RAF inhibitor re-challenge therapy in BRAF-aberrant pan-cancers: the RE-RAFFLE study
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ABSTRACT:
SUBMITTER: Nelson B
PROVIDER: S-EPMC10964523 | biostudies-literature | 2024 Jan
REPOSITORIES: biostudies-literature
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Project description:Structure-based optimization of a set of aryl urea RAF inhibitors has led to the identification of Type II pan-RAF inhibitor GNE-9815 (7), which features a unique pyrido[2,3-d]pyridazin-8(7H)-one hinge-binding motif. With minimal polar hinge contacts, the pyridopyridazinone hinge binder moiety affords exquisite kinase selectivity in a lipophilic efficient manner. The improved physicochemical properties of GNE-9815 provided a path for oral dosing without enabling formulations. In vivo evaluation of GNE-9815 in combination with the MEK inhibitor cobimetinib demonstrated synergistic MAPK pathway modulation in an HCT116 xenograft mouse model. To the best of our knowledge, GNE-9815 is among the most highly kinase-selective RAF inhibitors reported to date.
| S-EPMC8155235 | biostudies-literature
Project description:BRAF mutations occur in 10-15% of colorectal cancers (CRCs) and confer adverse outcome. While RAF inhibitors such as vemurafenib (PLX4032) have proven effective in BRAF mutant melanoma, they are surprisingly ineffective in BRAF mutant CRCs, and the reason for this disparity remains unclear. Compared to BRAF mutant melanoma cells, BRAF mutant CRC cells were less sensitive to vemurafenib, and P-ERK suppression was not sustained in response to treatment. Although transient inhibition of phospho-ERK by vemurafenib was observed in CRC, rapid ERK re-activation occurred through EGFR-mediated activation of RAS and CRAF. BRAF mutant CRCs expressed higher levels of phospho-EGFR than BRAF mutant melanomas, suggesting that CRCs are specifically poised for EGFR-mediated resistance. Combined RAF and EGFR inhibition blocked reactivation of MAPK signaling in BRAF mutant CRC cells and markedly improved efficacy in vitro and in vivo. These findings support evaluation of combined RAF and EGFR inhibition in BRAF mutant CRC patients.BRAF valine 600 (V600) mutations occur in 10% to 15% of colorectal cancers, yet these tumors show a surprisingly low clinical response rate (~5%) to selective RAF inhibitors such as vemurafenib, which have produced dramatic response rates (60%–80%) in melanomas harboring the identical BRAF V600 mutation. We found that EGFR-mediated MAPK pathway reactivation leads to resistance to vemurafenib in BRAF-mutant colorectal cancers and that combined RAF and EGFR inhibition can lead to sustained MAPK pathway suppression and improved efficacy in vitro and in tumor xenografts.
| S-EPMC3308191 | biostudies-literature
Project description:Activating BRAF mutants and fusions signal as RAS-independent constitutively active dimers with the exception of BRAF V600 mutant alleles which can function as active monomers1. Current RAF inhibitors are monomer selective, they potently inhibit BRAF V600 monomers but their inhibition of RAF dimers is limited by induction of negative cooperativity when bound to one site in the dimer1-3. Moreover, acquired resistance to these drugs is usually due to molecular lesions that cause V600 mutants to dimerize4-8. We show here that PLX8394, a new RAF inhibitor9, inhibits ERK signaling by specifically disrupting BRAF-containing dimers, including BRAF homodimers and BRAF-CRAF heterodimers, but not CRAF homodimers or ARAF-containing dimers. Differences in the amino acid residues in the amino (N)-terminal portion of the kinase domain of RAF isoforms are responsible for this differential vulnerability. As a BRAF-specific dimer breaker, PLX8394 selectively inhibits ERK signaling in tumors driven by dimeric BRAF mutants, including BRAF fusions and splice variants as well as BRAF V600 monomers, but spares RAF function in normal cells in which CRAF homodimers can drive signaling. Our work suggests that drugs with these properties will be safe and useful for treating tumors driven by activating BRAF mutants or fusions.
| S-EPMC6404779 | biostudies-literature
Project description:BRAF and MEK inhibition is efficient in patients with BRAF V600-mutated metastatic melanoma, but due to acquired resistance the duration of response (DoR) is often only short-lived. In this retrospective multicenter study with 60 patients suffering from inoperable or metastatic melanoma we evaluated the efficacy of re-challenge with a BRAF inhibitor (BRAF2) with or without MEK-inhibition after progressive disease upon previous treatment with a BRAF inhibitor (BRAF1) with or without MEK inhibition. Treatment with BRAF1 led to a disease control rate (DCR) of 90% with 12% complete responses (CR), 58% partial responses (PR) and 20% stable diseases (SD), the median progression-free survival (PFS) was 9.9 and DoR 10.7 months. BRAF2 with (68%) or without (32%) additional MEK inhibition was initiated after a median interval of 3.4 months. DCR after re-challenge with BRAF2 was 57%, 8% CR, 20% PR and 28% SD, median PFS was 5.0 and DoR 14.0 months. The duration of the treatment interval or the treatment in the interval did not influence the DCR or PFS to BRAF2. The only predictive factor for response to BRAF2 was previous response to BRAF1; all patients with CR to BRAF1 achieved disease control with BRAF2, but only 60% of the patients with PR to BRAF1 (p=0.002). Addition of MEK inhibition to BRAF2 after treatment with BRAF1 as monotherapy did not significantly increase the DCR or PFS compared to patients treated solely with mono- or combination therapy. In conclusion re-challenge with a BRAF inhibitor is a meaningful therapeutic option for patients with BRAF V600-mutated metastatic melanoma.
| S-EPMC6188134 | biostudies-literature
Project description:Expression of aberrantly spliced BRAF V600E isoforms (BRAF V600E ΔEx) mediates resistance in 13%-30% of melanoma patients progressing on RAF inhibitors. BRAF V600E ΔEx confers resistance, in part, through enhanced dimerization. Here, we uncoupled BRAF V600E ΔEx dimerization from maintenance of MEK-ERK1/2 signaling. Furthermore, we show BRAF V600E ΔEx association with its substrate, MEK, is enhanced and required for RAF inhibitor resistance. RAF inhibitor treatment increased phosphorylation at serine 729 (S729) in BRAF V600E ΔEx. Mutation of S729 to a non-phosphorylatable residue reduced BRAF V600E ΔEx-MEK interaction, reduced dimerization or oligomerization, and increased RAF inhibitor sensitivity. Conversely, mutation of the BRAF dimerization domain elicited partial effects on MEK association and RAF inhibitor sensitivity. Our data implicate BRAF S729 in resistance to RAF inhibitor and underscore the importance of substrate association with BRAF V600E ΔEx. These findings may provide opportunities to target resistance driven by aberrantly spliced forms of BRAF V600E.
| S-EPMC6261462 | biostudies-literature
Project description:The complex biochemical effects of RAF inhibitors account for both the effectiveness and mechanisms of resistance to these drugs, but a unified mechanistic model has been lacking. Here we show that RAF inhibitors exert their effects via two distinct allosteric mechanisms. Drug resistance due to dimerization is determined by the position of the αC helix stabilized by inhibitor, whereas inhibitor-induced RAF priming and dimerization are the result of inhibitor-induced formation of the RAF/RAS-GTP complex. The biochemical effect of RAF inhibitor in cells is the combined outcome of the two mechanisms. Therapeutic strategies including αC-helix-IN inhibitors are more effective in multiple mutant BRAF-driven tumor models, including colorectal and thyroid BRAF(V600E) cancers, in which first-generation RAF inhibitors have been ineffective.
| S-EPMC5021590 | biostudies-literature
Project description:Activated RAS promotes dimerization of members of the RAF kinase family. ATP-competitive RAF inhibitors activate ERK signalling by transactivating RAF dimers. In melanomas with mutant BRAF(V600E), levels of RAS activation are low and these drugs bind to BRAF(V600E) monomers and inhibit their activity. This tumour-specific inhibition of ERK signalling results in a broad therapeutic index and RAF inhibitors have remarkable clinical activity in patients with melanomas that harbour mutant BRAF(V600E). However, resistance invariably develops. Here, we identify a new resistance mechanism. We find that a subset of cells resistant to vemurafenib (PLX4032, RG7204) express a 61-kDa variant form of BRAF(V600E), p61BRAF(V600E), which lacks exons 4-8, a region that encompasses the RAS-binding domain. p61BRAF(V600E) shows enhanced dimerization in cells with low levels of RAS activation, as compared to full-length BRAF(V600E). In cells in which p61BRAF(V600E) is expressed endogenously or ectopically, ERK signalling is resistant to the RAF inhibitor. Moreover, a mutation that abolishes the dimerization of p61BRAF(V600E) restores its sensitivity to vemurafenib. Finally, we identified BRAF(V600E) splicing variants lacking the RAS-binding domain in the tumours of six of nineteen patients with acquired resistance to vemurafenib. These data support the model that inhibition of ERK signalling by RAF inhibitors is dependent on levels of RAS-GTP too low to support RAF dimerization and identify a novel mechanism of acquired resistance in patients: expression of splicing isoforms of BRAF(V600E) that dimerize in a RAS-independent manner.
| S-EPMC3266695 | biostudies-literature
Project description:Resistance to RAF inhibitors such as vemurafenib and dabrafenib is a major clinical problem in the treatment of melanoma. Patients with mutant BRAF melanoma that progress on RAF inhibitors have limited treatment options, and drug removal from resistant tumors may elicit multiple effects. A frequent mechanism of resistance to RAF inhibitors is caused by expression of mutant BRAF splice variants. RAF inhibitor-resistant cell lines, generated in vivo, were tested as to whether or not mutant BRAF splice variants confer a fitness advantage in the presence of RAF inhibitor. Critically, cells expressing distinct mutant BRAF splice variants grow more efficiently in vitro and in vivo in the presence of the vemurafenib analog, PLX4720, compared with in the absence of inhibitor. PLX4720-treated BRAF splice variant-expressing cells exhibited levels of phospho-extracellular signal-regulated kinase (ERK)1/2 comparable to untreated parental cells. In addition, a reduction in phospho-ERK1/2 levels following treatment with the MEK inhibitor, trametinib (GSK1120212) phenocopied the fitness benefit provided by PLX4720. These data indicate that mutant BRAF splice variant-expressing melanoma cells are benefited by defined concentrations of RAF inhibitors.This study provides evidence that RAF inhibitor-resistant melanoma cells benefit from continued therapy.
| S-EPMC4020954 | biostudies-literature
Project description:BackgroundCurrently, there are no available targeted therapy options for non-V600 BRAF mutated tumors. The aim of this study was to investigate the effects of RAF and MEK concurrent inhibition on tumor growth, migration, signaling and apoptosis induction in preclinical models of non-V600 BRAF mutant tumor cell lines.MethodsSix BRAF mutated human tumor cell lines CRL5885 (G466 V), WM3629 (D594G), WM3670 (G469E), MDAMB231 (G464 V), CRL5922 (L597 V) and A375 (V600E as control) were investigated. Pan-RAF inhibitor (sorafenib or AZ628) and MEK inhibitor (selumetinib) or their combination were used in in vitro viability, video microscopy, immunoblot, cell cycle and TUNEL assays. The in vivo effects of the drugs were assessed in an orthotopic NSG mouse breast cancer model.ResultsAll cell lines showed a significant growth inhibition with synergism in the sorafenib/AZ628 and selumetinib combination. Combination treatment resulted in higher Erk1/2 inhibition and in increased induction of apoptosis when compared to single agent treatments. However, single selumetinib treatment could cause adverse therapeutic effects, like increased cell migration in certain cells, selumetinib and sorafenib combination treatment lowered migratory capacity in all the cell lines. Importantly, combination resulted in significantly increased tumor growth inhibition in orthotropic xenografts of MDAMB231 cells when compared to sorafenib - but not to selumetinib - treatment.ConclusionsOur data suggests that combined blocking of RAF and MEK may achieve increased therapeutic response in non-V600 BRAF mutant tumors.
| S-EPMC5941622 | biostudies-literature
Project description:BackgroundCancers with non-V600 BRAF-activating alterations have no matched therapy. Preclinical data suggest that these tumors depend on ERK signaling; however, clinical response to MEK/ERK inhibitors has overall been low. We hypothesized that a narrow therapeutic index, driven by ERK inhibition in healthy (wild-type) tissues, limits the efficacy of these inhibitors. As these mutants signal as activated dimers, we further hypothesized that RAF inhibitors given concurrently would improve the therapeutic index by opposing ERK inhibition in normal tissues and not activate ERK in the already activated tumor.Materials and methodsUsing cell lines and patient-derived xenografts, we evaluated the effect of RAF inhibition, alone and in combination with MEK/ERK inhibitors. We then undertook a phase I/II clinical trial of a higher dose of the MEK inhibitor binimetinib combined with the RAF inhibitor encorafenib in patients with advanced cancer with activating non-V600 BRAF alterations.ResultsRAF inhibition led to modest inhibition of signaling and growth in activated non-V600 BRAF preclinical models and allowed higher dose of MEK/ERK inhibitors in vivo for more profound tumor regression. Fifteen patients received binimetinib 60 mg twice daily plus encorafenib 450 mg daily (6 gastrointestinal primaries, 6 genitourinary primaries, 3 melanoma, and 2 lung cancer; 7 BRAF mutations and 8 BRAF fusions). Treatment was well tolerated without dose-limiting toxicities. One patient had a confirmed partial response, 8 had stable disease, and 6 had radiographic or clinical progression as best response. On-treatment biopsies revealed incomplete ERK pathway inhibition.ConclusionCombined RAF and MEK inhibition does not sufficiently inhibit activated non-V600 BRAF-mutant tumors in patients.
| S-EPMC10769795 | biostudies-literature