plx-4720 and dabrafenib

While response rates to BRAF inhibitiors (BRAFi) are high, disease progression emerges quickly. One strategy to delay the onset of resistance is to target anti-apoptotic proteins such as BCL-2, known to be associated with a poor prognosis. We analyzed BCL-2 family member expression levels of 34 samples from 17 patients collected before and 10 to 14 days after treatment initiation with either vemurafenib or dabrafenib/trametinib combination. The observed changes in mRNA and protein levels with BRAFi treatment led us to hypothesize that combining BRAFi with a BCL-2 inhibitor (the BH3-mimetic navitoclax) would improve outcome. We tested this hypothesis in cell lines and in mice. Pretreatment mRNA levels of BCL-2 negatively correlated with maximal tumor regression. Early increases in mRNA levels were seen in BIM, BCL-XL, BID and BCL2-W, as were decreases in MCL-1 and BCL2A. No significant changes were observed with BCL-2. Using reverse phase protein array (RPPA), significant increases in protein levels were found in BIM and BID. No changes in mRNA or protein correlated with response. Concurrent BRAF (PLX4720) and BCL2 (navitoclax) inhibition synergistically reduced viability in BRAF mutant cell lines and correlated with down-modulation of MCL-1 and BIM induction after PLX4720 treatment. In xenograft models, navitoclax enhanced the efficacy of PLX4720. The combination of a selective BRAF inhibitor with a BH3-mimetic promises to be an important therapeutic strategy capable of enhancing the clinical efficacy of BRAF inhibition in many patients that might otherwise succumb quickly to de novo resistance. Trial registrations: ClinicalTrials.gov NCT01006980; ClinicalTrials.gov NCT01107418; ClinicalTrials.gov NCT01264380; ClinicalTrials.gov NCT01248936; ClinicalTrials.gov NCT00949702; ClinicalTrials.gov NCT01072175.

Topics: Adult; Aged; Aniline Compounds; Animals; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Drug Resistance, Neoplasm; Humans; Imidazoles; Indoles; Melanoma; Mice; Middle Aged; Mutation, Missense; Neoplasm Metastasis; Oximes; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-bcl-2; Pyridones; Pyrimidinones; Sulfonamides; Vemurafenib

Diverse pathways drive resistance to BRAF/MEK inhibitors in BRAF-mutant melanoma, suggesting that durable control of resistance will be a challenge. By combining statistical modeling of genomic data from matched pre-treatment and post-relapse patient tumors with functional interrogation of >20 in vitro and in vivo resistance models, we discovered that major pathways of resistance converge to activate the transcription factor, c-MYC (MYC). MYC expression and pathway gene signatures were suppressed following drug treatment, and then rebounded during progression. Critically, MYC activation was necessary and sufficient for resistance, and suppression of MYC activity using genetic approaches or BET bromodomain inhibition was sufficient to resensitize cells and delay BRAFi resistance. Finally, MYC-driven, BRAFi-resistant cells are hypersensitive to the inhibition of MYC synthetic lethal partners, including SRC family and c-KIT tyrosine kinases, as well as glucose, glutamine, and serine metabolic pathways. These insights enable the design of combination therapies that select against resistance evolution.

Topics: Antineoplastic Agents, Hormonal; Benzimidazoles; Cell Line, Tumor; Estradiol; Evolution, Molecular; Female; Fulvestrant; Humans; Imidazoles; Indoles; Male; Melanoma; Oximes; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-myc; Pyridones; Pyrimidinones; Quinolines; Signal Transduction; Sulfonamides

Topics: 9,10-Dimethyl-1,2-benzanthracene; Administration, Topical; Animals; Antineoplastic Agents; Carcinogens; Carcinoma, Squamous Cell; Cell Line; Cell Proliferation; Fluorouracil; Humans; Imidazoles; Indoles; Melanoma; Mice; Oximes; Proto-Oncogene Proteins B-raf; Skin Neoplasms; Sulfonamides; Tetradecanoylphorbol Acetate; Vemurafenib

To evaluate the effects of treatment with the potent mutant BRAF inhibitors GSK2118436 or vemurafenib (PLX4720) on immune responses to metastatic melanoma in tissues taken before and after treatment.. Thirty-seven tumor biopsies were collected from 15 patients with unresectable American Joint Committee on Cancer stage III or IV melanoma immediately before and approximately 7 days after the commencement of BRAF inhibitor treatment and at the time of tumor progression. Immunohistochemical staining was carried out on the biopsies using specific antibodies for CD8, CD4, CD20, CD1a, and Granzyme B.. Tumor infiltration by CD4(+) and CD8(+) lymphocytes increased markedly following BRAF inhibitor treatment (both ρ = 0.015). There was a correlation between the degree of tumor infiltration by CD8(+) and Granzyme B-expressing lymphocytes in post-BRAF inhibitor-treated biopsies (r = 0.690 and ρ = 0.013). Increased intratumoral CD8(+) lymphocyte expression was correlated with a reduction in tumor size and an increase in necrosis in posttreatment biopsies (r = -0.793, ρ = 0.011; and r = 0.761, ρ = 0.004, respectively).. The increase in tumor-infiltrating lymphocytes induced by treatment with BRAF inhibitors provides strong support for conducting trials that combine BRAF inhibitors with immunotherapy in the hope of prolonging clinical responses.

Topics: Adult; Aged; Female; Granzymes; Humans; Imidazoles; Indoles; Lymphocytes, Tumor-Infiltrating; Male; Melanoma; Middle Aged; Mutation; Neoplasm Metastasis; Neoplasm Staging; Oximes; Prognosis; Protein Kinase Inhibitors; Proto-Oncogene Proteins B-raf; Sulfonamides; T-Lymphocytes; Young Adult

Relatively little attention has been paid to the activity of selective BRAF inhibitors in the induction of apoptosis in melanoma, particularly in vivo. In the present study, we have isolated cultures from biopsies taken from four patients before and during the treatment of their melanoma. We report that the cell lines taken during treatment show varying degrees of upregulation of the proapoptotic BH3 protein Bim and its splice forms, downregulation of Mcl-1, and upregulation of the splicing factor SRp55 as reported in previous in-vitro studies. There was also evidence of ongoing apoptotic signaling despite the continued growth of the cultures. The cultures established during the treatment were largely resistant to the selective BRAF inhibitor PLX4720, consistent with the acquired resistance of melanoma in the treated patients. These results provide further insights into the mechanism of action of these agents against melanoma.

Topics: Adult; Aged; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Biopsy; Cell Proliferation; Cell Shape; Drug Resistance, Neoplasm; Extracellular Signal-Regulated MAP Kinases; Female; Humans; Imidazoles; Indoles; Male; Melanoma; Membrane Proteins; Middle Aged; Mitogen-Activated Protein Kinase Kinases; Nuclear Proteins; Oximes; Phosphoproteins; Protein Kinase Inhibitors; Proto-Oncogene Proteins; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; RNA-Binding Proteins; Serine-Arginine Splicing Factors; Skin Neoplasms; Sulfonamides; Time Factors; Treatment Outcome; Tumor Cells, Cultured; Up-Regulation