BRAF fusions identified in melanomas have variable treatment responses and phenotypes.

Oncogenic BRAF fusions have emerged as an alternate mechanism for BRAF activation in melanomas and other cancers. A number of BRAF fusions with different 5' gene partners and BRAF exon breakpoints have been described, but the effects of different partners and breakpoints on cancer phenotypes and treatment responses has not been well characterized. Targeted RNA sequencing was used to screen 60 melanoma patient-derived xenograft (PDX) models for BRAF fusions. We identified three unique BRAF fusions, including a novel SEPT3-BRAF fusion, occurring in four tumors (4/60, 6.7%), all of which were "pan-negative" (lacking other common mutations) (4/18, 22.2%). The BRAF fusion PDX models showed variable growth rates and responses to MAPK inhibitors in vivo. Overexpression of BRAF fusions identified in our study, as well as other BRAF fusions previously identified in melanomas, resulted in a high degree of variability in 2D proliferation and 3D invasion between the different fusions. While exogenously expressed BRAF fusions all responded to MAPK inhibition in vitro, we observed potential differences in signaling and feedback mechanisms. In summary, BRAF fusions are actionable therapeutic targets, however there are significant differences in phenotypes, treatment responses, and signaling which may be clinically relevant.

BH3 mimetics induce apoptosis independent of DRP-1 in melanoma.

Despite the recent advancement in treating melanoma, options are still limited for patients without BRAF mutations or in relapse from current treatments. BH3 mimetics against members of the BCL-2 family have gained excitement with the recent success in hematological malignancies. However, single drug BH3 mimetic therapy in melanoma has limited effectiveness due to escape by the anti-apoptotic protein MCL-1 and/or survival of melanoma-initiating cells (MICs). We tested the efficacy of the BH3 mimetic combination of A-1210477 (an MCL-1 inhibitor) and ABT-263 (a BCL-2/BCL-XL/BCL-W inhibitor) in killing melanoma, especially MICs. We also sought to better define Dynamin-Related Protein 1 (DRP-1)'s role in melanoma; DRP-1 is known to interact with members of the BCL-2 family and is a possible therapeutic target for melanoma treatment. We used multiple assays (cell viability, apoptosis, bright field, immunoblot, and sphere formation), as well as the CRISPR/Cas9 genome-editing techniques. For clinical relevance, we employed patient samples of different mutation status, including some relapsed from current treatments such as anti-PD-1 immunotherapy. We found the BH3 mimetic combination kill both the MICs and non-MICs (bulk of melanoma) in all cell lines and patient samples irrespective of the mutation status or relapsed state (p < 0.05). Unexpectedly, the major pro-apoptotic proteins, NOXA and BIM, are not necessary for the combination-induced cell death. Furthermore, the combination impedes the activation of DRP-1, and inhibition of DRP-1 further enhances apoptosis (p < 0.05). DRP-1 effects in melanoma differ from those seen in other cancer cells. These results provide new insights into BCL-2 family's regulation of the apoptotic pathway in melanoma, and suggest that inhibiting the major anti-apoptotic proteins is sufficient to induce cell death even without involvement from major pro-apoptotic proteins. Importantly, our study also indicates that DRP-1 inhibition is a promising adjuvant for BH3 mimetics in melanoma treatment.

ALK Inhibitor Response in Melanomas Expressing EML4-ALK Fusions and Alternate ALK Isoforms.

Oncogenic ALK fusions occur in several types of cancer and can be effectively treated with ALK inhibitors; however, ALK fusions and treatment response have not been characterized in malignant melanomas. Recently, a novel isoform of ALK (ALKATI ) was reported in 11% of melanomas but the response of melanomas expressing ALKATI to ALK inhibition has not been well characterized. We analyzed 45 melanoma patient-derived xenograft models for ALK mRNA and protein expression. ALK expression was identified in 11 of 45 (24.4%) melanomas. Ten melanomas express wild-type (wt) ALK and/or ALKATI and one mucosal melanoma expresses multiple novel EML4-ALK fusion variants. Melanoma cells expressing different ALK variants were tested for response to ALK inhibitors. Whereas the melanoma expressing EML4-ALK were sensitive to ALK inhibitors in vitro and in vivo, the melanomas expressing wt ALK or ALKATI were not sensitive to ALK inhibitors. In addition, a patient with mucosal melanoma expressing ALKATI was treated with an ALK/ROS1/TRK inhibitor (entrectinib) on a phase I trial but did not respond. Our results demonstrate ALK fusions occur in malignant melanomas and respond to targeted therapy, whereas melanomas expressing ALKATI do not respond to ALK inhibitors. Targeting ALK fusions is an effective therapeutic option for a subset of melanoma patients, but additional clinical studies are needed to determine the efficacy of targeted therapies in melanomas expressing wt ALK or ALKATIMol Cancer Ther; 17(1); 222-31. ©2017 AACR.

MIR29B regulates expression of MLLT11 (AF1Q), an MLL fusion partner, and low MIR29B expression associates with adverse cytogenetics and poor overall survival in AML.

MLLT11, an MLL fusion partner, is a poor prognostic biomarker for paediatric acute myeloid leukaemia (AML), adult normal cytogenetics AML, and adult myelodysplastic syndrome. MLLT11 is highly regulated during haematopoietic progenitor differentiation and development but its regulatory mechanisms have not been defined. In this study, we demonstrate by transfection experiments that MIR29B directly regulates MLLT11 expression in vitro. MIR29B expression level was also inversely related to MLLT11 expression in a cohort of 56 AML patients (P<0·05). AML patients with low MIR29B/elevated MLLT11 expression had poor overall survival (P=0·038). Therefore, MIR29B may be a potential prognostic biomarker for AML patients.