Melanoma Patients In Phase Research Articles

Abstract PL02-03: Combination with/of immunotherapies

Abstract Systemic treatment of melanoma has faced a revolution the last decade. After 30 years of disappointments and no clinical benefit for patients, targeted agents, like the BRAF inhibitors vemurafenib or dabrafenib, and checkpoint inhibitors of CTLA-4 or PD-1/PD-L1, like ipilimumab, pembrolizumab or nivolumab, have improved for the first time the overall survival of late stage melanoma patients in phase 3 trials. Addition of MEK inhibitors to the BRAF inhibitors and combination of CTLA-4 and PD-1 blockade were able to improve the response rate and progression free survival further. However, despite these promising data, a considerable percentage of patients will not benefit long-term from these treatments. This raises questions in two directions: 1) how can we identify the patients that benefit long-term from current treatment options, and 2) what are future therapies, that should be tested in patients that do not benefit from current standards. Based on recent analyses from mainly expanded access programs and safety trials we propose to characterize patients (‘ tumors) according to: a) Tumor foreignness, e.g. mutational load, b) General immune status, e.g. lymphocyte count, c) Infiltration capacity, e.g. intratumoral CD8 infiltration, d) Absence of locally expressed inhibitory factors, e.g. PD-L1, e) Absence of local soluble inhibitory factors, e.g. IL-6, as measured by surrogate markers like C-reactive protein (CRP) or erythrocyte sedimentation rate (ESR), f) Absence of inhibitory tumor metabolism, e.g. as measured by LDH or glucose uptake, and g) Sensitivity to effector mechanisms, like MHC expression. Most these markers have already been identified to be clinical relevant for the patient's outcome upon T cell checkpoint blockade, and been shown to be crucial mechanisms for anti-tumor immune response in preclinical models. They often have a prognostic and a predictive relevance for patients treated with checkpoint modulators. This often raised critic cannot be solved, particularly for immune modulatory therapies, as the immune system, as effector compartment of any of these therapies is upfront any intervention present in the cancer patients, and thus will have always also prognostic relevance. Nevertheless, these clinical markers are important tools to understand requirements for an effective anti-tumor immune response in any cancer, and can be used to develop a cancer immunogram for individual patients or malignancies. Based on this immunogram personalized immunotherapy can be applied using current/future combinations of immunotherapeutic approaches and agents used in targeted therapy. I will discuss several of such combination therapy options currently tested in early phase trials that address the different levels of the cancer immunogram. Citation Format: Christian Blank. Combination with/of immunotherapies. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr PL02-03.

Serum lactate dehydrogenase (LDH) as a prognostic selection criterion for ipilimumab treatment in metastatic melanoma.

3036 Background: Ipilimumab, a CTLA4 blocking antibody, has been shown to improve survival in metastatic melanoma patients in phase III trials. However, only a subset of patients experiences long-term survival benefit. Therefore, we analysed two independent retrospective sets of patients treated in ‘real world’ settings to identify prognostic factors that correlate with better outcome after ipilimumab therapy. Methods: Advanced melanoma patients, eligible for ipilimumab therapy, were treated in the Dutch and UK Expanded Access Programs (EAP) and after European licensing on the 3mg/kg schedule. Baseline characteristics and peripheral blood parameters were assessed and patients were monitored for the occurrence of adverse events and responses. Results: A total of 166 patients were enrolled in the Dutch EAP. Best overall response rate and disease control rate were (DCR) 17% and 35%. Median follow-up was 17.9 months, with a median progression free survival of 2.9 months. Median overall survival (OS) was 7.5 months, and OS at 1 year 37.8% and at 2 years 22.9%. Univariate analysis revealed that gender, age, Breslow thickness, baseline and week 6 lymphocyte count (ALC), and prior treatment had no influence on OS, while mWHO, M stage, baseline LDH, S100, erythrocyte sedimentation rate (ESR), and the slope of ALC did. In a multivariate model only baseline LDH and ESR remained as significant independent prognostic factors. The relevance of LDH was validated in an independent cohort of 68 patients from the UK. Stratifying the patients in the NL cohort according to LDH levels (≤upper limit normal (ULN), 54.4% of patients versus >2xULN, 16.9% of patients) separated into groups of patients observing DCR of 48% versus 14%, and median OS of 13.7 versus 3.1 months, while toxicity was similar in both groups (48% and 41%). None of the patients was alive at 15 months after treatment if baseline LDH was >2xULN in both the NL and UK cohorts. Conclusions: Overall survival upon ipilimumab treatment is lower in an expanded access population as compared to phase III trials. LDH >2xULN at baseline is a potential prognosticator in patients receiving ipilimumab and should be considered in guiding ipilimumab treatment initiation.

Abstract 4025: Data-driven computational modeling to identify biomarkers of response to lenvatinib (E7080) in melanoma.

Abstract Background: Lenvatinib is an oral tyrosine kinase inhibitor targeting VEGFR1-3, FGFR1-4, RET, KIT and PDGFRβ. Anti-tumor activity has been observed in melanoma patients in Phase I studies. We applied an integrative supercomputer-driven analysis approach to identify biomarkers of lenvatinib treatment response in melanoma patients. Methods: Clinical data sets including tumor response data (RECIST), progression free survival (PFS), pharmacokinetic parameters (PK) and molecular data sets including baseline tumor gene expression (Affymetrix U133Plus2) and BRAF and NRAS mutational status, were collected from 18 patients with metastatic melanoma who received lenvatinib 10 mg orally twice daily in 28-day cycles. These clinical and molecular data sets were used to generate computational models developed using REFS (Reverse Engineering and Forward Simulation) modeling platform, which utilizes Bayesian network inference and simulations. Simulations were performed to identify biomarkers of lenvatinib treatment response. These potential predictive biomarkers were then tested for their ability to predict response to lenvatinib in preclinical models. Results: Using a model comprising gene expression, mutational status and PK data, REFS identified a panel of 18 potential predictive biomarkers of lenvatinib treatment response. Identified biomarkers were able to predict up to 89% of the observed variance in the tumor response data. A total of 32 identified genes including 6 candidate predictive biomarkers (TARBP2, CACNA1, C7ORF, RAP2A, SHMT1, IL22RA2) were further validated in a preclinical melanoma model system (n=12) and tissue bank samples with matched normal adjacent tissue (n=21). Expression of 14 genes correlated with relative tumor volume (r>0.35 or r<-0.35) in the preclinical model and expression levels of 19 genes were found to be higher in tumor compared to normal tissue (FC>2). Conclusions: Potential predictive biomarkers of lenvatinib treatment response in melanoma patients were identified by computational modeling and validated in a preclinical model system and tumor tissue bank samples. The identified biomarkers will be tested for their predictive value in an ongoing Phase 2 trial. Citation Format: Tadashi Kadowaki, Yasuhiro Funahashi, Junji Matsui, Kumar Pavan, Pallavi Sachdev, Jim O'Brien, Heming Xing, Paul D. McDonagh, Iya Khalil, Razelle Kurzrock, David S. Hong, John Nemunaitis. Data-driven computational modeling to identify biomarkers of response to lenvatinib (E7080) in melanoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4025. doi:10.1158/1538-7445.AM2013-4025

Assessment of Prognostic Circulating Tumor Cells in a Phase III Trial of Adjuvant Immunotherapy After Complete Resection of Stage IV Melanoma

To verify circulating tumor cell (CTC) prognostic utility in stage IV resected melanoma patients in a prospective international phase III clinical trial. Our studies of melanoma patients in phase II clinical trials demonstrated prognostic significance for CTCs in patients with AJCC stage IV melanoma. CTCs were assessed to determine prognostic utility in follow-up of disease-free stage IV patients pre- and during treatment. After complete metastasectomy, patients were prospectively enrolled in a randomized trial of adjuvant therapy with a whole-cell melanoma vaccine, Canvaxin, plus Bacille Calmette-Guerin (BCG) versus placebo plus BCG. Blood specimens obtained pretreatment (n = 244) and during treatment (n = 214) were evaluated by quantitative real-time reverse-transcriptase polymerase chain reaction (qPCR) for expression of MART-1, MAGE-A3, and PAX3 mRNA biomarkers. Univariate and multivariate Cox analyses examined CTC biomarker expression with respect to clinicopathological variables. CTC biomarker(s) (≥ 1) was detected in 54% of patients pretreatment and in 86% of patients over the first 3 months. With a median follow-up of 21.9 months, 71% of patients recurred and 48% expired. CTC levels were not associated with known prognostic factors or treatment arm. In multivariate analysis, pretreatment CTC (> 0 vs. 0 biomarker) status was significantly associated with disease-free survival (DFS; HR 1.64, P = 0.002) and overall survival (OS; HR 1.53, P = 0.028). Serial CTC (>0 vs. 0 biomarker) status was also significantly associated with DFS (HR 1.91, P = 0.02) and OS (HR 2.57, P = 0.012). CTC assessment can provide prognostic discrimination before and during adjuvant treatment for resected stage IV melanoma patients.

Mechanism of antitumor activity of E7080, a selective VEGFR and FGFR tyrosine kinase inhibitor (TKI), in combination with selective mutant BRAF inhibition.

8567 Background: E7080 is a TKI targeting VEGFR1-3, FGFR1-4, KIT, RET and PDGFR. Anti-tumor activity has been observed in melanoma patients in phase I. The objective of this study was to examine the effect of E7080 alone and in combination with PLX-4032 on tumor response, gene expression and tumor microvasculature in BRAFmu melanoma. Methods: BRAF mutant (mu) cell lines were created by transfection of BRAF V600E constructs into BRAF wild type (wt) cell lines. Anti-tumor activity of E7080 alone or in combination with the selective BRAF inhibitor PLX-4032 was examined in xenografted models bearing mock and BRAFmu transfectants. Results: The ratio of ANGPT1/2 expression was significantly higher in BRAFmu/ PTENwt compared to BRAFwt/PTENwt and BRAFmu/PTENmu among 14 melanoma cell lines. ANGPT1 expression was up-regulated in BRAFmu by 330%, and ANGPT2 in PTENmu by 290% compared to wt. PLX-4032 decreased ANGPT1 expression and pericyte coverage of tumor vessels in BRAFmu/PTENwt A375 melanoma. Similarly, overexpression of BRAF V600E in BRAFwt SK-MEL-2 melanoma resulted in increased ANGPT1 expression along with increased pericyte coverage. E7080 at 3, 10, 30 to 100 mg/kg showed significant anti-tumor activity against mock transfectants, but only at 100 mg/kg against BRAF V600E transfectants. Combination of E7080 at 10 mg/kg with PLX-4032 at 100 mg/kg showed synergistic anti-tumor activity against A375 melanoma and caused tumor regression, not observed by either single agent E7080 or PLX-4032. E7080 alone increased pericyte coverage of tumor vessels, but in combination with PLX-4032 resulted in marked decreased in pericyte coverage and microvessel density. Conclusions: The BRAFmu/ PTENwt melanomas studied demonstrate high pericyte coverage and resistance to E7080. Combination of E7080 with PLX-4032 achieves synergistic antitumor effect with enhanced reduction of pericyte coverage. These data provide a mechanistic basis for clinical study of BRAF inhibitors in combination with VEGFR and FGFR targeted agents.

The effect of E7080, a VEGFR and FGFR tyrosine kinase inhibitor (TKI), on BRAF wild-type melanoma.

8566 Background:E7080 is a TKI targeting VEGFR1-3, FGFR1-4, KIT, RET and PDGFR. Anti-tumor activity has been observed in melanoma patients in phase I. This study examines the mode of action of E7080 in preclinical melanoma models Methods: Mutational status of 18 genes (9 RTKs, 7 oncogenes and 2 tumor suppressor genes) was determined in 14 human melanoma cell lines. Gene profile and biomarkers of anti-angiogenesis and anti-tumor activity of E7080 were investigated in vitro and in vivo by gene expression analysis in a melanoma cell line panel and in human melanoma tissues. Results: 18 types of mutation (mu) were found among 9 RTKs in ten of 14 melanoma cell lines. 2 mu of BRAF, 9 mu of TP53, 3 mu of PTEN, 2 mu of CDKN2a and 1 mu of NRAS were also found among 9, 9, 3, 2 and 1 melanoma of 14 cell lines, respectively. 4 of the 5 melanoma with BRAF wild type (wt) were the most sensitive to E7080 in mouse xenograft models, while 5 of the 6 with BRAFmu/PTENwt were the most resistant, suggesting 3 sub-groups for sensitivity to E7080 based on BRAF/PTEN status: 1; BRAFwt/PTENwt, 2; BRAFmu/PTENmu and 3; BRAFmu/PTENwt. % of pericyte-covered tumor vessels was 25%, 29% and 35% among 3 sub-groups, respectively. Anti-tumor activity of E7080 was correlated to % of pericyte coverage in groups 1 and 2, in which tumor vessels are sensitive to E7080. DNA microarray analysis demonstrated decreased FGFR1-4 expression in BRAFmu melanoma, while qPCR analysis showed that FGFR2 and FGFR3 expression were significantly higher in BRAFwt than BRAFmu in human melanoma tissues. E7080 inhibited in vitro proliferation of BRAFwt melanoma, which overexpressed FGFR3. Knockdown of FGFR3, FRS-2a, NRAS and AKT-3 genes with siRNA transfection inhibited proliferation of E7080-sensiteve melanoma. Conclusions: BRAF/PTEN mutation status was associated with pericyte coverage of tumor vessels and the expression of FGFR1-4 in a panel of melanoma cell lines. E7080 showed superior anti-tumor activity against BRAFwt melanoma, in which the type of tumor vessels was sensitive to angiogenesis inhibition by E7080 and a subgroup of which were dependent on FGFR3 signaling.