Purpose The emerging need for rational combination treatment approaches led us to test the concept that co-targeting MEK and CDK4/6 would prove efficacious in KRAS mutant (KRASmt) colorectal cancers, where upregulated CDK4 and hyperphosphorylated retinoblastoma (RB) typify the vast majority of tumors. Experimental Design Initial testing was carried out in the HCT-116 tumor model, which is known to harbor a KRAS mutation. Efficacy studies were then performed with five RB+ patient-derived colorectal xenograft models, genomically diverse with respect to KRAS, BRAF, and PIK3CA mutational status. Tolerance, efficacy, and pharmacodynamic evaluation of target modulation were evaluated in response to daily dosing with either agent alone or concurrent co-administration. Results Synergy was observed in vitro when HCT-116 cells were treated over a broad range of doses of trametinib and palbociclib. Subsequent in vivo evaluation of this model showed a higher degree of antitumor activity resulting from the combination compared to that achievable with single agent treatment. Testing of colorectal patient-derived xenograft (PDX) models further showed that combination of trametinib and palbociclib was well tolerated and resulted in objective responses in all KRASmt models tested. Stasis was observed in a KRAS/BRAF wild type and a BRAFmt model. Conclusions Combination of trametinib and palbociclib was well tolerated and highly efficacious in all three KRAS mutant CRC PDX models tested. Promising preclinical activity seen here supports clinical evaluation of this treatment approach to improve therapeutic outcome for metastatic colorectal cancer patients.

Rationale Basal and diet-induced differences in mesolimbic function, particularly within the nucleus accumbens (NAc), may contribute to human obesity; these differences may be more pronounced in susceptible populations. Objectives We determined whether there are differences in cocaine-induced behavioral plasticity in rats that are susceptible vs. resistant to diet-induced obesity, and basal differences in the striatal neuron function in adult and adolescent obesity-prone and obesity-resistant rats. Methods Susceptible and resistant outbred rats were identified based on “junk-food” diet-induced obesity. Then, the induction and expression of cocaine-induced locomotor sensitization, which is mediated by enhanced striatal function and is associated with increased motivation for rewards and reward-paired cues, were evaluated. Basal differences in mesolimbic function were examined in selectively bred obesity-prone and obesity-resistant rats (P70-80 and P30-40) using both cocaine induced locomotion and whole-cell patch clamping approaches in NAc core medium spiny neurons (MSNs). Results In rats that became obese after eating “junk-food”, the expression of locomotor sensitization was enhanced compared to non-obese rats, with similarly strong responses to 7.5 and 15 mg/kg cocaine. Without diet manipulation, obesity-prone rats were hyper-responsive to the acute locomotor-activating effects of cocaine, and the intrinsic excitability of NAc core MSNs was enhanced by ~60% at positive and negative potentials. These differences were present in adult, but not adolescent rats. Post-synaptic glutamatergic transmission was similar between groups. Conclusions Mesolimbic systems, particularly NAc MSNs, are hyper-responsive in obesity-prone individuals; and interactions between predisposition and experience influence neurobehavioral plasticity in ways that may promote weight gain and hamper weight loss in susceptible rats.

The ineffectiveness of chemotherapy in patients with pancreatic cancer highlights a critical unmet need in pancreatic cancer therapy. Two commonly mutated genes in pancreatic cancer, KRAS and CDKN2A, have an incidence exceeding 90%, supporting investigation of dual targeting of MEK and CDK4/6 as a potential therapeutic strategy for this patient population. An in vitro proliferation synergy screen was conducted to evaluate response of a panel of high passage and patient-derived pancreatic cancer models to the combination of trametinib and palbociclib to inhibit MEK and CDK4/6, respectively. Two adenosquamous carcinoma models, L3.6pl and UM59, stood out for their high synergy response. In vivo studies confirmed that this combination treatment approach was highly effective in subcutaneously implanted L3.6pl and UM59 tumor-bearing animals. Both models were refractory to single agent treatment. Reverse phase protein array analysis of L3.6pl tumors excised from treated animals revealed strong down regulation of cyclooxygenase-2 (COX-2) expression in response to combination treatment. Expression of COX-2 under a CMV-driven promoter and shRNA knockdown of COX-2 both led to resistance to combination treatment. Our findings suggest that COX-2 may be involved in the improved therapeutic outcome seen in some pancreatic tumors that fail to respond to MEK or CDK4/6 inhibitors alone but respond favorably to their combination.

Pancreatic cancer is one of the deadliest forms of cancer, with median 5-year survival rates less than 10%. This disease is recalcitrant to chemotherapeutic approaches and recently approved therapies afford only modest improvements in survival. KRAS is the most commonly mutated gene in pancreatic tumors with an incidence rate exceeding 90%. Despite intensive efforts, KRAS has remained undruggable. Kinases acting both upstream and downstream of RAS continue to be exploited for the development of novel agents to attenuate signaling through this critical oncogene. While EGFR and MEK inhibitor monotherapies have been evaluated in the clinic for the treatment of pancreatic cancer, efficacy has been modest despite their relevance to MAP kinase (MAPK) pathway-altered tumors. Novel therapies are urgently needed to address adaptive signaling mechanisms that diminish the effectiveness of kinase-targeted approaches. MTX-211 is a first-in-class dual inhibitor of PI3K and EGFR kinase with a promising pharmaceutical profile and proven ability to potentiate the effectiveness of MEK inhibitor therapy in KRAS mutant colorectal tumors. The present study was undertaken to extend our evaluation of MTX-211 to include pancreatic cancer model systems. We have found that MTX-211 exhibits low micromolar potency against an extensive panel of primary models of pancreatic cancer and is highly synergistic with the MEK inhibitor trametinib. Preliminary data suggest that this synergy is driven by the ability of MTX-211 to target compensatory transcriptional activation of HER3 that occurs in response to MEK inhibition. A genetically engineered KRAS and p53 mutant (KPC) mouse model was transduced with a lentiviral construct encoding a caspase reporter, which was used to show that the combination of MTX-211 and trametinib elicited a significant increase in apoptosis over single agent controls. Results from an animal study conducted with the KPC model further showed that the combination of MTX-211 and trametinib significantly slowed growth of these aggressive tumors. Employing both KPC and patient-derived xenograft models of pancreatic cancer, efforts are underway to optimize therapeutic outcome in response to MTX-211-based combination treatment regimens. Citation Format: Christy Frankowski-McGregor, Joel Maust, Elizabeth Ziemke, Rachel Mumby, Amy Delaney, Alnawaz Rehemtulla, Christopher Whitehead, Judith S. Sebolt-Leopold. MTX-211, a dual and selective inhibitor of EGFR and PI3 kinase, shows promising activity in combination with MEK inhibition in preclinical models of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2931.

Agents targeting epidermal growth factor receptor (EGFR) have met with limited success in the clinical management of colorectal cancer (CRC). Mutations in KRAS, BRAF, and PIK3CA are important drivers of resistance to EGFR-targeted therapy. Conversely, EGFR-mediated feedback mechanisms serve to mediate resistance to MEK inhibitor-based treatment of CRC by reactivating MAP kinase signaling. Our central hypothesis is that a dual small molecule inhibitor that potently and selectively targets only EGFR and PI3KA, when combined with a MEK inhibitor, will be highly efficacious against subpopulations of BRAF mutant or KRAS mutant colorectal cancers that are dependent upon these kinase molecules to drive tumor progression. Employing a computational modeling approach, we exploited the known binding modes of structurally related ATP binding site inhibitors of EGFR and PI3K to design small molecules that simultaneously inhibit both kinases in a selective manner. To the best of our knowledge, the lead compound MTX-211, whose binding mode is flipped in PI3K compared to EGFR, represents a first in class selective inhibitor of these two critical oncogenic kinases. MTX-211 exhibits a favorable pharmaceutical and selectivity profile, possessing sub- to low nanomolar potency against both targets, >70% oral bioavailability, strong pharmacodynamic modulation of both EGFR and PI3K signaling, and strong in vivo single agent efficacy against multiple BRAFmt and KRASmt colorectal cancer models, as evidenced by T/C values of 29 to 36% after 10-14 days of oral dosing of 50 mg/kg. A significantly higher degree of in vivo activity is seen when MTX-211 is co-administered with the MEK inhibitor trametinib (>400% increase in survival compared to single agent arms), lending support for this polypharmacology approach over triple drug combination strategies. Based on its promising therapeutic profile, MTX-211 is the focus of an ongoing mouse trial of a large panel of patient-derived xenograft BRAFmt and KRASmt CRC models to inform the design of future human clinical trials. Citation Format: Joel D. Maust, Elizabeth K. Ziemke, Christy L. Frankowski-McGregor, Jun Beom Ku, Rachel Mumby, Karin M. Hardiman, Christopher E. Whitehead, Judith S. Sebolt-Leopold. A dual and selective small molecule inhibitor of EGFR and PI3 kinase shows promising preclinical activity against KRAS and BRAF mutant colorectal tumors [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 148. doi:10.1158/1538-7445.AM2017-148