3053 Background: Circulating tumor DNA (ctDNA) is an emerging biomarker for assessing treatment outcomes and monitoring disease recurrence in melanoma. However, evidence supporting its clinical utility at very early on-treatment timepoints remains limited. We evaluated whether ctDNA dynamics relative to a single cycle of immune checkpoint blockade (ICB) are associated with treatment outcomes in patients (pts) with advanced melanoma. Methods: In this prospective, single-center cohort, pts with unresectable stage III or IV melanoma were enrolled. Treatment included anti-PD1 monotherapy (n=22, 40%) or anti-PD1+anti-CTLA-4 therapy (n=33, 60.0%). ctDNA analysis was performed using a personalized, tumor-informed assay (Signatera Genome, Natera, Inc.) designed from WGS of matched tumor and normal pairs. ctDNA (reported in mean tumor molecules per mL, MTM/mL) was subsequently tracked in the pts’ longitudinal blood samples, which were collected at baseline (T0) and prior to cycle 2 (T1; 3-4 weeks after ICB initiation). ctDNA dynamics were categorized as favorable (ctDNA clearance at T1 or >10-fold decrease from T0 to T1) or unfavorable (rising ctDNA or <10-fold decrease from T0 to T1). Associations between these categories and progression-free survival (PFS) and overall survival (OS) were assessed using Kaplan-Meier estimates and Cox proportional hazards models. Results: A total of 55 pts with unresectable stage III (n=12, 21.8%) or stage IV (n=43, 78.2%) melanoma were included. Median follow-up was 36.43 months (mo) (range, 31.47-41.39). ctDNA-positivity was 83.6% (46/55) at baseline and 80.0% (44/55) at T1. Table 1 summarizes survival outcomes per ctDNA profile. Pts with an unfavorable profile (UP) of ctDNA dynamics experienced worse PFS and OS in contrast to those with a favorable profile (FP). Median PFS was 2.1 mo in the UP group versus 28.8 mo in the FP group; median OS was 19.3 vs 37.8 mo, respectively. All pts (n=21, 100%) with progressive disease had an UP, including 52.3% (11/21) with rising ctDNA and 47.7% (10/21) with <10-fold decrease. Median ctDNA levels at T1 were also higher in pts without objective response compared with responders (56.69 vs 0.38 MTM/mL; p<0.001). Conclusions: ctDNA dynamics assessed by a personalized WGS-based assay after a single cycle of ICB were strongly associated with response and survival outcomes in advanced melanoma. These findings support the evaluation of treatment-adaptation strategies based on early ctDNA dynamics in pts receiving immunotherapy. ctDNA profile median PFS (mo) 6-mo PFS 12-mo PFS PFS HR (95%CI)p-value median OS (mo) 6-mo OS 12-mo OS OS HR (95%CI)p-value FavorableN=24 28.8 100% (24/24) 91.7% (22/24) Reference 37.8 100% (24/24) 100% (24/24) Reference UnfavorableN=31 2.1 22.6% (7/31) 22.6% (7/31) 5.29(2.39–11.75)p<0.001 19.3 80.6% (25/31) 67.7% (21/31) 9.96(2.66–37.37)p<0.001

Intravenous administration of anti-CTLA4 with anti-PD1 provides durable tumour responses but causes severe treatment-related adverse events in patients with cancer1. Intratumoural administration at lower doses but high local concentrations could enhance antitumour efficacy while minimizing systemic exposure and toxicity. Here we report the randomized multicentre phase 1b NIVIPIT trial (ClinicalTrials.gov: NCT02857569 ), which enrolled 61 patients with untreated metastatic melanoma, randomly assigned 2:1 to receive intravenous nivolumab (anti-PD1; 1 mg kg-1) combined with either intratumoural ipilimumab (anti-CTLA4; 0.3 mg kg-1) or intravenous ipilimumab (3 mg kg-1). The primary end-point was met with significantly lower incidence of grade 3 or 4 treatment-related adverse events at 6 months in the intratumoural versus intravenous arm (22.6% versus 57.1%), equivalent to anti-PD1 monotherapy. RECIST (response evaluation criteria in solid tumours) best objective response rate reached 65.7% for anti-CTLA4 injected lesions and 50% for uninjected lesions, confirming the relationship between intratumoural exposure to anti-CTLA4 and efficacy. Baseline tumour immune profiling revealed that protumoural activated regulatory T (Treg) cells and M2 macrophages predict durable clinical benefit, regardless of the anti-CTLA4 administration route. A decrease in activated intratumoural Treg cells occurred only in patients who showed durable clinical benefit, who also presented high intratumoural Fcγ receptor (FcγR) expression. Our results provide a rationale for intratumoural anti-CTLA4 strategies in oligometastatic and early-stage cancers and indicate that high intratumoural activated Treg cell and FcγR+ M2 macrophage numbers are prerequisites for efficacy of combined anti-CTLA4 and anti-PD1.

BACKGROUND:Current first-line treatment for patients with metastatic melanoma with BRAFV600E or BRAFV600K mutations includes immunotherapy with immune checkpoint inhibitors and targeted therapy; however, the optimal sequencing of these treatments is unclear. We aimed to investigate the use of a targeted-therapy induction regimen before treatment with immune checkpoint inhibitors. METHODS:This open-label, randomised, controlled, phase 2 trial (EBIN) was conducted at 37 centres in eight European countries. Eligible patients were 18 years or older and had previously untreated, unresectable, stage III or IV melanoma with BRAFV600E or BRAFV600K mutations and an Eastern Cooperative Oncology Group performance status of 0 or 1. Patients were randomly assigned (1:1) to one of two groups. Those in the induction group received targeted therapy (oral encorafenib 450 mg once a day plus oral binimetinib 45 mg twice a day for 12 weeks) followed by immune checkpoint inhibitors (intravenous nivolumab 3 mg/kg plus intravenous ipilimumab 1 mg/kg once every 3 weeks for four doses, followed by intravenous nivolumab 480 mg once every 4 weeks until unacceptable toxicity, disease progression, or 2 years of treatment). Patients in the control group received immune checkpoint inhibitors as above without any induction targeted therapy. Randomisation was conducted using a minimisation technique and was stratified by centre and a variable defined using stage and lactate dehydrogenase activity. The primary outcome was progression-free survival in the intention-to-treat population. Safety was assessed in all patients who initiated the protocol treatment. In this Article we report the primary analysis. The study is registered with ClinicalTrials.gov, NCT03235245, and is ongoing. FINDINGS:Between Nov 12, 2018, and July 11, 2022, 271 patients were randomly assigned: 136 to the induction group and 135 to the control group. 103 (38%) patients were female, 168 (62%) were male, and the median age was 55 years (IQR 43-66). The median follow-up time was 21 months (IQR 13-33). There was no evidence of a longer progression-free survival in the induction group than in the control group (hazard ratio 0·87, 90% CI 0·67-1·12; p=0·36). The median progression-free survival was 9 months (95% CI 7-13) in the induction group and 9 months (5-14) in the control group. Grade 3-5 treatment-related adverse events occurred in 57 (42%) of 136 patients who started treatment in the induction group and in 42 (32%) of 131 patients who started treatment in the control group. The most common grade 3-4 treatment-related adverse event was hepatitis (17 [13%] of 136 patients in the induction group and nine [7%] of 131 patients in the control group). Serious treatment-related adverse events occurred in 45 (33%) of 136 patients in the induction group and 33 (25%) of 131 patients in the control group. There were three treatment-related deaths: two from cardiac events (heart failure and arrhythmia) in the induction group and one from meningitis in the control group. INTERPRETATION:The targeted-therapy induction regimen did not improve progression-free survival compared with first-line treatment with immune checkpoint inhibitors in unselected patients with advanced melanoma with BRAFV600E or BRAFV600K mutations. FUNDING:Bristol Myers Squibb and Pierre Fabre.

Representativeness of Study Participants

Washout, response to protocol therapy, AEs

Immunohistology: Response at time of biopsy, tissue, date biopsies, previous CPI and protocol CPI, elapse times, TILs and macrophages phenotypes.

BACKGROUND:Immune checkpoint inhibitors (ICIs) have revolutionized melanoma treatment, with programmed cell death protein 1 (PD-1) inhibitors-alone or in combination with cytotoxic T-lymphocyte-associated protein 4 or lymphocyte-activation gene 3 inhibitors-demonstrating significant efficacy. However, there is a critical lack of robust data to determine the optimal sequencing of these therapies for individual patients. In particular, the role of relatlimab+nivolumab (rela/nivo) within treatment sequences remains poorly defined. Choosing the right sequence is strategic, as an inappropriate order may compromise the effectiveness of subsequent treatments and limit long-term benefits. METHODS:This multicenter retrospective and prospective study evaluated 190 patients across three treatment arms: rela/nivo followed by ipilimumab+nivolumab (ipi/nivo) (arm A, N=40), ipi/nivo followed by rela/nivo (arm B, N=71), and anti-PD-1 followed by rela/nivo (arm C, N=79). The study assessed the impact of treatment sequencing on outcomes including response rate, progression-free survival, and overall survival (OS). RESULTS:The overall response rate to second treatment was highest in arm C (30.4%), followed by arm B (28.1%) and arm A (17.5%). Patients with secondary resistance to first-treatment ICIs had better responses to second-treatment ICIs than those with primary resistance, particularly in arm B (p=0026). Median OS from date of first ICI treatment was significantly longer in arms B (40.9 months) and C (42.5 months) compared with arm A (16.3 months). CONCLUSIONS:Our findings indicate that rela/nivo may remain active following anti-PD-1 or ipi/nivo therapy. Additionally, our results suggest that sequencing ipi/nivo before rela/nivo may yield better outcomes than starting with rela/nivo. Patients who respond to the first combination regimen appear to derive greater benefit from the second. Further efforts are needed to optimize sequencing strategies in advanced melanoma, and future studies should consider the impact of prior treatment outcomes.

Age, localisation primary, stage, TNM

Previous therapies and outcome

LBA9503 Background: The benefit of an induction treatment with targeted therapy (TT) with BRAF+MEK inhibitors prior to a combined immunotherapy (IT) with ipilimumab (ipi) + nivolumab (nivo) in patients (pts) with advanced BRAF-V600E/K mutant melanoma is still unclear. Methods: EBIN is an international randomized controlled phase II trial comparing upfront IT (arm A: nivo [3mg/kg] + ipi [1mg/kg] q3w x4 followed by nivo 480 mg q4w) with the sequential approach (arm B: 3 months induction with TT with encorafenib 450 mg QD + binimetinib 45 mg BID orally, followed by IT using the same regimen as in arm A), total treatment [Tx] duration in both arms: 2 years. In arm B, pts were allowed to be rechallenged with TT after progression. Pts with measurable BRAF-V600E/K unresectable stage III/IV melanoma, except pts with uveal melanoma, untreated or symptomatic brain or leptomeningeal involvement were randomly assigned 1:1 to arm A or B. Prior Tx for advanced melanoma was not allowed but adjuvant Tx completed at least 6 months before randomization was permitted. The primary objective was to show superiority of arm B in progression-free survival (PFS) using the log-rank test stratified by stage and lactate dehydrogenase (LDH) with a 1-sided alpha error set at 5%. The study had a power of 80% to detect a HR of 0.65. The study planned to randomize 135 pts in each arm. Results: All 136 pts randomized to arm B and 131 out of 135 in arm A started protocol Tx. At baseline, 170 (63%) pts had stage M1c, 129 (48%) had LDH above upper limit normal (ULN), 74 (27%) had a liver metastasis, and 19 (7%) received adjuvant therapy. The median follow-up was 21 months. In arm B, 135 (99%) pts were free of progression at week 12, when the end of TT was scheduled. In the intention-to-treat population, there was no evidence of a longer PFS in arm B (HR = 0.87, 90% confidence interval [CI] 0.67-1.12, p = 0.36). In a prespecified subgroup analysis, the HR for arm B vs arm A was 2.09 (95% CI 0.96-4.53), 0.74 (95% CI 0.43-1.29), 0.86 (95% CI 0.54-1.37), and 0.46 (95% CI 0.21-1.03) in pts with stage III with LDH≤ULN or M1a, M1b/M1c with LDH≤ULN, ULN < LDH≤2ULN, and LDH > 2ULN, respectively (p-value for interaction 0.045). In a post-hoc subgroup analysis, pts with ≥3 metastatic sites or a sum of target lesions ≥10cm at baseline did not have a longer PFS in arm B but in pts with liver metastasis the Tx HR was 0.48 (95% CI 0.28-0.80, p-value for interaction 0.008). The objective response rate was 53% in arm B and 45% in arm A. Complete response rate was 12% in arm B and 10% in arm A. Grade ≥3 adverse events occurred in 58% of pts in arm B and 51% in arm A. Conclusion: The EBIN trial shows there is no difference in PFS between the two treatment arms for unselected patients but supports the hypothesis that patients with very high LDH and those with liver metastases benefit from the sequential approach. Clinical trial information: NCT03235245 .

Background: Immune checkpoint blockers (ICBs) can induce immune -related adverse events (irAEs) whose management is based on expert opinion and may require the prescription of steroids and/or immunosuppressants (ISs). Recent data suggest that these treatments can reduce the effectiveness of ICBs. Objective: To investigate the relationship between the use of steroids and/or ISs and overall survival (OS) and progression -free survival (PFS) among ICB-treated patients with an irAE. Methods: We prospectively collected data from the medical records of patients with solid tumors or lymphoma in the French REISAMIC cohort and who had been treated with ICBs between June 2014 and June 2020. Results: 184 ICB-treated patients experienced at least one Common Terminology Criteria for Adverse Events grade >= 2 irAE. 107 (58.2%) were treated with steroids alone, 20 (10.9%) with steroids plus IS, 57 (31.0%) not received steroids or IS. The median OS was significantly shorter for patients treated with steroids alone (25.2 months [95% confidence interval (CI): 22.3 -32.4] than for patients treated without steroids or IS (63 months [95%CI: 40.4 -NA]) and those receiving an IS with steroids (53.4 months [95%CI: 47.3 -NA]) (p < 0.001). The median PFS was significantly shorter for patients treated with steroids alone (17.0 months [95%CI: 11.7 -22.9]) than for patients treated without steroids or IS (33.9 months [95%CI: 18.0 -NA]) and those receiving an IS with steroids (41.1 months [95%CI: 26.2 -NA]) (p = 0.006). There were no significant intergroup differences in the hospital admission and infection rates. Conclusion: In a prospective cohort of ICB-treated patients, the use of IS was not associated with worse OS or PFS, contrasting with the use of steroids for the management of irAEs.

Results of PLA, CD8 and PDL1 (MELscore) in all the samples

e18542 Background: The management of rare and/or complex cancers between patients living in metropolitan France and those living overseas remains unequal. Herein, medical transfer to metropolitan France is required in about 20% of overseas-living patients and is often experienced as an additional source of distress, which requires multidisciplinary support (social, psychological, medical, and cultural). Therefore, we developed a dedicated “Overseas” consultation (OC) to provide personalized cancer care in this specific population. We aim to present the evaluation of the first 6 months experience. Methods: The OC was dedicated to patients from the South Pacific (French Polynesia or New Caledonia) requiring “highly specialized” cancer treatments in Gustave Roussy, including surgery, chemotherapy, or diagnostic procedures, which were considered non-achievable in the local hospital after multidisciplinary discussion by videoconference. Multiple interventions including medical, administrative and psychological support were provided. Results: Between July 15, 2022, and January 15, 2023, 25 patients with rare and/or complex cancer diagnosis / treatments benefited from an OC. Median age was 57 years (30-81), 19 patients (76%) were women, 18 (72%) came from French Polynesia and 7 (28%) from New Caledonia. Median number of OC for each patient was 1 (range 1-3), and median length of stay in metropolitan France was 23 days (range 4-95 days). At the time of analysis, 7 patients had not yet returned back home. The most common cancer locations were breast (52%), lung (16%) and gastrointestinal (16%). Patient presented localized disease in 52% of cases. At the first OC, 92% patients were ECOG Performance Status 0-1. The main reasons for metropolitan France medical transfer were an imaging procedure (64%) or a complex surgical or interventional radiology biopsy (24%). Seven patients (28%) had complex management (defined as two or more procedures) during their stay. Eleven patients (44%) benefited from molecular profiling, 7 (27%) from an oncogenetic consultation and 1 (4%) from inclusion in a clinical research trial. Conclusions: OC, set up 6 months ago in our institution, has optimized diagnostic workflow and disease management of these complex patients living overseas and has demonstrated its feasibility. However, it requires the involvement of multiple health-care professionals, multidisciplinary concertation, paramedical interventions such as social and psychological support and the overarching support from institutions.

Summary of cutaneous adverse events at the different time point for each patient included in this study. n/a: not available

Proximity Ligation Assay principle. Two primary antibodies recognize the two antigens BRAF and CRAF. Specific secondary antibodies (Probes) bind to the primary antibodies and are bound to a unique short DNA. When the PLA probes are in close proximity, the DNA strands can interact. After joining they are amplified via rolling circle amplification (RCA) using a polymerase. The signal is easily made visible as a distinct brown or fluorescent dot after enzymatic conversion of a substrate.

Abstract Introduction: High LDH activity is associated with a poor prognosis in many cancer types, especially in melanoma where it is the strongest indicator of poor outcome even with the most potent anticancer immunotherapies or targeted therapies. Although high LDH blood activity is often associated with high tumor burden, this is not always the case and there is no definitive demonstration that blood LDH activity is directly derived from tumors cells. Our objectives were 1) to explore the biological impact of LDHA and LDHB independently of their metabolic function 2) to explore the respective prognostic values of the 5 LDH isoforms as well as the relationship between tumor and blood LDH isoforms repartition in melanoma patients. Material, Patients, Methods: A375 melanoma cells cultured in glucose or galactose (to evaluate glycolytic independent effects) were silenced for LDHA or LDHB expression using si-RNAs. RNAseq analysis of total RNAs and polysomal RNAs was performed to explore the transcriptional and translational impacts of LDHA and LDHB expression. In melanoma patients treated with immunotherapy, the enzymatic activities of each of the 5 LDH isoforms (encoded by LDHA and LDHB) were measured in tumors and blood samples. The prognostic values of the distinct isoforms as well as the correlation between the isoforms distribution in blood and tumors were evaluated. Results: In vitro, independently of their metabolic effect, both LDHA and LDHB had distinct transcriptional and translational impacts on the control of several key oncogenic pathways (adhesion, survival, proliferation, immunity) with a more important effect of LDHA vs LDHB silencing. In vivo, among 64 melanoma patients including 25% with high LDH, high LDH1 (B4) activity was significantly associated with response and overall survival (OS) whereas high LDH4 (A3B1) was inversely associated with response and OS. LDH1 and LDH4 were associated with survival even among patients with normal LDH levels. For patients with paired (tumor and blood) isoforms analysis, there was no correlation between isoforms repartition suggesting that blood LDH activity is not directly related to tumor LDH. Conclusion: In addition to their metabolic activity, LDH enzymes, particularly LDHA, control several key oncogenic pathways at the transcriptional level but also at the translational level for a set of genes involved in cancer biology. LDH1 and LDH4 isoforms blood levels are differentially correlated with response to immunotherapy and survival. They are more robust prognostic biomarkers than total blood LDH level used in our daily practice. Thus, LDH is much more than just a marker of tumor burden in melanoma and the origins, the significance and the roles of blood LDH isoforms need to be revisited. These results highlight the need to develop anticancer drugs targeting LDH, and especially LDHA. Citation Format: Laura Soumoy, Virginie Quidville, Lisa Fredeau, Giuseppina Claps, Caroline Pradon, Ludovic Lacroix, Emilie Routier, Djaouida Belkadi, Séverine Roy, Feras Chehade, Stephan Vagner, Caroline Robert. Roles of lactate dehydrogenase (LDH) in melanoma: an underestimated prognostic biomarker [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2177.

PD-L1 immunohistochemistry. Positive PD-L1 staining in immune cells (chromogen: diaminobenzidine). PT: peritumoral area, CT: center of the tumor, arrows: areas with dense PD-L1+ immune infiltrate.