Addition Of Trametinib Research Articles

FDA Approval Summary: Dabrafenib and Trametinib for the Treatment of Metastatic Non-Small Cell Lung Cancers Harboring BRAF V600E Mutations.

The approvals of dabrafenib and trametinib, administered concurrently, provide a new regimen for the treatment of a rare subset of non-small cell lung cancer (NSCLC) and demonstrate how drugs active for treatment of BRAF-mutant tumors in one setting predict efficacy and can provide supportive evidence for approval in another setting. The FDA also approved the first next-generation sequencing oncology panel test for simultaneous assessment of multiple actionable mutations, which will facilitate selection of optimal, personalized therapy. The test was shown to accurately and reliably select patients with NSCLC with the BRAF V600E mutation for whom treatment with dabrafenib and trametinib is the optimal treatment.

Abstract LB-B29: Combined inhibition of MAPK and PI3K signaling augments standard chemotherapy response in preclinical models of pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) remains a highly lethal cancer with mortality closely paralleling incidence despite substantial advances in biological understanding of the disease. Nab-paclitaxel (NPT) plus Gemcitabine (Gem) represents the current standard for systemic therapy of advanced PDAC. More than 90% of PDACs harbor an activating mutation in the KRAS oncogene. Presently, no therapeutics exist that effectively target this oncogene, but alternative strategies focus on inhibition of downstream effectors of KRAS signaling pathways. The RAF-MEK-ERK (MAPK) and the AKT-PI3K signaling pathways are well-described mediators of KRAS induced transformation and tumorigenesis in several cancers including PDAC and represent potential targets for combination therapy. We evaluated combination treatment benefits of NPT+Gem with the MEK inhibitor trametinib (Tra) and the AKT inhibitor MK-2206 (MK) in preclinical models to evaluate their therapeutic potential against PDAC. An animal survival study was performed in peritoneal dissemination model in NOD/SCID mice. Tumor growth inhibition studies were performed in subcutaneous PDAC cell-line-derived xenografts and PDAC patient-derived xenografts in mice. Intratumoral mechanism of action was determined by immunohistochemistry (IHC) and Immunoblot analysis. Median animal survival in peritoneal dissemination PDAC xenografts in mice revealed that the median survival was 21 days in controls, which was significantly improved by the NPT+Gem combination (35 days, a 67% increase over controls). Median survival was further increased by addition of trametinib or MK-2206 to the NPT+Gem chemotherapy regimen: NPT+Gem+Tra (43 days, a 105% increase over controls), NPT+Gem+MK (39 days, a 86% increase over controls) and NPT+Gem+MK+Tra (48 days, a 129% increase over controls). In human subcutaneous xenografts using AsPC-1 PDAC cells, trametinib and MK-2206 were also able to enhance NPT+Gem effects. Compared to controls (100±34.8), the percent net local tumor growth in different therapy groups was 21.8±5.9 for NPT+Gem, 7.1±11.2 for NPT+Gem+Tra, 17.2±4.8 for NPT+Gem+MK and 5.7±9.1 for NPT+Gem+MK+Tra. Effects of therapy on intratumoral proliferation and apoptosis corresponded with tumor growth inhibition data. Furthermore, in vivo effects of trametinib and MK-2206 correlated with reduced expression of phospho-ERK and phospho-AKT in tumor samples. More importantly, in a recently completed patient-derived xenograft study, we observed that the addition of trametinib or MK-2206 again augmented the NPT+Gem tumor growth inhibition response. In this experiment, compared to controls (350 mm3), the net local tumor growth in different therapy groups was 145.4 mm3 for NPT+Gem, 122 mm3 for NPT+Gem+Tra, 20.4 mm3 for NPT+Gem+MK and -43.7 mm3 (tumor regression) for NPT+Gem+MK+Tra. These findings from multiple preclinical model settings suggest that the effects of NPT+Gem can be enhanced through combined inhibition of MAPK and PI3K signaling, which supports the potential of these targets for clinical PDAC therapy. Citation Format: Niranjan Awasthi, Sheena Monahan, Alexis Stefaniak, Margaret A. Schwarz, Roderich E. Schwarz. Combined inhibition of MAPK and PI3K signaling augments standard chemotherapy response in preclinical models of pancreatic cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B29.

Abstract 3199: Diverse non-FLT3 molecular mechanisms of crenolanib resistance

Abstract FMS-like tyrosine kinase 3 (FLT3) activating mutations are primary molecular targets for the treatment of acute myeloid leukemia (AML) due to high prevalence and unfavorable prognosis. Several type II FLT3 inhibitors have shown clinical benefits but acquisition of secondary FLT3 mutations was reported as a common mechanism of resistance. Previous studies showed that crenolanib, a type I FLT3 inhibitor, had clinical activity without acquisition of secondary FLT3 mutations. To identify the molecular mechanisms associated with crenolanib sensitivity and resistance, we performed exome sequencing on crenolanib treated patients with FLT3-mutant multiply relapsed or refractory AML. Baseline mutational analysis revealed distinct mutational profiles in patients with prior FLT3 inhibitor exposure, especially in the following pathways: NRAS, IDH1, WT1 and RUNX1. Further analysis of patients who had poor response to crenolanib showed mutations in other cell signaling genes such as NRAS and PTPN11. Variant allele frequency (VAF) analysis showed that these mutations sometimes occurred in subclones independent of the FLT3 mutation-bearing clone or sometimes were acquired by the FLT3 mutation-bearing clone. To characterize the influence of these mutations on crenolanib sensitivity, we transduced genes of interest into cell lines harboring FLT3 activating mutations and treated these cells with crenolanib at various concentrations. Significantly increased crenolanib IC50 and IC90 were observed in NRAS G12V MOLM14 and PTPN11 A72D/FLT3 D835 Ba/F3 cells relative to the respective control NRAS WT MOLM14 and PTPN11 WT/FLT3 D835 Ba/F3 cells. Notably, addition of trametinib restored crenolanib sensitivity and demonstrated synergistic cytotoxic effects on cells with FLT3 and NRAS or PTPN11 mutations. In addition, increases in TET2 nonsense/frameshift mutations were observed in patients who did not respond to crenolanib. VAF analysis demonstrated that TET2 mutations co-occurred with FLT3 mutations. We also observed that bone marrow cells from FLT3-ITD knock-in/TET2 knock-out mice are resistant to crenolanib at low concentrations, but remain sensitive to azacytidine at the same level as FLT3-ITD knock-in/TET2 WT cells. The remaining patients exhibited a diverse spectrum of secondary mutations associated with chromatin modifiers, cohesion, spliceosomes and transcription factors which mostly expanded during treatment, suggesting an elaborate genetic/epigenetic mechanism of resistance to crenolanib. Our data suggest that comprehensive sequencing should be carried out on patient samples prior to treatment to identify and pre-emptively target problematic clones. In addition, even with high VAF FLT3 mutations, although FLT3 inhibitor monotherapy provide some clinical benefit, combining agents targeting cooperative lesions will be imperative to eradicate both the dominant clone and resistant subclones and improve patient responses. Citation Format: Haijiao Zhang, Anna M. Schultz, Samantha Savage, Daniel Bottomly, Beth Wilmot, Shannon K. McWeeney, Christopher Eide, Hoang Ho, Yee L. Lam, Richard Sweat, Jaime Faulkner, Evan Lind, Jeffrey W. Tyner. Diverse non-FLT3 molecular mechanisms of crenolanib resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3199. doi:10.1158/1538-7445.AM2017-3199

CRLF2 Rearrangement Status in Ph-like ALL Predicts Intrinsic Glucocorticoid Resistance In Vitro that is Reversible with Targeted MAPK and PI3K Pathway Inhibition

Background:Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy and is most often of B-cell lineage (Roberts, et al. NEJM 2014). One subtype of B-cell ALL, Philadelphia chromosome-like ALL (Ph-like ALL), is BCR-ABL negative with a gene expression signature similar to that of BCR-ABL positive ALL and is prognostic for poor clinical outcomes (Roberts, et al. NEJM 2014). Ph-like ALL is often associated with rearrangements involving the cytokine receptor-like factor 2 (CRLF2) component of the thymic stromal lymphopoietin receptor (TSLPR) leading to its overexpression (Shochat, et al., JEM 2011). TSLPR is a heterodimer of CRLF2 and IL-7Rα that signals to promote the proliferation and differentiation of B-cell progenitors and acts to promote B-cell transformation in the context of Ph-like ALL (Maude, et al. Blood 2012). Glucocorticoid (GC) therapy plays a central role in the treatment of childhood ALL and resistance to GCs confers a poor prognosis (Piovan, et al. Cancer Cell, 2013). This study examined the role of TSLPR signaling in mediating primary GC resistance and the effects of downstream signal transduction inhibitors to confer GC sensitivity.Methods:Viably cryopreserved splenocytes were obtained from 19 patient-derived xenografts of Ph-like ALL banked in the Children’s Oncology Group or Children’s Hospital of Philadelphia leukemia biorepositories. Assays were also performed using the Mutz 5 Ph-like ALL cell line. Flow cytometry was used to assess the protein expression of TSLPR and GC receptor (GR). Levels of pERK and pAKT were measured by phosphoflow cytometry at baseline and following TSLP stimulation. Cells were cultured in the presence of 1µM dexamethasone (dex), a GC, with or without 1µM trametinib, a MEK1/2 inhibitor, or 1µM MK2206, a pan-AKT inhibitor, in the presence of TSLP and viability was assessed by Hoechst staining and flow cytometry at 48 hours.Results:Of the 19 Ph-like ALL samples in this study, 11 were CRLF2-rearranged (CRLF2R) and 8 were non-rearranged (CRLF2NR). CRLF2 rearrangements involved P2RY8 or IGH and 9 of 11 samples had concomitant activating mutations in JAK1 or JAK2. CRLF2NR samples expressed a variety of other translocations involving genes such as JAK2, PDGFR, and ABL1. CRLF2R cells were shown to have significantly greater TSLPR protein expression relative to CRLF2NR cells (p = 0.03). In the presence of TSLP, CRLF2 rearrangement status predicted responsiveness to dex, with CRLF2R samples demonstrating significant resistance to dex relative to CRLF2NR samples (p = 0.004). There was no significant reduction in cell viability following dex treatment in CRLF2R samples (p = 0.5), while dex effectively attenuated cell viability in CRLF2NR samples (p = 0.008). Importantly, there was no difference in GR expression between these two groups (p = 0.6). CRLF2R samples demonstrated hyperresponsiveness to TSLP stimulation, with a significant induction of pERK and pAKT that exceeded the response of CRLF2NR samples (p = 0.007 and p = 0.0005, respectively) despite no differences in basal phosphoprotein levels between the two sample groups. Inhibition of MAPK signaling with trametinib or of AKT signaling with MK2206 significantly sensitized CRLF2R cells to dex in the presence of TSLP when used in combination relative to dex alone (p = 0.0003 and p < 0.0001, respectively) and resulted in a significant reduction in cell viability relative to untreated cells (p < 0.0001 and p < 0.0001, respectively). The Mutz 5 cell line, which expresses both a CRLF2 rearrangement and a JAK2 activating mutation, was used to assess the effect of simultaneous pathway inhibition. Mutz 5 cells were treated with dex alone or dex in combination with trametinib and/or MK2206. While dex alone had no significant effect on cell viability, the addition of trametinib or MK2206 resulted in a 43% and 36% reduction in cell viability, respectively. Furthermore, combined treatment with dex, trametinib, and MK2206 resulted in a 72% reduction in cell viability, demonstrating the efficacy of simultaneous MAPK and PI3K pathway inhibition to confer dex sensitivity.Conclusion:MAPK and PI3K pathway signaling play a role in mediating primary GC resistance in CRLF2R Ph-like ALL, making these pathways potential therapeutic targets for enhancing the efficacy of GC therapy in this patient group. DisclosuresMaude:Novartis: Consultancy. Teachey:Novartis: Research Funding.

Abstract 276: Enhancing standard chemotherapy response by targeted inhibition of MAPK signaling pathways in experimental pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) has the worst prognosis of all solid tumors, with an overall survival rate of less than 6%. Gemcitabine (Gem) remained the standard of care agent over the past 16 years despite limited clinical benefits. Nab-paclitaxel (NPT), a water-soluble albumin-bound formulation of paclitaxel, has recently shown greater efficacy in advanced PDAC. Nab-paclitaxel combination with gemcitabine is now standard of care for advanced PDAC. More than 90% of PDAC tumors harbor an activating mutation in the KRAS oncogene. Since to date no therapeutics have effectively targeted this oncogene product, alternative strategies focus on inhibition of downstream effectors of KRAS signaling pathways. Trametinib (Tra) is a small molecule inhibitor of the RAF-MEK-ERK (MAPK) signaling pathway that is a well-described mediator of KRAS induced transformation and tumorigenesis. In the present preclinical study, we evaluated combination treatment benefits of the standard chemotherapeutics with trametinib to define a novel therapeutic strategy for PDAC. Median animal survival over controls (20 days) in human intraperitoneal PDAC xenografts was significantly improved by NPT (33 days, a 65% increase, p = 0.0004), Gem (26 days, a 30% increase, p = 0.002), NPT+Gem (39 days, a 95% increase, p = 0.0001) and Tra (31 days, a 55% increase, p = 0.0004) therapy. Survival was further increased by addition of trametinib to chemotherapy: NPT+Tra (median: 37 days, a 85% increase, p = 0.0001), Gem+Tra (34 days, a 70% increase, p = 0.0001) and NPT+Gem+Tra (49 days, a 145% increase, p&amp;lt;0.0001). Treatment of subcutaneous tumor-bearing mice with chemotherapy and trametinib resulted in significant tumor growth inhibition. Net tumor growth in different therapy groups over two weeks varied between controls (432.6 mm3), NPT (105.3 mm3), Tra (184 mm3), NPT+Tra (81 mm3), NPT+Gem (37.3 mm3), and NPT+Gem+Tra (-8.1 mm3). Mean tumor weight (in g) in different therapy groups was as follows: controls 0.38±0.06, NPT 0.23±0.06, Tra 0.31±0.03, NPT+Tra 0.23±0.06, NPT+Gem 0.15±0.05 and NPT+Gem+Tra 0.11±0.05. In vitro studies demonstrated inhibition in PDAC cell (AsPC-1, Panc-1, Mia PaCa-2, CFPAC) proliferation by treatment with NPT+Gem, trametinib, and combination. Immunoblot analysis revealed that trametinib effects were accompanied by decrease in phospho-ERK expression and increase in the expression of apoptosis-related cleaved caspase-3 protein. These findings suggest that the effects of the standard chemotherapeutic regimens can be enhanced through specific inhibition of downstream components of the MAPK signaling pathway, which clinically could lead to improved PDAC therapy effects. Citation Format: Niranjan Awasthi, Sheena Monahan, Margaret A. Schwarz, Roderich E. Schwarz. Enhancing standard chemotherapy response by targeted inhibition of MAPK signaling pathways in experimental pancreatic cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 276.

Combination dabrafenib and trametinib in the management of advanced melanoma with BRAFV600 mutations

ABSTRACTIntroduction: In the 40–50% of advanced melanoma patients with tumors harboring BRAF V600E and V600 K mutations, BRAF inhibitors such as dabrafenib are a highly effective treatment. However, most patients develop resistance after several months on treatment. The addition of a MEK inhibitor, such as trametinib, to BRAF inhibition mitigates one key pathway of resistance, further increasing response rates and improving survival.Areas covered: This article summarizes the mechanism of action of the combination of dabrafenib and trametinib, its evolution through Phase I, II and III clinical trials and discusses its current use in the management of patients with advanced melanoma.Expert opinion: Combination therapy with dabrafenib and trametinib improves response rate, progression-free survival and overall survival when compared to dabrafenib or vemurafenib alone. The addition of trametinib to dabrafenib changes the adverse event profile, making hyperkeratosis and cutaneous squamous cell carcinomas less common but side effects such as fever and nausea more common. How dabrafenib/trametinib is best sequenced with other effective treatments such as immune checkpoint blockade remains uncertain.

ERBB activation modulates sensitivity to MEK1/2 inhibition in a subset of driver-negative melanoma.

Melanomas are characterized by activating "driver" mutations in BRAF, NRAS, KIT, GNAQ, and GNA11. Resultant mitogen-activated protein kinase (MAPK) pathway signaling makes some melanomas susceptible to BRAF (BRAF V600 mutations), MEK1/2 (BRAF V600, L597, fusions; NRAS mutations), or other kinase inhibitors (KIT), respectively. Among driver-negative ("pan-negative") patients, an unexplained heterogeneity of response to MEK1/2 inhibitors has been observed. Analysis of 16 pan-negative melanoma cell lines revealed that 8 (50%; termed Class I) are sensitive to the MEK1/2 inhibitor, trametinib, similar to BRAF V600E melanomas. A second set (termed Class II) display reduced trametinib sensitivity, paradoxical activation of MEK1/2 and basal activation of ERBBs 1, 2, and 3 (4 lines, 25%). In 3 of these lines, PI3K/AKT and MAPK pathway signaling is abrogated using the ERBB inhibitor, afatinib, and proliferation is even further reduced upon the addition of trametinib. A potential mechanism of ERBB activation in Class II melanomas is minimal expression of the ERK1/2 phosphatase, DUSP4, as ectopic restoration of DUSP4 attenuated ERBB signaling through potential modulation of the ERBB ligand, amphiregulin (AREG). Consistent with these data, immunohistochemical analysis of patient melanomas revealed a trend towards lower overall DUSP4 expression in pan-negative versus BRAF- and NRAS-mutant tumors. This study is the first to demonstrate that differential ERBB activity in pan-negative melanoma may modulate sensitivity to clinically-available MEK1/2 inhibitors and provides rationale for the use of ERBB inhibitors, potentially in combination with MEK1/2 inhibitors, in subsets of this disease.

Improved antitumor activity of immunotherapy with BRAF and MEK inhibitors in BRAF(V600E) melanoma.

Combining immunotherapy and BRAF targeted therapy may result in improved antitumor activity with the high response rates of targeted therapy and the durability of responses with immunotherapy. However, the first clinical trial testing the combination of the BRAF inhibitor vemurafenib and the CTLA4 antibody ipilimumab was terminated early because of substantial liver toxicities. MEK [MAPK (mitogen-activated protein kinase) kinase] inhibitors can potentiate the MAPK inhibition in BRAF mutant cells while potentially alleviating the unwanted paradoxical MAPK activation in BRAF wild-type cells that lead to side effects when using BRAF inhibitors alone. However, there is the concern of MEK inhibitors being detrimental to T cell functionality. Using a mouse model of syngeneic BRAF(V600E)-driven melanoma, SM1, we tested whether addition of the MEK inhibitor trametinib would enhance the antitumor activity of combined immunotherapy with the BRAF inhibitor dabrafenib. Combination of dabrafenib and trametinib with pmel-1 adoptive cell transfer (ACT) showed complete tumor regression, increased T cell infiltration into tumors, and improved in vivo cytotoxicity. Single-agent dabrafenib increased tumor-associated macrophages and T regulatory cells (Tregs) in tumors, which decreased with the addition of trametinib. The triple combination therapy resulted in increased melanosomal antigen and major histocompatibility complex (MHC) expression and global immune-related gene up-regulation. Given the up-regulation of PD-L1 seen with dabrafenib and/or trametinib combined with antigen-specific ACT, we tested the combination of dabrafenib, trametinib, and anti-PD1 therapy in SM1 tumors, and observed superior antitumor effect. Our findings support the testing of triple combination therapy of BRAF and MEK inhibitors with immunotherapy in patients with BRAF(V600E) mutant metastatic melanoma.

A phase 1b study of trametinib, an oral Mitogen-activated protein kinase kinase (MEK) inhibitor, in combination with gemcitabine in advanced solid tumours

PurposeThis phase 1b study determined the safety, tolerability, and recommended phase 2 dose (RP2D) and schedule of trametinib in combination with gemcitabine. Secondary objectives included assessment of clinical activity and steady-state pharmacokinetics. MethodsAdults with advanced solid tumours, adequate organ function and Eastern Co-operative Oncology Group performance status (ECOG PS)⩽1 were eligible. Once-daily oral trametinib (1mg, 2mg, 2.5mg) was escalated in a 3+3 design with standard gemcitabine dosing (1000mg/m2 IV Days 1, 8, and 15 of 28-day cycles). During expansion, trametinib 2mg was combined with gemcitabine. Pharmacokinetics samples were collected on Day 15 pre-dose and 1, 2, 4 and 6h post-dose; tumour assessments were repeated every two cycles. ResultsBetween 8/2009 and 11/2010, 31 patients (pancreas=11, breast=6, non-small cell lung cancer (NSCLC)=4, other=10) were treated. Dose-limiting toxicities (DLTs) occurred in each cohort, and included febrile neutropenia, transaminase elevation and uveitis. The RP2D was declared as trametinib 2mg daily with standard gemcitabine dosing. Common grade 3/4 toxicities at the RP2D included: neutropenia (38%), thrombocytopenia (19%) and transaminase elevation (14%). Of 10 patients with measurable pancreatic cancer, three partial responses (30%) were documented; two additional patients achieved objective responses (breast, complete response (CR); salivary glands, partial response (PR)). Pharmacokinetics suggested no change in exposures of either drug in combination. ConclusionAdministration of trametinib at its full monotherapy dose of 2mg daily in combination with standard gemcitabine dosing (1000mg/m2 IV Days 1, 8, and 15 every 28 days) was feasible. Though most toxicities were manageable, the addition of trametinib may increase gemcitabine-associated myelosuppression. Future studies of this combination will require monitoring to maintain dose and schedule.

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.

291 Background: Trametinib, an oral MEK1/2 inhibitor, holds promise for tumors that frequently harbor RAS activating mutations, such as pancreatic cancer. Trametinib monotherapy or in combination with gemcitabine showed preliminary activity in patients (pts) with advanced pancreatic cancer. Methods: Eligible pts with untreated metastatic pancreatic cancer were randomized (double-blind, 1:1) to receive gemcitabine 1000 mg/m2 intravenously weekly x 7 for 8 weeks, then weekly x 3 every 4 weeks, plus either trametinib 2 mg or placebo daily. The primary endpoint was overall survival (OS); secondary endpoints included progression-free survival (PFS), overall response rate (ORR), and duration of response (DoR). OS and ORR were also analyzed based on baseline KRAS status as determined in plasma cell free DNA (cfDNA). Results: Baseline characteristics for the 160 randomized and treated pts were similar across arms. Skin related events (73% vs. 34%), diarrhea (54% vs. 28%), thrombocytopenia (40% vs. 28%), and stomatitis (36% vs. 8%) were more frequent with trametinib, as was grade 3/4 anemia (22% vs. 11%). However, rates of grade ≥ 3 thrombocytopenia, neutropenia and febrile neutropenia were similar between the arms. More pts on trametinib arm required dose reduction or interruption due to AEs (68% vs. 49% and 74% vs. 43%, respectively). Median OS was 8.4 months with trametinib compared to 6.7 months with placebo [HR 0.98 (95% CI: 0.67, 1.44, p=0.453)]. Median PFS was 16 weeks on trametinib and 15 weeks on placebo arm. ORRs and median DoRs were 22% and 23.9 weeks and 18% and 16.1 weeks on trametinib and placebo arm, respectively. The median OS and ORR in the subgroup of pts with KRAS mutations (n=143) was similar to OS and ORR for all randomized pts. Conclusions: This is first randomized, placebo-controlled trial evaluating the combination of gemcitabine with a MEK inhibitor. There was an increased incidence of skin, GI, and hematologic toxicities with trametinib compared to placebo. The addition of trametinib did not improve OS, PFS, or response rate. These outcomes remained independent of KRAS mutations based on cfDNA. Clinical trial information: NCT01231581.