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Purpose: The combination of talimogene laherparepvec (T-VEC) and pembrolizumab previously demonstrated an acceptable safety profile and an encouraging complete response rate (CRR) in patients with advanced melanoma in a phase Ib study. We report the efficacy and safety from a phase III, randomized, double-blind, multicenter, international study of T-VEC plus pembrolizumab (T-VEC-pembrolizumab) versus placebo plus pembrolizumab (placebo-pembrolizumab) in patients with advanced melanoma., Methods: Patients with stage IIIB-IVM1c unresectable melanoma, naïve to antiprogrammed cell death protein-1, were randomly assigned 1:1 to T-VEC-pembrolizumab or placebo-pembrolizumab. T-VEC was administered at ≤ 4 × 10 6 plaque-forming unit (PFU) followed by ≤ 4 × 10 8 PFU 3 weeks later and once every 2 weeks until dose 5 and once every 3 weeks thereafter. Pembrolizumab was administered intravenously 200 mg once every 3 weeks. The dual primary end points were progression-free survival (PFS) per modified RECIST 1.1 by blinded independent central review and overall survival (OS). Secondary end points included objective response rate per mRECIST, CRR, and safety. Here, we report the primary analysis for PFS, the second preplanned interim analysis for OS, and the final analysis., Results: Overall, 692 patients were randomly assigned (346 T-VEC-pembrolizumab and 346 placebo-pembrolizumab). T-VEC-pembrolizumab did not significantly improve PFS (hazard ratio, 0.86; 95% CI, 0.71 to 1.04; P = .13) or OS (hazard ratio, 0.96; 95% CI, 0.76 to 1.22; P = .74) compared with placebo-pembrolizumab. The objective response rate was 48.6% for T-VEC-pembrolizumab (CRR 17.9%) and 41.3% for placebo-pembrolizumab (CRR 11.6%); the durable response rate was 42.2% and 34.1% for the arms, respectively. Grade ≥ 3 treatment-related adverse events occurred in 20.7% of patients in the T-VEC-pembrolizumab arm and in 19.5% of patients in the placebo-pembrolizumab arm., Conclusion: T-VEC-pembrolizumab did not significantly improve PFS or OS compared with placebo-pembrolizumab. Safety results of the T-VEC-pembrolizumab combination were consistent with the safety profiles of each agent alone.

Purpose: Effective treatments are needed for melanoma that progresses on inhibitors of programmed cell death protein-1 (PD-1) or its ligand (PD-L1). We conducted the phase II LEAP-004 study to evaluate the combination of the multikinase inhibitor lenvatinib and the PD-1 inhibitor pembrolizumab in this population (ClinicalTrials.gov identifier: NCT03776136)., Methods: Eligible patients with unresectable stage III-IV melanoma with confirmed progressive disease (PD) within 12 weeks of the last dose of a PD-1/L1 inhibitor given alone or with other therapies, including cytotoxic T-cell lymphocyte-associated antigen 4 (CTLA-4) inhibitors, received lenvatinib 20 mg orally once daily plus ≤ 35 doses of pembrolizumab 200 mg intravenously once every 3 weeks until PD or unacceptable toxicity. The primary end point was objective response rate (ORR) per RECIST, version 1.1, by independent central review., Results: A total of 103 patients were enrolled and treated. The median study follow-up was 15.3 months. ORR in the total population was 21.4% (95% CI, 13.9 to 30.5), with three (2.9%) complete responses and 19 (18.4%) partial responses. The median duration of response was 8.3 months (range, 3.2-15.9+). ORR was 33.3% in the 30 patients with PD on prior anti-PD-1 plus anti-CTLA-4 therapy. The median progression-free survival and overall survival in the total population were 4.2 months (95% CI, 3.8 to 7.1) and 14.0 months (95% CI, 10.8 to not reached), respectively. Grade 3-5 treatment-related adverse events occurred in 47 (45.6%) patients, most commonly hypertension (21.4%); one patient died from a treatment-related event (decreased platelet count)., Conclusion: Lenvatinib plus pembrolizumab provides clinically meaningful, durable responses in patients with advanced melanoma with confirmed PD on prior PD-1/L1 inhibitor-based therapy, including those with PD on anti-PD-1 plus anti-CTLA-4 therapy. The safety profile was as expected. These data support lenvatinib plus pembrolizumab as a potential regimen for this population of high unmet need.

Melanoma can frequently metastasize to the brain with severe consequences. However, variation of melanoma brain metastases (MBM) development among populations is not well studied, and underlying mechanisms and risk factors for MBM development are not consistently documented. We conducted a systematic literature review (SLR) including a total of 39 articles to evaluate the proportion of melanoma patients who are diagnosed with, or develop, brain metastases, and summarize the risk factors of MBM. The average proportion of MBM was calculated and weighted by the sample size of each study. Meta-analyses were conducted for the selected risk factors using a random-effects model. The proportion of MBM at diagnosis was 33% (975 with MBM out of 2948 patients) among patients with cutaneous melanoma (excluding acral) and 23% (651/2875) among patients with cutaneous mixed with other types of melanoma. The proportion at diagnosis was lower among populations with mucosal (9/96, 9%) or uveal (4/184, 2%) melanoma and among populations outside the United States and Europe. Meta-analysis demonstrated that male vs. female gender and left-sided tumors vs. right-sided were significantly associated with increased risk of melanoma brain metastases. These data may help clinicians to assess an individual patient's risk of developing melanoma brain metastases.

Survival is generally poor for Chinese patients with advanced melanoma because of high rates of acral and mucosal melanoma and limited therapeutic options. The first analysis of the phase 1b KEYNOTE-151 study showed second-line pembrolizumab was well tolerated and had clinically meaningful antitumor activity in Chinese patients with advanced melanoma. Three-year follow-up is presented. Eligible patients were of Chinese descent and had unresectable stage III/IV melanoma that progressed after first-line therapy. Patients received pembrolizumab 2 mg/kg every 3 weeks for ≤35 cycles. Primary end points were safety and objective response rate (ORR). Secondary end points included duration of response (DOR), progression-free survival (PFS), and overall survival (OS). Response was assessed per RECIST v1.1 by blinded independent central review. Subgroup analyses were conducted by melanoma subtype and BRAF and PD-L1 status (acral melanoma only). 103 patients were enrolled; median follow-up duration (time from first dose to data cutoff [July 13, 2020]) was 44.6 months (IQR, 39.1-46.2). Any-grade treatment-related adverse events (TRAEs) occurred in 85.4% of patients, and grade 3/4 TRAEs in 12.6%. No grade 5 TRAEs occurred. Three patients discontinued pembrolizumab because of TRAEs (immune-mediated hepatitis, pneumonia, and arthritis). Immune-mediated AEs and infusion reactions occurred in 34.0% (grade 3/4, 2.9%). ORR was 17.6% (95% CI, 10.8-26.4; 1 complete response/17 partial responses), and median DOR was 13.8 months (range, 2.7-37.4+). Median PFS was 2.8 months (95% CI, 2.7-3.5) and 36-month PFS rate was 5.0%. Median OS was 13.2 months (95% CI, 10.4-16.5) and 36-month OS rate was 22.3%. Median OS for patients with known melanoma subtype was 14.8 months for acral, 13.5 months for nonacral cutaneous, and 7.4 months for mucosal melanoma. Among the acral subgroup, median OS was 22.8 months for PD-L1-positive disease, 8.4 months for PD-L1-negative disease, 18.5 months for BRAF wild-type disease, and 5.8 months for BRAF -mutant disease. Over 3 years' follow-up, second-line pembrolizumab continued to show manageable safety, clinically meaningful antitumor activity, and durable responses in Chinese patients with advanced melanoma. Subgroup analysis suggested particular benefit in PD-L1-positive and BRAF wild-type acral melanoma, although small subgroup sizes preclude definitive conclusions., Clinical Trial Registration: https://clinicaltrials.gov, identifier NCT02821000., Competing Interests: The study received funding from Merck Sharp and Dohme LLC, a subsidiary of Merck and Co., Inc., Rahway, NJ, USA. The funder had the following involvement with the study: the funder of the study collaborated with academic advisers in designing the study, gathering, analyzing, and interpreting the results, and payment of open access publication fees for the current manuscript. LS reports receiving honoraria from MSD, Roche, Juushi Bio, and Novartis. XW reports receiving grants to their institution from Oriengene. SD reports employment at Merck Sharp and Dohme LLC, a subsidiary of Merck and Co., Inc., Rahway, NJ, USA, and is a shareholder of Merck and Co., Inc., Rahway, NJ, USA. CN reports employment at MSD, China, and receiving honoraria from MSD, China. JFL reports employment at Merck Sharp and Dohme LLC, a subsidiary of Merck and Co., Inc., Rahway, NJ, USA. HD reports employment at MSD, China, and receiving honoraria from MSD, China. JG reports advisory/consultancy roles with MSD, Roche, Bayer, Novartis, Simcere Pharmaceutical Group, Shanghai Junshi Biosciences, and Oriengene. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Si, Zhang, Shu, Pan, Wu, Liu, Mao, Wang, Wen, Gu, Zhu, Lan, Cai, Diede, Dai, Niu, Li and Guo.)

Background: Up to 60% of melanoma patients develop melanoma brain metastases (MBM), which traditionally have a poor diagnosis. Current treatment strategies include immunotherapies (IO), targeted therapies (TT), and stereotactic radiosurgery (SRS), but there is considerable heterogeneity across worldwide consensus guidelines., Objective: To summarize current treatments and compare worldwide guidelines for the treatment of MBM., Methods: Review of global consensus treatment guidelines for MBM patients., Results: Substantial evidence supported that concurrent IO or TT plus SRS improves progression-free survival (PFS) and overall survival (OS). Guidelines are inconsistent with regards to recommendations for surgical resection of MBM, since surgical resection of symptomatic lesions alleviates neurological symptoms but does not improve OS. Whole-brain radiation therapy is not recommended by all guidelines due to negative effects on neurocognition but can be offered in rare palliative scenarios., Conclusion: Worldwide consensus guidelines consistently recommend up-front combination IO or TT with or without SRS for the treatment of MBM., Competing Interests: X-LT, ES, RJ, SJD, and IMS are employees of Merck & Co., Inc. JH and AL are employed by Integrative Precision Health LLC. ES was also employed by Seagen Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tan, Le, Lam, Scherrer, Kerr, Lau, Han, Jiang, Diede and Shui.)

Background: Pembrolizumab prolongs progression-free and overall survival among patients with advanced melanoma and recurrence-free survival in resected stage III disease. KEYNOTE-716 assessed pembrolizumab as adjuvant therapy in patients with completely resected, high-risk, stage II melanoma. We report results from the planned first and second interim analyses for recurrence-free survival., Methods: In this double-blind, randomised, placebo-controlled phase 3 study, involving 160 academic medical centres and hospitals in 16 countries (Australia, Belgium, Brazil, Canada, Chile, France, Germany, Israel, Italy, Japan, Poland, South Africa, Spain, Switzerland, the UK, and the USA), patients aged 12 years or older with newly diagnosed, completely resected stage IIB or IIC melanoma (TNM stage T3b or T4 with a negative sentinel lymph node biopsy) were recruited. Eligible patients were randomly assigned (1:1), in blocks of four and stratified by T-category (3b, 4a, and 4b) and paediatric status (age 12-17 years vs ≥18 years), using an interactive response technology system to intravenous pembrolizumab 200 mg (2 mg/kg in paediatric patients) or placebo every 3 weeks for 17 cycles or until disease recurrence or unacceptable toxicity. All patients, clinical investigators, and analysts were masked to treatment assignment. The primary endpoint was investigator-assessed recurrence-free survival (defined as time from randomisation to recurrence or death) in the intention-to-treat (ITT) population (ie, all patients randomly assigned to treatment). The primary endpoint was met if recurrence-free survival was significantly improved for pembrolizumab versus placebo at either the first interim analysis (after approximately 128 patients had events) or second interim analysis (after 179 patients had events) under multiplicity control. Safety was assessed in all patients randomly assigned to treatment who received at least one dose of study treatment. This study is registered with ClinicalTrials.gov, NCT03553836, and is closed to accrual., Findings: Between Sept 23, 2018, and Nov 4, 2020, 1182 patients were screened, of whom 976 were randomly assigned to pembrolizumab (n=487) or placebo (n=489; ITT population). The median age was 61 years (IQR 52-69) and 387 (40%) patients were female and 589 (60%) were male. 874 (90%) of 976 patients were White and 799 (82%) were not Hispanic or Latino. 483 (99%) of 487 patients in the pembrolizumab group and 486 (99%) of 489 in the placebo group received assigned treatment. At the first interim analysis (data cutoff on Dec 4, 2020; median follow-up of 14·4 months [IQR 10·2-18·7] in the pembrolizumab group and 14·3 months [10·1-18·7] in the placebo group), 54 (11%) of 487 patients in the pembrolizumab group and 82 (17%) of 489 in the placebo group had a first recurrence of disease or died (hazard ratio [HR] 0·65 [95% CI 0·46-0·92]; p=0·0066). At the second interim analysis (data cutoff on June 21, 2021; median follow-up of 20·9 months [16·7-25·3] in the pembrolizumab group and 20·9 months [16·6-25·3] in the placebo group), 72 (15%) patients in the pembrolizumab group and 115 (24%) in the placebo group had a first recurrence or died (HR 0·61 [95% CI 0·45-0·82]). Median recurrence-free survival was not reached in either group at either assessment timepoint. At the first interim analysis, grade 3-4 treatment-related adverse events occurred in 78 (16%) of 483 patients in the pembrolizumab groups versus 21 (4%) of 486 in the placebo group. At the first interim analysis, four patients died from an adverse event, all in the placebo group (one each due to pneumonia, COVID-19-related pneumonia, suicide, and recurrent cancer), and at the second interim analysis, one additional patient, who was in the pembrolizumab group, died from an adverse event (COVID-19-related pneumonia). No deaths due to study treatment occurred., Interpretation: Pembrolizumab as adjuvant therapy for up to approximately 1 year for stage IIB or IIC melanoma resulted in a significant reduction in the risk of disease recurrence or death versus placebo, with a manageable safety profile., Funding: Merck Sharp & Dohme, a subsidiary of Merck & Co, Kenilworth, NJ, USA., Competing Interests: Declaration of interests JJL reports research funding to their institution for clinical studies from Merck Sharp & Dohme (MSD), AbbVie, Agios, Array, Astellas, AstraZeneca, Bristol-Myers Squibb, Corvus, Day One, EMD Serono, Fstar, Genmab, Ikena, Immatics, Incyte, Kadmon, KAHR, Macrogenics, Moderna, Nektar, Next Cure, Numab, Palleon, Pfizer, Replimune, Rubius, Servier, Scholar Rock, Synlogic, Takeda, Trishula, Tizona, and Xencor; membership on data safety monitoring boards for AbbVie and Immutep; membership on scientific advisory boards with no stock ownership or stock options for 7 Hills, Bright Peak, Fstar, Inzen, RefleXion, Xilioo, and with stock for Actym, Alphamab Oncology, Arch Oncology, Kanaph, Mavu, NeoTx, Onc.AI, Pyxis, and Tempest; compensation for consultancy with AbbVie, Alnylam, Bayer, Bristol-Myers Squibb, CheckMate, Codiak, Crown, Cstone, Day One, Eisai, EMD Serono, Flame, Genentech, Gilead, HotSpot, Kadmon, KQS, Janssen Ikena, Immunocore, Incyte, Macrogenics, Merck, Mersana, Nektar, Novartis, Pfizer, Regeneron, Ribon, Rublus, Silicon, Synlogic, Synthekine, TRex, Werewold, and Xencor; and a provisional patent for cancer immunotherapy (PCT/US18/36052: Microbiome Biomarkers for anti-PD-1/PD-L1 responsiveness: diagnostic, prognostic and therapeutic uses thereof). PR reports research funding to their institution from MSD and Pfizer and honoraria for lectures and advisory board participation from MSD, Bristol Myers Squibb, Novartis, Pierre Fabre, Sanofi, Merck, Philogen, and Blueprint Medicines. PQ reports research finding to their institution for clinical studies from MSD, and honoraria as consultant or advisor role from Bristol Myers Squibb, MSD, Merck, Novartis, Pierre Fabre, Sun Pharma, Sanofi-Regeneron, and Roche. MDV reports research finding to their institution for clinical studies from MSD and honoraria as consultant or advisor for Novartis, Bristol Myers Squibb, MSD, and Pierre Fabre. JM reports research funding to their institution for clinical studies from MSD; honoraria from Bristol Myers Squibb, MSD, Roche, Novartis, and Pierre Fabre; and consultant or advisor roles for Bristol Myers Squibb and MSD. VC-S reports research funding to their institution for clinical studies from MSD; reimbursement for travel and accommodation for medical congress from Bristol Myers Squibb and Pierre Fabre; and honoraria for advisory board participation from Novartis and Pierre Fabre. LdlCM reports research funding to their institution for clinical studies from Celgene, MSD, and Roche; reimbursement for travel expenses from Roche; and consultant or advisor roles for Bristol Myers Squibb, MSD, Novartis, and Roche. MAK reports research funding to their institution for clinical studies from MSD and MSD Australia; fees as speaker from Bristol Myers Squibb, Novartis, Merck Serono, and MSD Australia; and reimbursement for travel from Bristol Myers Squibb and Roche. DS reports research funding to their institution for clinical studies from Bristol Myers Squibb, MSD, Novartis, Amgen, and Array; grants paid to their institution from Bristol Myers Squibb, MSD, Novartis, Merck-EMD, Philogen, Pfizer, Array, InflarX, OncoSec, Replimune, Neracare, Nektar, SunPharma, Sandoz, and UltimoVacs; reimbursement for travel from Bristol Myers Squibb, Merck, Novartis, Merck-EMD, Pfizer, Pierre Fabre, InflarX, NeraCare, and Nektar; and non-financial support from Bristol Myers Squibb, MSD, Novartis, Merck-EMD, Pierre-Fabre, InFlarX, Neracare, Nektar, SunPharma, and Sandoz. GVL reports research funding to their institution for clinical studies from MSD and fees as a consultant or advisor from Aduro Biotech, Amgen, Arrary Biopharma, Boehringer Ingelheim International GmbH, Bristol Myers Squibb, Evaxion Biotech A/S, Hexal AG, Highlight Therapeutics S L, MSD, Novartis Pharma AG, OncoSec, Pierre Fabre, QBiotics Group, Regeneron Pharmaceuticals, SkylineDX BV, and Specialised Therapeutics Australia. PAA reports research funding to their institution for clinical studies from MSD, Bristol Myers Squibb, Roche-Genentech, Sanofi, and Pfizer and fees as a consultant or advisor from Bristol Myers Squibb, Roche-Genentech, MSD, Novartis, Merck Serono, Pierre Fabre, AstraZeneca, Sun Pharma, Sanofi, Idera, Sandoz, Immunocore, 4SC, Nektar, Italfarmaco, Boehringer Ingelheim, Eisai, Regeneron, Daiichi Sankyo, Pfizer, Oncosec, Nouscom, Lunaphore, Seagen, and ITeos. MM reports research funding to their institution for clinical studies from MSD and personal fees from MSD, Pierre Fabre, Novartis, Bristol Myers Squibb, and Sanofi. FDG and J-JG report research funding to their institutions for clinical studies from MSD. AH reports research funding to their institution for clinical studies from MSD and fees for advisory board membership from Novartis, Bristol Myers Squibb, and MSD. RD reports research funding to their institution for clinical studies from MSD and consulting or advisory roles with MSD, Novartis, Roche, Bristol Myers Squibb, Amgen, Takeda, Pierre Fabre, Sun Pharma, Sanofi, Catalym, Second Genome, Regeneron, Alligator, T3 Pharma, MaxiVAX SA, Pfizer, and touchIME. CR reports research funding to their institution for clinical studies from MSD and fees as a consultant or advisor for Amgen, AstraZeneca, Bristol Myers Squibb, MSD, Novartis, Pfizer, Pierre Fabre, Roche, and Sanofi. MSC reports research funding to their institution for clinical studies for MSD and honoraria for consulting from Bristol Myers Squibb, Eisia, IDEAYA Biosciences, Merck Serono, MSD, Novartis, Oncosec, Pierre Fabre, Qbiotics, Roche, and Sanofi. PM reports research funding to their institution for clinical studies from MSD, Amgen, Johnson & Johnson, Merck Serono, Novartis, Pfizer, Sanofi, and Sun Pharma; honoraria from Amgen, Bristol Myers Squibb, Merck, MSD, Novartis, Pierre Fabre, Roche/Genentech, and Sanofi; consulting or advisory roles for Amgen, Bristol Myers Squibb, Merck, MSD, Novartis, Pierre Fabre, Roche, and Sanofi; fees for speakers bureau from Amgen, Bristol Myers Squibb, MSD, Novartis, Roche, and Sanofi; and reimbursement for travel, accommodation, and expenses from Amgen, Bristol Myers Squibb, MSD, Novartis, Pierre Fabre, Roche, Sanofi, and Sun Pharma. AP reports research funding to their institution for clinical studies from MSD; fees for consulting or advisory roles from Novartis; and fees for speakers bureau from Bristol Myers Squibb. VKS reports research funding to their institution for clinical studies from MSD and Neogene Therapeutic and fees for participation in independent data safety monitoring committees and participation in advisory boards from Bristol Myers Squibb, Eisai, Novartis, Merck, Regeneron, and Replimune. RAS reports research funding to their institution from MSD and fees for professional services from F Hoffmann-La Roche, Evaxion, Provectus Biopharmaceuticals Australia, Qbiotics, Novartis, MSD, NeraCare, AMGEN, Bristol Myers Squibb, Myriad Genetics, and GSK. RAS is supported by an Australian National Health and Medical Research Council Practitioner Fellowship (APP1141295). JMK reports research funding to their institution from MSD, Amgen, Bristol Myers Squibb, Checkmate Pharmaceuticals, Immunocore, Iovance Biotherapies, Novartis Pharmaceuticals, Castle Biosciences, and Merck; personal fees from Amgen, Ankyra Therapeutics, Axio Research/Instil Bio, Bristol Myers Squibb, Checkmate Pharmaceuticals, DermTech, Elsevier, Harbour BioMed, Immunocore, Iovance Biotherapies, Istari Oncology, Millenium Pharm/Takeda Pharm, Natera, Novartis Pharmaceutical, OncoCyte Corporation, OncoSec Medical, Pfizer, Scopus BioPharma, Merck, Becker Pharmaceutical Consulting, Fenix Group International, Intellisphere LLC/Cancer Network, IQVIA, Replimune, and SR One Capital Managment. KC, SJD, SA, and NI are employees of and own stock in MSD. AMME reports research funding to their institution for clinical studies from MSD; fees for advisory board membership for Merck, Agenus, Biocad, BioInvent, BioNTech, Bristol Myers Squibb, CatalYm, Ellipses, Galecto, GSK, IO Biotech, ISA Pharmaceuticals, Merck, MSD, Nektar, Novartis, Pfizer, Sellas, SkylineDX, TigaTx, and TxDiscovery; personal fees for independent data monitoring committee for Biocad, GSK, Pfizer, and Novartis; fees for participation in speakers bureau from Biocad, Bristol Myers Squibb, and Merck; and equity in IO Biotech, SkylineDX., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Objectives: Parts 4 and 5 of the phase 1/2 KEYNOTE-022 study investigated the maximum tolerated dose (MTD), safety, and efficacy of pembrolizumab plus trametinib in solid tumours and BRAF wild-type melanoma., Patients and Methods: Patients received intermittent or concurrent dosing of pembrolizumab plus trametinib. Concurrent dosing was 2 or 4 weeks of trametinib run-in followed by concurrent pembrolizumab every 3 weeks (Q3W) plus trametinib once daily (QD). Intermittent dosing was 2 weeks of trametinib run-in followed by pembrolizumab plus intermittent trametinib (1 week off/2 weeks on). A 3 + 3 dose escalation was used, followed by dose confirmation., Results: Forty-two patients were enrolled. No dose-limiting toxicities (DLTs) occurred at initial dose levels (DL). At subsequent DLs, 10 of 38 evaluable patients had DLTs. For concurrent dosing, MTD was pembrolizumab 200 mg Q3W plus trametinib 1.5 mg QD, with a 2-week trametinib 1.5 mg QD run-in (concurrent DL2a); in concurrent DL2a group, five (31%) patients had grade 3/4 treatment-related adverse events (TRAEs); the objective response rate (ORR) was 0%. ORR was 40% in concurrent DL1 and 0% in concurrent DL2b. For intermittent dosing, MTD was pembrolizumab 200 mg Q3W plus trametinib 2 mg QD with a 2-week trametinib 2 mg QD run-in (intermittent DL2); in the intermittent DL2 group, seven (47%) patients had grade 3/4 TRAEs; ORR was 27%. ORR in intermittent DL1 was 33%., Conclusions: MTDs for concurrent and intermittent dosing of pembrolizumab with trametinib were identified. The combination had limited antitumour activity, numerically higher ORR with intermittent versus concurrent dosing, and manageable safety. CLINICALTRIALS., Gov Identifier: NCT02130466., Competing Interests: Conflict of interest statement The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: M.M. reports serving as a consultant/adviser for Bristol Myers Squibb, AstraZeneca, Roche, Merck & Co., Inc., Kenilworth, NJ, USA, Amgen, Pierre Fabre, Alfasigma USA Inc., and Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA (MSD). M.S.C. reports serving as a consultant/adviser for MSD, Bristol Myers Squibb, Novartis, Amgen, Roche, Pierre Fabre, Sanofi, Merck Serono, Nektar, Regeneron, Eisai Co., Ltd., QBiotics, and Ideaya and has received honoraria from MSD, Bristol Myers Squibb, and Novartis. A.M.J. reports consultant/advisory role (recipient: his institution) for Neolukin, Janssen Oncology, Ipsen, AstraZeneca, Sanofi, Noxopharm, IQvia, Pfizer, Novartis, Bristol-Myers Squibb, Merck Serono, Eisai; has received research funding to his institution from Bristol-Myers Squibb, Janssen Oncology, MSD, Mayne Pharma, Roche/Genentech, Bayer, Macrogenics, Lilly, Pfizer, AstraZeneca, and Corvus Pharmaceuticals; and stock and other ownership interests in Pricilium Therapeutics. E.M. reports serving as on advisory boards for MSD, Bristol Myers Squibb, Novartis, EMD Serono, and Sanofi. A.R. has received honoraria for consulting from Amgen, Chugai, Novartis, MSD, Sanofi, Vedanta, and Nurix; is a Science Advisory Board member and stockholder in Arcus, Highlight, Compugen, Merus, Rgenix, PACT Pharma, Tango Therapeutics, Apricity, ImaginAb, Isoplexis, Lutris Pharma, MapKure, and 4C Biomed; has previously been a Science Advisory Board member and stockholder in CytomX, Five Prime, RAPT, Advaxis, and Kite-Gilead; and declares institutional research grants from Agilent and Bristol Myers Squibb. P.A.A. reports serving as a consultant/advisor for Bristol Myers Squibb, Roche-Genentech, MSD, Array, Novartis, Merck Serono, Pierre Fabre, Incyte, Medimmune, AstraZeneca, Syndax, Sun Pharma, Sanofi, Idera, Ultimovacs, Sandoz, Immunocore, 4SC, Alkermes, Italfarmaco, Boehringer-Ingelheim, Eisai Co., Ltd. and Regeneron; declares research funds from Bristol Myers Squibb, Roche-Genentech, and Array; and declares travel support from MSD. W.H.M. declares having received consultant fees from Bristol Myers Squibb, Merck Canada Inc., Kirkland, QC, Canada, Roche, Novartis, Amgen, and GlaxoSmithKline. M.O.B. reports serving as a consultant/advisor for MSD, Bristol Myers Squibb, Novartis, Sanofi, Pfizer, GlaxoSmithKline, Immunocore, and EMD Serono; serving as a Safety Review Board member for Adaptimmune, and GlaxoSmithKline; and receiving support for trials from MSD, Bristol Myers Squibb, Novartis, Adaptimmune, and Immunocore. P.F.F. reports serving on advisory boards for Bristol Myers Squibb, MSD, Novartis, Pierre Fabre, and Roche. R.Z. reports serving on advisory boards for Roche, Pfizer, and AstraZeneca; and receiving speaker payments from MSD and Bristol Myers Squibb. M.D.V. reports serving as a consultant/advisor for Bristol Myers Squibb, MSD, Novartis, Pierre Fabre, and Sanofi. E.G. reports holding stocks in and reports receiving payment from Novartis. R.G. is an employee of Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA. S.J.D. is an employee of Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA and a shareholder in Merck & Co., Inc., Kenilworth, NJ, USA. E.C. was an employee of Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA, at the time of the study. O.H. declares having received speaker fees from Array, Pfizer, Bristol Myers Squibb, Novartis, and Sanofi Regeneron; having received consulting fees from Adura, Akeso, Amgen, Array, BeiGene, Bristol Myers Squibb, Genentech, GlaxoSmithKline, Immunocore, Incyte, Janssen, Merck & Co., Inc., Kenilworth, NJ, USA, Nextcure, Novartis, Sanofi Regeneron, Seattle Genetics, Tempus, and Zelluna; and contracted research for his institution from Arcus, Aduro, Akeso, Amgen, Array, Bristol Myers Squibb, CytomX, Exelixis, Genentech, GlaxoSmithKline, Immunocore, Incyte, Iovance, Merck & Co., Inc., Kenilworth, NJ, USA, Moderna, Merck Serono, NextCure, Novartis, Sanofi Regeneron, Seattle Genetics, Torque, and Zelluna., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Objective: Patients with melanoma and early stable disease (SD) with pembrolizumab have unclear prognosis. We present post hoc analyses of long-term outcomes for patients with early SD, partial response (PR) or complete response (CR) with pembrolizumab., Patients and Methods: Patients who received pembrolizumab in the KEYNOTE-001 and KEYNOTE-006 studies and had SD, PR or CR at weeks 12 or 24 were included., Results: Of 294 patients in the week 12 analysis, 107 (36.4%) had SD at week 12, of whom 7 (6.5%) had a best overall response of CR, 43 (40.2%) had PR and 57 (53.3%) had SD. Forty-eight-month overall survival (OS) rates were 95.2%, 73.0% and 47.7%, respectively, for patients with CR, PR and SD at week 12. Similar results were observed in the 241 patients in the week 24 analysis. Forty-eight-month OS rates were 72.1% for patients with SD at week 12 followed by subsequent response and 75.0% for patients with PR at week 12 followed by no change in response or progression. Thirty-six-month and 48-month OS rates were 11.6% and not reached, respectively, for patients with SD at week 12 followed by progression before week 24., Conclusions: A substantial proportion of patients (46.7%) with early (week 12) SD with pembrolizumab achieved subsequent PR or CR. Patients with SD at week 12 and subsequent CR/PR had similar survival to those who maintained PR. In contrast, patients with SD at week 12 and subsequent progression had poor survival outcomes. These findings may guide treatment decisions for patients achieving early SD., Trial Registration: Clinicaltrials.gov: NCT01295827 (KEYNOTE-001); NCT01866319 (KEYNOTE-006)., Competing Interests: Conflict of interest statement The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: O.H. is a consultant advisor for Aduro, Akeso, Amgen, Array, BeiGene, BMS, Genentech, GSK, Immunocore, Incyte, Janssen, Merck, NextCure, Novartis, Sanofi, Regeneron, Seattle Genetics, Tempus and Zelluna. He is a speaker for Array, BMS, Novartis, Pfizer, Sanofi and Regeneron. He has contracted research for his institution from Aduro, Akeso, Amgen, Arcus, Array, BMS, CytomX, Exelixis, Genentech, GSK, Immunocore, Incyte, Io-vance, Merck, Merck Serono, Moderna, NextCure, Novartis, Regeneron, Sanofi, Seattle Genetics, Torque and Zelluna. S.J.D., B.H.M. and E.J. are employees of Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA, and S.J.D. owns stock in Merck & Co., Inc., Kenilworth, NJ, USA. A.A. is a consultant advisor for Amgen, BMS, Merck, Novartis, Pierre Fabre, Roche, Sanofi and Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, USA. M.S.C. is a consultant advisor for Amgen, BMS, Eisai, IDEAYA, MSD, Merck Serono, Nektar, Novartis, Pierre Fabre, QBiotics, OncoSec, Regeneron, Roche and Sanofi. He has also received honoraria from BMS, MSD and Novartis. T.M. has served on advisory boards for Merck, BMS and OncoSec. A.D. has received grants from BMS, Merck, Pfizer/Array and Xencor. He also served on an advisory board meeting for Xencor. J.J.G. has served on advisory boards for BMS, Merck, MSD, Novartis, Pfizer, Pierre Fabre, Roche, Sanofi and Sun Pharma. He has also received personal fees from BMS, MSD, Novartis and Pierre Fabre for travel as well as personal fees for serving as a speaker for MSD, Novartis and Pierre Fabre. P.H. has no disclosures to report. F.S.H. is a consultant advisor for Aduro, Bristol Myers Squibb, Eisai, EMD Serono, Genentech/Roche, Idera, Merck, Novartis, Pieris Pharmaceutical, Sanofi and Takeda. He reports grants and royalties paid to his institution by Bristol Myers Squibb and Novartis, as well as personal fees from Gossamer. He also holds equity in Apricity, Bicara, Pionyr and Torque. He has served on advisory boards for Apricity, Bicara, Bioentre, Checkpoint Therapeutics, Compass Therapeutics, Iovance, Pionyr, Surface and Torque. His pending patents include Methods for Treating MICA-Related Disorders (#20100111973; includes royalties), Angiopoietin-2 Biomarkers Predictive of Anti-Immune Checkpoint Response (#20170248603), Compositions and Methods for Identification, Assessment, Prevention, and Treatment of Melanoma Using PD-L1 Isoforms (#20160340407), Therapeutic Peptides (#20160046716, #20140004112, #20170022275, #20170008962), Methods of Using Pembrolizumab and Trebananib and Anti-Galectin Antibody Biomarkers Predictive of Anti-Immune Checkpoint and Anti-Angiogenesis Responses (#20170343552). He has been issued patents for Tumor Antigens and Uses Thereof (#7250291), Therapeutic Peptides (#9402905), Vaccine Compositions and Methods for Restoring NKG2D Pathway Function Against Cancers (#10279021) and Antibodies that Bind to MHC Class I Polypeptide-Related Sequence A (#10106611). A.M.J. has received institutional support from MSD. L.M. has no disclosures to report. A.R. is a consultant for Amgen, Chugai, Merck, Novartis, Nurix, Sanofi and Vedanta. He is a current scientific advisory board member and stockholder in 4C Biomed, Apricity, Arcus, Compugen, Highlight, ImaginAb, Isoplexis, Lutris Pharma, MapKure, Merus, PACT Pharma, Rgenix and Tango Therapeutics and is a past scientific advisory board member and stockholder in Advaxis, Cytomx, Five Prime, Kite-Gilead and RAPT. He has also received grants paid to his institution from Agilent and Bristol Myers Squibb. C.R. is a consultant advisor for Amgen, AstraZeneca, BMS, Merck, MSD, Novartis, Pierre Fabre, Roche and Sanofi. J.S. has no disclosures to report. J.S.W. has received grants paid to his institution from Merck. He has been issued a PD-1 biomarker patent with Biodesix unrelated to the submitted work. J.D.W. is a non-compensated consultant for Merck. He is also a consultant for Amgen, Apricity, Arsenal IO, Ascentage Pharma, Astellas, AstraZeneca, Bayer, BeiGene, Boehringer Ingelheim, Bristol Myers Squibb, Celgene, Chugai, Eli Lilly, Elucida, F Star, Georgiamune, Imvaq, Kyowa Hakko Kirin, Linneaus, Merck, Neon Therapeutics, Polynoma, Psioxus, Recepta, Sellas, Serametrix, Surface Oncology, Syndax, Syntalogic, Takara Bio, Tizona Pharmaceuticals, Trieza, Truvax and Werewolf Therapeutics. He reports equity in Adaptive Biotech, Apricity, Arsenal IO, Belgene, Georgiamune, Imvaq, Linneaus and Tizona Pharmaceuticals. He has received research grants from Bristol Myers Squibb and Sephora. He has licensed patents for xenogeneic DNA vaccines (Merial; includes royalties), a myeloid-derived suppressor cell (MDSC) assay (Serametrix; includes royalties), an anti–PD-1 antibody (Agenus), anti-CTLA4 antibodies (Agenus) and anti-GITR antibodies and method of use thereof (Agenus/Incyte). He has also been issued patents for alphavirus replicon particles expressing Newcastle disease viruses for cancer therapy, phosphatidylserine targeting agents and uses thereof for adoptive T-cell therapies, immunosuppressive follicular help-like T cells modulated by immune checkpoint blockade, identifying and treating subjects at risk for checkpoint blockade therapy–associated colitis, CAR+ T cells targeting differentiation antigens as means to treat cancer, anti-CD40 agonist mAB fused to monophosphoryl lipid A (MPL) for cancer therapy and engineered vaccinia viruses for cancer immunotherapy. G.V.L. is consultant advisor for Aduro Biotech Inc, Amgen Inc, Array Biopharma inc, Boehringer Ingelheim International GmbH, Bristol Myers Squibb, Evaxion Biotech A/S, Hexel AG, Highlight Therapeutics S.L., Merck Sharp & Dohme, Novartis Pharma AG, OncoSec, Pierre Fabre, QBiotics Group Limited, Regeneron Pharmaceuticals Inc, SkylineDX B.V., and Specialised Therapeutics Australia Pty Ltd., (Copyright © 2021 Merck Sharp & Dohme Corp., The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Objective: Long-term safety of pembrolizumab in melanoma was analyzed in KEYNOTE-001, KEYNOTE-002, and KEYNOTE-006., Patients and Methods: Analysis involved patients who received ≥1 pembrolizumab dose. Lead-time bias was addressed via landmark analyses in patients who were progression-free before day 147., Results: Adverse events (AEs) were analyzed for 1567 patients (median follow-up, 42.4 months). Most AEs were mild/moderate; grade 3/4 treatment-related AEs occurred in 17.7% of patients. Two pembrolizumab-related deaths occurred. Any-grade immune-mediated AEs (imAEs) occurred in 23.0%, most commonly hypothyroidism (9.1%), pneumonitis (3.3%), and hyperthyroidism (3.0%); grade 3/4 imAEs occurred in 6.9% of patients. Most imAEs occurred within 16 weeks of treatment. In landmark analysis, patients who did (n = 79) versus did not (n = 384) develop imAEs had similar objective response rates (ORRs) (64.6% versus 63.0%); median time to response (TTR), 5.6 months for both; median duration of response (DOR), 20.0 versus 25.3 months; median progression-free survival (PFS), 17.0 versus 17.7 months; median overall survival (OS), not reached (NR) versus 43 months (p = 0.1104). Patients who did (n = 17) versus did not (n = 62) receive systemic corticosteroids had similar ORRs (70.6% vs. 62.9%) and median TTR (6.4 vs. 5.6 months) but numerically shorter median PFS (9.9 vs. 17.0 months); median DOR, 14.2 months versus NR; median OS, NR for both., Conclusions: These results enhance the knowledge base for pembrolizumab in advanced melanoma, with no new toxicity signals after lengthy follow-up of a large population. In landmark analyses, pembrolizumab efficacy was similar regardless of imAEs or systemic corticosteroid use., Clinical Trial Registry: NCT01295827, NCT01704287, NCT01866319., Competing Interests: Conflict of interest statement C.R. has participated in advisory boards for Roche; Pierre Fabre; Merck; Novartis; Amgen; Bristol-Myers Squibb; Novartis; Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc., Kenilworth, NJ, U.S.A. (MSD); and Sanofi. O.H. reports personal fees from Merck for consulting during the conduct of the study; as well as contracted research for his institution from Arcus, Aduro, Akeso, Amgen, Array, Bristol-Myers Squibb, CytomX, Exelixis, Genentech, GSK, Immunocore, Incyte, Iovance, Merck, Moderna, Merck Serono, NextCure, Novartis, Regeneron, Roche, Seattle Genetics, Torque, and Zelluna outside the submitted work. A.R. has received personal fees as honoraria for consulting from Amgen, Chugai, Genentech-Roche, Novartis, and Merck; and personal fees, as a scientific advisory member and stockholder from Arcus, Bi-oncotech, Compugen, CytomX, Five Prime, FLX-Bio, Merus, Rgenix, PACT Pharma, and Tango Therapeutics, outside the submitted work. J.S.W. reports personal fees for honoraria for advisory boards and transportation from Merck, Bristol-Myers Squibb, Genentech, Celldex, Pfizer, and AstraZeneca, outside the submitted work. In addition. J.S.W. has a patent named on a PD-1 biomarker patent by Biodesix issued. I.D. reports grants from Merck, Bristol-Myers Squibb, Incyte, OncoSec, and Regeneron, and personal fees from Regeneron, during the conduct of the study. F.S.H. reports other from Merck to their institution for clinical trial support during the conduct of the study; grants, personal fees, and consulting from Bristol-Myers Squibb; personal fees from Merck, EMD Serono, Genentech/Roche, Bayer, Partners Therapeutics, Sanofi, Pfizer, and Kairos for consulting; grants and personal fees from Novartis for consulting; personal fees from Takeda, Surface, Compass Therapeutics, Verastem, and Rheos for advisory boards; personal fees from Apricity and Bicara for scientific advisory board and equity; personal fees from Aduro for advisor consulting; personal fees from Pionyr for advisory board and equity; personal fees from 7 Hills Pharma for advisor; other from Torque for scientific advisory board and equity; outside the submitted work. In addition, F.S.H. has a patent for Methods for Treating MICA-Related Disorders (#20100111973) with royalties paid, a patent for Tumor antigens and uses thereof issued (#7250291), a patent for Angiopoieten-2 Biomarkers Predictive of Anti-immune checkpoint response pending (#20170248603), a patent for Compositions and Methods for Identification, Assessment, Prevention, and Treatment of Melanoma using PD-L1 Isoforms pending (#20160340407), five patents for Therapeutic peptides pending (#20160046716, #20140004112, #20170022275, #20170008962, #9402905), and a patent for “methods of using pembrolizumab and trebananib.” J.D.W. reports personal fees for consulting from Adaptive Biotech, Amgen, Apricity, Ascentage Pharma, Astellas, AstraZeneca, Bayer, BeiGene, Bristol-Myers Squibb, Celgene, Chugai, Eli Lilly, F-star, Imvaq, Kyowa Hakko Kirin, Linnaeus, MedImmune, Merck, Neon Therapeutics, Ono, Polaris Pharma, Polynoma, PsiOxus, Puretech, Recepta, Takara Bio, Trieza, Truvax, Serametrix, Surface Oncology, Syndax, and Syntalogic; research support from Bristol-Myers Squibb and AstraZeneca; and equity in Potenza Therapeutics, Tizona Pharmaceuticals, Adaptive Biotechnologies, Imvaq, BeiGene, Trieza, and Linnaeus. T.C.M. reports personal fees for honorarium from Bristol-Myers Squibb, Aduro, Merck, and Incyte outside the submitted work. R.W.J. reports other from Bristol-Myers Squibb, and Gilead for consulting, outside the submitted work. C.B. reports personal fees from Bristol-Myers Squibb (board advisor), speaker fees from Merck, and travel fees from Amgen and Sandoz outside the submitted work. GVL is consultant advisor for Aduro Biotech Inc, Amgen Inc, Array Biopharma inc, Boehringer Ingelheim International GmbH, Bristol-Myers Squibb, Highlight Therapeutics S.L., MSD, Novartis Pharma AG, Pierre Fabre, QBiotics Group Limited, Regeneron Pharmaceuticals Inc, SkylineDX B.V. I.P. is a consultant for Amgen, Merck, and Iovance, outside the submitted work. R.Dummer reports intermittent, project focused consulting and/or advisory relationships with Novartis, MSD, Bristol-Myers Squibb, Roche, Amgen, Takeda, Pierre Fabre, Sun Pharma, Sanofi, Catalym, Second Genome, Regeneron, Alligator, MaxiVAX SA and touchIME outside the submitted work. J.L. reports employment at MSD, and GSK. S.J.D. reports personal fees as an employee of MSD, and shareholder inMerck & Co., Inc., Kenilworth, NJ, USA. M.S.C. has served on advisory boards for Bristol-Myers Squibb, MSD, Amgen, Novartis, Pierre Fabre, Roche, Sanofi, Merck, Ideaya, Regeneron, Nektar, Eisai, Oncosec and Qbiotics., outside the submitted work. A.J. reports consulting and/or advisory role for Neolukin, Janssen Oncology, Ipsen, AstraZeneca, Sanofi, Noxopharm, IQvia, Pfizer, Novartis, Bristol-Myers Squibb, and Merck Serono, outside the submitted work. A.J. also reports research funding from Bristol-myers Squibb, Janssen Oncology, MSD, Mayne Pharma, Roche/Genentech, Bayer, Macrogenics, Lilly, Pfizer, AstraZeneca, and Corvus Pharmaceuticals. All remaining authors have declared no conflicts of interest., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: In the KEYNOTE-022 study, pembrolizumab with dabrafenib and trametinib (triplet) improved progression-free survival (PFS) versus placebo with dabrafenib and trametinib (doublet) without reaching statistical significance. Mature results on PFS, duration of response (DOR), and overall survival (OS) are reported., Methods: The double-blind, phase 2 part of KEYNOTE-022 enrolled patients with previously untreated BRAF V600E/K -mutated advanced melanoma from 22 sites in seven countries. Patients were randomly assigned 1:1 to intravenous pembrolizumab (200 mg every 3 weeks) or placebo plus dabrafenib (150 mg orally two times per day) and trametinib (2 mg orally one time a day). Primary endpoint was PFS. Secondary endpoints were objective response rate, DOR, and OS. Efficacy was assessed in the intention-to-treat population, and safety was assessed in all patients who received at least one dose of study drug. This analysis was not specified in the protocol., Results: Between November 30, 2015 and April 24, 2017, 120 patients were randomly assigned to triplet (n=60) or doublet (n=60) therapy. With 36.6 months of follow-up, median PFS was 16.9 months (95% CI 11.3 to 27.9) with triplet and 10.7 months (95% CI 7.2 to 16.8) with doublet (HR 0.53; 95% CI 0.34 to 0.83). With triplet and doublet, respectively, PFS at 24 months was 41.0% (95% CI 27.4% to 54.2%) and 16.3% (95% CI 8.1% to 27.1%); median DOR was 25.1 months (95% CI 14.1 to not reached) and 12.1 months (95% CI 6.0 to 15.7), respectively. Median OS was not reached with triplet and was 26.3 months with doublet (HR 0.64; 95% CI 0.38 to 1.06). With triplet and doublet, respectively, OS at 24 months was 63.0% (95% CI 49.4% to 73.9%) and 51.7% (95% CI 38.4% to 63.4%). Grade 3-5 treatment-related adverse events (TRAEs) occurred in 35 patients (58%, including one death) receiving triplet and 15 patients (25%) receiving doublet., Conclusion: In BRAF V600E/K -mutant advanced melanoma, pembrolizumab plus dabrafenib and trametinib substantially improved PFS, DOR, and OS with a higher incidence of TRAEs. Interpretation of these results is limited by the post hoc nature of the analysis., Competing Interests: Competing interests: PFF has received personal fees from Novartis, Roche, Bristol-Myers Squibb, Pierre Fabre, and Merck Sharp & Dohme Corp., a subsidiary of Merck & Co., Kenilworth, New Jersey, USA (MSD). AMDG reports other from MSD during the conduct of the study; reports other from Incyte, GlaxoSmithKline, Pierre Fabre, and Sanofi outside the submitted work; and has served as an advisor and consultant for MSD, Incyte, GlaxoSmithKline, Pierre Fabre, and Sanofi. MDV has received personal fees from Novartis, MSD, and BMS outside the submitted work and has served as an advisor and consultant for MSD, BMS, Novartis, Pierre Fabre and Sanofi. VA has received personal fees and travel support from Bristol-Myers Squibb; personal fees from MSD, Merck Serono, Novartis, Pierre Fabre, Roche and Nektar; and travel support from OncoSec Medical, during the conduct of the study. HS received a research grant and personal fees from MSD and has received personal fees from Bristol-Myers Squibb, Incyte, Novartis, and Pierre Fabre outside the submitted work. PQ has received personal fees from Roche, Novartis, Bristol-Myers Squibb, MSD, Sanofi, and Pierre Fabre. GVL is consultant advisor for Aduro Biotech Inc, Amgen Inc, Array Biopharma inc, Boehringer Ingelheim International GmbH, Bristol-Myers Squibb, Highlight Therapeutics S.L., MSD, Novartis Pharma AG, Pierre Fabre, QBiotics Group Limited, Regeneron Pharmaceuticals Inc, SkylineDx B.V. RS reports personal fees from MSD New Zealand, outside the submitted work. IMS reports other (trial expenses) from MSD. IMS has received other funds from MSD during the conduct of the study. EG is an employee and stockholder of Novartis. RG is an employee of MSD. SJD is an employee and stockholder of MSD. ESC was an employee of MSD at the time of this study. AR has received personal fees from Amgen, Chugai, Novartis, Genentech-Roche, Sanofi, and Merck and serves as an advisor for, holds stock in, and has received personal fees from Arcus, Bioncotech, Compugen, CytomX, Five Prime, RAPT Therapeutics, Merus, Rgenix, PACT Pharma, and Tango Therapeutics outside the submitted work. PAA has received grants and personal fees from Bristol-Myers Squib, Roche-Genentech, and Array BioPharma; personal fees and other from MSD; and personal fees from Novartis, Merck Serono, Pierre Fabre, Incyte, Genmab, NewLink Genetics, Medimmune, AstraZeneca, Syndax, Sun Pharma, Sanofi, Idera, Ultimovacs, Sandoz, Immunocore, 4SC, Alkermes, Italfarmaco, Nektar, and Boehringer Ingelheim. JS, ML, and MA-A report no competing interests., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Pembrolizumab (PEMBRO) and ipilimumab + nivolumab (IPI + NIVO) are approved advanced melanoma (AM) immunotherapies. To address limited health-related quality of life (QoL) real-world evidence with immunotherapies in AM, we compared QoL in AM patients receiving either treatment in clinical practice., Methods: A prospective US observational study enrolled adult AM patients initiating first-line PEMBRO or IPI + NIVO between June 2017 and March 2018. Endpoints included the QLQ-C30 global health score (GHS) and EuroQol visual analog scale (EQ-VAS) scores. Mean changes were compared using repeated measures mixed-effects models and are presented covariate adjusted., Results: 225 PEMBRO and 187 IPI + NIVO patients were enrolled. From baseline through week 24, PEMBRO was associated with 3.2 mean GHS score increase (95% CI 0.5, 5.9; p = .02), while no change was observed with IPI + NIVO; 0.2 (95% CI - 2.6, 3.0; p = 0.87). Among objective treatment-responders, GHS scores associated with PEMBRO increased 6.0 (95% CI 3.1, 8.8; p < .0001); IPI + NIVO patients increased 3.8 (95% CI 0.8, 6.9; p = .01). In treatment non-responders, IPI + NIVO was associated with GHS/QoL deterioration of - 3.7 (95% CI - 6.8, - 0.6; p = .02), PEMBRO non-responders demonstrated no change; 0.7 (95% CI - 2.3, 3.7; p = 0.6). Between treatments, PEMBRO patients increased 2.6 greater in EQ-VAS (95% CI 0.6, 4.5; p = .01) vs IPI + NIVO at 24 weeks., Conclusions: PEMBRO was associated with better 24-week QoL compared to IPI + NIVO in actual clinical practice settings. Real-world data has known limitations, but with further confirmation these results may have implications for treatment selection.

The third multistakeholder Paediatric Strategy Forum organised by ACCELERATE and the European Medicines Agency focused on immune checkpoint inhibitors for use in combination therapy in children and adolescents. As immune checkpoint inhibitors, both as monotherapy and in combinations have shown impressive success in some adult malignancies and early phase trials in children of single agent checkpoint inhibitors have now been completed, it seemed an appropriate time to consider opportunities for paediatric studies of checkpoint inhibitors used in combination. Among paediatric patients, early clinical studies of checkpoint inhibitors used as monotherapy have demonstrated a high rate of activity, including complete responses, in Hodgkin lymphoma and hypermutant paediatric tumours. Activity has been very limited, however, in more common malignancies of childhood and adolescence. Furthermore, apart from tumour mutational burden, no other predictive biomarker for monotherapy activity in paediatric tumours has been identified. Based on these observations, there is collective agreement that there is no scientific rationale for children to be enrolled in new monotherapy trials of additional checkpoint inhibitors with the same mechanism of action of agents already studied (e.g. anti-PD1, anti-PDL1 anti-CTLA-4) unless additional scientific knowledge supporting a different approach becomes available. This shared perspective, based on scientific evidence and supported by paediatric oncology cooperative groups, should inform companies on whether a paediatric development plan is justified. This could then be proposed to regulators through the available regulatory tools. Generally, an academic-industry consensus on the scientific merits of a proposal before submission of a paediatric investigational plan would be of great benefit to determine which studies have the highest probability of generating new insights. There is already a rationale for the evaluation of combinations of checkpoint inhibitors with other agents in paediatric Hodgkin lymphoma and hypermutated tumours in view of the activity shown as single agents. In paediatric tumours where no single agent activity has been observed in multiple clinical trials of anti-PD1, anti-PDL1 and anti-CTLA-4 agents as monotherapy, combinations of checkpoint inhibitors with other treatment modalities should be explored when a scientific rationale indicates that they could be efficacious in paediatric cancers and not because these combinations are being evaluated in adults. Immunotherapy in the form of engineered proteins (e.g. monoclonal antibodies and T cell engaging agents) and cellular products (e.g. CAR T cells) has great therapeutic potential for benefit in paediatric cancer. The major challenge for developing checkpoint inhibitors for paediatric cancers is the lack of neoantigens (based on mutations) and corresponding antigen-specific T cells. Progress critically depends on understanding the immune macroenvironment and microenvironment and the ability of the adaptive immune system to recognise paediatric cancers in the absence of high neoantigen burden. Future clinical studies of checkpoint inhibitors in children need to build upon strong biological hypotheses that take into account the distinctive immunobiology of childhood cancers in comparison to that of checkpoint inhibitor responsive adult cancers., Competing Interests: Conflict of interest J.A. is medical and scientific director for TC Biopharm and holds stock and share options in TC Biopharm and Autolus Ltd; M.B. is an employee of Merck Group; CB. is an employee of Tesaro; EB. is an employee of Pfizer; R.C. is an employee of Novartis; H.C. is an employee of Roche; S.J.D. is an employee of Merck & Co; M.E. is an employee of Boehringer-Ingelheimm Pharma GmbH; B.G. attended at a Roche sponsored advisory board for atezolizumab; R.I. is an employee of Celgene; M.K. is an employee and owns stock in Bristol-Myers Squibb; I.L. is an employee of Regeneron; L.V.M. has participated in advisory boards for AstraZeneca, Merck, Tesaro, Bayer and Celgene; M.M. is an employee of Syndax Pharmaceuticals; C.O. is an employee of Sanofi; A.D.J.P. has participated in advisory boards for Novartis, Taked, Merck, Lilly and Celgene; V.G.R.P. is an employee and shareholder of Autolus Ltd; C.R. attended advisory boards for Amgen, Celgen, EUSA Pharma, Genentch, Novartis and Roche, speaker honaria for Bristol Myers Squibb, Pfizer and Roche; P.K.S. is an employee of AstraZeneka; P.S. is an employee of Immunicum AB; C.v.T. participated in advisory boards for Novartis and Bayer; T.Y. is an employee of Beigene., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Both pembrolizumab (PEMBRO) and ipilimumab + nivolumab (IPI + NIVO) are FDA-approved immunotherapy regimens for advanced melanoma (AM). Each regimen has different toxicity profiles potentially impacting healthcare resource utilization (HCRU). This study compared real-world hospitalization and emergency department (ED) utilization within 12 months of therapy initiation of each regimen. Methods: A retrospective cohort study was conducted in AM patients ≥18 years old initiating PEMBRO or IPI + NIVO between January 1, 2016-December 30, 2017. Patients were identified from 12 US-based academic and satellite centers. All-cause hospitalization ED visits were identified. These events were used to calculate rates per 1,000 patient months. Utilization between groups was compared using multivariate logistic regression. Results: In total, 400 patients were included (200 PEMBRO, 200 IPI + NIVO). PEMBRO vs IPI + NIVO patients had poorer Eastern Cooperative Group (ECOG) performance status, 29% 2-4, vs 12% ( p  < .001); more diabetes, 21% vs 13% ( p  = .045); were more often PD-L1 expression positive, 77% vs 63% ( p  = .011); and less likely BRAF mutant, 35% vs 50% ( p  = .003). The proportion with more than one hospitalization over 12 months was 17% PEMBRO vs 24% IPI + NIVO. Less than 2% had more than one admission and none had more than two. Unadjusted mean (SD) hospitalizations per 1,000 patient-months were 16 (37) and 20 (38), PEMBRO and IPI + NIVO, respectively. Adjusted odds ratio for hospitalization was 0.6 (95% CI = 0.3-0.9; p  = .027) for PEMBRO vs IPI + NIVO. ED visits occurred in 18% vs 21%, PEMBRO and IPI + NIVO, respectively, 0.7 ( p  = .186). Conclusions: PEMBRO patients had a significantly lower probability of hospitalization through 12 months vs IPI + NIVO. The probability of ED visits did not differ.

Background: Pembrolizumab is approved for the treatment of advanced cancer in adults; however, no information is available on safety and efficacy in paediatric patients. We aimed to establish the recommended phase 2 dose of pembrolizumab and its safety and antitumour activity in advanced paediatric cancer., Methods: KEYNOTE-051 is an ongoing phase 1-2 open-label trial. In this interim analysis, children aged 6 months to 17 years were recruited at 30 hospitals located in Australia, Brazil, Canada, France, Germany, Israel, Italy, South Korea, Sweden, the UK, and the USA. Patients with melanoma or a centrally confirmed, PD-L1-positive, relapsed or refractory solid tumour or lymphoma, and a Lansky Play/Karnofsky Performance status score of 50 or higher, received intravenous pembrolizumab at an initial dose of 2 mg/kg every 3 weeks. Pharmacokinetics and dose-limiting toxicities were used to establish the recommended phase 2 dose, and the safety and antitumour activity of this dose were assessed. Primary endpoints were determination of dose-limiting toxicities at the maximum administered dose, safety and tolerability, and the proportion of patients with objective response to pembrolizumab for each tumour type according to the Response Evaluation Criteria in Solid Tumours version 1.1 or the International Neuroblastoma Response Criteria. Safety and efficacy were assessed in all treated patients who received at least one dose of pembrolizumab. Separate reporting of the cohort of patients with relapsed or refractory classical Hodgkin lymphoma was a post-hoc decision. The data cutoff for this interim analysis was Sept 3, 2018. This trial is still enrolling patients and is registered with ClinicalTrials.gov, number NCT02332668., Findings: Of 863 patients screened between March 23, 2015, and Sept 3, 2018, 796 had tumours that were evaluable for PD-L1 expression (278 [35%] were PD-L1-positive); 155 eligible patients were enrolled and 154 had at least one dose of pembrolizumab. The median age of the enrolled patients was 13 years (IQR 8-15). Median follow-up was 8·6 months (IQR 2·5-16·4). No dose-limiting toxicities were reported in phase 1, and pembrolizumab plasma concentrations were consistent with those previously reported in adults; the recommended phase 2 dose was therefore established as 2 mg/kg every 3 weeks. Of the 154 patients treated, 69 (45%) experienced grade 3-5 adverse events, most commonly anaemia in 14 (9%) patients and decreased lymphocyte count in nine (6%) patients. 13 (8%) of the 154 patients had grade 3-5 treatment-related adverse events, most commonly decreased lymphocyte count in three (2%) patients and anaemia in two (1%) patients. 14 (9%) patients had serious treatment-related adverse events, most commonly pyrexia (four [3%]), and hypertension and pleural effusion (two [1%] each). Four patients (3%) discontinued treatment because of treatment-related adverse events, and two (1%) died (one due to pulmonary oedema and one due to pleural effusion and pneumonitis). Of 15 patients with relapsed or refractory Hodgkin lymphoma, two had complete and seven had partial responses; thus, nine patients achieved an objective response (60·0%; 95% CI 32·3-83·7). Of 136 patients with solid tumours and other lymphomas, eight had partial responses (two patients each with adrenocortical carcinoma and mesothelioma, and one patient each with malignant ganglioglioma, epithelioid sarcoma, lymphoepithelial carcinoma, and malignant rhabdoid tumour); the proportion of patients with an objective response was 5·9% (95% CI 2·6-11·3)., Interpretation: Pembrolizumab was well tolerated and showed encouraging antitumour activity in paediatric patients with relapsed or refractory Hodgkin lymphoma, consistent with experience in adult patients. Pembrolizumab had low antitumour activity in the majority of paediatric tumour types, and responses were observed in only a few rare PD-L1-positive tumour types, suggesting that PD-L1 expression alone is not sufficient as a biomarker for the selection of paediatric patients who are likely to respond to PD-1 checkpoint inhibitors. Final results of KEYNOTE-051, expected by September, 2022, with the possibility for extension, will report further on the activity of pembrolizumab in Hodgkin lymphoma, microsatellite instability-high tumours, and melanoma., Funding: Merck Sharp & Dohme, a subsidiary of Merck & Co., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Background: Immunotherapy combination treatments can improve patient outcomes. Epacadostat, an IDO1 selective inhibitor, and pembrolizumab, a PD-1 inhibitor, showed promising antitumour activity in the phase 1-2 ECHO-202/KEYNOTE-037 study in advanced melanoma. In this trial, we aimed to compare progression-free survival and overall survival in patients with unresectable stage III or IV melanoma receiving epacadostat plus pembrolizumab versus placebo plus pembrolizumab., Methods: In this international, randomised, placebo-controlled, double-blind, parallel-group, phase 3 trial, eligible participants were aged 18 years or older, with unresectable stage III or IV melanoma previously untreated with PD-1 or PD-L1 checkpoint inhibitors, an ECOG performance status of 0 or 1, and had a known BRAF V600 mutant status or consented to BRAF V600 mutation testing during screening. Patients were stratified by PD-L1 expression and BRAF V600 mutation status and randomly assigned (1:1) through a central interactive voice and integrated web response system to receive epacadostat 100 mg orally twice daily plus pembrolizumab 200 mg intravenously every 3 weeks or placebo plus pembrolizumab for up to 2 years. We used block randomisation with a block size of four in each stratum. Primary endpoints were progression-free survival and overall survival in the intention-to-treat population. The safety analysis population included randomly assigned patients who received at least one dose of study treatment. The study was stopped after the second interim analysis; follow-up for safety is ongoing. This study is registered with ClinicalTrials.gov, number NCT02752074., Findings: Between June 21, 2016, and Aug 7, 2017, 928 patients were screened and 706 patients were randomly assigned to receive epacadostat plus pembrolizumab (n=354) or placebo plus pembrolizumab (n=352). Median follow-up was 12·4 months (IQR 10·3-14·5). No significant differences were found between the treatment groups for progression-free survival (median 4·7 months, 95% CI 2·9-6·8, for epacadostat plus pembrolizumab vs 4·9 months, 2·9-6·8, for placebo plus pembrolizumab; hazard ratio [HR] 1·00, 95% CI 0·83-1·21; one-sided p=0·52) or overall survival (median not reached in either group; epacadostat plus pembrolizumab vs placebo plus pembrolizumab: HR 1·13, 0·86-1·49; one-sided p=0·81). The most common grade 3 or worse treatment-related adverse event was lipase increase, which occurred in 14 (4%) of 353 patients receiving epacadostat plus pembrolizumab and 11 (3%) of 352 patients receiving placebo plus pembrolizumab. Treatment-related serious adverse events were reported in 37 (10%) of 353 patients receiving epacadostat plus pembrolizumab and 32 (9%) of 352 patients receiving placebo plus pembrolizumab. There were no treatment-related deaths in either treatment group., Interpretation: Epacadostat 100 mg twice daily plus pembrolizumab did not improve progression-free survival or overall survival compared with placebo plus pembrolizumab in patients with unresectable or metastatic melanoma. The usefulness of IDO1 inhibition as a strategy to enhance anti-PD-1 therapy activity in cancer remains uncertain., Funding: Incyte Corporation, in collaboration with Merck Sharp & Dohme., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Pembrolizumab has been approved in the United States for treating advanced melanoma for >4 years. We examined real-world pembrolizumab use and associated outcomes in US oncology clinical practices, including patients who would not be eligible for clinical trials.Flatiron Health longitudinal database was used to identify adult patients with advanced melanoma initiating ≥1 dose of pembrolizumab from September 4, 2014, through December 31, 2016, with follow-up through December 31, 2017. Patients in any clinical trial during the study period were excluded. Overall survival (OS) and time on treatment from pembrolizumab initiation were analyzed using the Kaplan-Meier (KM) method. Subgroup analyses were conducted to examine OS for several patient characteristics including Eastern Cooperative Oncology Group (ECOG) performance status >1, brain metastases, and corticosteroids before pembrolizumab initiation.Pembrolizumab was administered to 315 (59%), 152 (29%), and 65 (12%) patients as first-, second-, and third-line/later therapy. Median age at pembrolizumab initiation was 68 years (range, 18-84); most patients were male (66%) and white (94%). Of those with available data, 38% had BRAF-mutant melanoma, 21% had elevated lactate dehydrogenase (LDH) level, and 23% had ECOG >1. Overall, 18% had brain metastases, and 23% were prescribed corticosteroids <3 months before initiating pembrolizumab. Median study follow-up was 12.9 months (range, 0.03-39.6). Median OS was 21.8 months (95% confidence interval [CI] 16.8-29.1); KM 1-year and 2-year survival rates were 61% and 48%, respectively; and median time on pembrolizumab treatment was 4.9 months (95% CI 3.7-5.5). Median OS for first-line pembrolizumab was not reached, and for second-line and third-line/later was 13.9 and 12.5 months, respectively (log-rank P = .0095). Significantly better OS (all P ≤.0014, log-rank test) was evident for patients with ECOG performance status (PS) of 0 to 1 (vs >1), normal (vs elevated) LDH level, and no (vs yes) corticosteroid prescription <3 months before. No difference was recorded in OS by brain metastases (log-rank P = .22) or BRAF mutation status (log-rank P = .90).These findings support effectiveness of pembrolizumab in the real-world clinical setting and provide important insights into patient characteristics and outcomes associated with pembrolizumab therapy for a heterogeneous patient population with advanced melanoma, including patients who would not be eligible for clinical trials.

Background: Pembrolizumab shows robust antitumor activity and favorable safety in metastatic melanoma. KEYNOTE-151 evaluated pembrolizumab in Chinese patients, who have more aggressive melanoma subtypes than other populations., Methods: Chinese patients aged ≥18years with advanced melanoma previously treated with one line of therapy received pembrolizumab 2 mg/kg every 3 weeks for 35 cycles or until confirmed disease progression, intolerable toxicity, or study withdrawal. Primary end points were objective response rate (ORR) per RECIST v1.1 by blinded independent central review and safety. Key secondary end points included duration of response (DOR) and progression-free survival (PFS) per RECIST v1.1 and overall survival (OS)., Results: Median age was 52 years (N=103); 37.9% had acral and 14.6% had mucosal melanoma. Median follow-up was 7.9months at data cutoff (December 27, 2017). ORR was 16.7% (95% CI, 10.0-25.3%) (1 complete, 16 partial responses). Disease control rate was 38.2%. ORR was 15.8% for acral, 13.3% for mucosal melanoma. Median DOR was 8.4months; 65.6% of patients had response duration ≥6months. Median PFS was 2.8months (95% CI, 2.7-3.5months); 6-month rate was 20.4%. Median OS was 12.1months (95% CI, 9.6months-not reached); 6-month rate, 75.7%; 12-month rate, 50.6%. Treatment-related AEs (TRAEs) occurred in 87 (84.5%) patients; 9 (8.7%) experienced grade 3/4 TRAE and 2 (1.9%) discontinued because of TRAE; none died. Two deaths occurred that were unrelated to treatment., Conclusions: Pembrolizumab was well tolerated and provided clinically meaningful antitumor activity as second-line therapy in Chinese patients with advanced melanoma., (Copyright © 2019. Published by Elsevier Inc.)

Aim: To describe recent evolution in treatment patterns and outcomes for advanced melanoma (AMel)., Methods: This retrospective observational study analyzed de-identified electronic health record data from the Flatiron Health database for 1140 adult patients who initiated first-line therapy for AMel from 1 January 2014 to 30 June 2016 with follow-up through 28 February 2017., Results: The most common first-line regimens were ipilimumab-based therapies (34%), anti-PD-1 monotherapy (26%) and BRAF/MEK inhibitor(s) (20%). First-line ipilimumab-based and BRAF inhibitor regimens decreased after the third quarter of 2014 (3Q2014), and by 2Q2016, 55 and 91% of BRAF-mutant and BRAF wild-type cohorts, respectively, received a first-line anti-PD-1 regimen. Median overall survival from first-line initiation for all patients was 18.8 months (95% CI: 16.3-23.3)., Conclusion: Results illustrate changing paradigms of therapy and real-world patient outcomes for AMel.

Aim: To explore factors associated with pembrolizumab (PEMBRO) versus ipilimumab + nivolumab (IPI+NIVO) selection in advanced melanoma., Materials & Methods: Total of 12 academic and satellite clinics contributed to this study. Descriptive and logistic regression analyses were conducted to explore associations between clinical characteristics and treatment choice.  Results: Total of 400 patients were included: 200 PEMBRO and 200 IPI+NIVO. Patients were significantly more likely to receive PEMBRO versus IPI+NIVO if they had poorer Eastern Cooperative Oncology Group score, 2-4 versus 0-1 (odds ratio [OR]: 6.6; 95% CI: 3.0-14.7), if they were PD-L1 positive (OR: 4.5; 95% CI: 1.9-10.4) or had BRAF wild-type tumor (OR: 2.2; 95% CI: 1.4-3.6)., Conclusion: Patient factors are significantly associated with treatment selection in advanced melanoma. Outcomes comparisons should take this into consideration.

Background: Vorinostat combined with retinoids produces additive antitumor effects in preclinical studies of neuroblastoma. Higher systemic exposures of vorinostat than achieved in pediatric phase I trials with continuous daily dosing are necessary for in vivo increased histone acetylation and cytotoxic activity. We conducted a phase I trial in children with relapsed/refractory neuroblastoma to determine the maximum tolerated dose (MTD) of vorinostat on an interrupted schedule, escalating beyond the previously identified pediatric MTD., Methods: Isotretinoin (cis-13-retinoic acid) 80 mg/m 2 /dose was administered by mouth twice daily on days 1-14 in combination with escalating doses of daily vorinostat up to 430 mg/m 2 /dose (days 1-4; 8-11) in each 28-day cycle using the standard 3 + 3 design. Vorinostat pharmacokinetic testing and histone acetylation assays were performed., Results: Twenty-nine patients with refractory or relapsed neuroblastoma were enrolled and 28 were evaluable for dose escalation decisions. Median number of cycles completed was two (range 1-15); 11 patients received four or more cycles. Three patients experienced cycle 1 dose-limiting toxicities. A total of 18 patients experienced grade 3/4 toxicities related to study therapy. The maximum intended dose of vorinostat (430 mg/m 2 /day, days 1-4; 8-11) was tolerable and led to increased histone acetylation in surrogate tissues when compared to lower doses of vorinostat (P = 0.009). No objective responses were seen., Conclusions: Increased dose vorinostat (430 mg/m 2 /day) on an interrupted schedule is tolerable in combination with isotretinoin. This dose led to increased vorinostat exposures and demonstrated increased histone acetylation. Prolonged stable disease in patients with minimal residual disease warrants further investigation., (© 2018 Wiley Periodicals, Inc.)

Purpose Pembrolizumab provides durable antitumor activity in metastatic melanoma, including complete response (CR) in about 15% of patients. Data are limited on potential predictors of CR and patient disposition after pembrolizumab discontinuation after CR. We describe baseline characteristics and long-term follow-up in patients who experienced CR with pembrolizumab in the KEYNOTE-001 study ( ClinicalTrials.gov identifier: NCT01295827). Patients and Methods Patients with ipilimumab-naive or -treated advanced/metastatic melanoma received one of three dose regimens of pembrolizumab. Eligible patients who received pembrolizumab for ≥ 6 months and at least two treatments beyond confirmed CR could discontinue therapy. Response was assessed every 12 weeks by central Response Evaluation Criteria in Solid Tumors version 1.1. For this analysis, CR was defined per investigator assessment, immune-related response criteria, and potential predictors of CR were evaluated using univariate and multivariate analyses. Results Of 655 treated patients, 105 (16.0%) achieved CR after median follow-up of 43 months. At data cutoff, 92 patients (87.6%) had CR, with median follow-up of 30 months from first CR. Fourteen (13.3%) patients continued to receive treatment for a median of ≥ 40 months. Pembrolizumab was discontinued by 91 patients (86.7%), including 67 (63.8%) who proceeded to observation without additional anticancer therapy. The 24-month disease-free survival rate from time of CR was 90.9% in all 105 patients with CR and 89.9% in the 67 patients who discontinued pembrolizumab after CR for observation. Tumor size and programmed death-ligand 1 status were among the baseline factors independently associated with CR by univariate analysis. Conclusion Patients with metastatic melanoma can have durable complete remission after discontinuation of pembrolizumab, and the low incidence of relapse after median follow-up of approximately 2 years from discontinuation provides hope for a cure for some patients. The mechanisms underlying durable CR require further investigation.

Naturally stalled replication forks are considered to cause structurally abnormal chromosomes in tumor cells. However, underlying mechanisms remain speculative, as capturing naturally stalled forks has been a challenge. Here, we captured naturally stalled forks in tumor cells and delineated molecular processes underlying the structural evolution of circular mini-chromosomes (double-minute chromosomes; DMs). Replication forks stalled on the DM by the co-directional collision with the transcription machinery for long non-coding RNA. RPA, BRCA2, and DNA polymerase eta (Polη) were recruited to the stalled forks. The recruitment of Polη was critical for replication to continue, as Polη knockdown resulted in DM loss. Rescued stalled forks were error-prone and switched replication templates repeatedly to create complex fusions of multiple short genomic segments. In mice, such complex fusions circularized the genomic region surrounding MYC to create a DM during tumorigenesis. Our results define a molecular path that guides stalled replication forks to complex chromosomal rearrangements., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Here we report a phase 1b clinical trial testing the impact of oncolytic virotherapy with talimogene laherparepvec on cytotoxic T cell infiltration and therapeutic efficacy of the anti-PD-1 antibody pembrolizumab. Twenty-one patients with advanced melanoma were treated with talimogene laherparepvec followed by combination therapy with pembrolizumab. Therapy was generally well tolerated, with fatigue, fevers, and chills as the most common adverse events. No dose-limiting toxicities occurred. Confirmed objective response rate was 62%, with a complete response rate of 33% per immune-related response criteria. Patients who responded to combination therapy had increased CD8 + T cells, elevated PD-L1 protein expression, as well as IFN-γ gene expression on several cell subsets in tumors after talimogene laherparepvec treatment. Response to combination therapy did not appear to be associated with baseline CD8 + T cell infiltration or baseline IFN-γ signature. These findings suggest that oncolytic virotherapy may improve the efficacy of anti-PD-1 therapy by changing the tumor microenvironment. VIDEO ABSTRACT., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Several interesting biological questions arise when thinking about the heterogeneous presentation of neuroblastoma, especially with regard to the molecular differences between very low- and high-risk disease. Why do some metastatic tumours spontaneously differentiate or regress entirely? Does the presence of disseminated disease always indicate metastases, or might some cases be better considered as multifocal disease?

Genetic and epigenetic alterations are essential for the initiation and progression of human cancer. We previously reported that primary human medulloblastomas showed extensive cancer-specific CpG island DNA hypermethylation in critical developmental pathways. To determine whether genetically engineered mouse models (GEMMs) of medulloblastoma have comparable epigenetic changes, we assessed genome-wide DNA methylation in three mouse models of medulloblastoma. In contrast to human samples, very few loci with cancer-specific DNA hypermethylation were detected, and in almost all cases the degree of methylation was relatively modest compared with the dense hypermethylation in the human cancers. To determine if this finding was common to other GEMMs, we examined a Burkitt lymphoma and breast cancer model and did not detect promoter CpG island DNA hypermethylation, suggesting that human cancers and at least some GEMMs are fundamentally different with respect to this epigenetic modification. These findings provide an opportunity to both better understand the mechanism of aberrant DNA methylation in human cancer and construct better GEMMs to serve as preclinical platforms for therapy development.

Breakage-fusion-bridge (BFB) cycle is a series of chromosome breaks and duplications that could lead to the increased copy number of a genomic segment (gene amplification). A critical step of BFB cycles leading to gene amplification is a palindromic fusion of sister chromatids following the rupture of a dicentric chromosome during mitosis. It is currently unknown how sister chromatid fusion is produced from a mitotic break. To delineate the process, we took an integrated genomic, cytogenetic and molecular approach for the recurrent MCL1 amplicon at chromosome 1 in human tumor cells. A newly developed next-generation sequencing-based approach identified a cluster of palindromic fusions within the amplicon at ∼50-kb intervals, indicating a series of breaks and fusions by BFB cycles. The physical location of the amplicon (at the end of a broken chromosome) further indicated BFB cycles as underlying processes. Three palindromic fusions were mediated by the homologies between two nearby inverted Alu repeats, whereas the other two fusions exhibited microhomology-mediated events. Such breakpoint sequences indicate that homology-mediated fold-back capping of broken ends followed by DNA replication is an underlying mechanism of sister chromatid fusion. Our results elucidate nucleotide-level events during BFB cycles and end processing for naturally occurring mitotic breaks.

Rhabdomyosarcoma is a pediatric tumor of skeletal muscle that expresses the myogenic basic helix-loop-helix protein MyoD but fails to undergo terminal differentiation. Prior work has determined that DNA binding by MyoD occurs in the tumor cells, but myogenic targets fail to activate. Using MyoD chromatin immunoprecipitation coupled to high-throughput sequencing and gene expression analysis in both primary human muscle cells and RD rhabdomyosarcoma cells, we demonstrate that MyoD binds in a similar genome-wide pattern in both tumor and normal cells but binds poorly at a subset of myogenic genes that fail to activate in the tumor cells. Binding differences are found both across genomic regions and locally at specific sites that are associated with binding motifs for RUNX1, MEF2C, JDP2, and NFIC. These factors are expressed at lower levels in RD cells than muscle cells and rescue myogenesis when expressed in RD cells. MEF2C is located in a genomic region that exhibits poor MyoD binding in RD cells, whereas JDP2 exhibits local DNA hypermethylation in its promoter in both RD cells and primary tumor samples. These results demonstrate that regional and local silencing of differentiation factors contributes to the differentiation defect in rhabdomyosarcomas.

Rhabdomyosarcoma is the most common soft-tissue sarcoma in children. While cytogenetic abnormalities have been well characterized in this disease, aberrant epigenetic events such as DNA hypermethylation have not been described in genome-wide studies. We have analyzed the methylation status of 25,500 promoters in normal skeletal muscle, and in cell lines and tumor samples of embryonal and alveolar rhabdomyosarcoma from pediatric patients. We identified over 1,900 CpG islands that are hypermethylated in rhabdomyosarcomas relative to skeletal muscle. Genes involved in tissue development, differentiation, and oncogenesis such as DNAJA4, HES5, IRX1, BMP8A, GATA4, GATA6, ALX3, and P4HTM were hypermethylated in both RMS cell lines and primary samples, implicating aberrant DNA methylation in the pathogenesis of rhabdomyosarcoma. Furthermore, cluster analysis revealed embryonal and alveolar subtypes had distinct DNA methylation patterns, with the alveolar subtype being enriched in DNA hypermethylation of polycomb target genes. These results suggest that DNA methylation signatures may aid in the diagnosis and risk stratification of pediatric rhabdomyosarcoma and help identify new targets for therapy.

DNA amplification, particularly of chromosomes 8 and 11, occurs frequently in breast cancer and is a key factor in tumorigenesis, often associated with poor prognosis. The mechanisms involved in the amplification of these regions are not fully understood. Studies from model systems have demonstrated that palindrome formation can be an early step in DNA amplification, most notably seen in the breakage-fusion-bridge (BFB) cycle. Therefore, palindromes might be associated with gene amplicons in breast cancer. To address this possibility, we coupled high-resolution palindrome profiling by the Genome-wide Analysis of Palindrome Formation (GAPF) assay with genome-wide copy-number analyses on a set of breast cancer cell lines and primary tumors to spatially associate palindromes and copy-number gains. We identified GAPF-positive regions distributed nonrandomly throughout cell line and tumor genomes, often in clusters, and associated with copy-number gains. Commonly amplified regions in breast cancer, chromosomes 8q and 11q, had GAPF-positive regions flanking and throughout the copy-number gains. We also identified amplification-associated GAPF-positive regions at similar locations in subsets of breast cancers with similar characteristics (e.g., ERBB2 amplification). These shared positive regions offer the potential to evaluate the utility of palindromes as prognostic markers, particularly in premalignant breast lesions. Our results implicate palindrome formation in the amplification of regions with key roles in breast tumorigenesis, particularly in subsets of breast cancers.

DNA methylation might have a significant role in preventing normal differentiation in pediatric cancers. We used a genomewide method for detecting regions of CpG methylation on the basis of the increased melting temperature of methylated DNA, termed denaturation analysis of methylation differences (DAMD). Using the DAMD assay, we find common regions of cancer-specific methylation changes in primary medulloblastomas in critical developmental regulatory pathways, including Sonic hedgehog (Shh), Wingless (Wnt), retinoic acid receptor (RAR), and bone morphogenetic protein (BMP). One of the commonly methylated loci is the PTCH1-1C promoter, a negative regulator of the Shh pathway that is methylated in both primary patient samples and human medulloblastoma cell lines. Treatment with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-aza-dC) increases the expression of PTCH1 and other methylated loci. Whereas genetic mutations in PTCH1 have previously been shown to lead to medulloblastoma, our study indicates that epigenetic silencing of PTCH1, and other critical developmental loci, by DNA methylation is a fundamental process of pediatric medulloblastoma formation. This finding warrants strong consideration for DNA demethylating agents in future clinical trials for children with this disease.

Rhabdomyosarcomas are characterized by expression of myogenic specification genes, such as MyoD and/or Myf5, and some muscle structural genes in a population of cells that continues to replicate. Because MyoD is sufficient to induce terminal differentiation in a variety of cell types, we have sought to determine the molecular mechanisms that prevent MyoD activity in human embryonal rhabdomyosarcoma cells. In this study, we show that a combination of inhibitory Musculin:E-protein complexes and a novel splice form of E2A compete with MyoD for the generation of active full-length E-protein:MyoD heterodimers. A forced heterodimer between MyoD and the full-length E12 robustly restores differentiation in rhabdomyosarcoma cells and broadly suppresses multiple inhibitory pathways. Our studies indicate that rhabdomyosarcomas represent an arrested progress through a normal transitional state that is regulated by the relative abundance of heterodimers between MyoD and the full-length E2A proteins. The demonstration that multiple inhibitory mechanisms can be suppressed and myogenic differentiation can be induced in the RD rhabdomyosarcomas by increasing the abundance of MyoD:E-protein heterodimers suggests a central integrating function that can be targeted to force differentiation in muscle cancer cells.

Spinocerebellar ataxia type 20 (SCA20) has been linked to chromosome 11q12, but the underlying genetic defect has yet to be identified. We applied single-nucleotide polymorphism genotyping to detect structural alterations in the genomic DNA of patients with SCA20. We found a 260 kb duplication within the previously linked SCA20 region, which was confirmed by quantitative polymerase chain reaction and fiber fluorescence in situ hybridization, the latter also showing its direct orientation. The duplication spans 10 known and 2 unknown genes, and is present in all affected individuals in the single reported SCA20 pedigree. While the mechanism whereby this duplication may be pathogenic remains to be established, we speculate that the critical gene within the duplicated segment may be DAGLA, the product of which is normally present at the base of Purkinje cell dendritic spines and contributes to the modulation of parallel fiber-Purkinje cell synapses.

During early divisions of the C. elegans embryo, many maternally supplied determinants accumulate asymmetrically, and this asymmetry is crucial for proper cell fate specification. SKN-1, a transcription factor whose message is maternally supplied to the embryo, specifies the mesendodermal cell fate. In the 2-cell embryo, SKN-1 is expressed at a higher level in the posterior cell. This asymmetry becomes more pronounced at the 4-cell stage, when SKN-1 is high in the posterior cell's daughters and low in the daughters of the anterior blastomere. To date, the direct mechanisms that control SKN-1 distribution remain unknown. In this report, we identify eel-1, which encodes a putative Hect E3 ubiquitin ligase that shares several domains of similarity to the mammalian E3 ligase Mule. EEL-1 binds SKN-1 and appears to target SKN-1 for degradation. EEL-1 has two functions in regulating SKN-1 during early embryogenesis. First, eel-1 promotes the spatial asymmetry of SKN-1 accumulation at the 2- and 4-cell stages. Second, eel-1 acts in all cells to downregulate SKN-1 from the 12- to the 28-cell stage. Although loss of eel-1 alone causes a reduction in SKN-1 asymmetry at the 2-cell stage, the function of eel-1 in both the spatial and temporal regulation of SKN-1 is redundant with the activities of other genes. These data strongly suggest that multiple, functionally redundant pathways cooperate to ensure precise control of SKN-1 asymmetry and persistence in the early embryo.

Cell fate specification in the early C. elegans embryo requires the activity of a family of proteins with CCCH zinc-finger motifs. Two members of the family, MEX-5 and MEX-6, are enriched in the anterior of the early embryo where they inhibit the accumulation of posterior proteins. Embryos from mex-5 single-mutant mothers are inviable due to the misexpression of SKN-1, a transcription factor that can specify mesoderm and endoderm. The aberrant expression of SKN-1 causes a loss of hypodermal and neuronal tissue and an excess of pharyngeal muscle, a Mex phenotype (muscle excess). POS-1, a third protein with CCCH motifs, is concentrated in the posterior of the embryo where it restricts the expression of at least one protein to the anterior. We discovered that reducing the dosage of pos-1(+) can suppress the Mex phenotype of mex-5(-) embryos and that POS-1 binds the 3'-UTR of mex-6. We propose that the suppression of the Mex phenotype by reducing pos-1(+) is due to decreased repression of mex-6 translation. Our detailed analyses of these protein functions reveal complex interactions among the CCCH finger proteins and suggest that their complementary expression patterns might be refined by antagonistic interactions among them.

The Saccharomyces cerevisiae Mre11p/Rad50p/Xrs2p (MRX) complex is evolutionarily conserved and functions in DNA repair and at telomeres [1-3]. In vivo, MRX is required for a 5' --> 3' exonuclease activity that mediates DNA recombination at double-strand breaks (DSBs). Paradoxically, abolition of this exonuclease activity in MRX mutants results in shortened telomeric DNA tracts. To further explore the role of MRX at telomeres, we analyzed MRX mutants in a de novo telomere addition assay in yeast cells [4]. We found that the MRX genes were absolutely required for telomerase-mediated addition in this assay. Furthermore, we found that Cdc13p, a single-stranded telomeric DNA binding protein essential for telomere DNA synthesis and protection [5], was unable to bind to the de novo telomeric DNA substrate in cells lacking Rad50p. Based on the results from this model system, we propose that the MRX complex helps to prepare telomeric DNA for the loading of Cdc13p, which then protects the chromosome from further degradation and recruits telomerase and other DNA replication components to synthesize telomeric DNA.

The telomerase enzyme lengthens telomeres, an activity essential for chromosome stability in most eukaryotes. The enzyme is composed of a specialized reverse transcriptase and a template RNA. In Saccharomyces cerevisiae, overexpression of TLC1, the telomerase RNA gene, disrupts telomeric structure. The result is both shortened telomere length and loss of a special chromatin structure that normally silences telomere-proximal genes. Because telomerase function is not required for telomeric silencing, we postulated that the dominant-negative effect caused by overexpression of TLC1 RNA originates in a normal interaction between the RNA and an unknown telomeric factor important for silencing; the overexpressed RNA presumably continues to bind the factor and compromises its function. Here we show that a 48-nt stem-loop structure within the 1.3-kb TLC1 RNA is necessary and sufficient for disrupting telomeric silencing and shortening telomeres. Moreover, this short RNA sequence appears to function through an interaction with the conserved DNA end-binding protein Ku. We propose that, in addition to its roles in telomeric silencing, homologous recombination and non-homologous end-joining (NHEJ), S. cerevisiae Ku also helps to recruit or activate telomerase at the telomere through an interaction with this stem-loop of TLC1 RNA.

To better understand the requirements for telomerase-mediated telomere addition in vivo, we developed an assay in S. cerevisiae that creates a chromosome end immediately adjacent to a short telomeric DNA tract. The de novo end acts as a telomere: it is protected from degradation in a CDC13-dependent manner, telomeric sequences are added efficiently, and addition occurs at a faster rate in mutant strains that have long telomeres. Telomere addition was detected in M phase arrested cells, which permitted us to determine that the essential DNA polymerases alpha and delta and DNA primase were required. This indicates that telomeric DNA synthesis by telomerase is tightly coregulated with the production of the opposite strand. Such coordination prevents telomerase from generating excessively long single-stranded tails, which may be deleterious to chromosome stability in S. cerevisiae.

Nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) are members of a family of trophic factors designated the neurotrophins, each of which can bind to the low-affinity NGF receptor (LNGFR). To investigate the mechanisms that regulate the expression of the neurotrophins and the LNGFR in the developing brain, we grew cells from the embryonic mouse septum and hippocampus in reaggregating cell culture and compared neurotrophin and LNGFR expression in developing reaggregates with that seen in the developing septum and hippocampus in situ. NGF, BDNF, NT-3 and LNGFR were each expressed in septal and hippocampal reaggregates as well as the native septum and hippocampus. Additionally, the temporal expression profiles observed in reaggregates were generally similar to those seen in the respective brain regions in situ. In order to determine whether NGF can modulate neurotrophin or LNGFR expression, reaggregates were cultured in the continual presence of either exogenous NGF or anti-NGF antibodies. NGF-treated septal cultures expressed twice the level of LNGFR mRNA as was seen in untreated septal cultures; on the other hand, septal cultures grown in the presence of anti-NGF antibodies, to neutralize endogenously synthesized NGF, displayed a 3-fold decrease in LNGFR mRNA expression compared to untreated cultures. No effects of NGF or anti-NGF were observed on LNGFR expression in hippocampal reaggregates, or on neurotrophin mRNA expression in either reaggregate type. These results suggest that regulatory mechanisms intrinsic to the septal and hippocampal regions control neurotrophin and LNGFR expression. NGF is likely to be one of these regulatory cues since it acts locally in septal reaggregates to control the developmental expression of LNGFR mRNA. The possible roles of locally synthesized NGF and other neurotrophins in the development of septal neurons are discussed.