Faculty Bibliography

T-cell landscape differences between cutaneous squamous cell carcinoma (cSCC) tumors in immune competent (SCC in IC) and immunocompromised organ transplant recipients (TSCC in OTR) are unclear. We developed an analytical method to define tumor infiltrating lymphocyte (TIL) phenotype in cSCC from immune competent and immune suppressed patients using single-cell TCR sequencing and gene expression data. TSCC exhibits reduced proportions of cytotoxic and naïve TILs and similar numbers of regulatory TILs. Fewer, more heterogeneous TCR clonotypes are observed in TIL from OTR. Most TCR sequences for top ten clonotypes correspond to known antigens, while 24% correspond to putative neoantigens. OTR show increased cSCC events over 12 months possibly due to reduced cytotoxic T-cells. Our novel method of barcoding CD8+ T-cells is the first providing gene expression and TCR sequences in cSCC. Knowledge regarding putative antigens recognized by TCRs with phenotypic function of T-cells bearing those TCRs could facilitate personalized cSCC treatments.

BACKGROUND:Cemiplimab, a high-affinity, potent human immunoglobulin G4 monoclonal antibody to programmed cell death-1 demonstrated antitumor activity in a Phase 1 advanced cutaneous squamous cell carcinoma (CSCC) expansion cohort (NCT02383212) and the pivotal Phase 2 study (NCT02760498). Here we report the primary analysis of fixed dose cemiplimab 350 mg intravenously every 3 weeks (Q3W) (Group 3) and provide a longer-term update after the primary analysis of weight-based cemiplimab 3 mg/kg intravenously every 2 weeks (Q2W) (Group 1) among metastatic CSCC (mCSCC) patients in the pivotal study (NCT02760498). METHODS:The primary objective for each group was objective response rate (ORR) per independent central review (ICR). Secondary endpoints included ORR by investigator review (INV), duration of response (DOR) per ICR and INV, and safety and tolerability. RESULTS:For Group 3 (n=56) and Group 1 (n=59), median follow-up was 8.1 (range, 0.6 to 14.1) and 16.5 (range, 1.1 to 26.6) months, respectively. ORR per ICR was 41.1% (95% CI, 28.1% to 55.0%) in Group 3, 49.2% (95% CI, 35.9% to 62.5%) in Group 1, and 45.2% (95% CI, 35.9% to 54.8%) in both groups combined. Per ICR, Kaplan-Meier estimate for DOR at 8 months was 95.0% (95% CI, 69.5% to 99. 3%) in responding patients in Group 3, and at 12 months was 88.9% (95% CI, 69.3% to 96.3%) in responding patients in Group 1. Per INV, ORR was 51.8% (95% CI, 38.0% to 65.3%) in Group 3, 49.2% (95% CI, 35.9% to 62.5%) in Group 1, and 50.4% (95% CI, 41.0% to 59.9%) in both groups combined. Overall, the most common adverse events regardless of attribution were fatigue (27.0%) and diarrhea (23.5%). CONCLUSION/CONCLUSIONS:In patients with mCSCC, cemiplimab 350 mg intravenously Q3W produced substantial antitumor activity with durable response and an acceptable safety profile. Follow-up data of cemiplimab 3 mg/kg intravenously Q2W demonstrate ongoing durability of responses. TRIAL REGISTRATION NUMBER/BACKGROUND:Clinicaltrials.gov, NCT02760498. Registered May 3, 2016, https://clinicaltrials.gov/ct2/show/NCT02760498.

Background: TERT promoter mutations in various reports have been associated with poor patient survival in early stage melanomas emphasizing it as a separate subset of melanoma. Thus far, no studies investigated whether the immune composition of the tumor microenvironment (TME) in TERT (HS) mutant melanomas differs from TERT WT melanomas. Furthermore, the mechanism underlying the worse outcome in early stage TPHS melanomas remains unclear. Herein, we aim to characterize the tumor microenvironment (TME) of TERT promoter hotspot (TPHS) mutant melanomas & compare them to TERT WT melanomas in a cohort of early and advanced stage melanomas. We also aim to elucidate the clinical significance of the TME composition.
Design(s): We analyzed tissue from a cohort of 93 melanoma patients. DNA and RNA were extracted from primary and metastatic tumor tissue, resected prior to treatment with immune checkpoint inhibitors. The extracted DNA was genotyped using a customized next generation sequencing high throughput panel that targets 580 cancer-related genes to determine TPHS mutation status. Gene expression analysis was performed on the RNA from 52 patients using a customized 770-gene expression panel combining markers 48 biologically significant signatures with the N-counter system. Differential gene expression (DGE) and Gene set enrichment analysis (GSEA) were performed using R package [(p<0.01; FDR<0.01; FDR<0.30 for GSEA] using TERT WT as a reference.
Result(s): Table 1 illustrates the clinicopathological characteristics of the cohort. TPHS mutant melanoma was associated with downregulation of melanoma-associated antigens (MAGES) and endothelial cells/angiogenesis signature (p<0.01). Notably, MAGEA4, MAGEA1, CTAG1B, PALMD & KDR were among the most downregulated genes in the (lg2fc= -3.2; -2.2; -2.66, -1.23; - 0.66, respectively). GSEA showed a significant enrichment for NOD-like receptor (NLR) signaling pathway including NFKB1, TNF & NLR3P in TPHS mutant melanoma (Figure 1). Within TPHS mutant melanoma, high endothelial cells/angiogenesis signature (score >3.85/median) was more prevalent in stage (I/II) melanomas (p=0.025). No significant association between TERT mutational status, outcome nor histologic subtype were noted. (Table presented)
Conclusion(s): Our findings show that TPHS mutant melanoma and TERT WT have distinct TME composition. The higher endothelial/angiogenesis signature seen in early stage TPHS mutant melanoma compared to stage III/IV TPHS mutant melanomas could contribute to the poor outcome reported in the former group

BACKGROUND AND OBJECTIVES/OBJECTIVE:Metastasectomy for melanoma provides durable disease control in carefully selected patients. Similarly, BRAF-targeted and immune checkpoint inhibition has improved median overall survival (OS) in metastatic patients. We hypothesized that there is an increasing role for metastasectomy in melanoma patients responding to these therapies. METHODS:Retrospective analysis of a prospectively maintained database identified 128 patients with stage IV melanoma who received targeted molecular and/or checkpoint inhibitors at an academic institution from 2006 to 2017. Records were reviewed to characterize clinicopathologic characteristics, response to treatment, and intent of surgery for those who underwent metastasectomy. OS was analyzed by the Kaplan-Meier method. RESULTS:Median OS from stage IV diagnosis was 31.3 months. A total of 81 patients received checkpoint inhibitors, 11 received targeted inhibitors, and 36 received both. A total of 73 patients underwent metastasectomy. Indications for surgery included the intent to render disease-free (54%), palliation (34%), and diagnostic confirmation (11%). Responders to systemic therapy who underwent metastasectomy had improved OS compared to responders who did not (84.3 vs. 42.9 months, P = .018). CONCLUSIONS:Metastasectomy for melanoma is associated with improved OS in patients that respond to targeted molecular or immunotherapy. Resection should be strongly considered in this cohort as multimodality treatment results in excellent OS.

There have been several significant advances in cancer treatment in the last decade that are applicable to the treatment of melanoma and advanced nonmelanoma skin cancers. Among these are the development of immune checkpoint inhibitors targeting the programmed death protein-1 (PD-1)/programmed death legand-1 (PDL-1) axis, as well as targeted inhibitors of the BRAF/MEK signaling cascade in melanoma, and the hedgehog signaling pathway in basal cell carcinoma (BCC). These immune-based and targeted therapies have dramatically changed the treatment options for locally advanced and metastatic melanoma, Merkel's cell carcinoma, cutaneous squamous cell carcinoma (cSCC), and BCC. In this article, we will briefly review the currently approved targeted and immunotherapy-based treatments for locally advanced and metastatic melanoma, Merkel's cell carcinoma, and cSCC and discuss various combinations of approved therapies, as well as emerging therapeutic candidates that are currently in clinical trials, including novel checkpoint inhibitors in development, intratumoral oncolytic agents (viral and nonviral), and various immune-based therapies such as toll-like receptor (TLR) agonists, adoptive T-cell therapy, T-cell costimulation, and innate immune cell therapy. For advanced BCC, we will discuss trials investigating the currently approved smoothened (SMO) inhibitors for neoadjuvant use, emerging SMO inhibitors in development, topical SMO inhibitors, alternative targets in the hedgehog signaling pathway, and the use of anti-PD-1 agents for advanced BCC both alone and in combination with SMO inhibitors.

In the past 10 years, immune checkpoint inhibitors (ICIs) have become an additional pillar of cancer therapy by activating the immune system to treat a number of different malignancies. Many patients receiving ICIs develop immune-related adverse events (irAEs) that mimic some features of classical autoimmune diseases. Unfortunately, patients with underlying autoimmune conditions, many of whom have an increased risk for malignancy, have been excluded from clinical trials of ICIs due to a concern that they will have an increased risk of irAEs. Retrospective data from patients with autoimmune diseases and concomitant malignancy treated with ICIs are encouraging and suggest that ICIs may be tolerated safely in patients with specific autoimmune diseases, but there are no prospective data to guide management. In this manuscript, we review the relationship between pre-existing autoimmune disease and irAEs from checkpoint inhibitors. In addition, we assess the likelihood of autoimmune disease exacerbations in patients with pre-existing autoimmunity receiving ICI.

Mutational heterogeneity can contribute to therapeutic resistance in solid cancers. In melanoma, the frequency of inter- and intra-tumoral heterogeneity is controversial. We examined mutational heterogeneity within individual melanoma patients using multi-platform analysis of commonly mutated driver and non-passenger genes. We analyzed paired primary and metastatic tumors from 60 patients, and multiple metastatic tumors from 39 patients whose primary tumors were unavailable (n=271 tumors). We used a combination of multiplex SNaPshot assays, Sanger Sequencing, Mutation-specific PCR, or droplet digital PCR to determine the presence of BRAF^V600, NRAS^Q61, and TERT^-124C>T and TERT^-146C>T mutations. Mutations were detected in BRAF (39%), NRAS (21%) and/or TERT (78%). Thirteen patients had TERT^mutant discordant tumors; seven of these had a single tumor with both TERT^-124C>T and TERT^-146C>T mutations present at different allele frequencies. Two patients had both BRAF and NRAS mutations; one in different tumors and the other had a single tumor with both mutations. One patient with a BRAF^mutant primary lacked mutant BRAF in least one of their metastases. Overall, we identified mutational heterogeneity in 18/99 (18%) patients. These results suggest that some primary melanomas may be comprised of subclones with differing mutational profiles. Such heterogeneity may be relevant to treatment responses and survival outcomes.

BACKGROUND:The American Joint Committee on Cancer (AJCC) maintains that the eighth edition of its Staging Manual (AJCC8) has improved accuracy compared to the seventh (AJCC7). However, there are concerns that implementation may disrupt analysis of active clinical trials for stage III patients. We used an independent cohort of melanoma patients to test the extent to which AJCC8 has improved prognostic accuracy compared to AJCC7. METHODS:We analyzed a cohort of 1,315 prospectively enrolled patients. We assessed primary tumor and nodal classification of stage I-III patients using AJCC7 and AJCC8 to assign disease stages at diagnosis. We compared recurrence-free (RFS) and overall survival (OS) using Kaplan-Meier curves and log-rank tests. We then compared concordance indices of discriminatory prognostic ability and area under the curve (AUC) of 5-year survival to predict RFS/OS. All statistical tests were two-sided. RESULTS:Stage IIC continued to have worse outcomes than those for stage IIIA patients, with 5-year RFS of 26.5% (95%CI=12.8-55.1%) vs. 56.0% (95%CI=37.0-84.7%) by AJCC8 (P = 0.002). For stage I, removing mitotic index as T classification factor decreased its prognostic value, although not statistically significantly (RFS C-index=0.63 [95%CI=0.56-0.69] to 0.56 [95%CI=0.49-0.63], P = 0.07; OS C-index=0.48 [95%CI=0.38-0.58] to 0.48 [95%CI=0.41-0.56], P = 0.90). For stage II, prognostication remained constant (RFS C-index=0.65 [95%CI=0.57-0.72]; OS C-index=0.61 [95%CI=0.50-0.72]), and. For stage III, AJCC8 yielded statistically significantly enhanced prognostication for RFS (C-index=0.65 [95%CI=0.60-0.70] to 0.70 [95%CI=0.66-0.75], P = 0.01). CONCLUSIONS:Compared with AJCC7, we demonstrate that AJCC8 enables more accurate prognosis for patients with stage III melanoma. Restaging a large cohort of patients can enhance the analysis of active clinical trials.

PURPOSE/OBJECTIVE:To report the 5-year overall survival (OS) landmark and the long-term safety profile of vemurafenib plus cobimetinib (BRAF plus MEK inhibition, respectively) in the BRIM7 study. PATIENTS AND METHODS/METHODS:= 66]. Patients in the dose-escalation phase received vemurafenib at 720 or 960 mg twice daily in combination with cobimetinib at 60, 80, or 100 mg/d for 14 days on/14 days off, 21 days on/7 days off, or continuously. Two regimens were selected for expansion: vemurafenib (720 and 960 mg twice daily) and cobimetinib (60 mg/d 21/7). RESULTS:Median OS was 31.8 months [95% confidence interval (CI), 24.5-not estimable] in the BRAFi-naïve cohort. The landmark OS rate plateaued at 39.2% at years 4 and 5 of follow-up. In the vemurafenib monotherapy-PD cohort, the median OS was 8.5 months (95% CI, 6.7-11.1), and the landmark OS rate plateaued at 14.0% from 3 years of follow-up. No increase was observed in the frequency and severity of adverse events with long-term follow-up. No new toxicities were detected, and there was no increase in the frequency of symptomatic MEK inhibitor class-effect adverse events. CONCLUSIONS:-mutated melanoma treated with a combination regimen of vemurafenib and cobimetinib achieve favorable long-term outcomes.

Given its ability to induce both humoral and cellular immune responses, NY-ESO-1 has been considered a suitable antigen for a cancer vaccine. Despite promising results from early-phase clinical studies in melanoma patients, NY-ESO-1 vaccine immunotherapy has not been widely investigated in larger trials; consequently, many questions remain as to the optimal vaccine formulation, predictive biomarkers, and sequencing and timing of vaccines in melanoma treatment. We conducted an adjuvant phase I/II clinical trial in high-risk resected melanoma to optimize the delivery of poly-ICLC, a TLR-3/MDA-5 agonist as a component of vaccine formulation. A phase I dose escalation part was undertaken to identify the maximum tolerated dose of poly-ICLC administered in combination with NY-ESO-1 and montanide. This was followed by a randomized phase II part investigating the maximum tolerated dose of poly-ICLC with NY-ESO-1 with or without montanide. The vaccine regimens were generally well-tolerated, with no treatment-related grade 3/4 adverse events. Both regimens induced integrated NY-ESO-1-specific CD4+ T-cell and humoral responses. CD8+ T-cell responses were mainly detected in patients receiving montanide. T-cell avidity towards NY-ESO-1 peptides was higher in patients vaccinated with montanide. In conclusion, NY-ESO-1 protein in combination with poly-ICLC is safe, well-tolerated, and capable of inducing integrated antibody and CD4+ T-cell responses in most patients. Combination with montanide enhances antigen specific T-cell avidity and CD8+ T-cell cross-priming in a fraction of patients, indicating that montanide contributes to the induction of specific CD8+ T-cell responses to NY-ESO-1.