Faculty Bibliography

Significant advancements, including development of immune checkpoint inhibitors and targeted therapies, have transformed outcomes in patients (pts) with unresectable or metastatic melanoma. However, pts who do not respond or who progress while receiving these regimens have limited options. Spartalizumab is a high-affinity, humanized monoclonal antibody blocking the programmed cell death-1 (PD-1) receptor. This randomized, open-label, 2-part, multicenter, open-platform, phase 2 study (NCT03484923) will evaluate safety and efficacy of spartalizumab combination treatment in pts with unresectable or metastatic melanoma progressing after prior anti-PD-1/ L1 therapy and, if the tumor harbors a BRAF V600 mutation, a BRAF inhibitor. The primary endpoint is objective response rate in each arm per RECIST v1.1; secondary endpoints include duration of response and assessment of paired tumor biopsies for biomarkers of antitumor T-cell activity. The first "selection" part will begin with 3 combination arms: (1) spartalizumab + LAG525 (LAG-3 antibody), (2) spartalizumab + capmatinib (c-MET inhibitor), and (3) spartalizumab + canakinumab (IL-1beta antagonist). An adaptive design will allow dropping arms for futility, adding new arms, and selecting >= 1 arm for expansion. Bayesian methodology will be used with specific probability criteria for futility and efficacy assessments at each interim analysis. Pts (= 60-85) will be stratified by baseline lactate dehydrogenase and randomized equally to all open arms during the selection part. In the second "expansion" part, efficacy and safety of treatment combination(s) selected during part 1 will be further investigated. Sample size for part 2 will be adaptive and based on predictive power calculations considering the results from part 1

Background Nivolumab and ipilimumab (ipi) combination immunotherapy has a ~60% response rate in metastatic melanoma patients. However, the impact of these therapies on immune cell phenotypes and the relationship of those changes to patient outcomes remains under-investigated. Methods High dimension flow cytometry on baseline and at week 13 (e.g. after initial nivo or ipi therapy) was performed for peripheral blood samples from 33 metastatic melanoma patients receiving sequential nivo-ipi or the reverse sequence. We used a novel computational approach to analyze the data through semi-comprehensive Boolean gating in which immune cell lineages (e.g. CD3+CD4+) were evaluated for all possible combinations for up to 15 markers. Results 3,844 measured immunophenotypes were significantly altered post-nivo, and 7,133 immunophenotypes were altered post-ipi. The frequency of 584 immunophenotypes were significantly changed in both treatments, with 59 of those changing in opposing directions. In the nivo-ipi cohort, 260 baseline and 662 post-nivo immunophenotypes were significantly associated with response and survival (outcomes). In the ipi-nivo cohort, 432 baseline and 668 post-ipi immunophenotypes were associated with outcomes. Two highly similar immunophenotypes associated with outcomes overlapped between the cohorts, CD14⁺CD11C+CD33+CD15-CD19-PDL1- PDL2+CD163+GAL9-CD80-CD86-41BBL+CD40+OX40L+ cells. While lower levels of these cells were associated with response and improved survival in nivo-ipi treated patients, lower levels were associated with better outcomes in the ipi-nivo treated patients. Of the 3,844 immunophenotypes altered post-nivolumab, 100 were also associated with ipi-nivo response. Of these 100, 97% were altered in a manner positively associated with response (e.g. upregulated by nivolumab and higher in responders). For example, nivo upregulated CD4+CD45RO-CCR7+ frequencies, which were associated with response and longer survival in ipi-nivo treated patients. Of the 7,133 immunophenotypes altered post-ipi, 110 were also associated with nivo-ipi response. Of these, 95% were altered in a manner negatively associated with response. This includes ipi associated upregulation of a population of CD4+CD38+CD39+CD127-GARP- cells that are negatively associated with coutcomes in nivo-ipi treated patients as well as downregulation of a CD4+CD127+CD45RO+CD95+CCR7+ population of cells positively associated with outcomes. Conclusions These results demonstrate that nivo and ipi altered the peripheral immune landscape in distinct ways. While immunophenotypic changes post-nivo favored response in ipi-nivo treated patients, changes associated with ipi treatment favored progression with ipi-nivo. These data suggest that the immunophenotypic impact of ipi alters the immune landscape in a manner that may impair a subsequent response to nivo. These data also highlight several novel immune cell populations that are associated with both treatment effects and patient outcomes

Background Therapies targeting T-cell co-inhibitory molecules (e.g. PD1) have demonstrated unprecedented efficacy in the treatment of metastatic melanoma. However, not all patients respond to checkpoint inhibition, so there is an unmet need to identify mechanisms of resistance/response. Methods Using IsoLight, a platform for assessing the secretion of 32 analytes at single-cell resolution, we evaluated previously frozen peripheral blood T-cells from six responding and six progressing patients (according to RECIST 1.1 criteria) treated with nivolumab. Baseline and week 13, post-treatment CD4+ and CD8+ T-cell samples were assessed for each patient. T-cells were stimulated with CD3 and CD28 activating antibodies overnight and subsequently placed on capture chips for 20 hours. Approximately 400 single-cell events were assessed for each sample. Results CD4+ T-cells from progressing patients, compared to those from responding patients, had significantly (p<0.05) higher mean production of IL-17F post-nivolumab and an increased proportion of cells secreting IL-13, RANTES, IL-6, soluble CD137, TNF, MIP1a and MIP1b relative to baseline. Using an elastic net machine learning algorithm with cross validation, we assessed the ability of a manually curated list of analytes to predict patient outcomes. Delta values (post-treatment minus baseline values) were used for 15 parameters. A receiver operating characteristic with an area under the curve of 0.898 was achieved. The most important features in these models were the percentage of CD4+ T-cells expressing IL-6, MIP1a and soluble CD137 along with the percentage of CD8+ T-cells expressing IL-13. Conclusions These results demonstrate the ability of single-cell, high-dimension technologies coupled with machine learning to reveal complex associations between immune cell function and clinical outcomes. Specifically, these data show that changes in secretome production potential of T-cells after treatment with the PD1 blocking antibody nivolumab are associated with metastatic melanoma patient outcomes. We identified a signature of secreted molecules that were associated with patient outcomes, providing rationale for targeting these molecules to increase the efficacy of nivolumab. Work is underway to validate the observed associations in an independent set of patient samples

Background Siglec-15 (S15) is a member of the Siglec family (Sialic acidbinding Immunoglobulin Lectins), a distinct subgroup of immunoglobulin (Ig) superfamily proteins involved in discriminating self and non-self-immune regulation [1]. Recent studies have shown that S15 mediates suppression of T cell proliferation and negatively regulates T cell function. The expression of S15 and PD-L1 are mutually exclusive in NSCLC and other cancers, which supports S15 as a potential target for the treatment of cancer patients who are refractory to PD-1 directed therapies. NC318 is a first-in-class monoclonal antibody that blocks S15-mediated immune suppression and prevents tumor growth by normalizing T cell function and restoring anti-tumor immunity in the tumor microenvironment [2]. Methods NC318-01 is a multi-center, first-in-human, phase 1/2, open-label, non-randomized study to determine the safety and tolerability, define the maximum tolerated dose or pharmacologically active dose, and assess the preliminary efficacy of NC318. The phase 1 component uses a 3+3 dose escalation design to determine the recommended phase 2 dose (RP2D) of NC318. Primary endpoints include safety and tolerability. Secondary endpoints include assessment of pharmacokinetics, response rate (measured every 8 weeks), progression-free survival, and overall survival. Exploratory analyses include investigation of biomarkers associated with treatment benefit. Results As of August 2019, 43 patients have been dosed across 6 dose cohorts (8mg - 800mg Q2W). The most common tumor types enrolled included: 10 NSCLC, 7 ovarian, 6 melanoma, 3 breast, and 3 CRC. NC318 has been well tolerated with no DLTs. The most common treatmentrelated AEs included diarrhea (4- grade 1 and 2-grade 2), pruritis (3- grade 1 and 1- grade 2), rashes (grade 1 and 2), arthralgias (2- grade 1), elevated amylase (grade 1, 2, and 3), and elevated lipase (grade 3 and 4). Tumor responses were evaluable in 32 patients; 11 patients have not reached their first assessment, and their efficacy data will be reported at the conference. Single agent activity has been seen in NSCLC including 1 CR (ongoing at 41 weeks), a PR (ongoing at 14 weeks), 1 stable disease with tumor reduction (ongoing for 26 weeks), and 2 with stable disease. NSCLC BORR: 2/7 or 29%, DCR: 5/7 or 71%. Conclusions NC318 has been well tolerated across multiple dose levels and has shown encouraging anti-tumor activity when administered as monotherapy. Single agent anti-tumor activity was observed in 5 of 7 NSCLC subjects refractory to PD-1 therapies

Background The evolving melanoma treatment algorithm has highlighted the role of immune checkpoint inhibitors in treating patients with this cancer. From the first drug approval in 2011 through today, many changes have occurred in treatment approaches. Selecting the most appropriate therapy with implementation of monitoring strategies for managing adverse events is vital. The goal of this study was to determine if participation in an educational activity can improve the proficiency of oncologists on the application of checkpoint inhibitors in adjuvant and metastatic melanoma. Methods An online continuing medical education (CME)-certified interactive, text-based activity developed by Medscape Education Oncology and the Society for Immunotherapy of Cancer included 2 patient cases, which served as the foundation for interactive questions. Educational design included a "test, then teach" approach to elicit cognitive dissonance, with evidence-based feedback provided following each learner response. A repeated pairs pre-/post-assessment study design with 3 case-based questions and 1 confidence question was used. A chi-square test assessed differences from pre-to post-assessment. P values .26 is extensive). The activity was launched online 2/25/19 and data were collected through 5/8/19. Results Participation in education resulted in statistically significant improvements and an extensive educational effect for oncologists (n=62; P < .001; V =.342). An average of 41% correctly responded to pre-assessment questions, increasing to 75% post-assessment. 40% of oncologists reported greater confidence prescribing immunotherapy in melanoma. Significant improvements in competency were observed in the following areas: * Selection of an evidence-based regimen for patients with melanoma in the adjuvant setting (24% vs 73%, P < 0.001, V = 0.484, Figure 1) * Identification of the most appropriate regimen for patients with metastatic melanoma following molecular testing (47% vs 74%, P <.01, V = 0.280, Figure 2) * Implementing team-based strategies for monitoring and managing irAEs (52% vs 77%, P <.01, V = 0.269, Figure 3) Conclusions This online interactive, case-based, CME-certified educational activity resulted in significant gains in oncologist competency in identifying and selecting appropriate treatments for patients with melanoma in the adjuvant and metastatic setting, and in implementing monitoring and management strategies (Figure 4). These results demonstrate the effectiveness of on-demand education in improving the translation of clinical knowledge into practice scenarios while improving clinician confidence. (Table Presented)

PURPOSE/OBJECTIVE:PD-1 blockade induces durable responses in patients with metastatic melanoma and prolongs relapse-free survival in patients with resected melanoma; however, current biomarkers do not consistently associate with patient responses. In this study we investigated the impact of nivolumab therapy on peripheral blood Tregs and its relation to patient outcomes. EXPERIMENTAL DESIGN/METHODS:Peripheral blood Tregs and conventional CD4+ T-cells from patients with resected high-risk melanoma treated with adjuvant nivolumab were assessed for gene expression changes by RNA-seq. Percentages of circulating Tregs and phosphorylated-STAT3 (pSTAT3) expression levels were assessed by flow cytometry and validated in an independent cohort of active disease patients. Suppressive function of Tregs was assessed in allogeneic mixed lymphocyte reactions. RESULTS:Tregs from non-relapse patients had increased expression of proliferation associated genes . An increase in the proportion of circulating Tregs and pSTAT3 expression and a reduction in Treg suppressive capacity were observed in non-relapsing, but not relapsing patient samples 13 weeks after starting treatment. In vitro blockade of PD-1 increased Treg percentages and pSTAT3 expression, and reduced Treg suppressive function. PD-1 blockade also led to IL-10 production by T-cells, resulting in higher Treg proliferation. The addition of a STAT3 inhibitor ameliorated the increase in Tregs, enhanced suppressive function, and decreased T-cell IL-10 production in vitro. CONCLUSIONS:These results demonstrate that induction of pSTAT3, reduced suppressive function, and a paradoxical increase in Treg proliferation are novel correlates of patient benefit from PD-1 blockade.

PURPOSE/OBJECTIVE:-mutant melanoma, although acquired resistance remains common. Preclinical studies demonstrated that resistance could be overcome using concurrent treatment with the HSP90 inhibitor XL888. METHODS:-mutant melanoma. Primary endpoints were safety and determination of a maximum tolerated dose. Correlative proteomic studies were performed to confirm HSP inhibitor activity. RESULTS:Objective responses were observed in 15/20 evaluable patients (75%; 95% CI: 51-91%), with 3 complete and 12 partial responses. Median progression-free and overall survival were 9.2 months (95% CI: 3.8-not reached) and 34.6 months (6.2-not reached), respectively. The most common grade 3/4 toxicities were skin toxicities such as rash (n=4, 19%) and cutaneous squamous cell carcinomas (n=3, 14%), along with diarrhea (n=3, 14%). Pharmacodynamic analysis of patients' PBMCs showed increased day 8 HSP70 expression compared to baseline in the three cohorts with XL888 doses ≥45 mg. Diverse effects of vemurafenib-XL888 upon intratumoral HSP-client protein expression were noted, with the expression of multiple proteins (including ERBB3 and BAD) modulated on therapy. CONCLUSION/CONCLUSIONS:-mutant melanoma.

In this perspective, we identify emerging frontiers in clinical and basic research of melanocyte biology and its associated biomedical disciplines. We describe challenges and opportunities in clinical and basic research of normal and diseased melanocytes that impact current approaches to research in melanoma and the dermatological sciences. We focus on four themes: (1) clinical melanoma research, (2) basic melanoma research, (3) clinical dermatology, and (4) basic pigment cell research, with the goal of outlining current highlights, challenges, and frontiers associated with pigmentation and melanocyte biology. Significantly, this document encapsulates important advances in melanocyte and melanoma research including emerging frontiers in melanoma immunotherapy, medical and surgical oncology, dermatology, vitiligo, albinism, genomics and systems biology, epidemiology, pigment biophysics and chemistry, and evolution.