Inhibitors In Preclinical Models Research Articles

Synergistic effect of reirradiation and PD-1 inhibitors in recurrent high-grade gliomas.

105 Background: To date, studies of single agent PD-1 or PDL-1 inhibitors in recurrent high-grade gliomas (HGG) have shown infrequent responses ( < 10%) and limited efficacy. Reirradiation is considered one of the standard salvage regimens for selected patients with recurrent gliomas but tumor responses are also infrequent ( < 5%). Radiotherapy counteracts the immunosuppressive tumor microenvironment by increasing MHC class I expression and enhancing tumor neoantigen presentation and has a significant synergistic effect with PD-1 inhibition in preclinical models of glioma. Methods: From December 2014 to June 2016, 20 patients (14 men, 6 women) with recurrent HGG were treated with the combination of reirradiation and PD-1 inhibitors. 18 patients had a glioblastoma, 1 anaplastic astrocytoma and 1 anaplastic oligodendroglioma. The median KPS at the start of this regimen was 70 (50 to 80). The median number of prior treatments for recurrent tumor was 2 (1-4), 100% had prior radaition, 95% prior temozolomide and 55% prior bevacizumab. 8 patients received pembrolizumab (2 mg/Kg every 3 weeks) and 12 patients received nivolumab (3 mg/Kg or 240 mg flat dose every 2 weeks). Median reirradiation dose was 35 Gy (12 Gy to 35 Gy). Results: There were 7 confirmed partial responses (35% objective response rate, ORR), 5 stable disease and 8 progressive disease. The median duration of response was 5 months (2.2 to 10+ months). Median PFS was 4 months and median OS was 10 months. Most common side effects were increased ALT (3 patients) and fatigue (2 patients). There was no obvious case of cerebral edema related to treatment. 5 patients required a mild increase of dexamethasone (2-4 mg) and all other 15 patients had decreased or stable dexamethasone dosage after the reirradiation. Conclusions: PD-1 inhibitors in combination with re-irradiation can be administered safely to patients with recurrent HGG. The ORR of 35% is an early signal of the potential synergist effect of the combination of PD-1 inhibitors with re-irradiation in HGG. This retrospective analysis of a heavily pre-treated population of recurrent HGG showed significantly higher ORR than either PD-1 inhibitors or re-irradiation alone. A clinical trial under development to test this hypothesis.

Inhibition of phosphodiesterase 4 (PDE4) reduces dermal fibrosis by interfering with the release of interleukin-6 from M2 macrophages

ObjectivesTo investigate the disease-modifying effects of phosphodiesterase 4 (PDE4) inhibition in preclinical models of systemic sclerosis (SSc).MethodsWe studied the effects of PDE4 inhibition in a prevention and a treatment model...

Mitogen-activated protein kinases as therapeutic targets for asthma.

Corticosteroid-resistant asthmatics, although comprising only a portion of the asthma population, account for most of the morbidity, mortality and economic burden associated with asthma. Moreover, corticosteroids are not effective inhibitors of airway remodeling changes, and their long-term use is associated with debilitating systemic side effects. Therefore, potent and safe novel therapeutic alternatives, targeting basic pathophysiological mechanisms responsible for the severe asthmatic phenotype are urgently needed. Mitogen-activated protein kinases (MAPKs) are ubiquitously expressed signaling enzymes that are involved in almost all aspects of the asthmatic inflammatory network; as such, they represent an emerging target for the treatment of asthma. This paper provides a rationale for targeting MAPKs in the treatment of asthma by reviewing the in vitro evidence of its relevance to asthma pathogenesis. This is followed by discussing the results of MAPK inhibition in pre-clinical models of asthma. Finally, the potential safety concerns regarding MAPK inhibition in human disease, as well as the future prospects for its clinical development are explored. In conclusion, this review underlines the promising results of MAPK inhibition in animal asthma models especially in restoring corticosteroid sensitivity, as well as recent clinical safety and efficacy evidence obtained from trials in similar disease areas such as COPD, and of course, the paucity of clinical evidence for targeting MAPKs in asthma. Based on this review, a more rigorous effort for clinical development of MAPK inhibitors in asthma is justified.

Abstract P1-07-04: Unique overlapping subtypes of triple-negative breast and ovarian cancers and sensitivity of “mesenchymal-like” cancers to HSP90 inhibition is revealed by integrated gene expression and drug sensitivity profiling

Abstract Next-generation sequencing and gene expression signature analysis of primary tumors from The Cancer Genome Atlas (TCGA) revealed striking similarities between triple-negative breast cancer (TNBC) and ovarian cancer (OVCA). We hypothesized that these similarities may reveal transcriptionally-identifiable subgroups within a mix of TNBC and OVCA that are uniquely sensitive to certain drugs. To test this hypothesis, gene expression profiles for TNBC and OVCA cell lines in the Cancer Cell Line Encyclopedia were analyzed by unsupervised hierarchical clustering, which revealed two major unique subgroups containing both TNBC and OVCA cell lines that clustered according to the previously defined Mesenchymal and Basal subclasses of TNBC. Differential gene expression between “Mesenchymal-like” and “Basal-like” TNBC/OVCA cell lines was used to generate a gene signature that was subsequently validated using gene expression data in the Genomics of Drug Sensitivity in Cancer (GDSC) database. Drug sensitivity data from GDSC was then utilized to profile differential sensitivity of “Mesenchymal-like” and “Basal-like” cell lines to the 99 anti-cancer drugs with coverage of >50 breast and ovarian cell lines. Mesenchymal-like TNBC/OVCA cells were uniquely sensitive to HSP90 inhibition compared to Basal-like TNBC/OVCA cells for both HSP90 inhibitors in GDSC: CCT018159 (p<0.001) and 17-AAG (p=0.012). Strikingly, Mesenchymal-like TNBC/OVCA cells were most sensitive to HSP90 inhibition among all 33 solid tumor cancer lineages in GDSC, as well as other subgroups of breast and ovarian cancers. Differential sensitivity of Mesenchymal-like and Basal-like TNBC/OVCA cells to HSP90 inhibition with 4 agents (CCT018159, 17-AAG, AT13387, PU-H71) was validated using growth assays for 12 cell lines (6 of each subgroup) previously characterized in GDSC. To further validate the predictive value of our gene signature, gene expression data for 6 TNBC/OVCA cell lines with uncharacterized drug sensitivity was used to classify cell lines as “Mesenchymal-like” (n=4) or “Basal-like” (n=2). Our gene signature successfully predicted differential sensitivity to HSP90 inhibition, with the 4 Mesenchymal-like cell lines displaying greater sensitivity to HSP90 inhibition. The gene signature has been used to select PDX models with predicted differential sensitivity to HSP90 inhibition, and treatment studies are ongoing. In summary, we developed a novel approach to generate a gene expression signature that robustly predicts sensitivity to HSP90 inhibitors in preclinical models of TNBC and OVCA. HSP90 thus represents a therapeutic opportunity in an overlapping subpopulation of TNBC and OVCA. Our approach to identify novel combinations of drugs and histology/lineage-independent cancer subgroups may be used to discover new therapeutic opportunities in other cancer types. Citation Format: Shee K, Ung MH, Cheng C, Miller TW. Unique overlapping subtypes of triple-negative breast and ovarian cancers and sensitivity of “mesenchymal-like” cancers to HSP90 inhibition is revealed by integrated gene expression and drug sensitivity profiling [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-07-04.

MA11.07 Improved Small Cell Lung Cancer (SCLC) Response Rates with Veliparib and Temozolomide: Results from a Phase II Trial

PARP1 is overexpressed in small cell lung cancer (SCLC) and represents a novel therapeutic target for this disease. Preclinical data indicates that combining veliparib (an oral PARP-1/2 inhibitor) and temozolomide (TMZ) results in synergistic tumor growth delay or regression. In this study, we investigated whether adding veliparib to TMZ would improve outcomes in patients with relapsed sensitive and refractory SCLCs. Candidate predictive biomarkers, including SLFN11, were then explored. SCLC patients previously treated with 1 or 2 prior regimens were enrolled in the trial and randomized 1:1 to receive oral TMZ 150-200mg/m2/day (D1-5) with either veliparib or placebo 40mg twice daily, orally (D1-7) (NCT01638546). Primary endpoint was 4-month progression free survival (PFS). Data were analyzed in patients with platinum sensitive (progression >60 days after 1st line therapy) or refractory disease (progression ≤60 days after 1st line therapy, or in need of 3rd line treatment). Archived tissue was available for 53 patients for biomarker analysis. 104 patients were enrolled and 100 patients were treated. Baseline characteristics were balanced between treatment arms: 52% female; median age 62.5 (range, 31-84); 59% refractory disease; 33% needing 3rd-line therapy. Progression free survival at 4-months was similar between the two arms, 36% vs. 27% (p=0.39). However, in 93 evaluable pts, response rate was significantly higher in pts treated with veliparib/TMZ compared to TMZ alone (39% vs 14%, p =0.016). Median overall survival: 8.2 mos (95% CI: 6.4-12.2) in veliparib arm and 7 mos (95% CI: 5.3-9.5) in placebo arm, p = 0.50. Grade 3/4 thrombocytopenia and neutropenia more commonly occurred in the veliparib/TMZ arm: 50% vs 9% and 31% vs 7%, respectively. Levels of SLFN11, a marker of SCLC response to PARP inhibition in preclinical models, were assessed by immunohistochemistry. High SLFN11 in patient tumors (obtained at original diagnosis) was associated with a trend towards better overall survival in the veliparib/TMZ arm, but no difference in outcome in the TMZ alone arm. Additional correlative studies are ongoing, including assessment of MGMT promoter methylation, and will be available at the time of presentation. The combination of veliparib/TMZ increased response rates significantly, compared to TMZ alone. Hematologic toxicities of the combination may have impacted PFS (which was not significantly different between the arms) by limiting dosing. Biomarkers such as SLFN11, ATM, or MGMT promoter methylation could potentially help guide patient selection in the SCLC population.

The Synergistic Effect of Melphalan and XPO1 Inhibition in Pre-Clinical Models of Multiple Myeloma

Introduction:Multiple myeloma (MM) is an incurable cancer of plasma cells. It accounts for approximately 10% of all hematologic malignancies. In the US, it is estimated that there will be approximately 30,330 new cases and 12,650 deaths in 2016. In the past decade, responses/survivals have been significantly increased by newer therapies. However, almost all of the patients will eventually die from multi-drug resistant disease.Materials and Methods:Weused XPO1 inhibitors (XPO1i) selinexor (300nM) or KPT-8602 (300nM) +/- melphalan (15 μM) to treat human MM parental RPMI8226 and U266 cells, and melphalan resistant LR5 and LR6 cell lines for 20 hours and then assayed for apoptosis and viability by flow cytometry. DNA damage was assayed by the comet assay and phospho-H2AX protein expression in H929 human myeloma cells. p53, NFkB, IKKα, FANCF, and FANCL were assayed by Western blot in H929 MM cells. We also treated cells from patients with newly diagnosed or relapsed/refractory MM with the XPO1i (300nM)/ melphalan (10μM) combination and assayed for apoptosis. In addition, selinexor/melphalan treated NOD/SCID-gamma mice with U226 MM tumors were assayed for tumor growth, survival, and toxicity.Results:Cell viability of all tested MM cell lines was decreased synergistically and apoptosis increased by XPO1i/melphalan treatment (selinexor/melphalan, P = 2.2x10E-6 to 0.0032, KPT-8602/ melphalan, P = 1.2X10E-7 to 0.0031). Comet assays showed that the XPO1i/ melphalan drug combination increased DNA damage more than single agent melphalan or XPO1i alone. Phospho-H2AX expression also was increased (selinexor/ melphalan, P = 0.005 and KPT-8602/melphalan, P = 0.001). Western blot analysis showed that XPO1i treatment can increase p53 and decrease NFkB, IKKα, FANCF, and FANCL in MM cells. Apoptosis assays showed that both melphalan-resistant and parental MM cell lines were sensitized to melphalan by XPO1i. In addition, CD138+/light chain+ MM cells from newly diagnosed and relapsed/refractory MM patients were sensitized (20-fold and 5 to10-fold respectively) by XPO1i to melphalan. XPO1i/melphalan combination treatment demonstrated a strong synergistic anti-tumor effect when compared to single-agent melphalan (selinexor, P = 0.0024 and KPT-8602, P = 0.0030) in NOD/SCID-gamma mice challenged with U266 MM tumors. XPO1i/ melphalan treated mice had increased survival and no significant toxicity.Conclusions:XPO1i’s can improve the response of human MM cell lines and patient MM cells to melphalan both in vitro and ex vivo. The mechanism of this synergy reversing melphalan resistance may be due to increased nuclear p53, in combination with decreased NFkB and IKKα, and decreased DNA repair proteins FANCL and FANCF of the Fanconi Anemia/BRCA pathway. Our preliminary data suggest that the synergistic cell kill may be because XPO1i’s increase melphalan-induced DNA damage and block the repair of the DNA damage. Thus using combination therapies of XPO1i, especially the clinical compounds selinexor and KPT-8602 +/- melphalan may have potential to improve the treatment outcomes of MM. Based on these promising pre-clinical data, we designed a phase 1/2 clinical trial evaluating the combination of selinexor and high-dose melphalan as a conditioning regimen for autologous hematopoietic cell transplantation in patients with multiple myeloma (NCT02780609). DisclosuresShain:Novartis: Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Signal Genetics: Research Funding; Takeda/Millennium: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen/Onyx: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Baloglu:Karyopharm Therapeutics Inc: Employment, Other: stockholder. Nishihori:Signal Genetics: Research Funding; Novartis: Research Funding.

164TiP Phase I study of LOXO-101, a selective TRK inhibitor, in pediatric patients with cancer

Background Neurotrophin ligands and their receptors TRKA, TRKB, and TRKC (encoded by NTRK1, NTRK2, and NTRK3) are important for growth regulation,differentiation and survival of neurons. Translocations involving the NTRK1/2/3 kinase domain, mutations involving the TRK ligand-binding site, amplifications of NTRK, TRK splice variants, and autocrine/paracrine signaling have been described in diverse tumor types and may contribute to tumorigenesis. A broad range of pediatric malignancies have been found to harbor NTRK fusions, including infantile fibrosarcoma (IFS), congenital mesoblastic nephroma (CMN), secretory breast cancer, pediatric papillary thyroid cancer, pediatric gliomas and Ph-like acute lymphoblastic leukemia. Additionally, TRK protein over-expression is common in neuroblastoma. LOXO-101isthefirstsmall-moleculeselectiveinhibitorofTRKA,-B,and-Cinclinical development and has demonstrated tumor inhibition in preclinical models and clinically meaningful responses in patients with TRK fusion cancers in an adult phase 1 trial. Trialdesign: Wehaveinitiatedanopen-label, multi-centerPhaseIdoseescalation/dose expansionstudywithLOXO-101inpediatricpatientswithsolidtumorsandprimary CNS tumors (NCT02637687). Patients with advanced cancer between the ages of 1 and 21yearsareeligible,aswellaspatientsasyoungas1-monthofagewithaprimary diagnosis ofIFSorCMNandadocumentedNTRKfusion. Twice-dailyoraldosingof LOXO-101 capsulesisadministeredon acontinuous28-dayschedule. LOXO-101is available in an oral liquid formulation for patients unable to swallow capsules. PK- directedintra-subjectdoseescalationispermitted,withtargetexposuresequivalentto the recommended Phase 2 dose in adults of 100 mg BID. Dose escalation utilizes a Rolling 6 design. The objective of the study is to determine the maximum tolerated dose andinitialevidenceoftheefficacyofLOXO-101indifferenttumortypes.Eligibilityfor the dose expansion cohorts will require patient tumor samples to have documented alterations of an NTRK gene or TRK protein. Molecular abnormalities will be characterized through the analysis of archival tissue. Enrollment began in December 2015 and is ongoing. Clinical trial indentification: NCT02637687 Legal entity responsible for the study: Loxo Oncology, Inc. Funding: Loxo Oncology, Inc. Disclosure: S. Smith, M. Reynolds, S. Cruickshank: Consultant fees paid by Loxo Oncology, Inc. L. Donahue: Employee and stockholder of Loxo Oncology, Inc. M.C. Cox: Employee and stock hold of Loxo Oncology, Inc. All other authors have declared no conflicts of interest.

353 - Synergistic action of first-in-class anti-tropomyosin compound, ATM-3507 (Anisina) and microtubule targeting inhibitors in pre-clinical models of non-small cell lung cancer (NSCLC)

353 - Synergistic action of first-in-class anti-tropomyosin compound, ATM-3507 (Anisina) and microtubule targeting inhibitors in pre-clinical models of non-small cell lung cancer (NSCLC)

The Potent ALK Inhibitor Brigatinib (AP26113) Overcomes Mechanisms of Resistance to First- and Second-Generation ALK Inhibitors in Preclinical Models.

Non-small cell lung cancers (NSCLCs) harboring ALK gene rearrangements (ALK+) typically become resistant to the first-generation anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitor (TKI) crizotinib through development of secondary resistance mutations in ALK or disease progression in the brain. Mutations that confer resistance to second-generation ALK TKIs ceritinib and alectinib have also been identified. Here, we report the structure and first comprehensive preclinical evaluation of the next-generation ALK TKI brigatinib. A kinase screen was performed to evaluate the selectivity profile of brigatinib. The cellular and in vivo activities of ALK TKIs were compared using engineered and cancer-derived cell lines. The brigatinib-ALK co-structure was determined. Brigatinib potently inhibits ALK and ROS1, with a high degree of selectivity over more than 250 kinases. Across a panel of ALK+ cell lines, brigatinib inhibited native ALK (IC50, 10 nmol/L) with 12-fold greater potency than crizotinib. Superior efficacy of brigatinib was also observed in mice with ALK+ tumors implanted subcutaneously or intracranially. Brigatinib maintained substantial activity against all 17 secondary ALK mutants tested in cellular assays and exhibited a superior inhibitory profile compared with crizotinib, ceritinib, and alectinib at clinically achievable concentrations. Brigatinib was the only TKI to maintain substantial activity against the most recalcitrant ALK resistance mutation, G1202R. The unique, potent, and pan-ALK mutant activity of brigatinib could be rationalized by structural analyses. Brigatinib is a highly potent and selective ALK inhibitor. These findings provide the molecular basis for the promising activity being observed in ALK+, crizotinib-resistant patients with NSCLC being treated with brigatinib in clinical trials. Clin Cancer Res; 22(22); 5527-38. ©2016 AACR.

Efficacy of NEDD8 Pathway Inhibition in Preclinical Models of Poorly Differentiated, Clinically Aggressive Colorectal Cancer.

The NEDD8 conjugation pathway modulates the ubiquitination and activity of a wide range of intracellular proteins, and its blockade by pevonedistat is emerging as a promising therapeutic approach in various cancer settings. However, systematic characterization of pevonedistat efficacy in specific tumor types and definition of response predictors are still missing. We investigated in vitro sensitivity to pevonedistat in 122 colorectal cancer (CRC) cell lines by an ATP-based proliferation assay and evaluated apoptosis and DNA content by flow cytometry. Associations between pevonedistat sensitivity and CRC molecular features were assessed by Student's t test. A 184-gene transcriptional predictor was generated in cell lines and applied to 87 metastatic CRC samples for which patient-derived xenografts (PDXs) were available. In vivo reponse to pevonedistat was assessed in PDX models (≥5 mice per group). All statistical tests were two-sided. Sixteen (13.1%) cell lines displayed a marked response to pevonedistat, featuring DNA re-replication, proliferative block, and increased apoptosis. Pevonedistat sensitivity did not statistically significantly correlate with microsatellite instability or mutations in KRAS or BRAF and was functionally associated with low EGFR pathway activity. While ineffective on predicted resistant PDXs, in vivo administration of pevonedistat statistically significantly impaired growth of five out of six predicted sensitive models (P < .01). In samples from CRC patients, transcriptional prediction of pevonedistat sensitivity was associated with poor prognosis after surgery (hazard ratio [HR] = 2.49, 95% confidence interval [CI] = 1.34 to 4.62, P = .003) and early progression under cetuximab treatment (HR = 3.59, 95% CI = 1.60 to 8.04, P < .001). Histological and immunohistochemical analyses revealed that the pevonedistat sensitivity signature captures transcriptional traits of poor differentiation and high-grade mucinous adenocarcinoma. These results highlight NEDD8-pathway inhibition by pevonedistat as a potentially effective treatment for poorly differentiated, clinically aggressive CRC.

804P - A phase 1b dose-escalation study of TRC105 (endoglin antibody) in combination with axitinib in patients with metastatic renal cell carcinoma (mRCC)

Resistance to VEGF-targeted therapy is a major challenge in contemporary treatment of mRCC, and endoglin (CD105) activation may be an important mechanism leading to resistance. Endoglin is an essential angiogenic receptor expressed on proliferating tumor vessels and mRCC cancer stem cells, and is upregulated following VEGF inhibition. TRC105 is an endoglin monoclonal antibody that potentiates the anti-tumor activity of bevacizumab and VEGF receptor tyrosine kinase inhibitors in preclinical models. Heavily pretreated mRCC pts with ECOG PS 0-1 and acceptable organ function were treated with TRC105 weekly (8mg/kg and then 10mg/kg) in combination with axitinib (initially at 5 mg BID and then escalated per patient tolerance to a maximum of 10 mg BID). Eighteen mRCC pts (median age = 61.5; M:F 16:2; median number of prior therapies = 3, including > 1 VEGFR TKI, clear cell = 13, prior axitinib = 1) were treated. TRC105 dose escalation proceeded from 8 mg/kg (n = 3) to 10 mg/kg (n = 15) without dose limiting toxicity. Adverse events characteristic of each drug were not increased in frequency or severity when the two drugs were administered concurrently, and most commonly included epistaxis, headache, fatigue, diarrhea, and gingival bleeding. TRC105 and axitinib demonstrated preliminary evidence of activity, including partial responses in 29% of patients by RECIST 1.1, and longer PFS than expected with axitinib as a single agent. The overall disease control rate (CR/PR/SD > 2 months) was 88% (15 of 17). Median PFS overall was 8.4 months, and was 9.6 months among patients with clear cell RCC. Tumor response will be correlated with baseline protein biomarkers. TRC105 pharmacokinetic parameters will be reported. TRC105 at 8 and 10 mg/kg was well tolerated in combination with axitinib, with encouraging evidence of activity in patients with mRCC. A multicenter randomized Phase 2 trial of axitinib +/- TRC105 is actively enrolling at this time (NCT01806064).

BET inhibitors in the treatment of hematologic malignancies: current insights and future prospects

The bromodomain and extra-terminal (BET) family of proteins are important epigenetic regulators involved in promoting gene expression of critical oncogenes. BET inhibitors have been demonstrated to repress c-Myc expression, and were initially shown to have efficacy in a number of c-Myc-dependent hematologic malignancies. Recent studies have now revealed a broader role for BET inhibitors in hematologic malignancies. In this review, we summarize the efficacy of BET inhibitors in preclinical models of acute leukemia, lymphoma, and multiple myeloma. We also summarize recent results of clinical trials utilizing BET inhibitors in hematologic malignancies, characterize potential resistance mechanisms to BET inhibitors, and discuss potential combination therapies with BET inhibitors in patients with hematologic malignancies.

Molecular regulation of apoptotic machinery and lipid metabolism by mTORC1/mTORC2 dual inhibitors in preclinical models of HER2+/PIK3CAmut breast cancer.

The mechanistic target of rapamycin (mTOR) is a rational target for cancer treatment. While the mTORC1-selective rapalogs have shown significant benefits in the clinic, antitumor response may be further improved by inhibiting both mTORC1 and mTORC2. Herein, we established target profile of a novel mTOR kinase inhibitor (mTOR-KI) MTI-31 and employed it to study new therapeutic mechanism in breast cancer. MTI-31 demonstrated a potent mTOR binding affinity with >5000 fold selectivity over the related PI3K family isoforms. MTI-31 inhibited mTORC1- and mTORC2 function at ≤120 nM in cellular assays or 5 mg/kg orally in tumor-bearing mice. In a panel of breast cancer lines, the antitumor efficacy of MTI-31 was dependent on HER2+ and/or PIK3CAmut (HER2+/PIK3CAmut) status of the tumors and required mTORC2-specific modulation of Bim, MCL-1 and GSK3. Inactivation of Bim or GSK3 each attenuated apoptotic death resulting in mTOR-KI resistance. The antitumor response also required a suppression of lipid metabolism in therapy-sensitive tumors. Treatment with MTI-31 or AZD8055 substantially reduced lipogenesis and acetyl-CoA homeostasis, which was mechanistically linked to a blockade of mTORC2-dependent glucose-to-lipid conversion rate. We also found that the basal levels of carnitine palmitoyltransferase 1A and lipid catabolism were elevated in HER2+/PIK3CAmut breast cells and were inhibited upon mTOR-KI treatment. A CPT1A inhibitor etomoxir mimicked MTI-31 action in selective downregulation of cellular lipid catabolism. Co-treatments with MTI-31 and etomoxir enhanced the suppression of cyclin D1, c-Myc and cell growth in HER2+/PIK3CAmut tumors. These new mechanistic findings provide a rationale for targeting mTORC1 and mTORC2 in HER2+/PIK3CAmut breast cancer.

Abstract 1010: Evaluation of small-molecule FASN inhibitors in preclinical models of colorectal cancer

Abstract Fatty Acid Synthase (FASN), a key enzyme of de novo lipid synthesis, is upregulated in many cancers including colorectal cancer (CRC); increased FASN activity is associated with decreased survival and increased disease recurrence. Recently, a first-in-class, oral FASN inhibitor (TVB-2640) entered a Phase I clinical trial (3V2640-CLIN-002) in solid tumor patients demonstrating a favorable tolerability profile with no significant adverse events; however, tumor characteristics that would indicate responsiveness to FASN inhibition are not fully understood. The purpose of our study was: (i) to determine the effect of novel, selective and reversible FASN inhibitors on proliferation of primary CRC cell cultures, established CRC cell lines, and CRC patient-derived xenografts (PDXs); and (ii) to identify potential biomarkers associated with CRC responsiveness to FASN inhibition. METHODS. The effect of TVB-3166, TVB-3664, and TVB-3693 (all developed by 3-V Biosciences) on the proliferation of primary cells (established from 1st generation PDX tumors) and CRC cell lines was assessed by cell count; apoptosis was assessed by Cell Death ELISA. In addition, tumor growth was assessed in PDX models established in NOD SCID gamma mice using freshly resected CRC specimens (either primary CRC or metastasis) from our patient population. Once the xenografts grew to ∼100 mm3, mice were randomized into two groups (n = 5) to receive either vehicle or TVB-3664 (3mg/kg) by gavage daily. Tumor volume and animal weights were measured weekly. Western blot analysis, immunohistochemistry and immunofluorescent staining were used to identify FASN-mediated changes in β-catenin, Akt and AMPK pathways. RESULTS. TVB compounds tested showed similar efficacy in primary and established CRC cells with a wide range of sensitivity to FASN inhibition. The 5 cell lines that were most responsive to FASN inhibition demonstrated a low basal level of pAMPK and pAkt as compared to the 5 least responsive cells. Moreover, we noted that increased FASN protein expression was also associated with increased sensitivity to FASN inhibition. Inhibition of proliferation by TVB compounds was associated with decreased expression of active β-catenin, c-MYC, pAkt, and survivin, while an increase in apoptosis was noted by induction of PARP cleavage. Consistent with our in vitro studies, TVB-3664 treatment significantly reduced tumor volume in vivo with no weight changes or toxicity observed. CONCLUSIONS. Our studies show that the novel FASN inhibitors, as a single agent, significantly inhibit CRC growth both in vitro and in vivo. Importantly, our results suggest that basal activation of AMPK and Akt may be predictive of responsiveness to FASN inhibition and may function as potential biomarkers to allow a more personalized treatment approach. Citation Format: Yekaterina Y. Zaytseva, Piotr G. Rychahou, Tianyan Gao, Eun Y. Lee, Heidi L. Weiss, Timothy S. Heuer, George Kemble, B. Mark Evers. Evaluation of small-molecule FASN inhibitors in preclinical models of colorectal 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 1010.

Abstract 4693: Mechanism-based combination strategies for BET inhibitors in NUT midline carcinoma

Abstract NUT midline carcinoma (NMC) is a highly aggressive squamous cell cancer that responds poorly to standard chemotherapuetic approaches. NMC is characterized by translocations involving the NUT (nuclear protein in testes) protein, which in a majority of cases is fused to the BET (bromodomain and extra-terminal) protein family members BRD3 or BRD4. BET proteins (BRD2, BRD3, BRD4, BRDT) are epigenetic readers that modulate expression of genes involved in cell growth and oncogenesis. Selective small molecule inhibitors of BET proteins, such as the GSK I-BETs (I-BET762, I-BET151), abrogate binding of BET proteins to acetylated chromatin and inhibit the expression of BET target genes. Here we describe the activity in I-BET762 and other BET inhibitors in pre-clinical models of NMC. Consistent with previous reports, we observe profound growth inhibition and cytotoxicity in NMC cell lines in vitro, as well as significant tumor growth inhibition or tumor regression in cell line xenografts of NMC. I-BET762 treatment in NMC cell lines results in transcriptional changes affecting MYC and other pathways critical for cancer cell growth. We explore the contribution of these changes to the anti-proliferative effects observed in NMC models, and identify rational combinations to improve upon the efficacy of I-BET762 as a monotherapy. Taken together, our data highlight novel mechanisms through which BET inhibitors impact NMC cell growth and survival, and suggest potential treatment strategies to improve response in this highly aggressive disease. All studies were conducted in accordance with the GSK Policy on the Care, Welfare and Treatment of Laboratory Animals and were reviewed by the Institutional Animal Care and Use Committee either at GSK or by the ethical review process at the institution where the work was performed. Citation Format: Anastasia Wyce, Peter Soden, Daniel J. Felitsky, Jeanne J. Matteo, Susan Korenchuk, Gary Thripp, Kathryn Keenan, Charles F. McHugh, Rab Prinjha, Christopher Carpenter, Nicholas Smithers, Olena Barbash. Mechanism-based combination strategies for BET inhibitors in NUT midline carcinoma. [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 4693.

Abstract 1475: Local immune activation resulting in tumor growth inhibition with MEDI9197 - an intratumorally administered TLR7/8 agonist

Abstract MEDI9197 (formerly 3M-052) is a sustained-release imidazoquinoline toll like receptor (TLR) TLR7/8 agonist designed with a lipid tail that, when injected, is retained within the tumor. This has been shown in pharmacokinetic studies in mice and rats injected with MEDI9197 via the subcutaneous (SC) or intratumoral (IT, mouse only) routes. Stimulation of TLR7 and TLR8 in primary human dendritic cells induces the release of interferon-alpha (IFN-a) from plasmacytoid dendritic cells (pDCs) and interleukin 12 (IL-12) from myeloid dendritic cells (mDCs). Intratumoral administration of MEDI9197 induces a local immune response characterized by upregulation of genes involved in activation of innate and adaptive immunity both from the injected tumor and tumor draining lymph node. Furthermore, flow cytometric analysis of tumor infiltrating lymphocytes (TILs) show increased expression of activation markers, such as CD69, on natural killer (NK) cells and CD8 cytotoxic T cells. This local stimulation of immune cells with MEDI9197 results in tumor growth inhibition as shown in the B16F10 luc syngeneic mouse tumor model using in vivo imaging system (IVIS) equipment. Additionally, combination of MEDI9197 with immune checkpoint inhibitors enhances the efficacy observed in syngeneic mouse tumor models. The data presented shows intratumoral administration of MEDI9197 induces local immune activation leading to tumor growth inhibition in preclinical models of cancer. MEDI9197 is currently being evaluated as a monotherapy for safety and efficacy in human clinical trials. Citation Format: Stefanie Mullins, Iwen Grigsby, Lester I. Harrison, Song Ren, Serguei Soukharev, Lesley Young, James M. Elvecrog, Robert W. Wilkinson, Mark A. Tomai, Ronald Herbst, John P. Vasilakos, Andrew J. Leishman. Local immune activation resulting in tumor growth inhibition with MEDI9197 - an intratumorally administered TLR7/8 agonist. [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 1475.

Multifunctional Fanconi proteins, inflammation and the Fanconi phenotype

Multifunctional Fanconi proteins, inflammation and the Fanconi phenotype

A phase I study of the EZH2 inhibitor tazemetostat in pediatric subjects with relapsed or refractory INI1-negative tumors or synovial sarcoma.

TPS10587Background: Tazemetostat is a potent and selective inhibitor of the histone methyltransferase enhancer of zeste-homolog 2 (EZH2), the catalytic subunit of the polycomb repressive complex 2 (PRC2). EZH2 is responsible for trimethylation of lysine 27 on histone H3, which results in chromatin remodelling and repressed transcription. Aberrant EZH2 activity has been implicated as an oncogenic driver in non-Hodgkin lymphoma and other malignancies. The SWI/SNF complex also remodels chromatin, activates transcription, and acts in opposition to PRC2. Oncogenesis from mutation and/or loss of the SWI/SNF subunit INI1 in cancers such as malignant rhabdoid tumor (MRT) or SMARCA4 loss in ovarian “small cell carcinoma, hypercalcemic type” is associated with anticancer activity of EZH2 inhibition in preclinical models. This has translated into objective responses of promising durability in subjects (subjs) whose solid tumors exhibited either INI1 or SMARCA4 loss in the tazemetostat phase I adult study. Methods: T...

The impact of obesity on outcomes in metastatic melanoma (MM) patients (pts) treated with dabrafenib and trametinib.

9566Background: Obesity is associated with worse outcomes in many malignancies, and metabolic signaling pathways (i.e. IGF1R) promote resistance to MAPK pathway inhibitors in preclinical models. Ho...

A Phase II Basket Study of the Oral TRK Inhibitor LOXO-101 in Adult Subjects with NTRK Fusion-Positive Tumors.

TPS2599Background: The TRK family of neurotrophin receptors, TRKA, TRKB, and TRKC (encoded by NTRK1, NTRK2, and NTRK3 genes, respectively) and their neurotrophin ligands regulate growth, differentiation and survival of neurons. Fusion of the kinase domain of the NTRK genes with various partners are oncogenic, and have been identified across many common tumor types, but generally at a frequency of 1000x selectivity over other kinases. LOXO-101 is the only selective pan-TRK inhibitor in clinical development and has demonstrated tumor inhibition in preclinical models and meaningful responses in an adult phase 1 trial. Methods: The phase II study of LOX...