstrate the utility of transposon mutagenesis in discovering clinically relevant mechanisms of drug resistance.

Brain Metastases in Pancreatic Ductal Adenocarcinoma: Assessment of Molecular Genotype-Phenotype Features-An Entity With an Increasing Incidence?

PURPOSE: To assess clinical characteristics of patients with metastatic pancreas ductal adenocarcinoma (PDAC) and brain metastases (BM), and to assess somatic and germ-line molecular profiles where performed. PATIENTS AND METHODS: Patients with PDAC and BM between January 1990 and January 2016 were identified. Molecular characteristics of somatic and germ-line testing where performed in the subset of patients who had provided informed consent. Somatic alterations were assessed by either MSK-IMPACT testing (>340 key cancer genes) or Sequenom testing (8-gene panel). Overall survival was calculated from date of diagnosis to either date of last follow-up or death. Survival after BM was calculated from date of diagnosis of BM by radiology or pathology to either date of last follow-up or death. RESULTS: From a total of 5824 patients with PDAC identified from January 2000 to January 2016, twenty-five patients (0.4%) had BM. Median age at PDAC diagnosis was 58 years. Median time to the development of BM from initial PDAC diagnosis was 17 months (range, 0-79 months). Median overall survival after BM diagnosis was 1.5 months (range, 1-31 months). Overall survival for patients who had craniotomy (n = 4) was 11 months (range, 1-31 months), with 2 long-term survivors at 21 and 31 months, respectively. Four patients had leptomeningeal disease. Six of 25 patients had germ-line testing, and 3 had BRCA mutations (2 BRCA1 and 1 BRCA2). Somatic profiling identified KRAS mutations in 100% (4 G12D, 2 G12V, and 1 Q61K). CONCLUSION: BM from PDAC is a rare event. We identified a speculative association of germ-line BRCA1/2 alterations with BM in PDAC, which requires corroboration. Survival after BM development is poor; prolonged survival occurred in selected patients via a multidisciplinary approach.

Prospective Evaluation of Germline Alterations in Patients With Exocrine Pancreatic Neoplasms.

Background: Identification of pathogenic germline alterations (PGAs) has important clinical and therapeutic implications in pancreas cancer. We performed comprehensive germline testing (GT) in an unselected prospective cohort of patients with exocrine pancreatic neoplasms with genotype and phenotype association to facilitate identification of prognostic and/or predictive biomarkers and examine potential therapeutic implications. Methods: Six hundred fifteen unselected patients with exocrine pancreatic neoplasms were prospectively consented for somatic tumor and matched sample profiling for 410-468 genes. GT for PGAs in 76 genes associated with cancer susceptibility was performed in an "identified" manner in 356 (57.9%) patients and in an "anonymized" manner in 259 (42.1%) patients, using an institutional review board-approved protocol. Detailed clinical and pathological features, response to platinum, and overall survival (OS) were collected for the identified cohort. OS was analyzed with Kaplan-Meier curves. Results: PGAs were present in 122 (19.8%) of 615 patients involving 24 different genes, including BRCA1/2, ATM, PALB2, and multiple additional genes associated with the DNA damage response pathway. Of 122 patients with germline alterations, 41.8% did not meet current guidelines for GT. The difference in median OS was not statistically significant between patients with and without PGA (50.8 months, 95% confidence interval = 34.5 to not reached, two-sided P = .94). Loss of heterozygosity was found in 60.0% of BRCA1/2. Conclusions: PGAs frequently occur in pancreas exocrine neoplasms and involve multiple genes beyond those previously associated with hereditary pancreatic cancer. These PGAs are therapeutically actionable in about 5% to 10% of patients. These data support routinely offering GT in all pancreatic ductal adenocarcimona patients with a broad panel of known hereditary cancer predisposition genes.

Case report: primary acinar cell carcinoma of the liver treated with multimodality therapy.

We describe a case of primary acinar cell carcinoma (ACC) originating in the liver in a 54-year-old female, diagnosed following persistent abnormal elevated liver function. Imaging revealed two masses, one dominant lesion in the right hepatic lobe and another in segment IVA. A right hepatectomy was performed to remove the larger lesion, while the mass in segment IVA was unresectable due to its proximity to the left hepatic vein. Immunohistochemical staining showed positivity for trypsin and chymotrypsin. Postoperatively the patient underwent hepatic arterial embolization of the other unresectable lesion followed by FOLFOX chemotherapy. At 20 months from diagnosis the patient is currently under observation with a decreasing necrotic mass and no other disease evident. Based on histology, immunohistochemistry and radiological findings a diagnosis of primary ACC of the liver was made. Genomic assessment of somatic mutations within the patient's tumor was also performed through next generation sequencing and findings were consistent with an acinar malignancy. This case highlights a rare tumor subtype treated with a combination of therapeutic modalities through a multidisciplinary approach.

Real-Time Genomic Profiling of Pancreatic Ductal Adenocarcinoma: Potential Actionability and Correlation with Clinical Phenotype.

Purpose: Molecular profiling in cancer has identified potential actionable drug targets that have prompted attempts to discover clinically validated biomarkers to guide therapeutic decision-making and enrollment to clinical trials. We evaluated whether comprehensive genetic analysis of patients with pancreatic adenocarcinoma is feasible within a clinically relevant timeframe and whether such analyses provide predictive and/or prognostic information along with identification of potential targets for therapy.Experimental Design: Archival or prospectively acquired FFPE samples and matched normal DNA from N = 336 patients with pancreatic cancer were analyzed using a hybridization capture-based, next-generation sequencing assay designed to perform targeted deep sequencing of all exons and selected introns of 410 key cancer-associated genes. Demographic and treatment data were prospectively collected with the goal of correlating treatment outcomes and drug response with molecular profiles.Results: The median time from protocol consent to reporting of the genomic results was 45 days with a median time from tissue delivery of 20 days. All genetic alterations identified were stratified based upon prior evidence that the mutation is a predictive biomarker of drug response using the MSKCC OncoKB classification. Three of 225 patients (1%) received a matched therapy based upon the sequencing results.Conclusions: The practical application of molecular results to guide individual patient treatment is currently limited in patients with pancreatic adenocarcinoma. Future prospective molecular profiling efforts should seek to incorporate routine germline genetic analysis and the identification of DNA profiles that predict for clinical benefit from agents that target DNA damage repair and or immunotherapy. Clin Cancer Res; 23(20); 6094-100. ©2017 AACR.

Prospective Comprehensive Molecular Characterization of Lung Adenocarcinomas for Efficient Patient Matching to Approved and Emerging Therapies.

Tumor genetic testing is standard of care for patients with advanced lung adenocarcinoma, but the fraction of patients who derive clinical benefit remains undefined. Here, we report the experience of 860 patients with metastatic lung adenocarcinoma analyzed prospectively for mutations in >300 cancer-associated genes. Potentially actionable genetic events were stratified into one of four levels based upon published clinical or laboratory evidence that the mutation in question confers increased sensitivity to standard or investigational therapies. Overall, 37.1% (319/860) of patients received a matched therapy guided by their tumor molecular profile. Excluding alterations associated with standard-of-care therapy, 14.4% (69/478) received matched therapy, with a clinical benefit of 52%. Use of matched therapy was strongly influenced by the level of preexistent clinical evidence that the mutation identified predicts for drug response. Analysis of genes mutated significantly more often in tumors without known actionable mutations nominated STK11 and KEAP1 as possible targetable mitogenic drivers.Significance: An increasing number of therapies that target molecular alterations required for tumor maintenance and progression have demonstrated clinical activity in patients with lung adenocarcinoma. The data reported here suggest that broader, early testing for molecular alterations that have not yet been recognized as standard-of-care predictive biomarkers of drug response could accelerate the development of targeted agents for rare mutational events and could result in improved clinical outcomes. Cancer Discov; 7(6); 596-609. ©2017 AACR.See related commentary by Liu et al., p. 555This article is highlighted in the In This Issue feature, p. 539.

Frequent somatic CDH1 loss-of-function mutations in plasmacytoid variant bladder cancer.

Plasmacytoid bladder cancer is an aggressive histologic variant with a high risk of disease-specific mortality. Using whole-exome and targeted sequencing, we find that truncating somatic alterations in the CDH1 gene occur in 84% of plasmacytoid carcinomas and are specific to this histologic variant. Consistent with the aggressive clinical behavior of plasmacytoid carcinomas, which frequently recur locally, CRISPR/Cas9-mediated knockout of CDH1 in bladder cancer cells enhanced cell migration.

Targeted therapy in advanced bladder cancer: what have we learned?

Despite advances in the treatment of other genitourinary malignancies, no novel therapies have been approved by the US Food and Drug Administration for urothelial carcinoma (UC) in the last 20 years. To date, no clinical trials of targeted agents in UC have led to improvements in survival compared with cytotoxic therapy. This article outlines representative trials of targeted therapies in UC and discusses the significance of genetic preselection in trial design as a method to optimize responses to these agents, thus, hopefully expanding the armamentarium of treatment options against this lethal disease.