IGFBP7 is not required for B-RAF-induced melanocyte senescence.

Induction of senescence permanently restricts cellular proliferation after oncogenic stimulation thereby acting as a potent barrier to tumor development. The relevant effector proteins may therefore be fundamental to cancer development. A recent study identified IGFBP7 as a secreted factor mediating melanocyte senescence induced by oncogenic B-RAF, which is found commonly in cutaneous nevi. In contrast to the previous report, we demonstrate that B-RAF signaling does not induce IGFBP7 expression, nor the expression of the IGFBP7 targets, BNIP3L, SMARCB1, or PEA15, in human melanocytes or fibroblasts. We also found no correlation between B-RAF mutational status and IGFBP7 protein expression levels in 22 melanoma cell lines, 90 melanomas, and 46 benign nevi. Furthermore, using a lentiviral silencing strategy we show that B-RAF induces senescence in melanocytes and fibroblasts, irrespective of the presence of IGFBP7. Therefore, we conclude that the secreted protein IGFBP7 is dispensable for B-RAF(V600E)-induced senescence in human melanocytes.

Whole-genome landscapes of major melanoma subtypes.

Melanoma of the skin is a common cancer only in Europeans, whereas it arises in internal body surfaces (mucosal sites) and on the hands and feet (acral sites) in people throughout the world. Here we report analysis of whole-genome sequences from cutaneous, acral and mucosal subtypes of melanoma. The heavily mutated landscape of coding and non-coding mutations in cutaneous melanoma resolved novel signatures of mutagenesis attributable to ultraviolet radiation. However, acral and mucosal melanomas were dominated by structural changes and mutation signatures of unknown aetiology, not previously identified in melanoma. The number of genes affected by recurrent mutations disrupting non-coding sequences was similar to that affected by recurrent mutations to coding sequences. Significantly mutated genes included BRAF, CDKN2A, NRAS and TP53 in cutaneous melanoma, BRAF, NRAS and NF1 in acral melanoma and SF3B1 in mucosal melanoma. Mutations affecting the TERT promoter were the most frequent of all; however, neither they nor ATRX mutations, which correlate with alternative telomere lengthening, were associated with greater telomere length. Most melanomas had potentially actionable mutations, most in components of the mitogen-activated protein kinase and phosphoinositol kinase pathways. The whole-genome mutation landscape of melanoma reveals diverse carcinogenic processes across its subtypes, some unrelated to sun exposure, and extends potential involvement of the non-coding genome in its pathogenesis.

Musculoskeletal immune-related adverse events with the use of checkpoint inhibitors in malignancy.

BACKGROUND: Immunotherapy has revolutionised the treatment of many malignancies. Along with their success, there have been inflammatory and immune-related adverse events (irAE). There is a paucity of literature describing the Australian experience of rheumatic irAE. AIMS: To describe and characterise the Royal North Shore Hospital and Westmead Hospital cohort with rheumatic irAE. METHODS: This case series reports on 17 patients with advanced cancer treated at two sites in Sydney, Australia who were referred for rheumatological evaluation from 2013 to 2016. Data were collected retrospectively and inspected for clinical signs, duration of immunotherapy prior to onset of symptoms, management strategies and cancer outcomes. RESULTS: Patients presented with arthralgias, myalgias, periarticular and systemic symptoms. Onset of rheumatological symptoms was variable, with a median of 4 months (range 0.2-24) for monotherapy and 5.05 months (range 0.2-6.9) for combination. The predominant findings were of tenosynovitis (23%) and large joint involvement (65%). All patients were seronegative for RF and anti-CCP. Most patients responded well to non-steroidal anti-inflammatory drugs or low-dose prednisone (59%) and remained on immunotherapy (77%). The majority (76%) of patients experienced concomitant irAE in other organ systems. Sixty-five percent of patients had complete response of their malignancy to immunotherapy. CONCLUSION: Rheumatic irAE are heterogenous clinical entities, which require further evaluation into classification, patient susceptibility and response. From our study, there was no clear clinical pattern. The present case series supports that rheumatic irAE may be associated with tumour response. However, there is still limited experience in rheumatic irAE management and outcomes.

First-in-human, phase I/IIa study of CRLX301, a nanoparticle drug conjugate containing docetaxel, in patients with advanced or metastatic solid malignancies.

Background This was a phase I/IIa study to investigate the tolerability, efficacy and pharmacokinetics (PK)/ pharmacodynamics (PD) of CRLX301, CDP-based nanoparticle formulation of docetaxel. Methods The study was conducted in two parts. In part 1, dose-escalation using a standard 3 + 3 design was performed in two dosing schedules (every week (QW) and every 3 weeks (Q3W)). Part 2 was comprised of a dose expansion at 75 mg/m2 Q3W. PK studies were performed on both dosing schedules. Results Forty-two patients were recruited onto the study with a median age of 64(range 38-76); median number of prior systemic therapies was 5(range 0-10). Grade 3/4 treatment-related toxicities included: neutropenia (21.4 %), infusion related reaction (11.9 %), anemia (7.1 %), fatigue (4.8 %), diarrhea (4.8 %), and peripheral neuropathy (4.8 %). The maximum tolerated dose was 75 mg/m2 given on the Q3W schedule and was not determined on the QW schedule. In this heavily pre-treated population, four patients (12.9 %) achieved stable disease (SD) ≥ 4 months and 2 patients (6.5 %) achieved partial response (PR) for a clinical benefit rate (CBR) of 19.4 % (6/31 patients). The PRs were seen in prostate and breast adenocarcinoma (one each). CRLX301 exhibited some PK advantages over docetaxel including higher retention of drug in plasma, slower clearance and controlled slow release of docetaxel from the carrier. Conclusions In this heavily pretreated patient population, the safety profile was acceptable for CRLX301 therapy. There was some evidence of preliminary tumor efficacy, but further work is necessary to find the optimal dose and schedule of this formulation.Clinicaltrials.gov trial registration number: NCT02380677 (Date of registration: March 2, 2015).

Efficacy of novel immunotherapy regimens in patients with metastatic melanoma with germline CDKN2A mutations.

BACKGROUND: Inherited CDKN2A mutation is a strong risk factor for cutaneous melanoma. Moreover, carriers have been found to have poor melanoma-specific survival. In this study, responses to novel immunotherapy agents in CDKN2A mutation carriers with metastatic melanoma were evaluated. METHODS: CDKN2A mutation carriers that have developed metastatic melanoma and undergone immunotherapy treatments were identified among carriers enrolled in follow-up studies for familial melanoma. The carriers' responses were compared with responses reported in phase III clinical trials for CTLA-4 and PD-1 inhibitors. From publicly available data sets, melanomas with somatic CDKN2A mutation were analysed for association with tumour mutational load. RESULTS: Eleven of 19 carriers (58%) responded to the therapy, a significantly higher frequency than observed in clinical trials (p=0.03, binomial test against an expected rate of 37%). Further, 6 of the 19 carriers (32%) had complete response, a significantly higher frequency than observed in clinical trials (p=0.01, binomial test against an expected rate of 7%). In 118 melanomas with somatic CDKN2A mutations, significantly higher total numbers of mutations were observed compared with 761 melanomas without CDKN2A mutation (Wilcoxon test, p<0.001). CONCLUSION: Patients with CDKN2A mutated melanoma may have improved immunotherapy responses due to increased tumour mutational load, resulting in more neoantigens and stronger antitumorous immune responses.

Adjuvant dabrafenib plus trametinib versus placebo in patients with resected, BRAFV600-mutant, stage III melanoma (COMBI-AD): exploratory biomarker analyses from a randomised, phase 3 trial.

BACKGROUND: Adjuvant dabrafenib plus trametinib reduced the risk of relapse versus placebo in patients with resected, BRAFV600-mutant, stage III melanoma in the phase 3 COMBI-AD trial. This prespecified exploratory biomarker analysis aimed to evaluate potential prognostic or predictive factors and mechanisms of resistance to adjuvant targeted therapy. METHODS: COMBI-AD is a randomised, double-blind, placebo-controlled, phase 3 trial comparing dabrafenib 150 mg orally twice daily plus trametinib 2 mg orally once daily versus two matched placebos. Study participants were at least 18 years of age and underwent complete resection of stage IIIA (lymph node metastases >1 mm), IIIB, or IIIC cutaneous melanoma, per American Joint Committee on Cancer 7th edition criteria, with a BRAFV600E or BRAFV600K mutation. Patients were randomly assigned (1:1) to the two treatment groups by an interactive voice response system, stratified by mutation type and disease stage. Patients, physicians, and the investigators who analysed data were masked to treatment allocation. The primary outcome was relapse-free survival, defined as the time from randomisation to disease recurrence or death from any cause. Biomarker assessment was a prespecified exploratory outcome of the trial. We assessed intrinsic tumour genomic features by use of next-generation DNA sequencing and characteristics of the tumour microenvironment by use of a NanoString RNA assay, which might provide prognostic and predictive information. This trial is registered with ClinicalTrials.gov, number NCT01682083, and is ongoing but no longer recruiting participants. FINDINGS: Between Jan 31, 2013, and Dec 11, 2014, 870 patients were enrolled in the trial. Median follow-up at data cutoff (April 30, 2018) was 44 months (IQR 38-49) in the dabrafenib plus trametinib group and 42 months (21-49) in the placebo group. Intrinsic tumour genomic features were assessed in 368 patients (DNA sequencing set) and tumour microenvironment characteristics were assessed in 507 patients (NanoString biomarker set). MAPK pathway genomic alterations at baseline did not affect treatment benefit or clinical outcome. An IFNγ gene expression signature higher than the median was prognostic for prolonged relapse-free survival in both treatment groups. Tumour mutational burden was independently prognostic for relapse-free survival in the placebo group (high TMB, top third; hazard ratio [HR] 0·56, 95% CI 0·37-0·85, p=0·0056), but not in the dabrafenib plus trametinib group (0·83, 95% CI 0·53-1·32, p=0·44). Patients with tumour mutational burden in the lower two terciles seem to derive a substantial long-term relapse-free survival benefit from targeted therapy (HR [versus placebo] 0·49, 95% CI 0·35-0·68, p<0·0001). However, patients with high tumour mutational burden seem to have a less pronounced benefit with targeted therapy (HR [versus placebo] 0·75, 95% CI 0·44-1·26, p=0·27), especially if they had an IFNγ signature lower than the median (HR 0·88 [95% CI 0·40-1·93], p=0·74). INTERPRETATION: Tumour mutational burden alone or in combination with IFNγ gene expression signature or other markers for an adaptive immune response might be of relevance for identifying patients with stage III melanoma who might derive clinical benefit from targeted therapy. Further validation in prospective clinical trials is warranted. FUNDING: Novartis Pharmaceuticals.

Adjuvant Dabrafenib plus Trametinib in Stage III BRAF-Mutated Melanoma.

BACKGROUND: Combination therapy with the BRAF inhibitor dabrafenib plus the MEK inhibitor trametinib improved survival in patients with advanced melanoma with BRAF V600 mutations. We sought to determine whether adjuvant dabrafenib plus trametinib would improve outcomes in patients with resected, stage III melanoma with BRAF V600 mutations. METHODS: In this double-blind, placebo-controlled, phase 3 trial, we randomly assigned 870 patients with completely resected, stage III melanoma with BRAF V600E or V600K mutations to receive oral dabrafenib at a dose of 150 mg twice daily plus trametinib at a dose of 2 mg once daily (combination therapy, 438 patients) or two matched placebo tablets (432 patients) for 12 months. The primary end point was relapse-free survival. Secondary end points included overall survival, distant metastasis-free survival, freedom from relapse, and safety. RESULTS: At a median follow-up of 2.8 years, the estimated 3-year rate of relapse-free survival was 58% in the combination-therapy group and 39% in the placebo group (hazard ratio for relapse or death, 0.47; 95% confidence interval [CI], 0.39 to 0.58; P<0.001). The 3-year overall survival rate was 86% in the combination-therapy group and 77% in the placebo group (hazard ratio for death, 0.57; 95% CI, 0.42 to 0.79; P=0.0006), but this level of improvement did not cross the prespecified interim analysis boundary of P=0.000019. Rates of distant metastasis-free survival and freedom from relapse were also higher in the combination-therapy group than in the placebo group. The safety profile of dabrafenib plus trametinib was consistent with that observed with the combination in patients with metastatic melanoma. CONCLUSIONS: Adjuvant use of combination therapy with dabrafenib plus trametinib resulted in a significantly lower risk of recurrence in patients with stage III melanoma with BRAF V600E or V600K mutations than the adjuvant use of placebo and was not associated with new toxic effects. (Funded by GlaxoSmithKline and Novartis; COMBI-AD ClinicalTrials.gov, NCT01682083 ; EudraCT number, 2012-001266-15 .).

bcGST-an interactive bias-correction method to identify over-represented gene-sets in boutique arrays.

MOTIVATION: Gene annotation and pathway databases such as Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes are important tools in Gene-Set Test (GST) that describe gene biological functions and associated pathways. GST aims to establish an association relationship between a gene-set of interest and an annotation. Importantly, GST tests for over-representation of genes in an annotation term. One implicit assumption of GST is that the gene expression platform captures the complete or a very large proportion of the genome. However, this assumption is neither satisfied for the increasingly popular boutique array nor the custom designed gene expression profiling platform. Specifically, conventional GST is no longer appropriate due to the gene-set selection bias induced during the construction of these platforms. RESULTS: We propose bcGST, a bias-corrected GST by introducing bias-correction terms in the contingency table needed for calculating the Fisher's Exact Test. The adjustment method works by estimating the proportion of genes captured on the array with respect to the genome in order to assist filtration of annotation terms that would otherwise be falsely included or excluded. We illustrate the practicality of bcGST and its stability through multiple differential gene expression analyses in melanoma and the Cancer Genome Atlas cancer studies. AVAILABILITY AND IMPLEMENTATION: The bcGST method is made available as a Shiny web application at http://shiny.maths.usyd.edu.au/bcGST/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Patient-reported outcomes in patients with resected, high-risk melanoma with BRAFV600E or BRAFV600K mutations treated with adjuvant dabrafenib plus trametinib (COMBI-AD): a randomised, placebo-controlled, phase 3 trial.

BACKGROUND: In the phase 3 COMBI-AD study, patients with resected, stage III melanoma with BRAFV600E or BRAFV600K mutations received adjuvant dabrafenib plus trametinib or placebo. The primary analysis showed that dabrafenib plus trametinib significantly improved relapse-free survival at 3 years. These results led to US Food and Drug Administration approval of dabrafenib plus trametinib as adjuvant treatment for patients with resected stage III melanoma with BRAFV600E or BRAFV600K mutations. Here, we report the patient-reported outcomes from COMBI-AD. METHODS: COMBI-AD was a randomised, double-blind, placebo-controlled, phase 3 study done at 169 sites in 25 countries. Study participants were aged 18 years or older and had complete resection of stage IIIA (lymph node metastases >1 mm), IIIB, or IIIC cutaneous melanoma as per American Joint Committee on Cancer 7th edition criteria, with BRAFV600E or BRAFV600K mutations, and an Eastern Cooperative Oncology Group performance status of 0 or 1. Patients were randomly assigned (1:1) via an interactive voice response system, stratified by mutation type and disease stage, to receive oral dabrafenib (150 mg twice daily) plus oral trametinib (2 mg once daily) or matching placebos for 12 months. Patients, physicians, and the investigators who analysed the data were masked to treatment allocation. The primary endpoint was relapse-free survival, reported elsewhere. Health-related quality of life, reported here, was a prespecified exploratory endpoint, and was assessed with the European Quality of Life 5-Dimensions 3-Levels (EQ-5D-3L) questionnaire in the intention-to-treat population. We used a mixed-model repeated-measures analysis to assess differences in health-related quality of life between groups. This study is registered with ClinicalTrials.gov, number NCT01682083. The trial is ongoing, but is no longer recruiting participants. FINDINGS: Between Jan 31, 2013, and Dec 11, 2014, 870 patients were enrolled and randomly assigned to receive dabrafenib plus trametinib (n=438) or matching placebos (n=432). Data were collected until the data cutoff for analyses of the primary endpoint (June 30, 2017). The median follow-up was 34 months (IQR 28-39) in the dabrafenib plus trametinib group and 33 months (20·5-39) in the placebo group. During the 12-month treatment phase, there were no significant or clinically meaningful changes from baseline between groups in EQ-5D-3L visual analogue scale (EQ-VAS) or utility scores. During treatment, there were no clinically meaningful differences in VAS scores or utility scores in the dabrafenib plus trametinib group between patients who did and did not experience the most common adverse events. During long-term follow-up (range 15-48 months), VAS and utility scores were similar between groups and did not differ from baseline scores. At recurrence, there were significant decreases in VAS scores in both the dabrafenib plus trametinib group (mean change -6·02, SD 20·57; p=0·0032) and the placebo group (-6·84, 20·86; p<0·0001); the mean change in utility score also differed significantly at recurrence for both groups (dabrafenib plus trametinib -0·0626, 0·1911, p<0·0001; placebo -0·0748, 0·2182, p<0·0001). INTERPRETATION: These findings show that dabrafenib plus trametinib did not affect patient-reported outcome scores during or after adjuvant treatment, and suggest that preventing or delaying relapse with adjuvant therapy could be beneficial in this setting. FUNDING: Novartis.

Neoadjuvant dabrafenib combined with trametinib for resectable, stage IIIB-C, BRAFV600 mutation-positive melanoma (NeoCombi): a single-arm, open-label, single-centre, phase 2 trial.

BACKGROUND: Adjuvant dabrafenib plus trametinib therapy improves relapse-free survival in patients with resected stage III melanoma. We aimed to ascertain the proportion of patients who would have a pathological response and a response according to Response Evaluation Criteria in Solid Tumors (RECIST) after neoadjuvant dabrafenib plus trametinib therapy for resectable clinical stage III melanoma. METHODS: NeoCombi was a single-arm, open-label, single-centre, phase 2 study done at Melanoma Institute Australia (Sydney, NSW, Australia). Eligible patients were adults (aged ≥18 years) with histologically confirmed, resectable, RECIST-measurable, clinical stage IIIB-C (American Joint Committee on Cancer [AJCC] 7th edition), BRAFV600-mutant melanoma, and had an Eastern Cooperative Oncology Group performance status of 1 or lower. Patients received 150 mg dabrafenib orally, twice daily, plus 2 mg trametinib orally, once daily, for 52 weeks (12 weeks of neoadjuvant therapy before complete resection of the pre-therapy tumour bed, and 40 weeks of adjuvant therapy thereafter). CT and PET scans were done at baseline and before resection. The primary outcomes were the proportion of patients achieving a complete pathological response and the proportion of patients achieving a response according to RECIST at week 12, analysed as per protocol. This trial is registered with ClinicalTrials.gov, NCT01972347, and follow-up of patients is ongoing. FINDINGS: Between Aug 20, 2014, and April 19, 2017, 40 patients were screened, of whom 35 eligible patients were enrolled, received neoadjuvant dabrafenib plus trametinib, and underwent resection. At the data cutoff (Sept 24, 2018), median follow-up was 27 months (IQR 21-36). At resection, 30 (86%) patients achieved a RECIST response; 16 (46%; 95% CI 29-63) had a complete response and 14 (40%; 24-58) had a partial response. Five patients (14%; 95% CI 5-30) had stable disease, and no patients progressed. After resection and pathological evaluation, all 35 patients achieved a pathological response, of whom 17 (49%; 95% CI 31-66) patients had a complete pathological response and 18 (51%; 95% CI 34-69) had a non-complete pathological response. Treatment-related serious adverse events occurred in six (17%) of 35 patients and grade 3-4 adverse events occurred in ten (29%) patients. No treatment-related deaths were reported. INTERPRETATION: Neoadjuvant dabrafenib plus trametinib therapy could be considered in the management of RECIST-measurable resectable stage III melanoma as it led to a high proportion of patients achieving a complete response according to RECIST and a high proportion of patients achieving a complete pathological response, with no progression during neoadjuvant therapy. FUNDING: GlaxoSmithKline; Novartis; National Health and Medical Research Council, Australia; and Melanoma Institute Australia.

Whole genome sequencing of melanomas in adolescent and young adults reveals distinct mutation landscapes and the potential role of germline variants in disease susceptibility.

Cutaneous melanoma accounts for at least >10% of all cancers in adolescents and young adults (AYA, 15-30 years of age) in Western countries. To date, little is known about the correlations between germline variants and somatic mutations and mutation signatures in AYA melanoma patients that might explain why they have developed a cancer predominantly affecting those over 65 years of age. We performed genomic analysis of 50 AYA melanoma patients (onset 10-30 years, median 20); 25 underwent whole genome sequencing (WGS) of both tumor and germline DNA, exome data were retrieved from 12 TCGA AYA cases, and targeted DNA sequencing was conducted on 13 cases. The AYA cases were compared with WGS data from 121 adult cutaneous melanomas. Similar to mature adult cutaneous melanomas, AYA melanomas showed a high mutation burden and mutation signatures of ultraviolet radiation (UVR) damage. The frequencies of somatic mutations in BRAF (96%) and PTEN (36%) in the AYA WGS cohort were double the rates observed in adult melanomas (Q < 6.0 × 10-6 and 0.028, respectively). Furthermore, AYA melanomas contained a higher proportion of non-UVR-related mutation signatures than mature adult melanomas as a proportion of total mutation burden (p = 2.0 × 10-4 ). Interestingly, these non-UVR mutation signatures relate to APOBEC or mismatch repair pathways, and germline variants in related genes were observed in some of these cases. We conclude that AYA melanomas harbor some of the same molecular aberrations and mutagenic insults occurring in older adults, but in different proportions. Germline variants that may have conferred disease susceptibility correlated with somatic mutation signatures in a subset of AYA melanomas.

Liquid biomarkers in melanoma: detection and discovery.

A vast array of tumor-derived genetic, proteomic and cellular components are constantly released into the circulation of cancer patients. These molecules including circulating tumor DNA and RNA, proteins, tumor and immune cells are emerging as convenient and accurate liquid biomarkers of cancer. Circulating cancer biomarkers provide invaluable information on cancer detection and diagnosis, prognosticate patient outcomes, and predict treatment response. In this era of effective molecular targeted treatments and immunotherapies, there is now an urgent need to implement use of these circulating biomarkers in the clinic to facilitate personalized therapy. In this review, we present recent findings in circulating melanoma biomarkers, examine the challenges and promise of evolving technologies used for liquid biomarker discovery, and discuss future directions and perspectives in melanoma biomarker research.

Standard-dose pembrolizumab in combination with reduced-dose ipilimumab for patients with advanced melanoma (KEYNOTE-029): an open-label, phase 1b trial.

BACKGROUND: Reduced-dose nivolumab in combination with standard-dose ipilimumab improves objective response and progression-free survival compared with standard-dose ipilimumab alone, but increases toxicity. We assessed the safety and anti-tumour activity of standard-dose pembrolizumab in combination with reduced-dose ipilimumab. METHODS: In this open-label, phase 1b trial, we recruited patients from 12 medical centres in Australia, New Zealand, and the USA. Eligible patients were aged at least 18 years, had advanced melanoma, had an Eastern Coooperative Oncology Group performance status of 0 or 1, had measurable disease according to the Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1, had adequate organ function, had resolution of toxic effects of the most recent previous chemotherapy to grade 1 or less, had no active autoimmune disease requiring systemic steroids or immunosuppressive agents, had no active non-infectious pneumonitis, had no uncontrolled thyroid dysfunction or diabetes, had no active brain metastases, and had not received previous immune checkpoint inhibitor therapy. Patients received intravenous pembrolizumab 2 mg/kg plus intravenous ipilimumab 1 mg/kg every 3 weeks for four doses, followed by intravenous pembrolizumab 2 mg/kg every 3 weeks for up to 2 years or disease progression, intolerable toxicity, withdrawal of consent, or investigator decision. The primary endpoint was safety and tolerability. The proportion of patients achieving an objective response assessed per RECIST version 1.1 by independent central review and overall survival were secondary endpoints. We also assessed progression-free survival. The primary endpoint was assessed in all patients who received at least one dose of combination therapy. Activity was assessed in all enrolled patients. This trial is registered with ClinicalTrials.gov, number NCT02089685. Enrolment into this cohort is closed, but patients are still being monitored for safety and anti-tumour activity. FINDINGS: Between Jan 13, 2015, and Sept 17, 2015, we enrolled and treated 153 patients. As of the Oct 17, 2016, cutoff date, median follow-up was 17·0 months (IQR 14·8-18·8). 110 (72%) of 153 patients received all four pembrolizumab plus ipilimumab doses; 64 (42%) remained on pembrolizumab monotherapy. 110 grade 3-4 treatment-related adverse events occurred in 69 (45%) patients. No treatment-related deaths occurred. Treatment-related adverse events led to discontinuation of pembrolizumab and ipilimumab in 22 (14%) patients, including 17 (11%) who discontinued both treatments for the same event and five (3%) who discontinued ipilimumab for one event and later discontinued pembrolizumab for another. 12 (8%) patients discontinued ipilimumab only and 14 (9%) discontinued pembrolizumab only because of treatment-related adverse events. 158 immune-mediated adverse events of any grade occurred in 92 (60%) patients, and 50 immune-mediated adverse events of grade 3-4 occurred in 42 (27%) patients; the most common immune-mediated adverse events were hypothyroidism (25 [16%]) and hyperthyroidism (17 [11%]). 93 (61% [95% CI 53-69]) patients achieved an objective response. Estimated 1 year progression-free survival was 69% (95% CI 60-75), and estimated 1 year overall survival was 89% (95% CI 83-93). INTERPRETATION: Standard-dose pembrolizumab given in combination with four doses of reduced-dose ipilimumab followed by standard-dose pembrolizumab has a manageable toxicity profile and provides robust anti-tumour activity in patients with advanced melanoma. These data suggest that standard-dose pembrolizumab plus reduced-dose ipilimumab might be a tolerable, efficacious treatment option for patients with advanced melanoma. A randomised phase 2 trial of alternative dosing strategies of this combination is underway. FUNDING: Merck & Co, Inc.

Efficacy of anti-PD-1 therapy in patients with melanoma brain metastases.

BACKGROUND: There is limited data on the efficacy of anti-programmed death 1 (PD-1) antibodies in patients (pts) with melanoma brain metastasis (BM), particularly those which are symptomatic. METHOD: We retrospectively assessed pts with melanoma BM treated with PD-1 antibodies, nivolumab and pembrolizumab. Clinicopathologic and treatment parameters were collected and outcomes determined for intracranial (IC) response rate (RR) using a modified RECIST criteria, with up to five IC target lesions used to determine IC response, disease control rate (DCR) and progression-free survival (PFS). RESULTS: A total of 66 pts were identified with a median follow up of 7.0 months (range 0.8-24.5 months). A total of 68% were male and 45% BRAF V600 mutation positive. At PD-1 antibody commencement, 50% had an elevated LDH; 64% had local therapy to BM prior to commencing anti-PD1, of which 5% had surgical resection, 14% stereotactic radiosurgery (SRS), 18% whole-brain radiotherapy (WBRT), 27% had surgery and radiotherapy. Twenty-one per cent started anti-PD-1 as first line systemic therapy. No pt had prior anti-PD-1 treatment. The IC overall RR was 21 and DCR 56%. Responses occurred in 21% of pts with symptomatic BM. The median OS was 9.9 months (95% CI 6.93-17.74). Pts with symptomatic BM had shorter PFS than those without symptoms (2.7 vs 7.4 months, P=0.035) and numerically shorter OS (5.7 vs 13.0 months, P=0.068). Pts requiring corticosteroids also had a numerically shorter PFS (3.2 vs 7.4 months, P=0.081) and OS (4.8 vs 13.1 months, P=0.039). CONCLUSIONS: IC responses to anti-PD-1 antibodies occur in pts with BM, including those with symptomatic BM requiring corticosteroids. Prospective trials evaluating anti-PD-1 therapy in pts with BM are underway.

A novel recurrent mutation in MITF predisposes to familial and sporadic melanoma.

So far, two genes associated with familial melanoma have been identified, accounting for a minority of genetic risk in families. Mutations in CDKN2A account for approximately 40% of familial cases, and predisposing mutations in CDK4 have been reported in a very small number of melanoma kindreds. Here we report the whole-genome sequencing of probands from several melanoma families, which we performed in order to identify other genes associated with familial melanoma. We identify one individual carrying a novel germline variant (coding DNA sequence c.G1075A; protein sequence p.E318K; rs149617956) in the melanoma-lineage-specific oncogene microphthalmia-associated transcription factor (MITF). Although the variant co-segregated with melanoma in some but not all cases in the family, linkage analysis of 31 families subsequently identified to carry the variant generated a log of odds (lod) score of 2.7 under a dominant model, indicating E318K as a possible intermediate risk variant. Consistent with this, the E318K variant was significantly associated with melanoma in a large Australian case-control sample. Likewise, it was similarly associated in an independent case-control sample from the United Kingdom. In the Australian sample, the variant allele was significantly over-represented in cases with a family history of melanoma, multiple primary melanomas, or both. The variant allele was also associated with increased naevus count and non-blue eye colour. Functional analysis of E318K showed that MITF encoded by the variant allele had impaired sumoylation and differentially regulated several MITF targets. These data indicate that MITF is a melanoma-predisposition gene and highlight the utility of whole-genome sequencing to identify novel rare variants associated with disease susceptibility.

Survey of practices around pharmaceutical company funding for continuing professional development among medical oncologists and trainees in Australia.

BACKGROUND: The completion of continuing professional development (CPD) is mandatory for medical oncologists and trainees (MO&T). Pharmaceutical companies may fund some CPD activities, but there is increasing debate about the potential for conflicts of interest (COI). AIM: To assess current practices around funding to attend CPD activities. METHODS: An electronic survey was distributed to Australian MO&T. The survey asked questions about current practices, institutional policies and perceptions about attending CPD funded by pharmaceutical companies. The design looked at comparing responses between MO&T as well as their understanding of and training around institutional and ethical process. RESULTS: A total of 157 of 653 (24%) responses was received, the majority from MO (76%). Most CPD activities attended by MO&T were self-funded (53%), followed by funding from institutions (19%), pharmaceutical companies (16%) and salary award (16%). Most institutions allowed MO&T to receive CPD funding from professional organisations (104/157, 66%) or pharmaceutical companies (90/157, 57%). A minority of respondents (13/157, 8%) reported that the process to use pharmaceutical funds had been considered by an ethics committee. Although 103/157 (66%) had received pharmaceutical funding for CPD, most (109/157, 69%) reported never receiving training about potential COI. The lack of education was more noticeable among trainees (odds ratio (OR) 8.61, P = 0.02). MO&T acknowledged the potential bias towards a pharmaceutical product (P = 0.05) but believed there was adequate separation between themselves and pharmaceutical companies (P < 0.01). CONCLUSION: Majority of CPD attended by MO&T is self-funded. There is lack of clarity in institutional policies regarding external funding support for CPD activities. Formal education about potential COI is lacking.

Cessation of targeted therapy after a complete response in BRAF-mutant advanced melanoma: a case series.

BACKGROUND: It is unknown whether melanoma patients achieving complete response (CR) with targeted therapy can safely discontinue treatment. METHODS: All patients treated with BRAF/MEK inhibitors achieving CR and ceasing treatment before progression were identified. Clinical data at treatment initiation, cessation and progression were examined. RESULTS: A total of 12 eligible patients were identified, with median follow-up of 16 months, of whom 6 (50%) recurred at a median of 6.6 months after treatment cessation. One patient lost to follow-up until presentation with symptomatic recurrence was the only relapser to die. At relapse, the remaining five patients had an LDH <1.2 times ULN, four were ECOG 0 and one ECOG 1. Baseline characteristics and time to CR and to discontinuation did not influence the rate of relapse. CONCLUSIONS: A large proportion of patients achieving CR with BRAF/MEK inhibitors relapse after treatment cessation. The optimal treatment duration in such patients is unclear, particularly where alternative treatments are available.

Safety and tumor responses with lambrolizumab (anti-PD-1) in melanoma.

BACKGROUND: The programmed death 1 (PD-1) receptor is a negative regulator of T-cell effector mechanisms that limits immune responses against cancer. We tested the anti-PD-1 antibody lambrolizumab (previously known as MK-3475) in patients with advanced melanoma. METHODS: We administered lambrolizumab intravenously at a dose of 10 mg per kilogram of body weight every 2 or 3 weeks or 2 mg per kilogram every 3 weeks in patients with advanced melanoma, both those who had received prior treatment with the immune checkpoint inhibitor ipilimumab and those who had not. Tumor responses were assessed every 12 weeks. RESULTS: A total of 135 patients with advanced melanoma were treated. Common adverse events attributed to treatment were fatigue, rash, pruritus, and diarrhea; most of the adverse events were low grade. The confirmed response rate across all dose cohorts, evaluated by central radiologic review according to the Response Evaluation Criteria in Solid Tumors (RECIST), version 1.1, was 38% (95% confidence interval [CI], 25 to 44), with the highest confirmed response rate observed in the cohort that received 10 mg per kilogram every 2 weeks (52%; 95% CI, 38 to 66). The response rate did not differ significantly between patients who had received prior ipilimumab treatment and those who had not (confirmed response rate, 38% [95% CI, 23 to 55] and 37% [95% CI, 26 to 49], respectively). Responses were durable in the majority of patients (median follow-up, 11 months among patients who had a response); 81% of the patients who had a response (42 of 52) were still receiving treatment at the time of analysis in March 2013. The overall median progression-free survival among the 135 patients was longer than 7 months. CONCLUSIONS: In patients with advanced melanoma, including those who had had disease progression while they had been receiving ipilimumab, treatment with lambrolizumab resulted in a high rate of sustained tumor regression, with mainly grade 1 or 2 toxic effects. (Funded by Merck Sharp and Dohme; ClinicalTrials.gov number, NCT01295827.).

Cutaneous adverse events (AEs) of anti-programmed cell death (PD)-1 therapy in patients with metastatic melanoma: A single-institution cohort.

BACKGROUND: Anti-programmed cell death (PD)-1 therapy is emerging as the backbone of new standard of care immunotherapy for metastatic melanoma. Immune-related cutaneous events are observed in these patients. OBJECTIVE: We sought to describe cutaneous adverse events observed in patients with metastatic melanoma on anti-PD-1 therapy. METHODS: We reviewed the clinical and histologic information of all patients treated with single-agent anti-PD-1 therapy for metastatic melanoma at Westmead Hospital, Sydney, Australia, from May 2012 to February 2015. RESULTS: Of the 82 patients included in the study, 34 had dermatology assessments. Forty (49%) developed a form of anti-PD-1-associated cutaneous adverse events. In all, 17% developed lichenoid reactions and eczema, and 15% developed vitiligo. An estimated 25% of patients were expected to develop their first lichenoid reactions within 8.3 months, and eczema and vitiligo within 10.3 months of therapy. These adverse events tend to appear together in patients on anti-PD-1 therapy. LIMITATIONS: The study was from a single center and clinical information was reviewed retrospectively in patients not referred to dermatology. CONCLUSION: Anti-PD-1 therapy is associated with the development of immune-related cutaneous events. Lichenoid reactions, eczema, and vitiligo are the 3 most prevalent lesions observed in our population. There is a tendency for lichenoid reactions and eczema to occur with vitiligo.

Dynamics of chemokine, cytokine, and growth factor serum levels in BRAF-mutant melanoma patients during BRAF inhibitor treatment.

The purpose of this study is to profile the changes in the serum levels of a range of chemokines, cytokines, and growth and angiogenic factors in MAPK inhibitor-treated metastatic melanoma patients and to correlate these changes with clinical outcome and changes in melanoma tissue biopsies taken from the same patients. Forty-two chemokine, cytokine, angiogenic, and growth factors were measured in the sera of 20 BRAF inhibitor-treated and four combination BRAF and MEK inhibitor-treated metastatic melanoma patients using a multiplex chemokine assay. The changes were correlated with Ki-67 and CD8(+) tumor-infiltrating lymphocytes in the tumor biopsies taken at the same time points, as well as clinical outcome, including response rate, progression-free survival, and overall survival. Serum levels of IFN-γ, CCL4, and TNF-α were significantly increased, whereas CXCL8 significantly decreased from pretreatment (PRE) to early during treatment (EDT) serum samples. The decrease in serum CXCL8 levels from PRE to EDT significantly correlated with decreases in markers of melanoma proliferation (Ki-67) and increases in cytotoxic tumor-infiltrating T cells in corresponding tumor biopsies. In addition, a greater fold reduction in CXCL8 serum levels from PRE to EDT serum samples was associated with decreased overall survival. These results suggest that BRAF inhibition causes decreased CXCL8 secretion from melanoma cells and induce an immune response against the tumor associated with increased IFN-γ, CCL4, and TNF-α. Further studies are needed to determine if CXCL8 is predictive of response and to confirm the functions of these chemokine and cytokine in BRAF-mutant melanoma under BRAF inhibition.