Keith T. Flaherty, MD
Director of Developmental Therapeutics
Massachusetts General Hospital
Cancer Center, Boston, MA
Systemic therapies for advanced melanoma have lagged behind treatments for most other cancers, both in the metastatic and earlier-stage, adjuvant setting. Metastatic melanoma continues to be largely refractory to available therapies, with a small percentage of patients responding to either chemotherapies or immunotherapies such as dacarbazine and interleukin-2. In the adjuvant setting, high-dose interferon remains the treatment standard. However, this therapy is not as effective at preventing recurrence as adjuvant therapies for several other cancer types, and it is associated with significant toxicity burdens.
The treatment of some other cancers has been revolutionized by evolving molecular understanding of signal transduction within cancer cells and of how mutations in key genes that encode signaling molecules underlie the development of these malignancies. The advent of targeted therapies that block the signaling function of such “oncogenes” has provided far more effective and tolerable therapy for patients with chronic myelogenous leukemia, gastrointestinal stromal tumors, non-small cell lung cancer, breast cancer, colon cancer, renal cell carcinoma, and glioblastoma multiforme. Several of these tumors share with melanoma the same resistance to conventional cytotoxic chemotherapy, yet have proven susceptible to oncogene-targeted therapy.
Great advances have recently been made in understanding the oncogenes and signal transduction pathways that contribute to melanoma formation and dissemination. The gene that has garnered most attention in recent years is BRAF, a constituent of the so-called MAP (mitogen-activated protein) kinase signaling pathway, in which activating mutations occur in 50-60 percent of melanomas.1 Laboratory evidence has consistently shown that targeting BRAF can be an effective strategy for slowing growth and inducing cell death in melanoma cells that harbor BRAF mutations, though these tumor cells also harbor a host of additional mutations in other key signaling molecules. However, it has taken seven years from the time of initial discovery of BRAF mutations to establish that a drug can successfully target BRAF in humans with metastatic melanoma.
Especially with the recent early success using PLX4032, we have new evidence that changes the landscape of clinical research in melanoma and will hopefully soon change the treatment paradigm for this traditionally refractory disease.
|Figure 1: MAP Kinase Pathway With and Without BRAF Mutation|
|The MAP kinase pathway normally links growth factor receptors to the nucleus by transmission of signals that promote growth and metastasis. The pathway is activated or inactivated in the presence or absence of growth factors on the cell surface. In the setting of BRAF mutation, the pathway remains constitutively activated without regard to growth factor activation of cell surface receptors.|
Raf kinases are components of the MAP kinase pathway, a signal transduction pathway that normally controls cell growth and division downstream of activated cell surface growth factor receptors. RAF1, formerly known as CRAF, is the most ubiquitously expressed of the RAF family members, and previously was the most extensively studied. However, activating mutations in RAF1 are known to be very rare. BRAF is expressed most highly in neuronal tissues and melanocytes (both of neural crest origin), as well as testis and hematopoietic cells. Based on several large genetic screening studies, BRAF appears to be mutated in approximately 7 percent of all cancers, but most commonly in melanoma. (See Figure 1.)
While there are numerous types of mutations in BRAF, the T1796A point mutation accounts for 97 percent of the mutations, resulting in substitution of glutamic acid for valine at the 600 position of the amino acid sequence (V600EBRAF). This mutation locks BRAF in the active signaling conformation so that it drives signaling through the MAP kinase pathway regardless of activation of receptor tyrosine kinases or other inputs. The first evidence that blocking V600EBRAF activity could have potential therapeutic value in melanoma came from laboratory experiments in which V600EBRAF was eliminated from cells using a sequence of nucleotides that neutralize the messenger RNA encoding BRAF. However, this is very different from demonstrating an effect with a drug administered systemically to humans. That milestone had to await the successful discovery and development of far more potent and specific inhibitors of BRAF.
BRAF Inhibitors in Clinical Trials
Sorafenib (BAY 43-9006, Nexavar™) was first selected for development as an inhibitor of RAF1, the ubiquitously expressed but rarely mutated Raf isoform. Sorafenib is tenfold less potent against BRAF, and apparently even less potent against V600EBRAF. In the laboratory setting, sorafenib is able to block V600EBRAF and downstream activation of the MAP kinase pathway. However, the concentrations of the drug needed to induce cell death also kill melanoma cells that lack a BRAF mutation. This is concerning in that BRAF appears to be an important component of MAP kinase pathway signaling and a cell survival factor only when it is mutated. Therefore, sorafenib’s effect on melanoma cells may not relate to BRAF inhibition. Furthermore, the concentrations required to kill melanoma cells were sufficiently high that similar concentrations may not be achievable in humans.
At the maximum tolerated dose, sorafenib was not found to have significant single-agent activity in melanoma.2,3 Mechanistic investigations have been limited, but suggest that sorafenib does not effectively inhibit BRAF in human tumors, as measured by ERK (extracellular signal-regulated kinases) activation before and during treatment, whereas significant changes in tumor vascular permeability (an established surrogate for VEGF signaling) were noted.3 Based on this evidence, it appeared that sorafenib was unable to exert a sufficient effect on BRAF to test the value of BRAF as a therapeutic target in melanoma. Several trials were conducted investigating the possibility that sorafenib might enhance the efficacy of chemotherapy. Although single-arm phase II trials suggested a benefit, randomized trials comparing the efficacy of chemotherapy alone versus chemotherapy with sorafenib did not show a significant contribution from sorafenib.4,5
RAF-265 and XL281
RAF-265 and XL281 are the leading examples of an emerging class of broad-spectrum kinase inhibitors that have greater potency and modestly improved selectivity for BRAF compared to sorafenib. Notably, these agents include receptor tyrosine kinases in their spectrum and are very potent inhibitors of RAF1. They are in early clinical development, and preliminary results in metastatic melanoma are awaited.
Several highly selective B-Raf inhibitors are being developed, with PLX4032 by far the most advanced in clinical studies.6 GSK2118436 is the second of this class to have entered clinical trials, with a phase I trial currently under way.[http://clinicaltrials.gov/ct2/show/NCT00880321] PLX4032 and its analogs have demonstrated selectivity between highly homologous wild-type BRAF and RAF1, and some selectivity for V600EBRAF compared to non-mutated BRAF.7 These compounds inhibit the MAP kinase pathway only in the context of a V600EBRAF mutation. Subsequent effects on proliferation and apoptosis are also entirely restricted to cells harboring BRAF mutations. In animal models, experimental efficacy also has been restricted to BRAF mutant tumors, confirming the lack of biologically significant effects on non-tumor, BRAF wild-type cells in the tumor microenvironment.
The preliminary results of the PLX4032 phase I trial were recently reported. In this study, 49 of 55 patients enrolled in the dose escalation portion of the study had metastatic melanoma. Furthermore, there was enrichment for patients with V600EBRAF mutations, as many patients’ tumors were prospectively evaluated prior to study entry, particularly those patients assigned to the higher dose levels. Toxicity was clearly related to dose, with increasing frequency and severity of rash, fatigue, and arthralgia at the highest doses. At the five highest dose levels evaluated, 21 melanoma patients were enrolled. Sixteen of the 21 had V600EBRAF mutations. Among these 16, tumor regression was observed in 14, with 9 of 16 (56 percent) having partial responses by RECIST criteria. As follow-up was very short and most patients were still being treated, progression-free survival has yet to be well-defined. Nonetheless, the preliminary estimate of six months is promising compared to available therapies.
Selected patients underwent biopsy of superficial tumors to confirm target inhibition. At the same doses with which significant tumor regression was noted in patients, the MAP kinase pathway appeared to be almost completely inhibited.8 It is remarkable that early responses to the drug are so commonly seen, despite the diversity of additional genetic alterations in these tumors. As patients’ tumors begin to progress following initial response, it will be of great interest to know what the drivers of treatment resistance might be, but at this time, no data are available to shed light on this question. In the context of the ongoing phase II and phase III trials with single-agent PLX4032, many more patients will be treated, and further insight into this issue should be gained.
Two clinical trials are now under way to further characterize the efficacy of PLX4032 in metastatic melanoma. Hopefully, these trials will serve as the basis for regulatory approval and incorporation of this highly targeted therapy as standard treatment for metastatic melanoma patients whose tumors harbor BRAF mutations. A single-arm phase II trial is being conducted to evaluate the objective response rate and duration of response in a larger cohort of V600EBRAF-mutated metastatic melanoma patients than were included in the original phase I/II trial. The second trial, a larger, randomized, phase III trial, is comparing PLX4032 to the reference standard therapy for metastatic melanoma, dacarbazine, in patients with previously untreated metastatic melanoma. The goal of this trial is to show an improvement in overall survival, providing definitive proof that this agent alters the natural history of metastatic disease, rather than merely inducing short-term regression.
There are numerous possibilities to consider in the effort to improve single-agent BRAF inhibition. The next critical goal is to enhance longevity of response or obtain complete responses. It is possible that the greatest disease control will come from sequencing targeted therapies aimed at intercepting the mechanism of escape/resistance following BRAF inhibition. Combination strategies are also being considered, since concomitant inhibition of signaling pathways known to be activated in concert with BRAF mutations is theorized to induce greater degrees of cell death and potentially delay the emergence of resistance.
BRAF appears to represent an important new target in melanoma. Melanoma has been the disease context in which the most extensive clinical evaluation has been conducted with RAF inhibitors. The clinical trials currently under way for metastatic melanoma represent the first opportunity for these therapies to be established as standard in the treatment of cancer. However, melanoma is an aggressive tumor, associated with a large array of genetic alterations beyond mutations in BRAF. We anticipate that some tumors will be more or less sensitive to treatment based on the constellation of other genetic changes, and these will be defined as predictive markers. Understanding the network of signal transduction pathways and how they may adapt to BRAF will point to the next generation of clinical trials investigating rational sequences of therapy and combination regimens.
1. Davies H, Bignell GR, Cox C, et al. Mutations of the BRAF gene in human cancer. Nature 2002; 417(6892):949-54.
2. Eisen T, Ahmad T, Flaherty KT, et al. Sorafenib in advanced melanoma: a Phase II randomised discontinuation trial analysis. Br J Cancer 2006; 95(5):581-6.
3. Flaherty KT, Redlinger M, Schuchter LM, et al. Phase I/II, pharmacokinetic and pharmacodynamic trial of BAY 43-9006 alone in patients with metastatic melanoma. Proc Am Soc Clin Oncol 2005; 23(16S pt 1): abstract 3037.
4. Hauschild A, Agarwala SS, Trefzer U, et al. Results of a phase III, randomized, placebo-controlled study of sorafenib in combination with carboplatin and paclitaxel as second-line treatment in patients with unresectable stage III or stage IV melanoma. J Clin Oncol 2009; 27(17):2823-30.
5. McDermott DF, Sosman JA, Gonzalez R, et al. Double-blind randomized phase II study of the combination of sorafenib and dacarbazine in patients with advanced melanoma: a report from the 11715 Study Group. J Clin Oncol 2008; 26(13):2178-85.
6. Flaherty KT, Puzanov I, Sosman J, et al. Phase I study of PLX4032: Proof of concept for V600E BRAF mutation as a therapeutic target in human cancer. J Clin Oncol 2009; 27(15S) (suppl: abstr 9000).
7. Tsai J, Lee JT, Wang W, et al. Discovery of a selective inhibitor of oncogenic B-Raf kinase with potent antimelanoma activity. Proc Natl Acad Sci USA 2008; 105(8):3041-6.
8. Puzanov IN, Nathanson KL, Chapman PB, et al. PLX4032, a highly selective V600EBRAF kinase inhibitor: Clinical correlation of activity with pharmacokinetic and pharmacodynamic parameters in a phase I trial. J Clin Oncol 2009; 27(15s) (May 20 suppl: abstract 9021).