Functional characterization of a PROTAC directed against BRAF mutant V600E.
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ABSTRACT:
SUBMITTER: Posternak G
PROVIDER: S-EPMC7862923 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The RAF family kinases function in the RAS-ERK pathway to transmit signals from activated RAS to the downstream kinases MEK and ERK. This pathway regulates cell proliferation, differentiation, and survival enabling mutations in RAS and RAF to act as potent drivers of human cancers. Drugs targeting the prevalent oncogenic mutant BRAFV600E have shown great efficacy in the clinic but long-term effectiveness is limited by resistance mechanisms that often exploit the dimerization-dependent process by which RAF kinases are activated. Here, we investigated a proteolysis targeting chimera (PROTAC) approach to BRAF inhibition. The most effective PROTAC termed P4B displayed superior specificity and inhibitory properties relative to non-PROTAC controls in BRAFV600E cell lines. In addition, P4B displayed utility in two cell lines harboring alternate BRAF mutations that impart resistance to conventional BRAF inhibitors. This work provides a rationale for optimizing the drug-like properties of P4B to enable proof of concept studies in vivo.
2020-05-25 | GSE148500 | GEO
Project description:Functional characterization of a PROTAC directed against BRAFV600E
| PRJNA624659 | ENA
Project description:The discovery of activating BRAF V600E mutations in 50% of all cutaneous melanomas has revolutionized the understanding of melanoma biology and provided new strategies for the therapeutic management of this deadly disease. Highly potent small molecule inhibitors of BRAF are now showing great promise as a novel therapeutic strategy for melanomas harboring activating BRAF V600E mutations and are associated with high levels of response. This commentary article discusses the latest data on the role of mutated BRAF in the development and progression of melanoma as the basis for understanding the mechanism of action of BRAF inhibitors in the preclinical and clinical settings. We further address the issue of BRAF inhibitor resistance and outline the latest insights into the mechanisms of therapeutic escape as well as describing approaches to prevent and abrogate the onset of both intrinsic and acquired drug resistance. It is likely that our evolving understanding of melanoma genetics and signaling will allow for the further personalization of melanoma therapy with the goal of improving clinical responses.
| S-EPMC4001781 | biostudies-literature
Project description:Opinion statementThe optimal management of advanced stage BRAF-mutated melanoma is widely debated and complicated by the availability of several different regimens that significantly improve outcomes but have not been directly compared. While there are many unanswered questions relevant to this patient population, the major uncertainty in current practice is the choice between BRAF/MEK inhibitors or immunotherapy for those with previously untreated metastatic or high-risk disease. Decisions regarding first line therapy should include consideration of patient preference as well as the presence of symptomatic metastatic disease and degree of comorbidity, particularly secondary to any history of severe auto-immune disorder.BRAF/MEK inhibitors have a high response rate and rapid onset and thus can be quickly introduced when patients are symptomatic. They have also produced long-term responses in a subset of patients with more favorable prognostic indicators. In addition, impressive survival benefits have also been observed in patients with resected stage 3 disease at high risk of recurrence. On the other hand, anti-PD-1 monotherapy is associated with high rates of clinical benefit (~45% response rate in the metastatic setting) and low rates of severe toxicity. In many patients with adverse prognostic features, we use combined anti-PD-1 and anti-CTLA-4 for metastatic disease. While associated with high rates of toxicity, adverse events are largely manageable with corticosteroids and treatment cessation, in which case patients may continue to benefit even after a limited duration of treatment.Multiple treatment options exist for patients with BRAF V600 mutant melanoma. Herein, we review the clinical data for safety and efficacy of these options.
| S-EPMC10176035 | biostudies-literature
Project description:Purpose: BRAF inhibitors are clinically active in patients with advanced BRAFV600-mutant melanoma, although acquired resistance remains common. Preclinical studies demonstrated that resistance could be overcome using concurrent treatment with the HSP90 inhibitor XL888.Patients and Methods: Vemurafenib (960 mg p.o. b.i.d.) combined with escalating doses of XL888 (30, 45, 90, or 135 mg p.o. twice weekly) was investigated in 21 patients with advanced BRAFV600-mutant melanoma. Primary endpoints were safety and determination of a maximum tolerated dose. Correlative proteomic studies were performed to confirm HSP inhibitor activity.Results: Objective responses were observed in 15 of 20 evaluable patients [75%; 95% confidence interval (CI), 51%-91%], with 3 complete and 12 partial responses. Median progression-free survival and overall survival were 9.2 months (95% CI, 3.8-not reached) and 34.6 months (6.2-not reached), respectively. The most common grade 3/4 toxicities were skin toxicities, such as rash (n = 4, 19%) and cutaneous squamous cell carcinomas (n = 3, 14%), along with diarrhea (n = 3, 14%). Pharmacodynamic analysis of patients' peripheral blood mononuclear cells (PBMC) showed increased day 8 HSP70 expression compared with baseline in the three cohorts with XL888 doses ?45 mg. Diverse effects of vemurafenib-XL888 upon intratumoral HSP client protein expression were noted, with the expression of multiple proteins (including ERBB3 and BAD) modulated on therapy.Conclusions: XL888 in combination with vemurafenib has clinical activity in patients with advanced BRAFV600-mutant melanoma, with a tolerable side-effect profile. HSP90 inhibitors warrant further evaluation in combination with current standard-of-care BRAF plus MEK inhibitors in BRAFV600-mutant melanoma. Clin Cancer Res; 24(22); 5516-24. ©2018 AACR See related commentary by Sullivan, p. 5496.
| S-EPMC6195480 | biostudies-literature
Project description:Radiation (RT) is critical to the treatment of high-grade gliomas (HGGs) but cures remain elusive. The BRAF mutation V600E is critical to the pathogenesis of 10-20% of pediatric gliomas, and a small proportion of adult HGGs. Here we aim to determine whether PLX4720, a specific BRAF V600E inhibitor, enhances the activity of RT in human HGGs in vitro and in vivo. Patient-derived HGG lines harboring wild-type BRAF or BRAF V600E were assessed in vitro to determine IC50 values, cell cycle arrest, apoptosis and senescence and elucidate mechanisms of combinatorial activity. A BRAF V600E HGG intracranial xenograft mouse model was used to evaluate in vivo combinatorial efficacy of PLX4720+RT. Tumors were harvested for immunohistochemistry to quantify cell cycle arrest and apoptosis. RT+PLX4720 exhibited greater anti-tumor effects than either monotherapy in BRAF V600E but not in BRAF WT lines. In vitro studies showed increased Annexin V and decreased S phase cells in BRAF V600E gliomas treated with PLX4720+RT, but no significant changes in β-galactosidase levels. In vivo, concurrent and sequential PLX4720+RT each significantly prolonged survival compared to monotherapies, in the BRAF V600E HGG model. Immunohistochemistry of in vivo tumors demonstrated that PLX4720+RT decreased Ki-67 and phospho-MAPK, and increased γH2AX and p21 compared to control mice. BRAF V600E inhibition enhances radiation-induced cytotoxicity in BRAF V600E-mutated HGGs, in vitro and in vivo, effects likely mediated by apoptosis and cell cycle, but not senescence. These studies provide the pre-clinical rationale for clinical trials of concurrent radiotherapy and BRAF V600E inhibitors.
| S-EPMC4907264 | biostudies-literature
Project description:BRAF is a serine/threonine kinase frequently mutated in human cancers. BRAFV600E mutated protein is targeted through the use of kinase inhibitors which are approved for the treatment of melanoma; however, their long-term efficacy is hampered by resistance mechanisms. The PROTAC-induced degradation of BRAFV600E has been proposed as an alternative strategy to avoid the onset of resistance. In this study, we designed a series of compounds where the BRAF kinase inhibitor encorafenib was conjugated to pomalidomide through different linkers. The synthesized compounds maintained their ability to inhibit the kinase activity of mutated BRAF with IC50 values in the 40-88 nM range. Selected compounds inhibited BRAFV600E signaling and cellular proliferation of A375 and Colo205 tumor cell lines. Compounds 10 and 11, the most active of the series, were not able to induce degradation of mutated BRAF. Docking and molecular dynamic studies, conducted in comparison with the efficient BRAF degrader P5B, suggest that a different orientation of the linker bearing the pomalidomide substructure, together with a decreased mobility of the solvent-exposed part of the conjugates, could explain this behavior.
| S-EPMC9736157 | biostudies-literature
Project description:BRAF-V600E (1799T > A) is one of the most frequently reported driver mutations in multiple types of cancers, and patients with such mutations could benefit from selectively inactivating the mutant allele. Near this mutation site, there are two TTTN and one NGG protospacer-adjacent motifs (PAMs) for Cpf1 and Cas9 CRISPR nucleases, respectively. The 1799T > A substitution also leads to the occurrence of a novel NGNG PAM for the EQR variant of Cas9. We examined the editing efficacy and selectivity of Cpf1, Cas9, and EQR variant to this mutation site. Only Cpf1 demonstrated robust activity to induce specific disruption of only mutant BRAF, not wild-type sequence. Cas9 recognized and cut both normal and mutant alleles, and no obvious gene editing events were observed using EQR variant. Our results support the potential applicability of Cpf1 in precision medicine through highly specific inactivation of many other gain-of-function mutations.
| S-EPMC5542387 | biostudies-literature
Project description:Background: The aim of this study was to describe the oncologic outcomes of patients with BRAFV600E-mutated anaplastic thyroid cancer (ATC) who had neoadjuvant BRAF-directed therapy with subsequent surgery. For context, we also reviewed patients who received BRAF-directed therapy after surgery, and those who did not have surgery after BRAF-directed therapy. Methods: This was a single-center retrospective cohort study conducted at a tertiary care cancer center in Texas from 2017 to 2021. Fifty-seven consecutive patients with BRAFV600E-mutated ATC and at least 1 month of BRAF-directed therapy were included. Primary outcomes were overall survival (OS) and progression-free survival (PFS). Results: All patients had stage IVB (35%) or IVC (65%) ATC. Approximately 70% of patients treated with BRAF-directed therapy ultimately had surgical resection of residual disease. Patients who had neoadjuvant BRAF-directed therapy followed by surgery (n = 32) had 12-month OS of 93.6% [confidence interval (CI) 84.9-100] and PFS of 84.4% [CI 71.8-96.7]. Patients who had surgery before BRAF-directed therapy (n = 12) had 12-month OS of 74.1% [CI 48.7-99.5] and PFS of 50% [CI 21.7-78.3]. Finally, patients who did not receive surgery after BRAF-directed therapy (n = 13) had 12-month OS of 38.5% [CI 12.1-64.9] and PFS of 15.4% [CI 0-35.0]. Neoadjuvant BRAF-directed therapy reduced tumor size, extent of surgery, and surgical morbidity score. Subgroup analysis suggested that any residual ATC in the surgical specimen was associated with significantly worse 12-month OS and PFS (OS = 83.3% [CI 62.6-100], PFS = 61.5% [CI 35.1-88]) compared with patients with pathologic ATC complete response (OS = 100%, PFS = 100%). Conclusions: We observed that neoadjuvant BRAF-directed therapy reduced extent of surgery and surgical morbidity. While acknowledging potential selection bias, the 12-month OS rate appeared higher in patients who had BRAF-directed therapy followed by surgery as compared with BRAF-directed therapy without surgery; yet, it was not significantly different from surgery followed by BRAF-directed therapy. PFS appeared higher in patients treated with neoadjuvant BRAF-directed therapy relative to patients in the other groups. These promising results of neoadjuvant BRAF-directed therapy followed by surgery for BRAF-mutated ATC should be confirmed in prospective clinical trials.
| S-EPMC10122263 | biostudies-literature
Project description:We recently reported that BRAF V600E is the principal oncogenic driver of papillary craniopharyngioma, a highly morbid intracranial tumor commonly refractory to treatment. Here, we describe our treatment of a man age 39 years with multiply recurrent BRAF V600E craniopharyngioma using dabrafenib (150mg, orally twice daily) and trametinib (2mg, orally twice daily). After 35 days of treatment, tumor volume was reduced by 85%. Mutations that commonly mediate resistance to MAPK pathway inhibition were not detected in a post-treatment sample by whole exome sequencing. A blood-based BRAF V600E assay detected circulating BRAF V600E in the patient's blood. Re-evaluation of the existing management paradigms for craniopharyngioma is warranted, as patient morbidity might be reduced by noninvasive mutation testing and neoadjuvant-targeted treatment.
| S-EPMC4862417 | biostudies-literature