Kimberly Elizabeth Fultz

Wild-Type APC Regulates Caveolin-1 Expression in Human Colon Adenocarcinoma Cell Lines Via FOXO1a and C-myc

Genetic evidence suggests that caveolin‐1, an essential component of membrane caveolae, acts as a tumor promoter in some, and a tumor suppressor in other cancers. The role of caveolin‐1 in colon carcinogenesis is controversial. We report here, for the first time, that caveolin‐1 is transcriptionally induced in colon cancer cells in response to conditional expression of a full length adenomatous polyposis coli (APC) gene. This induction of caveolin‐1 by APC is mediated by both FOXO1a, a member of the Forkhead family of transcription factor, and c‐myc. The FOXO1a protein, which is increased by wild‐type APC expression, induces caveolin‐1 promoter–reporter activity and binds directly to a FKHR consensus binding sequence in the caveolin‐1 promoter. The c‐myc protein, which is reduced in the presence of wild‐type APC, acts to repress caveolin‐1 expression by acting at non‐E‐box containing elements in the caveolin‐1 promoter. These data predict that caveolin‐1 protein expression would be decreased early in colonic carcinogenesis, which is associated with loss of wild‐type APC. Our results would be consistent with the interpretation that caveolin‐1 may have tumor suppressing functions during early stages of colon carcinogenesis.

Discovery and SAR exploration of a novel series of imidazo[4,5-b]pyrazin-2-ones as potent and selective mTOR kinase inhibitors.

We report here the discovery of a novel series of selective mTOR kinase inhibitors. A series of imidazo[4,5-b]pyrazin-2-ones, represented by screening hit 1, was developed into lead compounds with excellent mTOR potency and exquisite kinase selectivity. Potent compounds from this series show >1000-fold selectivity over the related PI3Kα lipid kinase. Further, compounds such as 2 achieve mTOR pathway inhibition, blocking both mTORC1 and mTORC2 signaling, in PC3 cancer cells as measured by inhibition of pS6 and pAkt (S473).

CC-223, a Potent and Selective Inhibitor of mTOR Kinase: In Vitro and In Vivo Characterization.

mTOR is a serine/threonine kinase that regulates cell growth, metabolism, proliferation, and survival. mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2) are critical mediators of the PI3K–AKT pathway, which is frequently mutated in many cancers, leading to hyperactivation of mTOR signaling. Although rapamycin analogues, allosteric inhibitors that target only the mTORC1 complex, have shown some clinical activity, it is hypothesized that mTOR kinase inhibitors, blocking both mTORC1 and mTORC2 signaling, will have expanded therapeutic potential. Here, we describe the preclinical characterization of CC-223. CC-223 is a potent, selective, and orally bioavailable inhibitor of mTOR kinase, demonstrating inhibition of mTORC1 (pS6RP and p4EBP1) and mTORC2 [pAKT(S473)] in cellular systems. Growth inhibitory activity was demonstrated in hematologic and solid tumor cell lines. mTOR kinase inhibition in cells, by CC-223, resulted in more complete inhibition of the mTOR pathway biomarkers and improved antiproliferative activity as compared with rapamycin. Growth inhibitory activity and apoptosis was demonstrated in a panel of hematologic cancer cell lines. Correlative analysis revealed that IRF4 expression level associates with resistance, whereas mTOR pathway activation seems to associate with sensitivity. Treatment with CC-223 afforded in vivo tumor biomarker inhibition in tumor-bearing mice, after a single oral dose. CC-223 exhibited dose-dependent tumor growth inhibition in multiple solid tumor xenografts. Significant inhibition of mTOR pathway markers pS6RP and pAKT in CC-223–treated tumors suggests that the observed antitumor activity of CC-223 was mediated through inhibition of both mTORC1 and mTORC2. CC-223 is currently in phase I clinical trials. Mol Cancer Ther; 14(6); 1295–305. ©2015 AACR.

Optimization of a Series of Triazole Containing Mammalian Target of Rapamycin (mTOR) Kinase Inhibitors and the Discovery of CC-115

We report here the synthesis and structure-activity relationship (SAR) of a novel series of triazole containing mammalian target of rapamycin (mTOR) kinase inhibitors. SAR studies examining the potency, selectivity, and PK parameters for a series of triazole containing 4,6- or 1,7-disubstituted-3,4-dihydropyrazino[2,3-b]pyrazine-2(1H)-ones resulted in the identification of triazole containing mTOR kinase inhibitors with improved PK properties. Potent compounds from this series were found to block both mTORC1(pS6) and mTORC2(pAktS473) signaling in PC-3 cancer cells, in vitro and in vivo. When assessed in efficacy models, analogs exhibited dose-dependent efficacy in tumor xenograft models. This work resulted in the selection of CC-115 for clinical development.

Use of core modification in the discovery of CC214-2, an orally available, selective inhibitor of mTOR kinase

We report here the discovery of a novel series of selective mTOR kinase inhibitors and the identification of CC214-2, a compound with demonstrated anti-tumor activity upon oral dosing in a PC3 prostate cancer xenograft model. A series of 4,6-disubstituted-3,4-dihydropyrazino[2,3-b]pyrazine-2(1H)-ones were discovered through a core modification of our original compound series. Analogs from this series have excellent mTOR potency and maintain selectivity over the related PI3Kα lipid kinase. Compounds such as CC214-2 were found to block both mTORC1(pS6) and mTORC2(pAktS473) signaling in PC3 cancer cells, in vitro and in vivo.

Discovery of Mammalian Target of Rapamycin (mTOR) Kinase Inhibitor CC-223

We report here the synthesis and structure-activity relationship (SAR) of a novel series of mammalian target of rapamycin (mTOR) kinase inhibitors. A series of 4,6- or 1,7-disubstituted-3,4-dihydropyrazino[2,3-b]pyrazine-2(1H)-ones were optimized for in vivo efficacy. These efforts resulted in the identification of compounds with excellent mTOR kinase inhibitory potency, with exquisite kinase selectivity over the related lipid kinase PI3K. The improved PK properties of this series allowed for exploration of in vivo efficacy and ultimately the selection of CC-223 for clinical development.

Abstract A165: Antitumor activity of mTOR kinase inhibitor CC-223 in a mouse model of prostate cancer.

CC-223 is an orally active selective mammalian target of rapamycin (mTOR) kinase inhibitor which is currently in clinical development. In biochemical assays CC-223 was highly selective to mTOR over 249 kinases. In cellular assays CC-223 inhibited both mTORC1 and mTORC2 as evidenced by inhibition of pS6RP(Ser 235/236) and pAkt(Ser 473). The current study was aimed to determine the pharmacokinetic/pharmacodynamic (PK/PD) relationship, antitumor activity, and mechanism of action of CC-223 in mice with PC3 human prostate cancer tumors. In PK/PD studies with PC3 tumor-bearing mice, CC-223 exhibited a dose-dependent inhibition mTOR pathway biomarkers, pS6RP, and pAkt, which correlated with plasma exposure. Antitumor activity of CC-223 was tested in mice with PC3 tumors of approximately 100-150 mm3 in size. CC-223 consistently exhibited dose-dependent tumor growth inhibition. The minimum dose required to obtain >65% tumor volume reduction compared with vehicle control was 5 mg/kg BID. In addition to showing antitumor activity in smaller tumors, CC-223 also caused the regression or growth delay of larger PC3 tumors. At 10 mg/kg BID, CC-223 caused the regression of tumors, whereas 5 and 1 mg/kg BID slowed tumor growth compared to the vehicle control. The mechanism of action of CC-223 on PC3 tumors was characterized by analyzing tumors for functional markers (proliferation, apoptosis, and blood vessel markers). The number of proliferating cells in CC-223-treated tumors was significantly reduced compared to vehicle control. A significant increase in the number of apoptotic cells and a significant decrease in blood vessel density was observed in CC-223-treated tumors. In conclusion, treatment with the mTOR kinase inhibitor CC-223 significantly inhibited PC3 prostate tumor growth. Significant inhibition of mTOR pathway markers pS6RP and pAkt in CC-223-treated tumors suggests that the observed antitumor activity of CC-223 was mediated through inhibition of both mTORC1 and mTORC2. Maintenance of >80% and >60% biomarker inhibition for pS6 and pAkt, respectively, and plasma levels greater than 0.2 μM through 8 hours twice daily confers good antitumor efficacy in PC3 tumors. Further immunohistochemical data demonstrated that the antitumor activity of CC-223 was not only due to the inhibition of tumor cell proliferation but also due to the increase in apoptosis and antiangiogenic activity of CC-223. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A165. Citation Format: Rama Krishna Narla, Sophie Peng, Jim Gamez, Jason Katz, Julius Apuy, Mehran Moghaddam, Kimberly E. Fultz, Sabita Sankar, Deborah S. Mortensen, Heather K. Raymon. Antitumor activity of mTOR kinase inhibitor CC-223 in a mouse model of prostate cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A165.