Emerging Targeted Therapies for KRAS-Mutated Non–Small Cell Lung Cancer

Published OnlineFirst June 3, 2014; DOI: 10.1158/1078-0432.CCR-13-1762

Clinical Cancer Review Research

A RAS Renaissance: Emerging Targeted Therapies for KRAS-Mutated Non–Small Cell Lung Cancer

Neil Vasan1, Julie L. Boyer2, and Roy S. Herbst3

Abstract Of the numerous oncogenes implicated in human cancer, the most common and perhaps the most elusive to target pharmacologically is RAS. Since the discovery of RAS in the 1960s, numerous studies have elucidated the mechanism of activity, regulation, and intracellular trafficking of the RAS gene products, and of its regulatory pathways. These pathways yielded druggable targets, such as farnesyltransferase, during the 1980s to 1990s. Unfortunately, early clinical trials investigating farnesyltransferase inhibitors yielded disappointing results, and subsequent interest by pharmaceutical companies in targeting RAS waned. However, recent advances including the identification of novel regulatory enzymes (e.g., Rce1, Icmt, Pded), siRNA-based synthetic lethality screens, and fragment-based small-molecule screens, have resulted in a "Ras renaissance," signified by new Ras and Ras pathway–targeted therapies that have led to new clinical trials of patients with Ras-driven cancers. This review gives an overview of KRas signaling pathways with an emphasis on novel targets and targeted therapies, using non–small cell lung cancer as a case example. Clin Cancer Res; 20(15); 3921–30. 2014 AACR.

Introduction sary for GTP hydrolysis (6), whereas G12 and G13 mutants Three RAS genes encode four proteins: HRas, KRas4a, prevent binding of Ras to its GAP and interfere with the KRas4b, and NRas (1). These proteins are GTPases, which orientation of Q61. These mutants result in Ras-GTP in an function as molecular switches: "on" when bound to GTP "on" state, driving oncogenesis (7; Fig. 1B). and "off" when bound to GDP. Ras-GTP can bind to The Ras proteins are important mediators of cell signal- numerous partner proteins, termed "effectors," and these ing. There is a wide range of Ras effector proteins, notably Ras-effector interactions lead to a cascade of downstream Raf (MAP kinase pathway), PI3K (Akt/mTOR pathway), signaling events (2). In normal cells, Ras signaling is crucial and RalGDS (Ral pathway). These effectors (which repre- for proliferation, differentiation, and survival (3). sent only a subset of downstream Ras signaling nodes) are The hydrolysis of GTP to GDP by Ras is a slow process, highly complex with numerous redundancies and interac- and therefore Ras cycles between these states with the aid of tions between pathways (8). Dysregulated Ras signaling regulatory proteins. GTPase-activating proteins (GAP) cat- results in increased proliferation, decreased apoptosis, dis- alyze the hydrolysis of GTP to GDP ("on to off"), whereas rupted cellular metabolism, and increased angiogenesis, all guanine nucleotide exchange factors (GEF) catalyze the seminal hallmarks of cancer (9; Fig. 1B). dissociation of GDP, with GTP binding afterward due to Ras Mutations: Differences from Isoform to its high concentration in cells ("off to on"; ref. 4; Fig. 1A). Amino Acid However, this pathway is co-opted by oncogenic mutations in Ras. Among the four Ras isoforms, the most RAS is the most commonly mutated oncogene in cancer (8), common mutations are at amino acid positions G12, with distinct Ras isoforms detected in various cancers (10). G13, and Q61 (5). Crystal structures of Ras proteins have KRas is the most commonly mutated isoform. Listed in order modeled these mutants’ mechanisms of activation. Q61 of percentage of cases, KRas mutations are most common in mutants prevent coordination of a water molecule neces- cancers of the pancreas, colon, biliary tract, and lung (the majority of which are adenocarcinomas); NRas mutations are most common in cancers of the skin (malignant melanoma) Authors' Afﬁliations: 1Department of Internal Medicine, Massachusetts and hematopoietic system (acute myeloid leukemia, AML); General Hospital, Boston, Massachusetts; 2The Sandra and Edward Meyer HRas mutations are most common in cancers of the head and Cancer Center at Weill Cornell Medical College, New York, New York; and 3Yale Cancer Center and Smilow Cancer Hospital at Yale-New Haven, New neck (squamous cell carcinoma) andurinarytract(transitional Haven, Connecticut cell carcinoma). Ras mutations are much less common in Corresponding Author: Roy S. Herbst, Yale University, 333 Cedar Street, cancers of the breast, central nervous system, or prostate WWW-221, New Haven, CT 06520. Phone: 203-785-6879; Fax: 203-737- (5; Fig. 2A). Why certain cancers seem to be driven preferen- 5698; E-mail: [email protected] tially by speciﬁc isoforms remains an outstanding question. doi: 10.1158/1078-0432.CCR-13-1762 Another unsettled issue in oncogenesis is the differential 2014 American Association for Cancer Research. role, if any, among different Ras-activating point mutations.

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RAS-GDP RAS-GDP GDP Pi GDP Pi

GEF GAP GEF GAP

GTP RAS-GTP GTP RAS*-GTP

Figure 1. A, normal Ras signaling. B, oncogenic Ras signaling. When Effector RAF PI3K RALGDS Ras is mutated, it is constitutively bound to GTP such that its GAP cannot bind. The activated Ras signals through a multitude of effectors and downstream Normal signaling MEK AKT RAL signaling pathways, a subset of which is shown here.

ERK mTOR RLIP

Oncogenic signaling

In lung cancer, the most common mutations are KRas Together, the in vitro, in vivo, and patient data suggest a G12C, G12V, and G12D (11). Other KRas-driven cancers greater oncogenic potential for KRas G12V (present in have different mutational frequencies: in the colon G12D, 20% of KRas-mutated lung cancers) compared with other G12V, and G13D; in the pancreas G12D, G12V, and G12R; mutations (18). and in the biliary tract G12D, G12V, and G12S (Fig. 2B). Unlike in HRas and NRas, KRas Q61 oncogenic mutations are very rare (7). Inhibiting Ras Membrane Association In vitro, KRas G12V and G12R have greater transform- A series of enzymes (Fig. 3), beginning with farnesyl- ing ability, as shown by soft agar colony formation (12). transferase (FTase), acts posttranslationally on the C-termi- Unexpectedly, there is no correlation between the GTPase nal C-A-A-X motif of Ras, resulting in the attachment to activity of the mutant and its propensity to transform membranes through cysteine prenylation (19). Next, Ras (13). However, once transformed, certain mutations are traffics to the endoplasmic reticulum, where its last three more aggressive than others. Mice with KRas G12V, amino acids are proteolyzed by Ras-converting enzyme G12R, and G12D had higher-stage lung tumors com- (RCE1), and then its C-terminus is methylated by isopre- pared with KRas G12C or wild-type (14). In patients nylcysteine carboxyl methyltransferase (ICMT; ref. 20). In with lung cancer, KRas G12C resulted in increased sen- the Golgi, HRas, NRas, and KRas4A are palmitoylated, and sitivity to pemetrexed and paclitaxel compared with the fully processed Ras then traffics to its final plasma G12V and G12D, although G12D patients were more membrane location (21). KRas4b is not palmitoylated but likely to respond to sorafenib (15). rather associates electrostatically with the membrane A recent retrospective analysis of the Biomarker-integrat- through a polybasic stretch in its C-terminus (22). Thus, ed Approaches of Targeted Therapy for Lung Cancer Elim- two modes of membrane association poise Ras isoforms for ination (BATTLE) clinical trial (16; discussed below) found activation and signaling. worse progression-free survival (PFS) for the group of patients with either KRas G12C or G12V, compared with FTase other KRas mutants, or wild-type [1.84 months vs. 3.35 Initial attempts to inhibit Ras focused on FTase (23). months (P ¼ 0.046), vs. 1.95 months; ref. 17]. KRas G12C These FTase inhibitors (e.g., lonafarnib, tipifarnib; ref. 24) and G12V had increased signaling through Ral and were oral medications, well tolerated, specific for FTase, and decreased signaling through Akt. This study suggests that were effective against HRas-transformed cells and HRas- targeted treatments and clinical trials in non–small cell lung driven murine tumors (25). However, these drugs did not cancer (NSCLC) may need to consider the specific KRas increase survival in clinical trials of patients with KRas- point mutation. mutated pancreatic cancer (26). Later studies found that

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Figure 2. A, graph showing the percentage of cancers with Ras A mutations in different organ types, 60 arranged in descending KRAS frequency. KRas, NRas, and 50 HRas-driven cancers are denoted NRAS by color. The frequencies of the 40 HRAS predominant histology of that organ-speciﬁcKRas-mutated 30 cancer are listed below. For example, 17% of all lung cancers 20 have KRas mutations; of these with RAS mutations

KRas-mutated lung cancers, 53% of cancers Percentage 10 are adenocarcinomas. These frequencies are likely 0 underestimates as many samples Pancreas Colorectal Biliary Lung Skin Hematologic Urinary Head and deposited into the COSMIC tract tract neck database are listed as a Frequency of 78% 91% 72% 53% 66% 42% 39% 69% "nonspecific" histology. Data were predominant Adenocarcinoma Melanoma AML Transitional Squamous accessed on May 15, 2013. B, pie histology cell carcinoma charts showing frequencies of different KRas mutations in KRas- B mutated lung, colorectal, pancreatic, and biliary tract G12V cancers. In all cancers, G12D and Lung Colorectal Pancreatic Biliary tract G12D G12V mutations are common; G12C however, each cancer displays a G12R different "KRas profile." In lung G12S cancer, G12C is the most common mutation, followed by G12V and G12A G12D. In colorectal, pancreatic, G13D and biliary tract cancers, the most common mutations are G12D and G12V; the third most common mutations are G13D, G12R, and © 2014 American Association for Cancer Research G12S, respectively, for these organ subtypes. CCR Reviews with inhibition of FTase, KRas could be alternatively pre- status or other mutations in the Ras signaling pathway nylated by geranylgeranyltransferase I (GGTase I; ref. 25). (34).Thissuggestsalternativemechanismsofresistance Moreover, dual inhibition of FTase and GGTase I did not such as Ras gene amplification, which has been observed decrease levels of prenylated KRas (27). Notably, tipifarnib asaresistancemechanismtoMETtyrosinekinaseinhi- has shown antitumor activity against AML, chronic mye- bitors (35) or off-target effects, which could be mediated logenous leukemia, and myeloproliferative disorders (28; by the large number (>55) of prenylated substrates (e.g., often driven by NRas) and in breast cancer, (29) which other small GTPases; ref. 36). This may explain why warrants further study. there is not an adequate single biomarker, such as dec- Another class of FTase inhibitors has been developed reased Ras prenylation, for monitoring FTase inhibitor (e.g., salirasib; ref. 30) containing farnesylcysteine, thought effects. to compete for membrane-docking proteins that bind far- Nevertheless, many current clinical trials are investigat- nesyl moieties (Fig. 3). In vitro, salirasib inhibits all Ras ing combination therapies of FTase inhibitors with other isoforms (31). However, it failed to induce radiographic cytotoxic and targeted therapies (37). The failure of FT response or increase survival in a phase II NSCLC trial (32). inhibition as a general strategy for targeting KRas has Together, these studies revealed several inherent pro- spurred preclinical studies of the other enzymes in the Ras blems in targeting Ras prenylation. First, there is alternative processing pathway: Rce1, Icmt, and Pded. prenylation with dissimilarities among isoforms—KRas and NRas are prenylated by GGTase I (33)—previously RCE1, ICMT, PDEd thought to be identical to that of HRas. Although HRas Disruption of the RCE1 gene slows mouse fibroblast cell mutations are infrequent compared with KRas or NRas, growth (38); conversely, in another study, deficiency of FTase inhibitors would target HRas because HRas does not RCE1 in a genetically engineered mouse model (GEMM) of undergo alternative prenylation by GGTase I (33). Such KRas-driven myeloproliferative disease actually increased findings could be exploited in a clinical trial of FTase disease progression (39). These paradoxical results may inhibitors for patients with HRas-driven urothelial cancers. reflect cell-specific differences in KRas signaling. Several In addition, any observed decreases in tumor size due inhibitors against yeast and human Rce1 have been devel- to FTase inhibition do not correlate with KRas mutation oped (40); however, these bind with only low micromolar

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Tipifarnib ER SH S S S

CAAX FTASE CAAX CAAX CAAX RAS RAS RAS RAS GGTASE RCE1 ICMT

Cysmethynil, Figure 3. Ras processing, Inhibitors fi methotrexate traf cking, membrane association, and inhibition. Ras is first farnesylated by FTase. In the endoplasmic reticulum (ER), it is PDE6d Palmitoyl proteolyzed by RCE1 and Benzimidazole transferases methylated by ICMT. HRas, NRas, compounds and KRas4a traffic to the Golgi, KRAS4b PDE6d (CO)OMe RAS (CO)OMe where they are palmitoylated. C C +++ Then they traffic to and associate S S with the plasma membrane. After RAS (CO)OMe depalmitoylation, these isoforms C Depalmitoylation Golgi are chaperoned by PDE6d and reaccumulate at the Golgi for S another round of palmitoylation. KRas4b is chaperoned by PDE6d Salirasib and associates with the plasma membrane through its farnesyl group and electrostatically through a positively charged Plasma membrane patch. Membrane association of Ras isoforms is aided by farnesyl- binding membrane docking RAS All RAS isoforms +++ Positively charged patch proteins. Relevant potential inhibitors are indicated in the fi Palmitoyl moiety gure. KRAS4b KRAS4b Farnesyl moiety

Non-KRAS4b isoforms Farnesyl-binding membrane RAS (KRAS4a, HRAS, NRAS) docking protein

affinity so future optimization must occur before these HRas, NRas, and KRas4a at the Golgi (45; Fig. 3). Impor- drugs enter clinical trials. tantly, downmodulation of Pde6d decreases oncogenic Inactivation of ICMT in the presence of activated KRas in KRas signaling. A recent article reports benzimidazole fibroblasts leads to decreased cell growth and xenograft small-molecule compounds that inhibit the mammalian tumor formation (41). Concordantly, inactivation of ICMT Pded–KRas interaction with nanomolar affinity, suppres- in a GEMM of KRas-driven myeloproliferative disease also sing oncogenic signaling in vitro in KRAS-mutated pancre- decreased lung tumor formation (42). Several small-mol- atic ductal adenocarcinoma cells and most tantalizingly in ecule inhibitors of Icmt have been developed including vivo in a pancreatic cancer mouse xenograft model (46). cysmethynil, which reduces anchorage-independent cell These findings likely will herald phase I trials of these novel growth in colon cancer cells (43). Of note, inhibition of inhibitors. Icmt has been shown to be an off-target effect of methotrexate, possibly through an increase in S-adenosylhomo- cysteine, a methyltransferase inhibitor (44). Direct Inhibition of the Ras Protein Another recently discovered intracellular trafficking tar- The affinity of GTP for Ras is extremely potent, in the get is Pded (phosphodiesterase delta), a subunit of the cyclic picomolar range (47); thus, attempts to inhibit Ras com- GMP phosphodiesterase complex. Pded also functions as a petitively would be difficult. Several previous reports have chaperone protein that binds to farnesylated Ras and been given of Ras small-molecule inhibition—nucleotide enhances the presence of fully processed KRas4b at the exchange inhibitors (48) and Ras–Raf inhibitors (49)—but plasma membrane, and the presence of depalmitoylated in the absence of clear structural data, it is difficult to know if

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these effects are direct or indirect. This has led to a search for and combination approaches of targeted therapies with allosteric inhibitors of Ras. conventional cytotoxic chemotherapy. On the basis of the crystal structure of SOS (a Ras GEF) bound to Ras, Patgiri and colleagues designed a cell-per- Identification of synthetic lethal targets meable peptide to inhibit Ras activation (50). As shown by Synthetic lethality is a phenomenon through which per- nuclear magnetic resonance (NMR) spectroscopy, the pep- turbation in either one of two different genes does not cause tide binds Ras with micromolar affinity at the same location cell death, but perturbation of both leads to cell death (54). In as the analogous region of SOS, but does so about twice as cancer, this initial perturbation could be a mutated oncogene avidly. This results in inhibition of downstream MAP kinase or tumor suppressor, making this an attractive strategy for signaling. targeting cancer cells because normal cells would not have the Two recent groups (51, 52) have used NMR fragment- mutation of interest and would theoretically be unaffected by based lead discovery and structure-based drug design to find a targeted treatment. This is also a strategy to develop ways to novel small-molecule allosteric inhibitors of KRas. Both target oncogenes that are not readily "druggable" (e.g., Ras, groups’ inhibitors bind at a hydrophobic pocket between transcription factors) or tumor suppressors in which protein the switch 2 and core b sheet region of the protein, with target levels are decreased. The most successful example of micromolar affinity. The binding site is distinct from but this approach in oncology is the use of mTOR and VEGF partially overlapping with the SOS binding site such that inhibitors that indirectly target HIF1a in von Hippel-Lindau– SOS is unable to activate KRas. mutated clear cell renal cell carcinoma (55). Several FDA-approved tyrosine kinase inhibitors (e.g., Although the Ras signaling pathway is complex, it is afatinib, ibrutinib) take advantage of irreversible binding thought that there may be critical nodes in the pathway to a cysteine amino acid residue close to the active site. This that could be exploited. In KRas-mutated NSCLC, several approach inspired the development of irreversible inhibitors small-molecule synthetic lethality screens have yielded lead of KRas G12C, a specific mutant in which the glycine at the compounds including lanperisone, which induces oxida- twelfth position of the KRas protein is mutated to cysteine tive stress in KRas-mutated cells (56), and oncrasin, which is (53). With submicromolar affinity, these inhibitors block synthetically lethal between KRas and PKCi (protein kinase SOS-mediated nucleotide exchange, favoring the binding C iota) and functions through inhibition of RNA poly- of GDP instead of GTP, and rendering the KRas protein in merase II (57). Moreover, the rheumatoid arthritis drug an "off" state. When bound to KRas G12C, the compounds aurothiomalate, which inhibits PKCi signaling, is currently create a new binding surface mostly involving the switch 2 in phase I clinical trials in NSCLC (58). More recently, RNA region. Importantly, they decreased viability and activated silencing technologies have facilitated the identification of apoptosis in a KRas G12C–specific lung cancer cell line. new targets that, when deleted, are synthetically lethal with Although these approaches are appealing, more potent KRas mutations; this can be exploited clinically if the newly drugs that bind with nanomolar affinity would be needed identified target has a known inhibitor. For example, in for a viable drug. Nevertheless, these studies provide novel KRas-driven colon cancer, previous RNAi studies have lead compounds for future optimization. shown the importance of TAK1 (TGFb activated kinase- 1), a MAP kinase kinase kinase, and Polo-like kinase 1, which functions at the mitotic spindle (59). Here, we focus Inhibiting Downstream Ras Signaling on novel targets in preclinical development elucidated Many classes of inhibitors exist against components of through synthetic lethality studies of NSCLC (Table 1). the canonical Ras signaling pathway, including Raf, mTOR, PI3K, PI3K/mTOR, Akt, and MEK. Clinical trials with these TBK1 agents are ongoing, but to date, these drugs have not been Using an shRNA screen targeting genes encoding drug- shown to be effective against Ras-driven cancers as single gable proteins—kinases, phosphatases, and oncogenes— agents. This has led to the identification of new targets, Barbie and colleagues identified TANK-binding kinase 1 combination approaches of multiple targeted therapies, (TBK1) as essential in KRas-driven cancers (60). These

Table 1. Genes that are synthetically lethal with KRas in KRas-mutated lung adenocarcinoma cells, their functions, and inhibitor or inhibitor combinations

Gene Function Inhibitors Reference TBK1 Kinase 6-aminopyrazolopyrimidine derivatives 60, 61, 62 WT1 Transcription factor None 64, 65 CDK4 Kinase PD-0332991 66 GATA-2 Transcription factor Bortezomib þ fasudil 67, 68 BCL-XL Antiapoptotic factor Selumetinib þ navitoclax (ABT-263) 69

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authors validated initially identified candidate genes onto NF-kB signaling, which contributes to KRas-driven through a higher-stringency secondary screen in lung ade- oncogenesis. nocarcinoma cells, with TBK1 knockdown leading to the Because transcriptional factors are notoriously difficult greatest amount of cell death, after KRas itself. TBK1 is to target, the authors relied on approved drugs that target linked to KRas through RalB, a small GTPase downstream these newly delineated components of the GATA-2 path- of KRas that is part of the Ral signaling pathway, and is way: bortezomib, which inhibits the proteasome and NF-kB activated by RalB and Sec5, a component of the exocyst, an indirectly, and fasudil, which inhibits Rho/ROCK signaling. intracellular tethering complex (61). Activated TBK1 leads Together, this combination therapy resulted in tumor to increased antiapoptotic NFkB signaling through B-cell regression in a KRas-driven lung cancer GEMM. This non- lymphoma-extra large (BCL-XL) and the c-Rel proto-onco- oncogene addiction to GATA-2 was also confirmed inde- gene. TBK1 inhibitors with nanomolar affinity have been pendently by a separate group (68). developed (62), confirming pharmacologic tractability. BCL-XL WT1 With the goal of finding genes whose inhibition may Targeting genes from a previously identified KRas tran- cooperate with the MEK inhibitor selumetinib in KRas- scriptional signature (63), as well as transcriptional regu- mutated cells, Corcoran and colleagues (69) designed a lators, Vicent and colleagues identified the Wilms tumor 1 pooled shRNA drug screen of "druggable" genes and iden- transcription factor (WT1) as critical for oncogenic KRas tified BCL-XL, an antiapoptotic gene. Using navitoclax, a signaling (64). Mechanistically, loss of WT1 decreases pro- BCL-XL small-molecule inhibitor, in combination with liferation and increases cell senescence in KRas-driven can- selumetinib, they showed increased apoptosis in KRas- cers and was confirmed in mouse cell lines, GEMMs, and in driven lung adenocarcinoma cell lines. Alone, selumetinib human cell lines. Moreover, the authors were able to cor- increases the amount of BIM protein, a proapoptotic factor, relate WT1 expression levels with prognosis in patients with but also increases the amount of BIM bound to BCL-XL; BIM KRas-driven lung cancer, strengthening this connection. must be unbound to induce apoptosis, so its levels induced WT1 is a well-known tumor suppressor in Wilms tumor, by selumetinib are insufficient for this effect. but recent data have implicated it as a possible oncogene However, the combination of selumetinib with navito- with overexpression in lung cancer (65). Although WT1 is clax increases the total amount of BIM, and decreases the currently not druggable, further examination of WT1’s role amount of BIM/BCL-XL complex. Some of this free BIM in cell senescence may yield future therapies. forms a complex with MCL-1, and this complex is able to drive apoptosis. Thus, the authors proposed that the com- CDK4 bination therapy "frees up" more BIM to sufficient levels to By querying the role of individual cyclin-dependent drive apoptosis. Demonstrating the robustness and gener- kinases (CDK) in KRas-driven NSCLC, Puyol and colleagues alizability of this combination therapy, they show increased found a synthetic lethal interaction by disrupting CDK4, but apoptosis in a large percentage of KRas-driven colorectal, not other related CDKs, causing cellular senescence (66). lung, and pancreatic cancer cell lines; in KRas-mutant This effect was recapitulated in mouse embryonic fibro- xenografts; and in a KRas-driven lung cancer GEMM. blasts and in a KRas-driven GEMM. Interestingly, the neces- sity of CDK4 in KRas-driven cancer was observed only in the lung, and not in the colon or pancreas, pointing to possible Clinical Trials Targeting KRas-Driven NSCLC tissue-specific dependencies in the Ras pathway. In addi- Patients with KRas-mutated NSCLC derive less benefit tion, these authors showed that small-molecule inhibition from clinical trials compared with their KRas-wild-type of CDK4 (using a CDK4/CDK6 dual antagonist) resulted in patient counterparts (70, 71). Several targeted therapy clin- tumor regression, with decreased CDK4-mediated phos- ical trials specifically address patients with KRas-mutated phorylation; however, this inhibition did not cause the NSCLC. These trials represent the translational extension of previously observed senescence, pointing to the need for the many decades of basic science research on the Ras more potent and specific CDK4 inhibitors as well as using pathway (Table 2). them in combination therapies. BATTLE and BATTLE-2 GATA-2 As we have seen, biomarkers (e.g., mutations, overexpres- Screening KRas-driven human NSCLC cell lines, Kumar sion) do not always correlate with effects of a targeted and colleagues (67) discovered the transcription factor therapy. Moreover, as patients receive multiple treatment GATA-2 as necessary for cell viability, in vitro and in vivo modalities, these markers may change even though the in a lung cancer xenograft model. Gene expression analysis therapy is often dictated by the pretreatment tumor geno- revealed multiple upregulated pathways: the proteasome, type. The phase II BATTLE trial addressed this issue in a IL1 signaling, and Rho/ROCK signaling. The authors novel manner (16). Ninety-seven patients with chemore- showed that GATA-2 normally upregulates the proteasome fractory NSCLC were randomized into 4 treatment groups through Nrf1, IL1 signaling through TRAF6, and Rho sig- based on biomarker analysis of prospectively biopsied naling through STAT5. Together, these pathways converge tumors: KRas/BRaf mutation, treated with sorafenib; EGFR

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Table 2. Past and present clinical trials targeting KRas-mutated NSCLC

KRas-mutant patient response Drug combination Target (vs. KRas wild-type patients) Reference Sorafenib Raf, VEGFR Increased DCR (61% vs. 32%, P ¼ 0.11) NCT00409968 Selumetinib þ docetaxel MEK 1/2, Increased PFS (5.3 months vs. 2.1 months, NCT00890825 microtubules 80% CI, 0.42–0.79; one-sided P ¼ 0.014) Increased OS (9.4 months vs. 5.2 months, 80% CI, 0.56–1.14; one-sided P ¼ 0.21) Selumetinib þ MK2206 MEK 1/2, Akt Ongoing NCT01248247 Bortezomib Proteasome Ongoing NCT01833143 Retaspimycin þ everolimus Hsp90, mTOR Ongoing NCT01427946 Selumetinib þ erlotinib versus MEK 1/2, EGFR Ongoing NCT01229150 selumetinib alone

NOTE: Drug combinations, targets, patient responses, and references are tabulated. DCR (disease control rate) ¼ complete response, partial response, or stabilized disease.

mutation/copy number, treated with erlotinib; VEGF/ Selumetinib/docetaxel combination therapy VEGFR2 expression, treated with vandetanib; and RXRs/ A recent phase II clinical trial tested a combination of Cyclin D1 expression and CCND1 copy number, treated MEK inhibition with cytotoxic chemotherapy (74). J€anne with bexarotene and erlotinib. and colleagues randomized 87 patients with pretreated On the basis of the cumulative real-time results of the initial KRas-mutant NSCLC to receive either docetaxel alone or 97 patients over 8 weeks of treatment, 158 additional patients in combination with selumetinib (74); selumetinib alone were adaptively randomized to receive the most effective did not increase overall survival in NSCLC (75). Combi- therapy for their particular biomarker profile. In other words, nation therapy increased overall survival, however, without if a patient from the first group with a KRas mutation had statistical significance [9.4 months vs. 5.2 months; HR, an adequate clinical response to sorafenib, there would be a 0.80; 80% confidence interval (CI), 0.56–1.14; one-sided greater than 25% chance that another patient with a KRas P ¼ 0.21]. Median PFS was significantly increased (5.3 mutation would be placed into that treatment arm. months vs. 2.1 months; HR, 058; 80% CI, 0.42–0.79; In total, 20% of the 255 patients randomized had KRas one-sided P ¼ 0.014). Of note, 37% (n ¼ 16) of patients mutations. The overall 8-week disease control rate (DCR) given the combination therapy had an objective response, was 46%; patients in the sorafenib arm had the highest DCR as measured by a decrease in tumor burden; however, 82% at 58%. Post hoc biomarker analysis showed that relative to (n ¼ 36) of the combination group had adverse events of the other treatments, sorafenib had a significantly higher grade 3 or 4, mostly neutropenia, febrile neutropenia, or DCR in EGFR wild-type patients (64% vs. 33%, P < 0.001), asthenia. and higher but nonstatistically significant DCR in KRas- Waterfall plots (graphs that depict the continuum of mutant patients (61% vs. 32%, P ¼ 0.11). Of note, in the tumor growth, positive to negative for all patients in a KRas/BRaf group, sorafenib gave a higher DCR than erlo- study) of response to this therapy were widespread, with tinib (79% vs. 14%, P ¼ 0.016), with the G12C/G12V group 5 patients in the combination group having a >20% increase associated with decreased PFS compared with other KRas in tumor size (74). It would be interesting to know if these mutants or KRas wild-type (1.84 months vs. 3.55 months patients also had mutations in the Lkb1 tumor suppressor, vs. 2.83 months, P ¼ 0.026). These findings are similar to which has been shown to potentiate resistance to selume- findings from a post hoc biomarker analysis of the MISSION tinib/docetaxel combination therapy in a KRas-mutant phase III trial, in which EGFR mutation, but not KRas GEMM (76). It will also be important to understand how mutation, predicted increased overall survival with sorafe- MEK signaling cooperates with the microtubule depolymer- nib as third- or fourth-line monotherapy (72). izing activity of docetaxel and if KRas is necessary for this Together, the findings from this trial show the feasibility functional interaction. of real-time biomarker analysis and adaptive randomiza- The results from these two clinical trials represent an tion, and provides a rationalization for further targeted important step forward in targeting KRas. They show that therapy clinical trials, especially combination regimens, for patients with KRas mutations have a small response to patients with KRas mutations. Of note, the BATTLE-2 trial, targeted therapies. However, KRas mutation status is likely currently ongoing, is testing four treatment arms—erloti- not the only marker involved because therapeutic responses nib, erlotinib þ MK2206 (an Akt inhibitor), selumetinib þ in KRas-mutant patients were not sufficiently robust, and MK2206, and sorafenib—with multiple biomarkers includ- because KRas wild-type patients also had a response. Also ing KRas (73). remaining to be seen are the effects of other gene mutations,

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such as those induced by smoking, on KRas, and how this this process may seem slow and unforgiving, we should be may influence therapy. reminded that paradigm changes take time—the renais- Other ongoing clinical trials aimed at targeting KRas- sance lasted almost three centuries—and that continued mutant NSCLC include a phase II trial of bortezomib studies of the Ras pathway will likely reveal novel therapies (77), a phase Ib/II trial of retaspimycin (an Hsp90 inhib- for this subset of patients, providing personalized medicine itor) in combination with everolimus (78), and a phase II against this most common oncogene. trial of selumetinib þ erlotinib versus selumetinib alone (79; Table 2). Disclosure of Potential Conflicts of Interest No potential conflicts of interest were disclosed. New Directions Grant Support The difficulties of targeting KRas-mutated NSCLC remain This study was supported by the NIH under award number MSTP daunting, but a renaissance of discoveries—novel modes of TG 2T32GM07205 (to N. Vasan) and 5R01CA155196-04, Personalizing inhibition, Ras regulatory proteins, and Ras-dependent NSCLC Therapy: Exploiting KRAS activated pathways (to R. Herbst). targets—is defining a new battery of drugs. In turn, these Received November 19, 2013; revised April 11, 2014; accepted April 15, drugs are informing a new wave of clinical trials. Although 2014; published OnlineFirst June 3, 2014.

References 1. Barbacid M. Ras genes. Annu Rev Biochem 1987;56:779–827. 19. Reiss Y, Goldstein JL, Seabra MC, Casey PJ, Brown MS. Inhibition of 2. Mitin N, Rossman KL, Der CJ. Signaling interplay in Ras superfamily purified p21ras farnesyl: protein transferase by Cys-AAX tetrapep- function. Curr Biol CB 2005;15:R563–74. tides. Cell 1990;62:81–8. 3. Bourne HR, Sanders DA, McCormick F. The GTPase superfamily: a 20. Ashby MN. CaaX converting enzymes. Curr Opin Lipidol 1998;9: conserved switch for diverse cell functions. Nature 1990;348: 99–102. 125–32. 21. Mor A, Philips MR. Compartmentalized Ras/MAPK signaling. Annu 4. Bos JL, Rehmann H, Wittinghofer A. GEFs and GAPs: critical elements Rev Immunol 2006;24:771–800. in the control of small G proteins. Cell 2007;129:865–77. 22. Hancock JF, Paterson H, Marshall CJ. A polybasic domain or palmi- 5. Forbes SA, Bindal N, Bamford S, Cole C, Kok CY, Beare D, et al. toylation is required in addition to the CAAX motif to localize p21ras to COSMIC: mining complete cancer genomes in the Catalogue of the plasma membrane. Cell 1990;63:133–9. Somatic Mutations in Cancer. Nucleic Acids Res 2011;39:D945–50. 23. Rowinsky EK. Lately, it occurs to me what a long, strange trip it's 6. Scheidig AJ, Burmester C, Goody RS. The pre-hydrolysis state of p21 been for the farnesyltransferase inhibitors. J Clin Oncol 2006;24: (ras) in complex with GTP: new insights into the role of water molecules 2981–4. in the GTP hydrolysis reaction of ras-like proteins. Structure 1999; 24. Basso AD, Kirschmeier P, Bishop WR. Lipid posttranslational mod- 7:1311–24. ifications. Farnesyl transferase inhibitors. J Lipid Res 2006;47: 7. Pylayeva-Gupta Y, Grabocka E, Bar-Sagi D. RAS oncogenes: weaving 15–31. a tumorigenic web. Nat Rev Cancer 2011;11:761–74. 25. Kohl NE, Omer CA, Conner MW, Anthony NJ, Davide JP, Desolms SJ, 8. Downward J. Targeting RAS signalling pathways in cancer therapy. et al. Inhibition of farnesyltransferase induces regression of mammary Nat Rev Cancer 2003;3:11–22. and salivary carcinomas in ras transgenic mice. Nat Med 1995;1: 9. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. 792–7. Cell 2011;144:646–74. 26. Van Cutsem E, van de Velde H, Karasek P, Oettle H, Vervenne WL, 10. Karnoub AE, Weinberg RA. Ras oncogenes: split personalities. Nat Szawlowski A, et al. Phase III trial of gemcitabine plus tipifarnib Rev Mol Cell Biol 2008;9:517–31. compared with gemcitabine plus placebo in advanced pancreatic 11. Riely GJ, Marks J, Pao W. KRAS mutations in non–small cell lung cancer. J Clin Oncol 2004;22:1430–8. cancer. Proc Am Thorac Soc 2009;6:201–5. 27. Lobell RB, Liu D, Buser CA, Davide JP, DePuy E, Hamilton K, et al. 12. Seeburg PH, Colby WW, Capon DJ, Goeddel DV, Levinson AD. Preclinical and clinical pharmacodynamic assessment of L-778,123, a Biological properties of human c-Ha-ras1 genes mutated at codon dual inhibitor of farnesyl:protein transferase and geranylgeranyl:pro- 12. Nature 1984;312:71–5. tein transferase type-I. Mol Cancer Ther 2002;1:747–58. 13. Der CJ, Finkel T, Cooper GM. Biological and biochemical properties of 28. Gotlib J. Farnesyltransferase inhibitor therapy in acute myelogenous human rasH genes mutated at codon 61. Cell 1986;44:167–76. leukemia. Curr Hematol Rep 2005;4:77–84. 14. Leone-Kabler S, Wessner LL, McEntee MF, D'Agostino RB Jr, Miller 29. Johnston SRD, Hickish T, Ellis P, Houston S, Kelland L, Dowsett M, MS. Ki-ras mutations are an early event and correlate with tumor stage et al. Phase II study of the efficacy and tolerability of two dosing in transplacentally-induced murine lung tumors. Carcinogenesis regimens of the farnesyl transferase inhibitor, R115777, in advanced 1997;18:1163–8. breast cancer. J Clin Oncol 2003;21:2492–9. 15. Garassino MC, Marabese M, Rusconi P, Rulli E, Martelli O, Farina G, 30. Blum R, Cox AD, Kloog Y. Inhibitors of chronically active ras: potential et al. Different types of K-Ras mutations could affect drug sensitivity for treatment of human malignancies. Recent Patents Anticancer Drug and tumour behaviour in non-small-cell lung cancer. Ann Oncol Discov 2008;3:31–47. 2011;22:235–7. 31. Weisz B, Giehl K, Gana-Weisz M, Egozi Y, Ben-Baruch G, Marciano D, 16. Kim ES, Herbst RS, Wistuba II, Lee JJ, Blumenschein GR Jr, Tsao A, et al. A new functional Ras antagonist inhibits human pancreatic tumor et al. The BATTLE trial: personalizing therapy for lung cancer. Cancer growth in nude mice. Oncogene 1999;18:2579–88. Discov 2011;1:44–53. 32. Riely GJ, Johnson ML, Medina C, Rizvi NA, Miller VA, Kris MG, et al. A 17. Ihle NT, Byers LA, Kim ES, Saintigny P, Lee JJ, Blumenschein GR, et al. phase II trial of Salirasib in patients with lung adenocarcinomas with Effect of KRAS oncogene substitutions on protein behavior: implica- KRAS mutations. J Thorac Oncol 2011;6:1435–7. tions for signaling and clinical outcome. J Natl Cancer Inst 2012;104: 33. Whyte DB, Kirschmeier P, Hockenberry TN, Nunez-Oliva I, James L, 228–39. Catino JJ, et al. K- and N-Ras are geranylgeranylated in cells treated 18. Miller MS, Miller LD. RAS mutations and oncogenesis: not all RAS with farnesyl protein transferase inhibitors. J Biol Chem 1997;272: mutations are created equally. Front Genet 2011;2:100. 14459–64.

3928 Clin Cancer Res; 20(15) August 1, 2014 Clinical Cancer Research

A RAS Renaissance

34. Tsimberidou AM, Chandhasin C, Kurzrock R. Farnesyltransferase 56. Shaw AT, Winslow MM, Magendantz M, Ouyang C, Dowdle J, Sub- inhibitors: where are we now? Expert Opin Investig Drugs 2010;19: ramanian A, et al. Selective killing of K-ras mutant cancer cells by small 1569–80. molecule inducers of oxidative stress. Proc Natl Acad Sci U S A 35. Cepero V, Sierra JR, Corso S, Ghiso E, Casorzo L, Perera T, et al. MET 2011;108:8773–8. and KRAS gene amplification mediates acquired resistance to MET 57. Guo W, Wu S, Wang L, Wei X, Liu X, Wang J, et al. Antitumor activity of a tyrosine kinase inhibitors. Cancer Res 2010;70:7580–90. novel oncrasin analogue is mediated by JNK activation and STAT3 36. Sebti SM, Der CJ. Opinion: Searching for the elusive targets of inhibition. PLoS ONE 2011;6. farnesyltransferase inhibitors. Nat Rev Cancer 2003;3:945–51. 58. Mansfield AS, Fields AP, Jatoi A, Qi Y, Adjei AA, Erlichman C, et al. 37. https://clinicaltrials.gov/ct2/results?term=farnesyltransferase [Inter- Phase I dose escalation study of the protein kinase C iota inhibitor net]. [cited 2014 Jun 16]. Available from: http://www.clinicaltrials. aurothiomalate for advanced non-small cell lung cancer, ovarian gov/. cancer, and pancreatic cancer. Anticancer Drugs 2013;24:1079–83. 38. Bergo MO, Ambroziak P, Gregory C, George A, Otto JC, Kim E, et al. 59. Luo J, Emanuele MJ, Li D, Creighton CJ, Schlabach MR, Westbrook Absence of the CAAX endoprotease Rce1: effects on cell growth and TF, et al. A genome-wide RNAi screen identifies multiple synthetic transformation. Mol Cell Biol 2002;22:171–81. lethal interactions with the Ras oncogene. Cell 2009;137:835–48. 39. Wahlstrom AM, Cutts BA, Karlsson C, Andersson KME, Liu M, Sjogren 60. Barbie DA, Tamayo P, Boehm JS, Kim SY, Moody SE, Dunn IF, et al. A-KM, et al. Rce1 deficiency accelerates the development of K-RAS- Systematic RNA interference reveals that oncogenic KRAS-driven induced myeloproliferative disease. Blood 2007;109:763–8. cancers require TBK1. Nature 2009;462:108–12. 40. Manandhar SP, Hildebrandt ER, Schmidt WK. Small-molecule inhi- 61. Chien Y, Kim S, Bumeister R, Loo Y-M, Kwon SW, Johnson CL, et al. bitors of the Rce1p CaaX protease. J Biomol Screen 2007;12: RalB GTPase-mediated activation of the IkappaB family kinase TBK1 983–93. couples innate immune signaling to tumor cell survival. Cell 2006; 41. Bergo MO, Gavino BJ, Hong C, Beigneux AP, McMahon M, Casey PJ, 127:157–70. et al. Inactivation of Icmt inhibits transformation by oncogenic K-Ras 62. Ou Y-H, Torres M, Ram R, Formstecher E, Roland C, Cheng T, et al. and B-Raf. J Clin Invest 2004;113:539–50. TBK1 directly engages Akt/PKB survival signaling to support onco- 42. Wahlstrom AM, Cutts BA, Liu M, Lindskog A, Karlsson C, Sjogren genic transformation. Mol Cell 2011;41:458–70. A-KM, et al. Inactivating Icmt ameliorates K-RAS-induced myelopro- 63. Sweet-Cordero A, Mukherjee S, Subramanian A, You H, Roix JJ, Ladd- liferative disease. Blood 2008;112:1357–65. Acosta C, et al. An oncogenic KRAS2 expression signature identified 43. Winter-Vann AM, Baron RA, Wong W, dela Cruz J, York JD, Gooden by cross-species gene-expression analysis. Nat Genet 2005;37:48–55. DM, et al. A small-molecule inhibitor of isoprenylcysteine carboxyl 64. Vicent S, Chen R, Sayles LC, Lin C, Walker RG, Gillespie AK, et al. methyltransferase with antitumor activity in cancer cells. Proc Natl Wilms tumor 1 (WT1) regulates KRAS-driven oncogenesis and senes- Acad Sci U S A 2005;102:4336–41. cence in mouse and human models. J Clin Invest 2010;120:3940–52. 44. Winter-Vann AM, Kamen BA, Bergo MO, Young SG, Melnyk S, James 65. Oji Y, Miyoshi S, Maeda H, Hayashi S, Tamaki H, Nakatsuka S-I, et al. SJ, et al. Targeting Ras signaling through inhibition of carboxyl meth- Overexpression of the Wilms' tumor gene WT1 in de novo lung ylation: an unexpected property of methotrexate. Proc Natl Acad Sci cancers. Int J Cancer J Int Cancer 2002;100:297–303. U S A 2003;100:6529–34. 66. Puyol M, Martín A, Dubus P, Mulero F, Pizcueta P, Khan G, et al. A 45. Chandra A, Grecco HE, Pisupati V, Perera D, Cassidy L, Skoulidis F, synthetic lethal interaction between K-Ras oncogenes and Cdk4 et al. The GDI-like solubilizing factor PDEd sustains the spatial orga- unveils a therapeutic strategy for non-small cell lung carcinoma. nization and signalling of Ras family proteins. Nat Cell Biol Cancer Cell 2010;18:63–73. 2012;14:148–58. 67. Kumar MS, Hancock DC, Molina-Arcas M, Steckel M, East P, Die- 46. Zimmermann G, Papke B, Ismail S, Vartak N, Chandra A, Hoffmann M, fenbacher M, et al. The GATA2 transcriptional network is requisite et al. Small molecule inhibition of the KRAS-PDEd interaction impairs for RAS oncogene-driven non-small cell lung cancer. Cell 2012;149: oncogenic KRAS signalling. Nature 2013;497:638–42. 642–55. 47. Gysin S, Salt M, Young A, McCormick F. Therapeutic strategies for 68. Steckel M, Molina-Arcas M, Weigelt B, Marani M, Warne PH, Kuznet- targeting Ras proteins. Genes Cancer 2011;2:359–72. sov H, et al. Determination of synthetic lethal interactions in KRAS 48. Palmioli A, Sacco E, Abraham S, Thomas CJ, Di Domizio A, De Gioia L, oncogene-dependent cancer cells reveals novel therapeutic targeting et al. First experimental identification of Ras-inhibitor binding interface strategies. Cell Res 2012;22:1227–45. using a water-soluble Ras ligand. Bioorg Med Chem Lett 2009;19: 69. Corcoran RB, Cheng KA, Hata AN, Faber AC, Ebi H, Coffee EM, et al. 4217–22. Synthetic lethal interaction of combined BCL-XL and MEK inhibition 49. Rosnizeck IC, Graf T, Spoerner M, Trankle€ J, Filchtinski D, Herrmann C, promotes tumor regressions in KRAS mutant cancer models. Cancer et al. Stabilizing a weak binding state for effectors in the human Cell 2013;23:121–8. ras protein by cyclin complexes. Angew Chem Int Ed Engl 2010;49: 70. Winton T, Livingston R, Johnson D, Rigas J, Johnston M, Butts C, et al. 3830–3. Vinorelbine plus cisplatin vs. observation in resected non-small-cell 50. Patgiri A, Yadav KK, Arora PS, Bar-Sagi D. An orthosteric inhibitor of lung cancer. N Engl J Med 2005;352:2589–97. the Ras-Sos interaction. Nat Chem Biol 2011;7:585–7. 71. Eberhard DA, Johnson BE, Amler LC, Goddard AD, Heldens SL, Herbst 51. Maurer T, Garrenton LS, Oh A, Pitts K, Anderson DJ, Skelton NJ, et al. RS, et al. Mutations in the epidermal growth factor receptor and in Small-molecule ligands bind to a distinct pocket in Ras and inhibit KRAS are predictive and prognostic indicators in patients with non- SOS-mediated nucleotide exchange activity. Proc Natl Acad Sci U S A small-cell lung cancer treated with chemotherapy alone and in com- 2012;109:5299–304. bination with erlotinib. J Clin Oncol 2005;23:5900–9. 52. Sun Q, Burke JP, Phan J, Burns MC, Olejniczak ET, Waterson AG, et al. 72. Mok TSK, Paz-Ares L, Wu Y-L, Novello S, Juhasz E, Aren O, et al. Discovery of small molecules that bind to K-Ras and inhibit Sos- Association between tumor EGFR and KRas mutation status and mediated activation. Angew Chem Int Ed Engl 2012;51:6140–3. clinical outcomes in NSCLC patients randomized to sorafenib plus 53. Ostrem JM, Peters U, Sos ML, Wells JA, Shokat KM. K-Ras(G12C) best supportive care (BSC) or BSC alone: subanalysis of the Phase III inhibitors allosterically control GTP affinity and effector interactions. MISSION Trial. Ann Oncol 2012;23:ixe1–ixe30. Nature 2013;503:548–51. 73. BATTLE-2 program: a biomarker-integrated targeted therapy study - 54. Brough R, Frankum JR, Costa-Cabral S, Lord CJ, Ashworth A. Search- ClinicalTrials.gov [Internet]. [cited 2014 Jun 16]. Available from: http:// ing for synthetic lethality in cancer. Curr Opin Genet Dev 2011;21: www.clinicaltrials.gov/ct2/show/NCT01248247. 34–41. 74. Janne€ PA, Shaw AT, Pereira JR, Jeannin G, Vansteenkiste J, Barrios C, 55. Patel PH, Chadalavada RSV, Chaganti RSK, Motzer RJ. Targeting von et al. Selumetinib plus docetaxel for KRAS-mutant advanced non- Hippel-Lindau pathway in renal cell carcinoma. Clin Cancer Res small-cell lung cancer: a randomised, multicentre, placebo-controlled, 2006;12:7215–20. phase 2 study. Lancet Oncol 2013;14:38–47.

www.aacrjournals.org Clin Cancer Res; 20(15) August 1, 2014 3929

Vasan et al.

75. Hainsworth JD, Cebotaru CL, Kanarev V, Ciuleanu TE, Damyanov D, [cited 2014 Jun 16]. Available from: http://www.clinicaltrials.gov/ct2/ Stella P, et al. A phase II, open-label, randomized study to assess the show/NCT01833143. efficacy and safety of AZD6244 (ARRY-142886) versus pemetrexed in 78. Phase 1b/2 study of retaspimycin HCl (IPI-504) in combination with patients with non-small cell lung cancer who have failed one or two everolimus in KRAS mutant non-small cell lung cancer - ClinicalTrials. prior chemotherapeutic regimens. J Thorac Oncol Off Publ Int Assoc gov [Internet]. [cited 2014 Jun 16]. Available from: http://www. Study Lung Cancer 2010;5:1630–6. clinicaltrials.gov/ct2/show/NCT01427946. 76. Chen Z, Cheng K, Walton Z, Wang Y, Ebi H, Shimamura T, et al. A 79. Randomized phase II study of AZD6244 (MEK inhibitor) with erlotinib in murine lung cancer co-clinical trial identifies genetic modifiers of KRAS wild type advanced NSCLC and a randomized phase II study of therapeutic response. Nature 2012;483:613–7. AZD6244 with erlotinib in mutant KRAS advanced NSCLC - Clinical- 77. Bortezomib in KRAS-mutant non-small cell lung cancer in never Trials.gov [Internet]. [cited 2014 Jun 16]. Available from: http://www. smokers or those with KRAS G12D - ClinicalTrials.gov [Internet]. clinicaltrials.gov/ct2/show/NCT01229150.

3930 Clin Cancer Res; 20(15) August 1, 2014 Clinical Cancer Research

A RAS Renaissance: Emerging Targeted Therapies for KRAS-Mutated Non−Small Cell Lung Cancer

Neil Vasan, Julie L. Boyer and Roy S. Herbst

Clin Cancer Res 2014;20:3921-3930. Published OnlineFirst June 3, 2014.

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