Eric Michael Robinson

The Hippo effector YAP promotes resistance to RAF- and MEK-targeted cancer therapies

Resistance to RAF- and MEK-targeted therapy is a major clinical challenge. RAF and MEK inhibitors are initially but only transiently effective in some but not all patients with BRAF gene mutation and are largely ineffective in those with RAS gene mutation because of resistance. Through a genetic screen in BRAF-mutant tumor cells, we show that the Hippo pathway effector YAP (encoded by YAP1) acts as a parallel survival input to promote resistance to RAF and MEK inhibitor therapy. Combined YAP and RAF or MEK inhibition was synthetically lethal not only in several BRAF-mutant tumor types but also in RAS-mutant tumors. Increased YAP in tumors harboring BRAF V600E was a biomarker of worse initial response to RAF and MEK inhibition in patients, establishing the clinical relevance of our findings. Our data identify YAP as a new mechanism of resistance to RAF- and MEK-targeted therapy. The findings unveil the synthetic lethality of combined suppression of YAP and RAF or MEK as a promising strategy to enhance treatment response and patient survival.

Sensitivity of plasma BRAFmutant and NRASmutant cell-free DNA assays to detect metastatic melanoma in patients with low RECIST scores and non-RECIST disease progression

Melanoma lacks a clinically useful blood‐based biomarker of disease activity to help guide patient management. To determine whether measurements of circulating, cell‐free, tumor‐associated BRAFmutant and NRASmutant DNA (ctDNA) have a higher sensitivity than LDH to detect metastatic disease prior to treatment initiation and upon disease progression we studied patients with unresectable stage IIIC/IV metastatic melanoma receiving treatment with BRAF inhibitor therapy or immune checkpoint blockade and at least 3 plasma samples obtained during their treatment course. Levels of BRAFmutant and NRASmutant ctDNA were determined using droplet digital PCR (ddPCR) assays. Among patients with samples available prior to treatment initiation ctDNA and LDH levels were elevated in 12/15 (80%) and 6/20 (30%) (p = 0.006) patients respectively. In patients with RECIST scores <5 cm prior to treatment initiation, ctDNA levels were elevated in 5/7 (71%) patients compared to LDH which was elevated in 1/13 (8%) patients (p = 0.007). Among all disease progression events the modified bootstrapped sensitivities for ctDNA and LDH were 82% and 40% respectively, with a median difference in sensitivity of 42% (95% confidence interval, 27%–58%; P < 0.001). In addition, ctDNA levels were elevated in 13/16 (81%) instances of non‐RECIST disease progression, including 10/12 (83%) instances of new brain metastases. In comparison LDH was elevated 8/16 (50%) instances of non‐RECIST disease progression, including 6/12 (50%) instances of new brain metastases. Overall, ctDNA had a higher sensitivity than LDH to detect disease progression, including non‐RECIST progression events. ctDNA has the potential to be a useful biomarker for monitoring melanoma disease activity.

Association between Ki-67 expression and clinical outcomes among patients with clinically node-negative, thick primary melanoma who underwent nodal staging

Patients with thick primary melanomas (≥4 mm) have highly variable survival outcomes. Cell proliferation marker Ki‐67 has been identified as promising biomarker in thick melanoma but has not been evaluated since the wide spread adoption of sentinel lymph node biopsy. We revisit its prognostic relevance in the sentinel node era.

Outcomes in Melanoma Patients Treated with BRAF/MEK-Directed Therapy or Immune Checkpoint Inhibition Stratified by Clinical Trial versus Standard of Care

Objectives: Since 2011, metastatic melanoma treatment has evolved with commercial approval of BRAF- and MEK-targeted therapy and CTLA-4- and PD-1-blocking antibodies (immune checkpoint inhibitors, ICI). While novel therapies have demonstrated improved prognosis in clinical trials, few studies have examined the evolution of prognosis and toxicity of these drugs among an unselected population. We assess whether survival and toxicity reported in trials, which typically exclude most patients with brain metastases and poor performance status, are recapitulated within a commercial access population. Methods: 182 patients diagnosed with stage IV melanoma from July 2006 to December 2013 and treated with BRAF- and/or MEK-targeted therapy or ICI were prospectively studied. Outcomes and clinicopathologic differences between trial and commercial cohorts were assessed. Results: Patients receiving commercial therapy (vs. on trial) had poorer prognostic features (i.e., brain metastases) and lower median overall survival (mOS) when assessed across all treatments (9.2 vs. 17.5 months, p = 0.0027). While toxicity within trial and commercial cohorts did not differ, patients who experienced toxicity had increased mOS (p < 0.001), irrespective of stratification by trial status or therapy. Conclusion: Metastatic melanoma patients receiving commercial treatment may represent a different clinical population with poor prognostic features compared to trial patients. Toxicity may prognosticate treatment benefit.

Abstract 2164: Revisiting the mechanisms of PTEN loss in melanoma

Introduction Inactivation of the PTEN tumor suppressor, a negative regulator of oncogenic PI3K/Akt signaling, occurs in a subset of melanomas and is correlated with disease progression, treatment resistance and reduced patient survival. PTEN loss of function in melanoma is thought to result largely from gene deletion, mutation and/or methylation. However, the low frequency of these genomic alterations cannot explain the higher rate of loss of PTEN protein expression that is observed in melanoma. In this study, we tested the hypothesis that loss of PTEN expression in melanoma might be the consequence of predominantly non-genomic mechanisms, including post-transcriptional and post-translational dysregulation. Specifically, we postulated that PTEN loss in some melanomas might result from aberrant expression of PTENP1, a PTEN pseudogene-derived long non-coding RNA, and from increased PTEN ubiquitination and hence protein degradation. Experimental Procedures We performed an integrated analysis of data obtained from The Cancer Genome Atlas (TCGA; n = 276) and metastatic melanoma human specimens from patients enrolled at NYU (n = 48). We examined the frequency of PTEN copy number loss using genomic qPCR, PTEN mutations using targeted sequencing and qPCR mutation arrays, and PTEN methylation status was determined in tumors lacking PTEN deletion or mutation with bisulfite treatment and pyrosequencing. Furthermore, we assessed PTEN and PTENP1 expression in tumor samples by RT-qPCR, western blotting and immunohistochemistry. Results Complete PTEN deletions were found in 8% of TCGA cases and in 10% of NYU cases, while partial PTEN deletions were detected in 27% of NYU cases. Somatic PTEN mutations were found in 8% of TCGA cases and 3% of NYU cases. The frequency of PTEN methylation at five CpG islands ranged between 0% and 28% and did not predict PTEN protein expression, thus identifying a subset of tumors that exhibit PTEN protein loss despite a lack of detectable PTEN deletions, mutations or methylation. Integration of PTEN and PTENP1 expression data from TCGA and the NYU cohort revealed a strong positive correlation between PTEN and PTENP1 levels in melanoma. Analysis of a role for aberrant PTEN ubiquitination in promoting PTEN loss in melanoma is ongoing. Conclusions Our data challenge the existing model in which PTEN loss in melanoma is thought to occur primarily as a result of gene deletion, mutation or methylation. Rather, we find that these are uncommon events in melanoma and that PTEN loss may instead be attributed to post-transcriptional or post-translational mechanisms, such as dysregulation by loss of the PTENP1 long non-coding RNA. A comprehensive understanding of these non-genomic processes could identify novel approaches (“PTEN-restoring therapy”) to block tumor progression and improve treatment responses and patient survival, suggesting that suppression of PTEN levels in melanoma might be reversible and thus could be exploited clinically. Citation Format: Keith Giles, Yang Li, Amel Salhi, Jinhua Wang, Eric Robinson, Iman Osman. Revisiting the mechanisms of PTEN loss in melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2164. doi:10.1158/1538-7445.AM2015-2164