MEK Inhibitors

Dr. Alex A. Adjei from Roswell Park Cancer Institute

Alex A. Adjei, MD, PhD

Professor and Chair, Department of Medicine

Senior Vice President of Clinical Research

The Katherine Anne Gioia Chair in Cancer Medicine

Roswell Park Cancer Institute

Abstract

The RAS-RAF-MEK-extracellular signal—regulated kinase 1 and 2 (ERK1/2) pathway is the most mutated pathway in human cancer. Thus, components of this pathway have been seen as promising targets for cancer therapy. MEK inhibitors have been in clinical evaluation for more than a decade. While low-level clinical activity has been documented with virtually all MEK inhibitors tested in clinical trials, dramatic responses have been few. Several inhibitors are at various stages of clinical evaluation. Singleagent activity has been detected in tumors harboring RAS/RAF mutations such as melanoma, and one inhibitor, trametinib, has been approved by the US Food and Drug Administration (FDA) for the treatment of melanoma. Combinations of MEK inhibitors with cytotoxic chemotherapy or other targeted agents are being investigated.

Introduction

Three well-characterized subfamilies of mitogenactivated protein kinases (MAPKs) control many physiologic processes in humans. Because of its role in cell proliferation and carcinogenesis, the most characterized MAP kinase pathway is the RAS-RAF- MEK-ERK pathway. This is the most frequently dysregulated signal transduction pathway in human cancer. Common aberrations are gain-of-function mutations ofRASandRAFfamily members.

Mutations inKRAShave been found in 90% of pancreatic, in 20% of non-small cell lung cancer (NSCLC), and in up to 50% of colorectal and thyroid cancers,¹while mutations ofBRAFhave been identified in more than 50% of melanoma and in 40% to 60% of papillary thyroid cancers.^2-4Although mitogen- activated protein kinase 1/2 (MEK1/2) is rarely mutated, constitutively active MEK has been found in more than 30% of primary tumor cell lines tested.⁵

The RAS-RAF-MEK-ERK pathway is activated by a wide array of growth factors and cytokines such as epidermal growth factor (EGF), insulin-like growth factor (IGF), and transforming growth factor (TGF), which initially bind to, and activate, transmembrane receptors located on the cell surface. Through sequential interactions with adapter proteins and nucleotide exchange proteins, these growth factors activate RAS through a conversion from the inactive guanosine diphosphate (GDP)-bound form to the active guanosine triphosphate (GTP)-bound form.

Activated RAS recruits RAF kinase to the membrane, where it is activated by multiple phosphorylation events. Activated RAF phosphorylates and activates MEK kinase. MEK kinase in turn phosphorylates and activates ERK kinase. Phosphorylated ERK can translocate to the nucleus, where it phosphorylates and activates various transcription factors.^6-8This leads to altered gene transcription and cellular proliferation.^9-12

Clinical Pearls

• Most MEK inhibitors in clinical trials are allosteric inhibitors.
• Single-agent activity has been seen consistently only in tumors withBRAFmutations andNRASmutations, such as melanoma.
• Mechanism-based toxicities with these agents include ocular toxicities, skin rash, peripheral edema, left ventricular dysfunction, creatine phosphokinase elevation, and central nervous system abnormalities such as confusion and hallucinations.
• Combinations of MEK inhibitors with BRAF inhibitors have demonstrated increased efficacy accompanied with reduced toxicity.
• Trametinib is the first MEK inhibitor to be approved by the FDA for the treatment of melanoma, alone and in combination with the BRAF inhibitor, dabrafenib.
• MEK inhibitors, when used as single agents, show no activity inBRAF-mutant melanoma that has become refractory to BRAF inhibitors and should not be used in this setting.

History of MEK Inhibitors in Clinical Studies

Notably, the RAF family consists of A-RAF, B-RAF, and RAF-1, all sharing RAS as a common upstream activator, and MEK1/2 as principal kinase effectors.¹³MEK1/2 are dual-specificity kinases, catalyzing the phosphorylation of both tyrosine and threonine on ERK1 and ERK2, their only known physiologic substrates.¹⁴In contrast to RAF and MEK1/2, which have narrow substrate specificity, activated ERK1/2 phosphorylates numerous cytoplasmic and nuclear substrates. This process regulates many cellular responses, such as angiogenesis, cell differentiation, embryogenesis, metastasis, metabolism, and apoptosis (Figure 1).^15-22The structure of the MEK1/2 protein makes it ideal for targeting. It possesses a pocket structure adjacent to the adenoside triphosphate (ATP)-binding site that is only conserved in MEK proteins. Upon inhibitor binding, several conformational changes occur and lock the unphosphorylated MEK1/2 into a catalytically inactive state. Since this ATP-noncompetitive mechanism does not exert inhibitory effects through the highly conserved ATP pocket, it potentially avoids undesired, off-target side effects associated with inadvertent inhibition of other protein kinases, and the challenge of competing with millimolar intracellular concentrations of ATP.^23,24Several compounds with highly potent, specific and allosteric inhibitory activity of MEK1/2 have been developed and tested in clinical studies.

Figure 1. Simplified Schematic of MEK Signaling

The first MEK inhibitor, CI-1040, entered phase I clinical trials in 2000. Since then, only one MEK inhibitor, trametinib, has been approved for clinical use. This is because these agents have failed to demonstrate significant clinical activity in most tumor types. One potential mechanism underlying this modest activity has been elucidated preclinically. An autoregulatory negative feedback loop between ERK and RAF that mediates sensitivity to MEK inhibitors has been described. Activated ERK releases tonic inhibition of RAF kinases, thereby leading to activated RAF, which activates anti-apoptotic downstream RAF targets, thus abrogating the cytotoxic activity of MEK inhibitors (Figure 2). Tumors harboringBRAF^V600Emutations lack this negative feedback loop and are sensitive to MEK inhibitors.²⁵These data predicted that a combination of MEK inhibitors and RAF inhibitors would be synergistic. This prediction has been confirmed in the clinic, as described in the trametinib section later on in this article.

Class Toxicities of MEK Inhibitors

Since 2000, more than 10 MEK inhibitors have entered clinical trials, providing a large body of data on mechanism-based toxicities of these compounds. Some toxicities, such as fatigue, diarrhea, and skin rash, are common to many small-molecule kinase inhibitors. Others are relatively unique to MEK inhibitors, and include ocular toxicities manifest as blurred vision and loss of visual acuity; retinal vein occlusions²⁶; and the most common toxicity, central serous retinopathy²⁷(Figure 3). This potentially serious toxicity involves subretinal fluid accumulation, which generally subsides after drug interruption followed by dose reduction. Peripheral edema, particularly periorbital edema, occurs as dose elevation of creatine phosphokinase without any associated troponin abnormalities, evidence of rhabdomyolysis, or any underlying pathology.

Selumetinib (AZD6244, ARRY-142886)

Rare cases of left ventricular dysfunction have been reported. Also, central nervous system (CNS) effects have occurred, such as hallucinations and confusion (presumably with the subset of agents with good CNS penetration).Table 1illustrates selected MEK inhibitors that are in clinical trials. Selumetinib is the MEK inhibitor with the largest amount of published data. Trametinib has received FDA approval alone and in combination with dabrafenib. These two compounds are discussed in the sections that follow. A brief description of other selected MEK inhibitors is also provided.Selumetinib, a second-generation MEK inhibitor, was introduced in the clinic together with PD- 0325901 after the development of CI-1040 was discontinued because of limited potency and poor pharmacology.²⁴In its initial phase I study, skin rash was the most frequent and dose-limiting toxicity (DLT). Inhibition of ERK phosphorylation was established as a pharmacodynamic biomarker. Prolonged disease stabilization was achieved in one medullary thyroid cancer patient and in one patient with uveal melanoma (out of 34 evaluable patients).²⁸A solid oral capsule formulation of this compound was subsequently developed with improved pharmacologic properties. A prolonged complete response (CR) in a patient with melanoma bearing aBRAF^V600Emutation was observed with this formulation in a phase I study.²⁹

Figure 2. Feedback inhibition of ERK on RAF is abrogated by MEK inhibitors, leading to activation of MEK independent downstream RAF targets and resistance to apoptosis. This may explain the lack of activity of single-agent MEK inhibitors in a number of tumors. In addition, this figure may explain the synergy between MEK inhibitors and BRAF inhibitors.

Single-agent selumetinib has been evaluated in multiple phase II studies in a variety of solid tumors as well as in hematologic malignancies.^30-34Treatment with single-agent selumetinib in 28 patients with metastatic biliary cancers yielded 3 objective responses (ORs) (12%) and 14 stable diseases (SDs) lasting for more than 4 months, whereas no significant antitumor activity was observed in either papillary thyroid carcinoma or hepatocellular carcinoma. In aBRAF^V600E/K-mutated melanoma study, no antitumor activity was observed in the 10 patients with high phosphorylated AKT (pAKT) levels, while 3 of 5 patients with low pAKT levels achieved tumor regression, suggesting a potential role of PI3/AKT activation in MEK inhibitor resistance. In a phase II study, 47 patients with relapsed/refractory acute myeloid leukemia (AML) or who were 60 years or older with untreated AML were stratified byFLT3-internal tandem duplication (ITD) mutation status and were treated with selumetinib. In theFLT3wildtype cohort, 6 of 36 patients (17%) had a response (1 partial response, 3 minor responses, 2 unconfirmed minor responses [uMR]). No patient withFLT3-ITD responded.NRASandKRASmutations were detected in 7% and 2% of patients, respectively. The sole patient withKRASmutation had uMR with hematologic improvement in platelets. Baseline p-ERK activation was observed in 85% of patients analyzed, but this activation did not correlate with a response. A single-nucleotide polymorphism (SNP), rs3733542 in exon 18 of theKITgene, was detected in a significantly higher number of patients with response/ stable disease compared with nonresponders (60% vs 23%;P= .027).³⁴

Selumetinib has also been evaluated in combination with other anticancer agents. In a phase I study combining selumetinib with the AKT inhibitor MK- 2206, DLTs were rash, stomatitis, grade 2 detached retinal pigment epithelium, diarrhea, grade 4 lipase elevation, bilateral posterior, grade 1 subcapsular cataracts, and fatigue. In this 51-patient trial, 1 patient withKRAS-mutated NSCLC and 1 patient withKRAS-mutated ovarian cancer achieved durable confirmed partial responses (PR), and 1 unconfirmed PR was seen in a patient with pancreatic cancer.³⁵In a phase I study assessing selumetinib and cetuximab in solid tumors andKRAS-mutated colorectal cancer (CRC), the most common toxicities reported were acneiform rash, fatigue, nausea/vomiting, and diarrhea, while the DLT was grade 4 hypomagnesemia.³⁶Among 13 evaluable patients treated in the dose-escalation cohort, 2 PRs were observed in patients with CRC, and SD was achieved in 1 patient with tonsillar squamous cell carcinoma, 1 patient with NSCLC, and 2 patients with CRC. Results in theKRAS-mutated CRC expansion cohort have not been reported.

Figure 3. Central serous retinopathy from a MEK inhibitor. Retinal picture and OCT stain pretreatment (A, B). Fluid collection behind retina after 2 weeks of treatment on OCT stain (C, D).

Randomized phase II studies comparing selumetinib versus temozolomide in chemotherapynaïve melanoma, selumetinib versus pemetrexed in NSCLC beyond first/second-line therapies, selumetinib versus capecitabine in pancreatic cancer after failing gemcitabine, and selumetinib versus capecitabine in CRC beyond first/second-line therapies did not demonstrate superiority, even though antitumor activity as single agent was observed in each study.^37-40

In previously treated NSCLC withKRASmutation, the addition of selumetinib to docetaxel did not yield a statistically significantly improved median overall survival (OS) (9.4 months vs 5.2 months; hazard ratio [HR] 0.80; 80% confidence interval [CI]: 0.56- 1.14; 1-sidedP= .21), although improvement in median progression-free survival (PFS) (5.3 months vs 2.1 months; HR = 0.58; 80% CI: 0.42-0.79; 1-sidedP= .014) and OR rate (37% vs 0%; P <.0001) were seen in this study. A higher incidence of myelosuppression and fatigue was found in the selumetinib group.40 Based on the high OR rate of this combination, a phase III study has been started. Selumetinib plus dacarbazine versus placebo plus dacarbazine was compared in patients withBRAF-mutant melanoma as first-line treatment in a phase II study. Although significantly improved PFS was observed with the addition of selumetinib, no OS benefit was demonstrated.⁴¹

Finally, the effect of selumetinib on radioiodine sensitivity in radioiodine-resistant differentiated thyroid cancers was tested in a 24-patient pilot study. A 4-week treatment of selumetinib restored sensitivity to radioiodine that exceeded the threshold for radioiodine treatment in 8 patients, allowing administration of therapeutic radioiodine; PR and SD were achieved in 5 and 3 patients, respectively.

Trametinib (GSK1120212,JTP-74057)

In this small study, selumetinib appeared to convert a higher percentage ofNRAS-mutant tumors to radioiodine sensitivity thanBRAF-mutated tumors.⁴²Phase I and II trials combining selumetinib with vandetanib, cixutumumab, gemcitabine, and irinotecan are under way or have been completed.^40,43Trametinib, a third-generation MEK inhibitor, was evaluated in a 206-patient, phase I study.²⁷The most common adverse events were rash and diarrhea, and DLTs were rash, diarrhea, and central serous retinopathy. The effective half-life of trametinib was found to be about 4 days. Although a dose level of 3 mg/day was determined to be the maximum tolerated dose (MTD), because of poor tolerance beyond the first cycle of treatment, the recommended phase II dose was 2 mg/day. Twenty-one (10%) ORs were noted at all dose levels, with the most sensitive population beingBRAF-mutant melanoma.²⁷Subanalysis of participants withBRAF-mutant melanoma revealed a 33% response rate among 30 patients who were BRAF inhibitor-na&iuml;ve. This result was further confirmed in a phase II study, in which a 25% OR rate was achieved in patients who were BRAF inhibitor- na&iuml;ve, while only minimal clinical activity was observed in patients with BRAF inhibitor-resistant disease.⁴⁴

Table 1. MEK Inhibitors in Clinical Development

Drug

Manufacturer

Stage of Clinical Development

AZD8330 (ARRY- 424704)

AstraZeneca, Array Biopharma

Phase II

Rafametinib (BAY 86- 9766 RDEA119)

Bayer, Ardea Bioscience

Phase II

CI-1040 (PD 184352)

Pfizer

Discontinued

Cobimetinib (GDC-0973; XL-518, RG7421)

Genentech

Phase III

E6201

Eisai Inc.

Phase II

MEK162 (ARRY-438162)

Array Biopharma, Norvartis

Phase III

PD-0325901

Pfizer

Phase II

Pimasertib (AS703026, MSC1936369B)

EMD Serono

Phase II

RO4987655 (CH4987655)

Hoffmann-La Roche

Phase II

RO5126766

Hoffmann-La Roche

Phase II

Selumetinib (AZD6244, ARRY-142886)

AstraZeneca

Phase III

TAK-733

Takeda

Phase I

Trametinib (GSK1120212)

GlaxoSmithKline

FDA approved

WX-554

Wilex

Phase I

Trametinib was subsequently advanced to a phase III study. Patients withBRAF^V600E- orBRAF^V600K-mutant melanoma who were not previously treated with a BRAF or MEK inhibitor or with ipilimumab were randomized to receive either trametinib or chemotherapy (dacarbazine or paclitaxel). The trametinib arm had a median PFS of 4.8 months, compared with 1.5 months in the chemotherapy group. Median OS has not been reached.⁴⁵However, based on these results, the FDA granted approval of trametinib as a single agent for the treatment ofBRAF^V600EorBRAF^V600Kmutation—positive unresectable or metastatic melanoma. The preclinical data suggest that the efficacy of MEK inhibitors may be enhanced by combination with a RAF inhibitor.²⁵In addition, activation of the MAPK pathway has been postulated as a potential resistance mechanism to RAF inhibitors in melanoma. Simultaneous inhibition of RAF and MEK is therefore a rational approach to overcoming resistance to BRAF inhibitors. Trametinib was combined with dabrafenib, a BRAF inhibitor, in a phase I/II trial to determine tolerability, and to compare single-agent dabrafenib to the combination. Cutaneous squamous cell carcinoma, a BRAF inhibitor-associated adverse event presumed to be due to paradoxical MAPK pathway activation, was lower in the combination arm. A significant improvement in PFS was observed in the combination arm (HR, 0.39; 95% CI: 0.25-0.62;P<.001), indicating the potential of MAPK inhibition to delay resistance to BRAF inhibition. This study was further extended into a phase III trial (COMBI-D) in 162 patients with histologically confirmed stage IIIC or IV melanoma with aBRAF^V600E(85%) orBRAF^V600K(15%) mutation. Only one prior chemotherapy regimen and/or interleukin-2 therapy was permitted. Patients with prior exposure to BRAF inhibitors or MEK inhibitors were ineligible. Fifty-four patients with performance status (PS) 0 to 1 each were assigned to 3 arms; trametinib (1 mg in one arm, 2 mg in the other arm) orally, once daily in combination with dabrafenib 150 mg orally, twice daily, with single-agent dabrafenib as the control arm. In this study, 67% of patients had M1c disease, and 81% had not received prior anticancer therapy for unresectable or metastatic disease. Objective response rates and response durations were 76% (95% CI: 62-87) and 10.5 months (95% CI: 7-15), respectively, in the trametinib 2-mgplus- dabrafenib combination arm and 54% (95% CI: 40-67) and 5.6 months (95% CI: 5-7), respectively, in the single-agent dabrafenib arm. Objective response rates were similar in subgroups defined byBRAF^V600mutation subtype,BRAF^V600EandBRAFV600K.The trial&rsquo;s primary safety endpoint, the incidence of cutaneous squamous cell carcinoma (including squamous cell carcinomas of the skin and keratoacanthomas), was 7% (95% CI: 2-18) in the trametinib 2 mg plus dabrafenib combination arm compared with 19% (95% CI: 9-32) in the single-agent dabrafenib arm. The most frequent (&ge;20% incidence) adverse reactions from trametinib in combination with dabrafenib were fever, chills, fatigue, rash, nausea, vomiting, diarrhea, abdominal pain, peripheral edema, cough, headache, arthralgia, night sweats, decreased appetite, constipation, and myalgia. The most frequent grades 3 and 4 adverse events (at least 5% incidence) were acute renal failure, fever, hemorrhage, and back pain. Serious but less common adverse drug reactions occurring in patients taking trametinib in combination with dabrafenib were hemorrhage, venous thromboembolism, new primary malignancy, febrile reactions, cardiomyopathy, skin toxicity, and eye disorders such as retinal pigmented epithelial detachments. Based on these data, the FDA granted accelerated approval to trametinib in January 2014 for use in combination with dabrafenib in the treatment of patients with unresectable or metastatic melanoma with aBRAF^V600Eor aBRAF^V600Kmutation.⁴⁶The trametinib/dabrafenib combination is also being evaluated in the adjuvant setting after surgical resection (COMBI-AD), and in a phase III study comparing this combination with another RAF inhibitor, vemurafenib (COMBI-v).

It must be noted, however, that trametinib and other MEK inhibitors when used as a single agent have no activity againstBRAF-mutant melanoma that has progressed on a BRAF inhibitor such as vemurafenib. In fact, anecdotal data suggest that such treatment may lead to a disease flare and a rapid demise of patients, and should be avoided.

Cobimetinib (GDC-0973, XL-518, RG7421)

Trametinib combinations have also been evaluated in several other tumor types. In a randomized, placebo-controlled study, the addition of trametinib to gemcitabine failed to improve efficacy parameters in 160 patients with metastatic pancreatic cancer irrespective of theirKRASmutation status.⁴⁷Other phase I/Ib studies evaluating combinations of trametinib with various targeted therapeutic and conventional chemotherapy agents are under study, such as everolimus, pazopanib, dabrafenib, the AKT inhibitor GSK2141795, the PI3K inhibitor BKM120, erlotinib, 5-fluorouracil, and radiation therapy.^48-51A second third-generation MEK inhibitor, cobimetinib, is also undergoing phase III clinical trials. Cobimetinib was well tolerated in phase I trials, but there were no ORs.⁵²Cobimetinib was further evaluated in a phase Ib study in combination with the PI3-kinase inhibitor, GDC-0941, in advanced solid tumors. The most common adverse events of this combination were diarrhea, fatigue, nausea, and rash. Among the 46 evaluable patients, PRs were observed in 1 patient withBRAF-mutated melanoma, 1 withBRAF-mutated pancreatic cancer, and 1 withKRAS-mutated endometrial cancer.^53,54Cobimetinib was also combined with vemurafenib inBRAF^V600E-mutated melanoma in another phase Ib study. Tumor shrinkage was observed in all 8 evaluable patients who were vemurafenib na&iuml;ve. Notably, of the 44 subjects, only 1 developed cutaneous squamous cell carcinoma.⁵⁴On the basis of these results, a phase III study comparing this combination with vemurafenib has been initiated. Phase II studies combining cobimetinib with other targeted agents, including GDC-0941 and the AKT inhibitor GDC-0068, are ongoing.

Rafametinib (BAY 86-9766, RDEA119)

Pimasertib (AS703026, MSC1936369B)

Other MEK Inhibitors

Conclusions

Other MEK inhibitors are in phase Ib, phase I/II, and phase II clinical trials and are briefly described in the sections that follow.In a phase I study of rafametinib, 1 patient with CRC of a total of 53 evaluable patients achieved a PR.⁵⁵Based on preclinical data⁵⁶and results of a phase I study, rafametinib was combined with sorafenib as first-line systemic treatment for patients with hepatocellular carcinoma in a phase II study. Among 70 subjects, 3 had confirmed PR and 25 had prolonged (>4 months) SD. However, there were 4 grade 5—related adverse events reported, and almost all patients required dose modifications because of adverse events.⁵⁷A phase I/II study evaluating rafametinib in combination with gemcitabine in patients with metastatic pancreatic cancer is ongoing.⁵⁸Phase I studies of pimasertib found no ORs in a total of 180 patients. However, 5 patients with eitherBRAForNRASmutations had confirmed tumor shrinkage, but not enough to meet OR criteria.^59,60A combination study with 5-fluorouracil, leucovorin, and irinotecan (FOLFIRI) as second-line treatment inKRAS-mutated metastatic CRC was discontinued because of toxicity.⁶¹Studies combining pimasertib with a number of agents are ongoing. These include the PI3K/mTOR inhibitor SAR245409 (phase Ib) and temsirolimus (phase I). There are also phase II studies comparing pimasertib and dacarbazine in NRASmutated melanoma, pimasertib combined with gemcitabine versus gemcitabine alone in pancreatic adenocarcinoma, and a phase II study of pimasertib in patients with advanced hematologic malignancies.Phase I studies evaluating AZD8330, WX-554, E6201, RO4987655, TAK-733, and RO5126766, either alone or in multiple combinations are ongoing or have been recently completed.^62-67A randomized, phase III study with MEK162 compared with dacarbazine in patients with advanced unresectable or metastaticNRASmutation-positive melanoma is currently enrolling patients.⁶⁸A phase III, randomized 3-arm open-label study of MEK162 plus LGX818 and LGX818 monotherapy compared with vemurafenib in patients with unresectable or metastaticBRAF^V600-mutant melaoma is also currently enrolling patients.⁶⁹Despite the prevalence of aberrant MEK signaling in human cancers, highly potent and specific MEK inhibitors have failed to demonstrate striking singleagent activity in most solid tumors and hematologic malignancies. Apart fromBRAF-mutant and NRASmutant melanoma, clinical activity has been reported in biliary cancers, serous ovarian cancer, and uveal melanoma. While these agents are not active inBRAF-mutant tumors that develop resistance to BRAF inhibitors, activity has been demonstrated in combination with BRAF inhibitors. This combination is superior to single-agent BRAF inhibitors in terms of response rate and duration of response. Combinations with other targeted agents may also improve efficacy and overcome resistance. However, toxicity of a number of these combinations (AKT inhibitors, PI3-kinase inhibitors) has caused problems. Combinations with classical cytotoxic agents may also be efficacious, and results of the ongoing phase III trial of the combination of selumetinib with docetaxel are awaited with interest.

References

1. Jose DG, De Kretser T. Oncogenes of human tumour cells. A review of advances in virology and molecular biology of cancer.Australas Radiol. 1984;28(1):43-50.
2. Cohen Y, Xing M, Mambo E, et al. BRAF mutation in papillary thyroid carcinoma.J Natl Cancer Inst. 2003;95(8):625-627.
3. Davies H, Bignell GR, Cox C, et al. Mutations of the BRAF gene in human cancer.Nature. 2002;417(6892):949-954.
4. Xu X, Quiros RM, Gattuso P, et al. High prevalence of BRAF gene mutation in papillary thyroid carcinomas and thyroid tumor cell lines.Cancer Res. 2003.63(15):4561-4567.
5. Hoshino R, Chatani Y, Yamori T, et al. Constitutive activation of the 41- /43-kDa mitogen-activated protein kinase signaling pathway in human tumors.Oncogene. 1999;18(3):813-822.
6. Weinstein-Oppenheimer CR, Blalock WL, Steelman LS, et al. The Raf signal transduction cascade as a target for chemotherapeutic intervention in growth factor-responsive tumors.Pharmacol Ther. 2000;88(3): 229-279.
7. Stefanovsky VY, Pelletier G, Hannan R, et al. An immediate response of ribosomal transcription to growth factor stimulation in mammals is mediated by ERK phosphorylation of UBF.Mol Cell. 2001;8(5):1063-1073.
8. Cowley S, Paterson H, Kemp P, Marshall CJ. Activation of MAP kinase kinase is necessary and sufficient for PC12 differentiation and for transformation of NIH 3T3 cells.Cell. 1994;77(6):841-852.
9. Wasylyk B, Hagman J, Gutierrez-Hartmann A. Ets transcription factors: nuclear effectors of the Ras-MAP-kinase signaling pathway.Trends Biochem Sci. 1998;23(6):213-216.
10. Kolch W, Heidecker G, Lloyd P, Rapp UR. Raf-1 protein kinase is required for growth of induced NIH/3T3 cells.Nature. 1991;349(6308):426-428.
11. Moodie SA, Willumsen BM, Weber MJ, Wolfman A. Complexes of Ras. GTP with Raf-1 and mitogen-activated protein kinase kinase.Science. 1993;260(5114):1658-1661.
12. Marshall CJ. Cell signalling. Raf gets it together.Nature. 1996;383 (6596):127-128.
13. Hagemann C, Rapp UR. Isotype-specific functions of Raf kinases.Exp Cell Res. 1999;253(1):34-46.
14. Roskoski R Jr. ERK1/2 MAP kinases: structure, function, and regulation.Pharmacol Res. 2012;66(2):105-143.
15. Meloche S, Pouyssegur J. The ERK1/2 mitogen-activated protein kinase pathway as a master regulator of the G1- to S-phase transition.Oncogene. 2007;26(22):3227-3239.
16. Shaul YD, Seger R. The MEK/ERK cascade: from signaling specificity to diverse functions.Biochim Biophys Acta. 2007;1773(8):1213-1226.
17. Lewis TS, Shapiro PS, Ahn NG. Signal transduction through MAP kinase cascades.Adv Cancer Res. 1998;74:49-139.
18. Alberola-Ila J, Forbush KA, Seger R, Krebs EG, Perlmutter RM. Selective requirement for MAP kinase activation in thymocyte differentiation.Nature. 1995;373(6515):620-623.
19. Johnson GL, Vaillancourt RR. Sequential protein kinase reactions controlling cell growth and differentiation.Curr Opin Cell Biol. 1994;6(2): 230-238.
20. D&rsquo;Angelo G, Struman I, Martial J, Weiner RI. Activation of mitogenactivated protein kinases by vascular endothelial growth factor and basic fibroblast growth factor in capillary endothelial cells is inhibited by the antiangiogenic factor 16-kDa N-terminal fragment of prolactin.Proc Natl Acad Sci U S A. 1995;92(14):6374-6378.
21. Na J, Furue MK, Andrews PW. Inhibition of ERK1/2 prevents neural and mesendodermal differentiation and promotes human embryonic stem cell self-renewal.Stem Cell Res. 2010;5(2):157-169.
22. Raman M, Chen W, Cobb MH. Differential regulation and properties of MAPKs.Oncogene. 2007;26(22):3100-3112.
23. Ohren JF, Chen H, Pavlovsky A, et al. Structures of human MAP kinase kinase 1 (MEK1) and MEK2 describe novel noncompetitive kinase inhibition.Nat Struct Mol Biol. 2004;11(12):1192-1197.
24. Friday BB, Adjei AA. Advances in targeting the Ras/Raf/MEK/Erk mitogen-activated protein kinase cascade with MEK inhibitors for cancer therapy.Clin Cancer Res. 2008;14(2):342-346.
25. Friday BB,Yu C, Dy GK, et al. BRAF V600E disrupts AZD6244-induced abrogation of negative feedback pathways between extracellular signalregulated kinase and Raf proteins.Cancer Res. 2008;68(15):6145-6153.
26. LoRusso PM, Adjei AA, Varterasian M, et al. Phase I pharmacokinetic and pharmacodynamic study of the oral MAPK/ERK kinase inhibitor PD- 0325901 in patients with advanced cancers.Clin Cancer Res. 2010; 16(6):1924-1937.
27. Falchook GS, Lewis KD, Infante JR, et al. Activity of the oral MEK inhibitor trametinib in patients with advanced melanoma: a phase 1 doseescalation trial.Lancet Oncol. 2012;13(8):782-789.
28. Adjei AA, Cohen RB, Frankin W, et al. Phase I pharmacokinetic and pharmacodynamic study of the oral, small-molecule mitogen-activated protein kinase kinase 1/2 inhibitor AZD6244 (ARRY-142886) in patients with advanced cancers.J Clin Oncol. 2008;26(13):2139-2146.
29. Banerji U, Camidge DR, Verheul HM, et al. The first-in-human study of the hydrogen sulfate (Hyd-sulfate) capsule of the MEK1/2 inhibitor AZD6244 (ARRY-142886): a phase I open-label multicenter trial in patients with advanced cancer.Clin Cancer Res. 2010;16(5):1613-1623.
30. Catalanotti F, Solit DB, Pulitzer MP, et al. Phase II trial of MEK inhibitor selumetinib (AZD6244) in patients with BRAFV600E/K-mutated melanoma.Clin Cancer Res. 2013;19(8):2257-2264.
31. Hayes DN, Lucas AS, Tanvetyanon T, et al. Phase II efficacy and pharmacogenomic study of selumetinib (AZD6244; ARRY-142886) in iodine-131 refractory papillary thyroid carcinoma with or without follicular elements.Clin Cancer Res. 2012;18(7):2056-2065.
32. O&rsquo;Neil BH, Goff LW, Kauh JS, et al. Phase II study of the mitogenactivated protein kinase 1/2 inhibitor selumetinib in patients with advanced hepatocellular carcinoma.J Clin Oncol. 2011;29(17):2350- 2356.
33. Bekaii-Saab T, Phelps MA, Li X, et al. Multi-institutional phase II study of selumetinib in patients with metastatic biliary cancers. J Clin Oncol. 2011;29(17):2357-2363.
34. Jain N, Curran E, Iyenger NM, Diaz-Flores E, Kunnavakkam R. Phase II study of the oral MEK inhibitor selumetinib in advanced acute myelogenous leukemia: a University of Chicago phase II consortium trial.Clin Cancer Res. 2014;20:490-498.
35. Hayat K, Khan LY, Mezynski J, et al. A phase I dose escalation study of oral MK-2206 (allosteric Akt inhibitor) with oral selumetinib (AZD6244; ARRY-142866) (MEK 1/2 inhibitor) in patients with advanced or metastatic solid tumors.J Clin Oncol. 2012;28(15s):e13599.
36. Deming DA, Sam WRS, Lubner J, et al. A phase I study of selumetinib (AZD6244/ARRY-142866) in combination with cetuximab (cet) in refractory solid tumors and KRAS mutant colorectal cancer (CRC).J Clin Oncol. 2012;30(suppl):Abstract 3103.
37. Kirkwood JM, Bastholt L, Robert C, et al. Phase II, open-label, randomized trial of the MEK1/2 inhibitor selumetinib as monotherapy versus temozolomide in patients with advanced melanoma.Clin Cancer Res. 2012;18(2):555-567.
38. Hainsworth JD, Cebotaru CL, Kanarev V, et al. A phase II, open-label, randomized study to assess the efficacy and safety of AZD6244 (ARRY- 142886) versus pemetrexed in patients with non-small cell lung cancer who have failed one or two prior chemotherapeutic regimens.J Thorac Oncol. 2010;5(10):1630-1636.
39. Bodoky G, Timcheva C, Spigel DR, et al. A phase II open-label randomized study to assess the efficacy and safety of selumetinib (AZD6244 [ARRY-142886]) versus capecitabine in patients with advanced or metastatic pancreatic cancer who have failed first-line gemcitabine therapy.Invest New Drugs. 2012;30(3):1216-1223. Published correction appears in:Invest New Drugs. 2012;30(3):1272-1273.
40. Bennouna J, Lang I, Valladares-Ayerbes M, et al. A phase II, open-label, randomised study to assess the efficacy and safety of the MEK1/2 inhibitor AZD6244 (ARRY-142886) versus capecitabine monotherapy in patients with colorectal cancer who have failed one or two prior chemotherapeutic regimens.Invest New Drugs.2011;29(5):1021-1028.
41. Robert C, Dummer R, Gutzmer R, et al. Selumetinib plus dacarbazine versus placebo plus dacarbazine as first-line treatment for BRAF-mutant metastatic melanoma: a phase 2 double-blind randomised study.Lancet Oncol. 2013;14(8):733-740.
42. Ho AL, Grewal RK, Leboeuf R, et al. Selumetinib-enhanced radioiodine uptake in advanced thyroid cancer.N Engl J Med. 2013;368(7):623-632.
43. ClinicalTrial.gov. 2013 [cited November 24, 2013]. http://clinicaltrials .gov/ct2/results?term=selumetinib&Search=Search.
44. Kim KB, Kefford R, Pavlick AC, et al. Phase II study of the MEK1/MEK2 inhibitor trametinib in patients with metastatic BRAF-mutant cutaneous melanoma previously treated with or without a BRAF inhibitor.J Clin Oncol. 2013;31:482-489.
45. Flaherty KT, Robert C, Hersey P, et al. Improved survival with MEK inhibition in BRAF-mutated melanoma. N Engl J Med. 2012;367:107-114.
46. National Cancer Institute. http://www.NCI.gov. Accessed on January 24, 2014.
47. Jeffrey R, Infante BGS, Joon Oh Park, et al. A randomized, double-blind, placebo-controlled trial of trametinib, a MEK inhibitor, in combination with gemcitabine for patients with untreated metastatic adenocarcinoma of the pancreas.J Clin Oncol. 30;2012-2013;(suppl 34):Abstract 291.
48. Becerra CR, Garbo LE, Gordon MS, et al. A five-arm, open-label, phase I/ lb study to assess safety and tolerability of the oral MEK1/MEK2 inhibitor trametinib (GSK1120212) in combination with chemotherapy or erlotinib in patients with advanced solid tumors.J Clin Oncol. 2012;28:15s, 30(suppl):Abstract 3023.
49. Ahmed SR, Ball DW, Rudek MA, et al. A phase I study determining the safety and tolerability of combination therapy with pazopanib, a VEGFR/ PDGFR/raf inhibitor, and GSK1120212, a MEK inhibitor, in advanced solid tumors enriched with patients with advanced differentiated thyroid cancer.J Clin Oncol. 2012;30(suppl):Abstract TPS3117.
50. Jeffrey R, Infante AP, Jones SF, et al. A phase IB study of the MEK inhibitor GSK1120212 combined with everolimus in patients with solid tumors: Interim results.Molecular Cancer Therapeutics. 2011;10(11 suppl 1).
51. Kurzrock R, Rosenstein L, Fu S, et al. Phase I dose-escalation of the oral MEK1/2 inhibitor GSK11202125 (GSK212) dosed in combination with the oral AKT inhibitor GSK2141795 (GSK795).J Clin Oncol. 2011;29(suppl):Abstract 3085.
52. Shapiro G, Kwak JM, Cleary L, et al. Clinical combination of the MEK inhibitor GDC-0973 and the PI3K inhibitor GDC-0941: a first-in-human phase Ib study testing daily and intermittent dosing schedules in patients with advanced solid tumors.J Clin Oncol. 2011;29(suppl):Abstract 3005.
53. LoRusso P, Pandya SS, Kwak EL, et al. A first-in-human phase Ib study to evaluate the MEK inhibitor GDC-0973, combined with the pan-PI3K inhibitor GDC-0941, in patients with advanced solid tumors.J Clin Oncol. 2012;28:15s, 30(suppl ):Abstract 2566.
54. Gonzalez R, Daud A, Pavlick TF, et al. Phase Ib study of vemurafenib in combination with the MEK inhibitor, GDC-0973, in patients (pts) with unresectable or metastatic BRAFv600 mutated melanoma (Brim7). ESMO. 2012—Melanoma and other skin tumors.
55. Weekes CD, Von Hoff DD, Adjei AA, et al. Multicenter phase I trial of the mitogen-activated protein kinase 1/2 inhibitor BAY 86-9766 in patients with advanced cancer.Clin Cancer Res. 2013;19(5):1232-1243.
56. Schmieder R, Puehler F, Neuhaus R, et al. Allosteric MEK1/2 inhibitor refametinib (BAY 86-9766) in combination with sorafenib exhibits antitumor activity in preclinical murine and rat models of hepatocellular carcinoma.Neoplasia. 2013;10:1161-1171.
57. Lim HY, Tak WY, Heo J, et al. A phase II trial of MEK inhibitor BAY 86- 9766 in combination with sorafenib as first-line systemic treatment for patients with unresectable hepatocellular carcinoma (HCC).J Clin Oncol. 2012;(suppl):Abstract 4103.
58. Van Laethem JL, Martens UM, Jassem J, et al. A phase I/II study of the MEK inhibitor BAY 86-9766 (BAY) in combination with gemcitabine (GEM) in patients with nonresectable, locally advanced or metastatic pancreatic cancer (PC): phase I dose-escalation results.J Clin Oncol. 2012;30(suppl):Abstract 4050.
59. Delord J, Awada A, Taamma A, et al. First-in-human phase I safety, pharmacokinetic (PK), and pharmacodynamic (PD) analysis of the oral MEK-inhibitor AS703026 (two regimens [R]) in patients (pts) with advanced solid tumors.J Clin Oncol. 2010;28:15s(suppl):Abstract 2504.
60. Houede N, Awada A, Raymond E, et al. Safety and evidence of activity of MSC1936369, an oral MEK1/2 inhibitor, in patients with advanced malignancies.J Clin Oncol. 2011;29(suppl):Abstract 3019.
61. Macarulla T, Cervantes A, Rosell&oacute; S, et al. Phase I/Ii study of folfiri plus the Mek1/2 inhibitor pimasertib (MSC1936369b) as second-line treatment for KRAS mutated metastatic colorectal cancer.Ann Oncol. 2012;23(suppl 4):iv19-iv30.
62. Cohen RB, Aamdal S, Nyakas M, et al. A phase I dose-finding, safety and tolerability study of AZD8330 in patients with advanced malignancies.Eur J Cancer. 2013;49(7):1521-1529.
63. Leijen S, Middleton MR, Tresca P, et al. Phase I dose-escalation study of the safety, pharmacokinetics, and pharmacodynamics of the MEK inhibitor RO4987655 (CH4987655) in patients with advanced solid tumors.Clin Cancer Res. 2012;18(17):4794-4805.
64. Martinez-Garcia M, Banerji U, Albanell J, et al. First-in-human, phase I dose-escalation study of the safety, pharmacokinetics, and pharmacodynamics of RO5126766, a first-in-class dual MEK/RAF inhibitor in patients with solid tumors.Clin Cancer Res. 2012;18(17): 4806-4819.
65. Borad MJ, Ramanathan RK, Northfelt DW, et al. Phase I dose-escalation study of E6201, a MEK-1 inhibitor, in advanced solid tumors.J Clin Oncol. 2010;28(15s):Abstract 2505.
66. Mala C, Haindl E, Buergle M, et al. A phase I, first-in-human single ascending dose study of the MEK inhibitor WX-554 given to healthy male subjects.J Clin Oncol. 2010;28(suppl):Abstract e13666.
67. Sosman JA, LoRusso P, Michael SA, et al. First-in-human, multicenter, dose-escalation, phase I study of the investigational drug TAK-733, an oral MEK inhibitor, in patients (pts) with advanced nonhematologic malignancies and melanoma.J Clin Oncol. 2011;29(suppl):Abstract TPS145.
68. NIH Clinical Trials Registry. www.ClinicalTrials.gov. Identifier NCT01763164.
69. NIH Clinical Trials Registry. www.ClinicalTrials.gov. Identifier NCT01909453.