Domingo‐Musibay, Evidio

9503 Background: Neoadjuvant ± adjuvant immunotherapy improves event-free survival relative to adjuvant immunotherapy alone for patients with high-risk resectable stage III melanoma. However, the optimal regimen balancing efficacy and tolerability is not known. T-cell immunoglobulin and ITIM domain (TIGIT) is a promising immune checkpoint but its therapeutic potential in stage III melanoma is underexplored. Methods: In this phase II trial, patients with resectable, macroscopic stage III melanoma received four 21-day neoadjuvant cycles of 1200mg IV atezolizumab (atezo, anti-PD-L1) + 600mg IV tiragolumab (tira, anti-TIGIT), followed by therapeutic lymph node dissection (TLND) and eight 21-day adjuvant cycles of 1200mg IV atezo. Primary endpoints were pathologic response (of all patients initiating neoadjuvant therapy) and recurrence-free survival (RFS) from the time of TLND in patients who were operated on per protocol and received adjuvant therapy. Secondary endpoint was adverse events (AEs); exploratory endpoints included event-free survival (EFS) and distant metastasis-free survival (DMFS). Results: Thirty-four patients, median age 59 years, were accrued and initiated neoadjuvant atezo/tira. 76.5% had >1 metastatic lymph node involved at baseline and 73.5% presented with Stage IIIC disease. All 34 patients were evaluable for AEs, pathologic response and EFS. Four patients were diagnosed with metastatic disease during neoadjuvant treatment, while 30 had TLND per protocol and 28 received adjuvant treatment and were evaluable for RFS and DMFS. Major pathologic responses (MPR, ≤10% viable tumor) were observed in 16/34 patients (47.1%, Table). With 19.9 months median follow-up from registration, 12-month EFS was 72.0% (95% CI 57.9 to 89.5%). With 16 months median follow-up from operation, 12-month RFS was 73.3% (n=28, 95% CI 56.9 to 94.5%), while 12-month DMFS was 86.0% (n=28, 95% CI 72.2 to 100%). In the 16 patients with an MPR, 2 did not receive adjuvant treatment and were followed for 33.3 and 10.8 months without recurrence/death. In the remaining 14 patients, 12-month RFS and 12-month DMFS were both 91.7% (95% CI 77.3 to 100%). 5 patients (14.7%) experienced any grade 3+ AE with 2 (5.9%) at least possibly related to the neoadjuvant regimen. Conclusions: Among patients with high-risk resectable stage III melanoma, neoadjuvant atezo/tira was a promising regimen with a favorable safety profile and warrants further study. Identification of predictive biomarkers will allow for optimization of neoadjuvant therapy for individual patients. Clinical trial information: NCT03554083 . Pathologic response. N=34 Major Pathologic Response 16 (47.1%) Pathologic complete response (no viable tumor) 13 (38.2%) Near-pathologic complete response (0.1−10% viable tumor) 3 (8.8%) Pathologic partial response (>10.0−50% viable tumor) 2 (5.9%) Pathologic non-response (>50% viable tumor) 12 (35.3%) No per protocol operation 4 (11.8%)

Background Neoadjuvant treatment has become standard for patients with high-risk operable stage III melanoma, but the optimal regimen is unknown. Targeted therapy approaches yield high pathological response rates, while immunotherapy regimens show favorable recurrence-free survival (RFS). NeoACTIVATE was designed to address whether a neoadjuvant combination of both targeted therapy and immunotherapy might leverage the benefits of each. Methods We tested neoadjuvant treatment with 12 weeks of vemurafenib, cobimetinib, and atezolizumab for patients with BRAF-mutated (BRAFm) melanoma (cohort A) and cobimetinib and atezolizumab for patients with BRAF-wild-type (BRAFwt) melanoma (cohort B), regimens which we have shown generate a substantial major pathological response. After therapeutic lymph node dissection, patients received 24 weeks of adjuvant atezolizumab. Here, we report survival outcomes and their association with biomarkers assayed among the gut microbiome and peripheral blood immune subsets. Results With 49 months median follow-up, the median RFS was not reached for cohort A and was 40.8 months for cohort B. At 24 months after operation, 2 of 14 cohort A patients and 4 of 13 cohort B patients had experienced distant relapse. Key findings from correlative analyses included diversity, taxonomic and functional metagenomic gut microbiome signals associated with distant metastasis-free survival at 2 years. Notably, we observed a strong correlation between low microbial arginine biosynthesis (required for T-cell activation and effector function) and early distant recurrence (p=0.0005), which correlated with taxonomic differential abundance findings. Peripheral blood immune monitoring revealed increased double-positive (CD4+CD8+) T cells in patients with early recurrence. Conclusions Neoadjuvant treatment with cobimetinib and atezolizumab±vemurafenib was associated with a low rate of distant metastasis in patients with high-risk stage III melanoma. Freedom from early distant metastasis was highly associated with taxonomic differences in gut microbiome structure and with functional pathway alterations known to modulate T cell immunity. Identification of predictive biomarkers will permit optimization of neoadjuvant therapy regimens for individual patients. Trial registration number NCT03554083.

Abstract Purpose: Mucosal melanoma is a rare, aggressive form of melanoma with extremely high recurrence rates despite definitive surgical resection with curative intent. Currently there is no consensus on adjuvant therapy. Data on checkpoint inhibitors for adjuvant therapy are lacking. Patients and Methods: We performed a single-arm, multicenter clinical trial using “flip dose” ipilimumab (1 mg/kg q3w × 4 cycles), and nivolumab (3 mg/kg q3w × 4 cycles), then nivolumab 480 mg q4w × 11 cycles to complete a year of adjuvant therapy. Participants must have had R0/R1 resection ≤90 days before registration, no prior systemic therapy (adjuvant radiotherapy allowed), ECOG 0/1, and no uncontrolled autoimmune disease or other invasive cancer. Patients were recruited through the Midwest Melanoma Partnership/Hoosier Oncology Network. Results: From September 2017 to August 2021, 35 patients were enrolled. Of these, 29 (83%) had R0 resections, and 7 (20%) received adjuvant radiotherapy. Median age was 67 years, 21 (60.0%) female. Recurrence-free survival (RFS) rates at 1 and 2 years were 50% [95% confidence interval (CI), 31%–66%] and 37% (95% CI, 19%–55%), respectively. Overall survival rates at 1 and 2 years were 87% (95% CI, 68%–95%) and 68% (95% CI, 46%–83%), respectively. Median RFS was 10.3 months (95% CI, 5.7–25.8). Most common grade 3 toxicities were diarrhea (14%), hypertension (14%), and hyponatremia (11%), with no grade 4/5 toxicities. Conclusions: Flip-dose ipilimumab and nivolumab after resection of mucosal melanoma is associated with outcomes improved over that of surgical resection alone. Long-term follow-up, subgroup analyses and correlative studies are ongoing.

Abstract Background: Uveal melanoma (UM) is a rare tumor of the eye which is biologically distinct from cutaneous melanoma (CM) and carries a poor prognosis once metastatic. We aimed to examine a multi-omics profile of UM as compared to CM. Methods: A total of 277 UM and 1,297 CM metastatic or primary samples were analyzed using next-generation sequencing (NextSeq, 592 genes and WES, NovaSeq, >700 genes) and whole transcriptome sequencing (WTS; NovaSeq) (Caris Life Sciences). Tumor mutational burden (TMB) was measured by totaling somatic mutations per tumor (TMB-high cutoff >10 mutations per MB). Immune cell fraction was calculated by QuantiSeq. T- cell inflamed score was calculated using gene expression data. Pathway gene enrichment analysis using WTS data was performed using GSEA. Biostatistical significance was determined using chi-square/Fisher-Exact or Wilcoxon rank sum test and adjusted for multiple comparisons. Results: For all cases (primary and metastatic), UM compared to CM had a higher rate of GNAQ/11 (93.8% vs. 2.1% p≤0.001), SF3B1 (20.4% vs 3% p≤0.001), and BAP1 (52.9% vs. 1.5% p≤0.001) mutations. UM also had higher levels of MYC (7% vs 0.9% p≤0.001), NCOA2 (3.1% vs 0.1% p≤0.001), and RUNX1T1 genomic amplifications (2.3% vs 0.2% p≤0.001). Assessing only samples of liver metastases, UM was shown to have higher abundance of M2 macrophages (P≤0.0001) and less abundance of M1 macrophages(P≤0.0001), monocytes(P≤0.0001), CD4 T-cells(P≤0.0001), and regulatory T-cells (p = 0.0003) as compared to CM. GSEA comparison between liver metastases of UM and CM samples showed G2M checkpoint (NES score, 1.60 p≤0.0001) and E2F targets (NES score, 1.58 p=0.002) to be higher in metastatic CM. When comparing T-cell inflamed, intermediate, and non-inflamed tumors from any site, CM or UM, the average Z scores for both G2M and E2F were highest for inflamed and lowest for non-inflamed, P≤0.0001. A similar trend of higher Z score was seen for TMB high vs TMB low tumors, p≤0.0001. Conclusions: UM and CM are characterized by significantly different genomic and transcriptomic alterations, and cellular composition of TME. Further understanding of these characteristics may lead to better treatment strategies for metastatic UM. Citation Format: Justin C. Moser, Yusra F. Shao, Joanne Xiu, Yasmine Baca, Takami Sato, Lauren A. Dalvin, Sourat Darabi, Michael Korn, Burton Eisenberg, Geoff Gibney, Evidio Domingo-Musibay, Gino In, Dave Hoon, Michael S. Gordon. Multi-omics and immune cell profiling of metastatic uveal melanomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 50.

9573 Background: Mucosal melanoma is a rare, highly aggressive form of melanoma with extremely high recurrence rates, despite definitive surgical resection. Median RFS has been reported to be 5.4m, with RFS rates at 1 and 2 years of 10%, and 0%, respectively (Lian B, Si L, Cui C, et al. Phase II Randomized Trial Comparing High-Dose IFN-α2b with Temozolomide Plus Cisplatin as Systemic Adjuvant Therapy for Resected Mucosal Melanoma. Clinical Cancer Research 2013, 19(16):4488-4498). Currently there is no consensus on recommendations for adjuvant therapy. Data on the use of immune checkpoint inhibitors (ICI) adjuvantly is lacking. Methods: We performed a single arm, multicenter clinical trial using “flip dose” ipilimumab (1mg/kg q3w x4 cycles),and nivolumab (3 mg.kg q3w x4 cycles), then Nivolumab 480 mg q4w x 11 cycles to complete a year of adjuvant therapy. The primary endpoint was recurrence-free survival (RFS), and the study had 85% power to detect an improvement in RFS between 5.5 and 9.5 months using a one-sided log rank test. Participants must have had R0/R1 resection <90 days prior to registration, and no prior systemic therapy (adjuvant radiation allowed), ECOG 0/1, no uncontrolled significant autoimmune disease or other invasive cancer. Patients were recruited through the Midwest Melanoma Partnership/Hoosier Oncology Network. Results: From 9/17 to 8/21, 44 patients were approached at 6 centers. Of these 9 were ineligible, and 35 were enrolled. Of these, 29 (83%) had R0 resections, and 7 (20%) had adjuvant radiation prior to enrollment. As of Dec 2021, 31 patients have completed the treatment phase. Of the 35 patients treated on study, 20 patients have recurred (7 local, 5 distant, 3 regional, 5 sites unconfirmed), 6 stopped therapy due to adverse effects, and 8 have died. The mean age of patients was 65.8 years and 21 (60.0%) were female. The primary site of disease was vulvovaginal N=12 (32.4%) patients, sinonasal N= 11 (29.7%), anorectal N= 9 (24.3%) and other site N= 5 (13.5%). Adjuvant radiation had been given in 7 pts. Driver mutations were rare, with only 3 (8.6%) patients having a KIT mutation, and one patient (2.9%) each having a NRAS or BRAF mutation. RFS rates at 1 and 2 years were 50% (95% CI 31-66%) and 37% (95% CI 19-55%), with OS rates at 1 and 2 years of 87% (95% CI 68-95%) and 68% (95% CI 46-83%). Median RFS was 10.3 m (95% CI5.7-25.8). Most common grade 3 adverse events were diarrhea (14%), hypertension (14%), hyponatremia (11%), with no grade 4/5 toxicities. Conclusions: Flip dose ipilimumab and nivolumab after resection is associated with outcomes improved over previously reported outcomes in the absence of therapy. Long term follow up is ongoing as are subgroup analyses and correlative studies. Clinical trial information: NCT03241186.

10006 Background: Treatment options are limited for patients with advanced melanoma who have progressed on checkpoint inhibitors and targeted therapies. Adoptive cell therapy using tumor-infiltrating lymphocytes (TIL) leverages and enhances the body’s natural defense against cancer and has shown durable responses in heavily pretreated melanoma patients. Methods: C-144-01 is a global Phase 2 open-label, multicenter study of efficacy & safety of lifileucel in patients with unresectable metastatic melanoma who have progressed on checkpoint inhibitors and BRAF/MEK inhibitors, if BRAFv600 mutant. We report on Cohort 2 (N = 66) patients who have received TIL. Tumors were resected at local institutions, processed in central GMP facilities for TIL production, manufactured, cryopreserved & shipped back to sites in a 22-day process. Therapy consisted of one week of lymphodepletion, a single lifileucel infusion, and up to 6 IL-2 doses. ORR was based on RECIST v1.1 by investigator assessment. Data cutoff was Feb 2, 2020. Results: Baseline characteristics: 3.3 mean prior therapies (anti-PD1 100%; anti-CTLA-4 80%; BRAF/MEK inhibitor 23%), high baseline tumor burden (106 mm mean target lesion sum of diameters), 44% liver/brain lesions, 40.9% LDH > ULN. ORR by investigator was 36.4% (2 CR, 22 PR) and DCR was 80.3%. Mean time to response was 1.9 months (range: 1.3-5.6). After a median study follow-up of 17.0 months, median DOR (mDOR) was still not reached. Six responders have progressed, 2 have died and 2 started other anti-cancer therapy without progression. The adverse event profile was consistent with the underlying advanced disease and the lymphodepletion and IL-2 regimens. Additional follow-up data will be available for presentation. Conclusions: Lifileucel treatment results in a 36.4% ORR and mDOR was not reached at 17.0 months of median study follow up in a heavily pretreated metastatic melanoma patients with high baseline disease burden who progressed on multiple prior therapies, including anti-PD1 and BRAF/MEK inhibitors, if BRAFv600 mutant. Clinical trial information: NCT02360579.

TPS65 Background: Mucosal melanoma is a rare and, therefore, poorly studied malignancy. Patients with resected primary tumors have a median time to recurrence of 5.5 months. While no one adjuvant therapy is of proven benefit for patients, immunotherapy has shown activity in metastatic mucosal melanoma. Methods: We are performing a single arm, phase II clinical trial through the Midwest Melanoma Partnership/Hoosier Cancer Research Network (6 sites) for resected mucosal melanoma. The primary endpoint is recurrence free survival (RFS), and it will include 36 subjects. Patients must have had an R0 or R1 resection of a mucosal melanoma (sinonasal, anorectal, vulvar, or other), and register within 90 days of surgery. Adjuvant radiation is allowed prior to registration but not required. Therapy consists of ipilimumab (1 mg/kg) and nivolumab (3 mg/kg) IV q3w x 4, then 480 mg nivolumab x 1 year. Statistical power is calculated to be 85% to detect a change in RFS from 5.5 months to 9.5 months, with a one sided alpha of 0.05. Enrollment began Sept 2017, with current participation from 6 large volume centers in the United States, with 17 patients enrolled as of Sept 2019. Full accrual is anticipated to complete by year end 2020. Patients will be followed for RFS and OS. Translational studies will include mutational burden, c-kit, BRAF, NRAS status, and serum markers of immunity -- including soluble PD-L1 and Bim. Clinical trial information: NCT03241186.

26 Background: NKTR-262 is a novel intratumoral prodrug designed to promote tumor antigen release, an immune stimulatory environment, and antigen presentation. NKTR-214 increases CD8+ T cells and NK cells in the tumor microenvironment. The combination of NKTR-262 with NKTR-214 accesses innate and adaptive immunity to provide an abscopal response and systemic tumor immunity. The Phase 1b/2 REVEAL trial is enrolling. Methods: In a 3+3 Phase 1 design, pts who are relapsed/refractory to ≥1 CPI receive escalating doses of intratumoral (IT) NKTR-262 followed by combined q3w treatment of IT NKTR-262 and fixed-dose intravenous (IV) NKTR-214 (0.006 mg/kg). Phase 1 will evaluate PK, safety, efficacy, and determine recommended Phase 2 dose. Scans are conducted every 9 wks (± 1 wk). Peripheral blood (PB) and tumor tissue are collected to measure biomarkers of immune activation. In Phase 2 expansion, NKTR-262 + NKTR-214 ± IV nivolumab (360 mg) will be assessed in specific tumor cohorts of IO-naïve and experienced pts. Results: As of 06 Nov 2018, 11 pts had received at least one cycle of NKTR-262 + NKTR-214 therapy. 7/11 pts were efficacy evaluable with at least one on-treatment scan (4 MEL, 2 SARC, 1 CRC). 2/4 R/R melanoma pts had uPRs with 100% (scan 2) and 40% (scan 1) reductions in target lesions per RECIST 1.1, respectively. Both patients remain on treatment. Two SARC pts had stable disease at scan 1. No pts experienced G3 or higher TRAEs or immune-related adverse events (AEs). Most common G1-2 AEs were transient flu-like symptoms. Gene expression analysis of tumor and PB show dose-dependent induction (4 to 16-fold) of TLR target genes. Consistent with the mechanism of action for NKTR-214, PB flow cytometry shows an average 13-fold increase from baseline in Ki67+ CD8+T cells in 6 pts. Conclusions: NKTR-262 + NKTR-214 engage the immune activation cascade from local tumor antigen production to anti-tumor T cell response. Initial dose levels of NKTR-262 with fixed-dose NKTR-214 were well-tolerated with early evidence of clinical activity. These data support continued evaluation of the combination +/- nivolumab. Clinical trial information: NCT03435640.

Abstract Lessons Learned Percutaneous thermal ablation combined with in situ granulocyte-macrophage colony-stimulating factor cytokine therapy was technically feasible and well tolerated. No significant clinical or immunologic responses were seen. Background Melanoma tumor-derived heat-shock proteins (HSPs) and HSP-peptide complexes can elicit protective antitumor responses. The granulocyte-macrophage colony-stimulating factor (GM-CSF) chemokine can also promote uptake and processing by professional antigen presenting cells (APCs). On this basis, we designed a pilot study of percutaneous thermal ablation as a means to induce heat-shock protein vaccination plus GM-CSF to determine safety and preliminary antitumor activity of this combination. Materials and Methods This study was designed to assess overall safety of percutaneous ablation combined with GM-CSF for unresectable, metastatic melanoma including uveal and mucosal types. All patients received heat-shock therapy (42°C for 30 minutes), then received one of three treatments: (a) intralesional GM-CSF (500 mcg standard dose); (b) radiofrequency ablation (RFA) + GM-CSF; or (c) cryoablation plus GM-CSF. The primary endpoint of the study was the induction of endogenous HSP70 and melanoma-specific cytotoxic T lymphocytes (CTL). Results Nine patients (three per study arm) were enrolled. No dose-limiting toxicity was observed as specified per protocol. All patients developed progressive disease and went on to receive alternative therapy. Median overall survival (OS) was 8.2 months (95% confidence interval [CI] 2–17.2). The study was not powered to detect a difference in clinical outcome among treatment groups. Conclusion Percutaneous thermal ablation plus GM-CSF was well tolerated, technically feasible, and demonstrated an acceptable adverse event profile comparable to conventional RFA and cryoablation. While HSP70 was induced following therapy, the degree of HSP70 elevation was not associated with clinical outcome or induced CTL responses. While percutaneous thermal ablation plus GM-CSF combinations including checkpoint inhibitors could be considered in future studies, the use of GM-CSF remains experimental and for use in the context of clinical trials.

Background:Myoepithelioma of the soft tissues is a rare entity and little is known about how best to manage locally recurrent and high-grade disease. Here, we retrospectively examined outcomes of surgery, chemotherapy, and radiation therapy (RT) for treatment of low-grade and high-grade myoepithelioma of soft tissues.Methods:We reviewed 20 cases of myoepithelioma of soft tissues seen at Mayo Clinic between 1994 and 2014. The effect of histologic grade and therapies received on relapse and survival were assessed.Results:We identified 13 patients with low-grade disease and 7 patients with high-grade disease. We found that low-grade disease was frequently effectively managed with surgical resection alone, whereas high-grade disease frequently metastasized and was often fatal. The 5-year event-free survival was 88% (confidence interval, 46%-98%) for low-grade disease versus 36% (confidence interval, 7%-75%;P=0.04) for high-grade disease. The relapse rate in low-grade disease was 29% at 5 years versus 64% (P=0.04) in high-grade disease. No significant responses to chemotherapy were noted, however, excellent responses to perioperative RT were seen.Conclusions:Surgery continues as the primary modality of treatment for myoepithelioma of soft tissues. Our study did not show a clear benefit of chemotherapy in the metastatic disease setting, but supports the use of perioperative RT in the management of high-grade disease; further investigation is warranted.