BRAF Emerges as Exploitable Target in NSCLC

After a decade of success in targeting molecular alterations in non—small cell lung cancer (NSCLC), researchers are focusing on small molecules that attack the BRAF cell-signaling network directly or in combination with other inhibitors.

Numerous studies have now confirmed that BRAF mutations represent a 1% to 4% slice of the genetic pie of NSCLC. These mutations are most renowned for their role in melanoma development and researchers are hoping to translate the success of BRAF-targeted therapies in this cancer type to NSCLC.

A Model of Personalized Cancer Therapy

Several of these drugs have demonstrated impressive efficacy in BRAF-mutant NSCLC patient populations and have been awarded breakthrough therapy designations from the FDA. The types of BRAF mutations in NSCLC may differ from those observed in melanoma, and a greater understanding of these nuances and their sensitivity to currently available drugs is a central focus of ongoing research.Thanks to advances in genomic sequencing, a targetable molecular alteration can now be identified in more than half of all patients with NSCLC, the most common histological subtype of lung cancer. The ability to detect these drivers with diagnostic tests and to pair them with targeted therapies has revolutionized the treatment of NSCLC.

The best studied drivers are the epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK), and the success of small molecule inhibitors targeting these tyrosine kinase receptors has served as a model of targeted drug development in oncology.

BRAF's Place in Signaling Network

Growth factors that bind to their receptors (EGFR, MET, ERBB3, ERBB2, and FGFR) promote RAS activity, which in turn sets off a cell-signaling cascade that includes BRAF activation.

Nussinov R, Jang H, Tsai CJ. The structural basis for cancer treatment. Oncotarget. 2014;5(17):7285-302.

A number of other kinases involved in cell signaling pathways have been implicated in the development of NSCLC, among them the BRAF protein, a core component of the mitogen- activated protein kinase (MAPK) signaling pathway that is central to driving the runaway proliferation of cancer cells.

Besides a predominance in the adenocarcinoma subtype of NSCLC, no other defining clinical features of BRAF-mutant NSCLC have yet been carved out. Perpetuating Growth Signals BRAF is one of three members of the RAF family of proteins that serve as kinase enzymes, which phosphorylate key serine or threonine residues within the amino acid sequence of other proteins. In this way, they propagate signals from the cell membrane to the nucleus through the MAPK pathway. One of the first steps in the MAPK pathway involves the activation of the RAS protein, a small GTPase that switches between a guanosine diphosphate (GDP)- bound “off” state and a guanosine triphosphate (GTP)-bound "on" state. RAS can be activated by signals generated from growth factors, cytokines, and hormones binding to their respective receptors on the cell surface.

Targeting BRAF

Once RAS is switched on, it recruits RAF to the cell membrane, where it is also activated, allowing it to subsequently phosphorylate and activate the next kinase in the chain. Ultimately, the cascade of phosphorylation events that make up the MAPK pathway leads to the ERK protein being shifted into the nucleus, where it stimulates the transcription of a number of different gene targets that orchestrate the cellular responses to the initial signal received at the membrane.BRAF is a proto-oncogene because its key role in the MAPK pathway makes it vulnerable to mutations that cause a change in the DNA sequence leading to the production of an altered protein, which impacts the normal function of the cell and can contribute to cancer development.

Studies suggest that somatic mutations in BRAF occur in just under 10% of all human cancers. Most notably, BRAF mutations are found in around half of malignant melanomas. In the vast majority of cases, the BRAF mutations observed in melanoma result in a single amino acid change within the kinase domain, in which a valine is substituted for a glutamine at position 600 (V600E).

This mutation occurs with the kinase domain and the insertion of a negatively charged amino acid is thought to mimic phosphorylation, resulting in increased kinase activity. Since BRAF is a central player in the MAPK pathway, the pathway becomes more highly activated in the presence of these activating mutations.

The identification of BRAF V600E as a key driver of melanoma growth served as the stimulus to develop small-molecule inhibitors to target the mutant protein, which have proved very successful in the management of BRAF-mutant melanoma. The FDA has approved two BRAF inhibitors in this setting; vemurafenib (Zelboraf) and dabrafenib (Tafinlar). The multitargeted tyrosine kinase inhibitors sorafenib (Nexavar) and regorafenib (Stivarga) also inhibit BRAF; however, these are classed as type II inhibitors, with a different mechanism of action whereby they bind BRAF in its inactive state.

BRAF-Targeting Strategies Under Development in NSCLC

^aTrial is ongoing, but not actively recruiting participants.

^bRecruitment has been suspended due to drug supply issues.

Although not commonly mutated itself, the mitogen- activated protein kinase kinase (MEK) protein sits immediately downstream of BRAF in the MAPK pathway and is the only known substrate of BRAF and, therefore, the direct target of its oncogenic activation. Small-molecule inhibitors of MEK have also enjoyed some success in melanoma, particularly in the context of combination therapy, when added to BRAF inhibitor regimens.

Trametinib (Mekinist) was originally approved as monotherapy in BRAF-mutant patients who had previously been treated with a BRAF inhibitor and was subsequently approved as first-line therapy in combination with dabrafenib, while cobimetinib (Cotellic) is approved in combination with vemurafenib in the front-line setting. Both BRAF inhibitors have an impressive response rate of around 50% in patients with BRAF V600E mutations, and that improves to upwards of 65% with the use of BRAF and MEK inhibitor combinations.

Currently available BRAF inhibitors are not only less effective in patients with wild-type BRAF, they can paradoxically activate the MAPK pathway, particularly in the presence of activating RAS mutations. Therefore, identifying BRAF mutations in patients to be treated with these drugs is of paramount importance.

The FDA has approved two companion diagnostics that help to identify patients with BRAF V600E mutations. Roche developed the cobas 4800 BRAF V600 Mutation Test, a polymerase chain reaction (PCR)-based test that is faster and more sensitive than traditional Sanger sequencing.

A second diagnostic test, the THxID BRAF kit, is a real-time PCR test developed in conjunction with trametinib and dabrafenib. BRAF Inhibitors in NSCLC Given the availability of FDA-approved BRAF inhibitors, the identification of BRAF mutations in patients with NSCLC has sparked interest in applying these drugs to the lung cancer setting. Preclinical studies suggested that vemurafenib and trametinib were effective as single agents in V600E-mutant NSCLC. Sorafenib and several MEK inhibitors have also been reported to be active in preclinical models of BRAF-mutant NSCLC. Both dabrafenib and vemurafenib have been examined as monotherapy for the treatment of patients with BRAF-mutated NSCLC.

The results of a phase II “basket study” of vemurafenib were recently published in The New England Journal of Medicine. Patients with BRAF V600E mutations, regardless of their specific type of cancer, were enrolled in the trial and received vemurafenib. In a cohort of patients with NSCLC, the response rate was 42% and the median progression-free survival (PFS) was 7.3 months. A recent preclinical study also hinted at the efficacy of a combination of vemurafenib and trametinib in NSCLC.

Dabrafenib monotherapy was evaluated in a single-arm, multicenter, open-label phase II trial (NCT01336634). The recently published data demonstrated a response rate of 33% among 78 previously treated patients with BRAF V600E-positive NSCLC who received oral dabrafenib 150 mg twice daily. On the basis of this study, dabrafenib was awarded breakthrough therapy designation for BRAF-mutant NSCLC in 2014.

The most exciting data were presented at the 2016 ASCO Annual Meeting in June. David Planchard MD, PhD, of the Institute Gustav Roussy in France, reported interim results from the ongoing phase II BRF113928 trial of the combination of dabrafenib (150 mg twice daily) and trametinib (2 mg once daily) in patients with previously treated BRAF V600E-mutant NSCLC.

Among 52 patients evaluable for efficacy, the overall response rate was 63%, the disease control rate was 79%, and the median duration of response was 9 months. The most common adverse events included pyrexia, nausea, vomiting, and diarrhea. The combination of dabrafenib and trametinib in NSCLC also has received breakthrough therapy designation now from the FDA.

Exploring Differences

Researchers are continuing to explore the potential of BRAF and MEK inhibitors as monotherapy and in combination with each other and several other types of targeted therapies. Although the majority of studies conducted thus far have involved patients with refractory disease, a small number of patients with treatment-naïve NSCLC were enrolled in the phase II trial of dabrafenib monotherapy and four of the six patients had an objective response.Emerging insights from studies of BRAF mutations in NSCLC suggest that there are some important distinctions from melanoma that could have implications for the use of BRAF- and MEK-targeted therapies in this patient population. While 90% of BRAF-mutant melanomas display the V600E mutation, the most common BRAF mutations in NSCLC are in fact non-V600 mutations, present in just over half of all cases. Among the non-V600 mutations are other activating mutations within the kinase domain, such as K601N, L597Q and G469V, as well as mutations that inactivate BRAF’s kinase activity such as D594 mutations.

BRAF inhibitors were designed to target V600-mutant forms of the BRAF protein and it is currently unclear how effective they are in patients with other kinds of BRAF mutations, even if the mutation is within the kinase domain. If these mutations are sensitive to BRAF inhibition, then a substantial number of patients with NSCLC are being excluded from an effective therapy by restricting their use to patients with V600 mutations.

A number of studies are examining both the clinical features of non-V600 mutations compared with V600 mutations and wild-type BRAF, and the sensitivity of these mutations to BRAF inhibition.

So far, although it has been suggested that non-V600 mutations may not be as responsive to currently available BRAF inhibitors and that these patients have poorer outcomes, the relatively small numbers of non-V600 mutation carriers enrolled have made it difficult to draw any solid conclusions. Indeed, efficacy may depend upon the specific type of mutation.

Conversely, BRAF inactivating mutations are not expected to respond to BRAF inhibitors. However, a study presented at the 2015 ASCO Annual Meeting suggested that these mutants might still activate the MAPK pathway through a different member of the RAF family—CRAF—and that they may be sensitive to combined BRAF and MEK inhibition. A previous study suggested that inactivating mutations in BRAF may be sensitive to the multitargeted kinase inhibitor dasatinib and a clinical trial was initiated. The trial has since been terminated due to lack of efficacy and poor enrollment.

Jane de Lartigue, PhD, is a freelance medical writer and editor based in New Haven, Connecticut.

Key Research

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