Published OnlineFirst February 15, 2019; DOI: 10.1158/2159-8290.CD-18-1321 REVIEW Targeting Alterations in the RAF–MEK Pathway Rona Yaeger1 and Ryan B. Corcoran2 ABSTRACT The MAPK pathway is one of the most commonly mutated oncogenic pathways in cancer. Although RAS mutations are the most frequent MAPK alterations, less fre- quent alterations in downstream components of the pathway, including the RAF and MEK genes, offer promising therapeutic opportunities. In addition to BRAFV600 mutations, for which several approved therapeutic regimens exist, other alterations in the RAF and MEK genes may provide more rare, but tractable, targets. However, recent studies have illustrated the complexity of MAPK signaling and high- lighted that distinct alterations in these genes may have strikingly different properties. Understanding the unique functional characteristics of specific RAF and MEK alterations, reviewed herein, will be criti- cal for developing effective therapeutic approaches for these targets. Significance: Alterations in the RAF and MEK genes represent promising therapeutic targets in multi- ple cancer types. However, given the unique and complex signaling biology of the MAPK pathway, the diverse array of RAF and MEK alterations observed in cancer can possess distinct functional character- istics. As outlined in this review, understanding the key functional properties of different RAF and MEK alterations is fundamental to selecting the optimal therapeutic approach. INTRODUCTION RAS to its inactive GDP-bound state (2). In its active, GTP- bound state, a conformational change occurs in the switch The MAPK signaling pathway is critically involved in I and II regions of RAS, which facilitate interactions with a many important cellular processes. Its dysregulation leads variety of downstream effectors, including the RAF family to uncontrolled cellular proliferation, survival, and dediffer- of kinases (ARAF, BRAF, and CRAF, the latter of which is entiation. As a consequence, the MAPK pathway is altered or encoded by the RAF1 gene; refs. 3–5). The association of RAF inappropriately activated in a majority of cancers. proteins with activated RAS through their conserved RAS- Under physiologic conditions, MAPK signaling is trig- binding domains leads to the formation of RAF homodimers gered through activation of RAS proteins (KRAS, NRAS, and (i.e., CRAF–CRAF) or heterodimers (i.e., BRAF–CRAF) with HRAS), a family of small guanine triphosphatases (GTPases) activated RAF kinase activity. For example, prior to binding that integrate signals from a variety of upstream sources, activated RAS, BRAF is in an autoinhibited conformation in most commonly from activated receptor tyrosine kinases which a short α helix in its activation loop associates and dis- (RTK; ref. 1). These upstream signals lead to activation of places the critical αC helix in the kinase domain in an inactive guanine nucleotide exchange factors, such as son-of-sevenless “out” state. BRAF dimerization and activation loop phospho- (SOS), which catalyze the exchange of RAS-bound guanine rylation destabilizes this autoinhibitory interaction to move diphosphate (GDP) for guanine triphosphate (GTP). RAS the BRAF kinase into the helix “αC-in” active conformation activity is negatively regulated by GTPase-activating proteins, (6). Once activated, RAF kinases phosphorylate and activate such as neurofibromin 1 (NF1), which augment the GTPase MEK kinases (MEK1 and MEK2, encoded by the MAP2K1 activity of RAS to hydrolyze GTP to GDP, thus reverting and MAP2K2 genes, respectively), which in turn phosphoryl- ate and activate ERK kinases (ERK1 and ERK2). The activated 1Memorial Sloan Kettering Cancer Center, New York, New York. ERK kinases then phosphorylate a host of critical substrates 2 Massachusetts General Hospital Cancer Center and Department of that regulate key cellular processes. Medicine, Harvard Medical School, Boston, Massachusetts. Given the many important roles of MAPK pathway signal- Corresponding Authors: Ryan B. Corcoran, Harvard Medical School, ing, activation of the pathway is tightly regulated. There are 149 13th Street, 7th Floor, Boston, MA 02129. Phone: 617-726-8599; Fax: 617-643-0798; E-mail: [email protected]; and Rona Yaeger, several levels of negative feedback controls that limit physi- Memorial Sloan Kettering Cancer Center, 300 East 66th Street, 10th Floor, ologic activation of MAPK signaling. For instance, negative New York, NY 10065. Phone: 646-888-5109; Fax: 646-888-4254; E-mail: feedback loops from ERK include direct inhibitory phospho- [email protected] rylation of CRAF and BRAF and induction of expression of doi: 10.1158/2159-8290.CD-18-1321 multiple MAPK phosphatases, such as DUSPs (7). ERK also ©2019 American Association for Cancer Research. inhibits the activation of RAS by RTKs by phosphorylating MARCH 2019 CANCER DISCOVERY | OF1 Downloaded from cancerdiscovery.aacrjournals.org on October 1, 2021. © 2019 American Association for Cancer Research. Published OnlineFirst February 15, 2019; DOI: 10.1158/2159-8290.CD-18-1321 REVIEW Yaeger and Corcoran Mutant activator of signaling Mutant amplifier of signaling RTK/RAS RTK/RAS signaling signaling Mutant Mutant ERK ERK Figure 1. Function of activator and amplifier mutations in the MAPK signaling pathway. Schema showing the effect of “activator” versus “amplifier” mutants in the MAPK pathway. Activator alterations lead to constitutive MAPK signaling through ERK activation that is independent of upstream path- way activity. Activator mutants strongly activate ERK and lead to negative feedback suppression of upstream signaling. Amplifier mutants augment the downstream signal to ERK and commonly co-occur with other activating mutations upstream in the pathway. They lead to modest activation of ERK and consequently cause minimal negative feedback inhibition of upstream signaling. SOS and a variety of RTKs and by inducing the expression characterized into two groups: activators and amplifiers of members of the Sprouty family of proteins (8). These (Fig. 1). “Activator” alterations lead to constitutive MAPK feedback signals modulate the output of oncogenic altera- pathway signaling through ERK activation that is independ- tions within the pathway, affecting the spectrum of recurrent ent of upstream pathway activity. Conversely, “amplifier” oncogenic alterations at each level of the pathway with impli- alterations are dependent on upstream activity and augment cations for targeted therapy response and drug resistance. the downstream signal to ERK. Activating alterations strongly The frequency of genomic alterations in the MAPK pathway activate ERK and are usually mutually exclusive with each decreases in incidence as one moves further downstream in the other, whereas amplifying alterations commonly co-occur pathway: across human tumors, RAS mutations occur in 22%, with other activating mutations upstream in the pathway. BRAF in 7%, and MEK in <1% of cases, and ERK mutations Interestingly, the incidence of activating mutations decreases are exceptionally rare. The degree of ERK activation produced further downstream in the pathway, and the proportion of by alterations upstream in the pathway (e.g., RAS mutations) amplifying alterations increases; activating alterations in is often susceptible to constraint by negative feedback signals, downstream components of the pathway, such as MEK or whereas those further downstream escape negative feedback ERK, would lead to very high levels of output as they evade regulation and can lead to more profound activation of path- feedback signals and may thus have a selective disadvantage. way output. In papillary thyroid cancers, where expression of Understanding the unique signaling properties of specific ERK-responsive genes important in iodide transport can be RAF and MEK alterations is key to devising strategies to over- readily assayed, differences in expression of ERK-responsive come them. Here, we summarize our current understanding genes are seen between BRAFV600E mutants (strongly activat- of recurrent alterations in RAF and MEK and their effects on ing) and RAS mutants (less activating; ref. 9). signaling, targeted therapy response, and drug resistance. On RAS mutations are by far the most common MAPK altera- the basis of this mechanistic framework, we outline rational tions observed in human cancer, and RAS signaling and strategies to target these specific alterations. strategies for targeting RAS have been reviewed extensively elsewhere (10). This review focuses on downstream altera- tions in the MAPK pathway, including alterations in the RAF ALTERATIONS RAF and MEK genes. Given the unique signaling biology BRAF is by far the most frequently altered gene in the of this pathway, recent studies have suggested that down- MAPK pathway downstream of RAS, altered in 7% to 10% of stream alterations in the MAPK pathway can be broadly all cancers (11). Point mutations are the most common mode OF2 | CANCER DISCOVERY MARCH 2019 www.aacrjournals.org Downloaded from cancerdiscovery.aacrjournals.org on October 1, 2021. © 2019 American Association for Cancer Research. Published OnlineFirst February 15, 2019; DOI: 10.1158/2159-8290.CD-18-1321 Targeting RAF and MEK Alterations REVIEW Class I Class II Class III RTK RASmut NF1 GDP RAS GDP RAS RAS RAS GDP GDP GTP RAS GTP RAS GDP RAS GDP RAS GTP RAS GTP RAS GTP RAS GTP RAS GT GT GT GT P P P P RA RA RA RA S S S B S mut Bmut Bmut Bmut C Bmut C MEK/ERK MEK/ERK MEK/ERK MEK/ERK V600E/K/D/R/M P367L/S D287H G464V/E V459L G469A/V/R G466A/E/V