Summary: BRAF fusions and mutations are the most frequent genetic alterations in pediatric low-grade gliomas. The work from Wang and colleagues identifies an acquired secondary BRAF mutation that confers resistance to pharmacologic BRAF inhibition in a BRAFV600E glioma. The authors demonstrate that the mutation results in increased BRAF homodimerization, which in turn is targetable with second-generation BRAF inhibitors. Cancer Discov; 8(9); 1064–5. ©2018 AACR.
See related article by Wang et al., p. 1130.
Alterations of the MAPK signaling pathway are some of the most studied oncogenic mechanisms across all cancers. The advent of BRAF inhibitors (BRAFi) and MEK inhibitors (MEKi) has changed the first-line therapeutic strategy for BRAF-mutant melanoma and has shown promising outcomes in several other cancer types, including colorectal and thyroid cancers. BRAF alterations are harbored in the majority of low-grade pediatric tumors, and although they mainly consist of BRAF fusions (50%–70% depending on the glioma subtype), another 15% to 20% bear a BRAFV600E mutation (1, 2). BRAF fusions exhibit resistance to first-generation BRAF inhibitors due to increased BRAF homodimerization, eventually resulting in paradoxical MAPK pathway activation (3). BRAFV600E cases are characterized by a worse prognosis and treatment response, with therapeutic options limited to excision surgery, radiotherapy, and chemotherapy. Even with relatively high response rates to these approaches, the percentage of patients with permanent cerebral damage is significant. For these reasons, there is an urgent need for targeted therapeutic approaches based on tumor molecular features for pediatric glioma.
In this issue of Cancer Discovery, Wang and colleagues (4) report a case from a patient who was enrolled in clinical trial NCT01677741 (phase I/II) for the testing of a BRAFi (dabrafenib) in pediatric tumors. The patient had a previous history of low-grade astrocytoma in infancy that relapsed 10 years later as an anaplastic ganglioglioma. After a first-line chemotherapeutic approach with temozolomide, vincristine, and lomustine, the patient showed signs of a third recurrence that was identified by clinical sequencing to bear a BRAFV600E mutation. The patient was then enrolled in the clinical trial, showing impressive results after only 8 weeks of treatment. A subsequent complete radiographic response was achieved, until a further relapse was noted at 40 weeks after treatment start. Whole-exome and RNA sequencing revealed that the newly relapsed tumor did not show signs of massive genetic evolution and, contrarily, was rather similar to the pre-dabrafenib ganglioglioma.
Because no obvious resistance-conferring mutations were detected postprogression, the authors focused on new genetic candidates, and an acquired BRAF mutation (L514V) stood out. L514V occurs in the ATP-binding pocket of BRAF, and the authors detected it in 30% of the post-dabrafenib cancer cells. Interestingly, L514V was described to be in cis with V600E and never detected independently from the latter, implying outgrowth from an initially BRAFi-sensitive tumor cell population. Indeed, high-sensitivity detection assays confirmed that L514V was not present in pretreatment blood or tissue specimens. This is the first study to report a secondary mutation of BRAF as a mechanism of BRAFi resistance, paralleling secondary resistance mutations in RTKs such as EGFRT790M gatekeeper mutations in response to EGFRi (5).
Functional studies performed by the authors showed that the coexpression of V600E and L514V was able to sustain phospho-ERK (pERK) levels even after dabrafenib administration, in several in vitro models. Short- and long-term assays demonstrated that residual pERK levels promoted by the double mutation were able to sustain cancer cell proliferation. Detailed and elegant molecular analyses then shed light on the mechanism of sustained pERK signaling: L514V and V600E co-occurrence favored BRAF dimerization much more efficiently than V600E alone. Dabrafenib is a first-generation BRAFi and, as a result, is not efficient in targeting dimerized BRAF. Contrarily, when second-generation, dimerization-targeting BRAFi were administered to double-mutant cells, sensitivity to BRAFi was restored and pERK levels were impaired. In contrast, MEKi were able to abolish pERK in the short term but did not achieve the same long-term effects as second-generation BRAFi. Interestingly, ERK inhibitors (ERKi), a newly introduced drug class that targets further downstream in the MAPK pathway, elicited therapeutic and molecular effects superimposable with second-generation BRAFi.
The identification of BRAFL514V as a novel BRAFi resistance mechanism has several clinical implications. First, it emphasizes that in both BRAF fusion and nonfusion pediatric gliomas, the mechanism of BRAFi resistance appears to converge on the increased dimerization of BRAF, positioning dimerization-targeted second-generation BRAFi as a potential novel first-line therapy. Second, it underlines the relevance of cell lineage in targeted therapy, as in contrast to the findings of this study, cumulative analysis of clinical BRAFi resistance in hundreds of melanomas has not identified any secondary BRAF mutations (6). This parallels the identification of EGFR as a lineage-restricted mechanism of intrinsic BRAFi resistance in BRAF-mutant colorectal cancer and emphasizes the need for cancer type–specific analyses of drug resistance (7, 8). Finally, the similar efficacy of ERKi as dimerization-targeted BRAFi implies the possibility of using ERKi alone or, possibly, in combination with dimerization-targeted BRAFi for the treatment of pediatric low-grade gliomas.
Indeed, following the paradigm of BRAFi + MEKi in melanoma, combination therapies for pediatric glioma may be less likely to cause up-front resistance and may even show decreased toxicity (9, 10). Drug combinations might also help to overcome the problem of tumor heterogeneity that can potentially arise from a treatment with dimerization-targeted BRAFi alone. The authors show, in fact, that dimerization-targeted BRAFi are equally efficient in inhibiting V600E and L514V, but they are in general less efficient than first-generation BRAFi in inhibiting V600E-only tumor cells. Future preclinical and clinical testing will be necessary to identify such combined therapies that can have a positive impact on patients' prognoses. One intriguing question for such studies will be whether dimerization-targeted BRAFi would be more effective as a first-line therapy or as a backup to first-generation BRAFi for BRAFV600E pediatric gliomas.
Previous work has emphasized the importance of RAF dimerization in multiple MAPK-driven cancers (11), and the latest findings extend the applicability of dimerization-targeted inhibitors not only to pediatric gliomas, but also to a novel mechanism of resistance: a second-site BRAF mutation. It remains to be seen how well these second-generation RAF inhibitors work in the clinic, given the existence of multiple non-RAF dimerization–dependent resistance mechanisms (e.g., PI3K pathway activation). Nevertheless, the implications here that they could form a cornerstone for combination therapies that can effectively forestall multiple mechanisms of drug resistance are highly promising. Overall, the work of Wang and colleagues is well-framed in the fundamental effort of translational medicine to bring efficacious therapeutic approaches to the bedside and to leverage benchtop experiments to further improve and develop such therapies.
Disclosure of Potential Conflicts of Interest
L.N. Kwong reports receiving a commercial research grant from Array Biopharma and is a consultant/advisory board member for Vuja De. No potential conflicts of interest were disclosed by the other author.