Targeted therapies directed at commonly overexpressed pathways in melanoma have clinical activity in numerous trials.,Little is known about how these therapies influence microRNA (miRNA) expression, particularly with combination regimens.,Knowledge of miRNAs altered with treatment may contribute to understanding mechanisms of therapeutic effects, as well as mechanisms of tumor escape from therapy.,We analyzed miRNA expression in metastatic melanoma tissue samples treated with a novel combination regimen of Temsirolimus and Bevacizumab.,Given the preliminary clinical activity observed with this combination regimen, we hypothesized that we would see significant changes in miRNA expression with combination treatment.,Using microarray analysis we analyzed miRNA expression levels in melanoma samples from a Cancer Therapy Evaluation Program-sponsored phase II trial of combination Temsirolimus and Bevacizumab in advanced melanoma, which elicited clinical benefit in a subset of patients.,Pre-treatment and post-treatment miRNA levels were compared using paired t-tests between sample groups (patients), using a p-value < 0.01 for significance.,microRNA expression remained unchanged with Temsirolimus alone; however, expression of 15 microRNAs was significantly upregulated (1.4 to 2.5-fold) with combination treatment, compared to pre-treatment levels.,Interestingly, twelve of these fifteen miRNAs possess tumor suppressor capabilities.,We identified 15 putative oncogenes as potential targets of the 12 tumor suppressor miRNAs, based on published experimental evidence.,For 15 of 25 miRNA-target mRNA pairings, changes in gene expression from pre-treatment to post-combination treatment samples were inversely correlated with changes in miRNA expression, supporting a functional effect of those miRNA changes.,Clustering analyses based on selected miRNAs suggest preliminary signatures characteristic of clinical response to combination treatment and of tumor BRAF mutational status.,To our knowledge, this is the first study analyzing miRNA expression in pre-treatment and post-treatment human metastatic melanoma tissue samples.,This preliminary investigation suggests miRNAs that may be involved in the mechanism of action of combination Temsirolimus and Bevacizumab in metastatic melanoma, possibly through inhibition of oncogenic pathways, and provides the preliminary basis for further functional studies of these miRNAs.
Despite limited clinical efficacy, treatment with dacarbazine or temozolomide (TMZ) remains the standard therapy for metastatic melanoma.,In glioblastoma, promoter methylation of the counteracting DNA repair enzyme O6-methylguanine-DNA-methyltransferase (MGMT) correlates with survival of patients exposed to TMZ in combination with radiotherapy.,For melanoma, data are limited and controversial.,Biopsy samples from 122 patients with metastatic melanoma being treated with TMZ in two multicenter studies of the Dermatologic Cooperative Oncology Group were investigated for MGMT promoter methylation.,We used the COBRA (combined bisulphite restriction analysis) technique to determine aberrant methylation of CpG islands in small amounts of genomic DNA isolated from paraffin-embedded tissue sections.,To detect aberrant methylation, bisulphite-treated DNA was amplified by PCR, enzyme restricted, and visualised by gel electrophoresis.,Correlation with clinical data from 117 evaluable patients in a best-response evaluation indicated no statistically significant association between MGMT promoter methylation status and response.,A methylated MGMT promoter was observed in 34.8% of responders and 23.4% of non-responders (P=0.29).,In addition, no survival advantage for patients with a methylated MGMT promoter was detectable (P=0.79).,Interestingly, we found a significant correlation between MGMT methylation and tolerance of therapy.,Patients with a methylated MGMT promoter had more severe adverse events, requiring more TMZ dose reductions or discontinuations (P=0.007; OR 2.7 (95% CI: 1.32-5.7)).,Analysis of MGMT promoter methylation comparing primaries and different metastases over the clinical course revealed no statistical difference (P=0.49).,In advanced melanoma MGMT promoter, methylation correlates with tolerance of therapy, but not with clinical outcome.