Abstract 1019: Cytotoxicity in vitro, pharmacokinetics and tissue distribution of Link-N3 and Link-F3, two bivalent small molecules inhibitors of c-Myc/Max interactions in C.B-17 SCID mice bearing Daudi Burkitt's lymphoma xenografts.
Authors: Guo Jianxia , Clausen Dana M. , Prochownik Edward V. , Metallo Steven J. , Parise Robert A. , Beumer Jan H. , Eiseman Julie L. 
Published: August 2014

Abstract
Introduction: c-Myc plays an important role in cellular proliferation, differentiation, apoptosis and cell cycle progression and its overexpression is associated with aggressiveness and poor prognosis in many cancers. In order to be transcriptionally active, c-Myc must heterodimerize via its bHLH-ZIP domain with its obligatory partner Max. Two small molecules that prevent c-Myc/Max heterodimerization and inhibit c-Myc function are 10074-G5 and 10058-F4. Analogues of these small molecule c-Myc inhibitors have been linked together by a flexible bridge to form Link-N3 and Link-F3. We have characterized the ability of these Link analogues to inhibit the in vitro growth of a c-Myc-overexpressing human cell line, Daudi Burkitt's lymphoma. We evaluated their pharmacokinetics, metabolism and efficacy in mice bearing Daudi xenografts.
Methods: Inhibition of Daudi cell growth by Link-N3 and Link-F3 was assessed by MTT assay after 72 h of incubation. Accumulation of Link-N3 and Link-F3 by Daudi cells incubated with 10 μM of Link-N3 and Link-F3 was measured by HPLC. For pharmacokinetic and metabolic studies, C.B-17 SCID mice bearing Daudi xenografts were treated with 10 mg/kg Link-N3 or 5mg/kg Link-F3 IV. Plasma and tissues were collected between 5 and 1440 min. Urine was collected at 360 and 1440 min. Concentrations of Link-N3 and Link-F3 were determined by HPLC-UV, and metabolites were identified by LC-MS/MS. For efficacy compounds were administered qdx4 at the same doses.
Results: The IC50’s for Link-N3 and Link-F3 against Daudi cells could not be determined because compounds precipitated at concentrations above 30 μM. Daudi cells accumulated Link-N3 and Link-F3 ∼19-fold and ∼17-fold, respectively, with peak intracellular concentrations between 3 and 6 h of incubation. Neither compound resulted in tumor growth inhibition following IV dosing. Following a single IV dose of 10 mg/kg Link-N3 or 5 mg/kg Link-F3, peak plasma concentrations at 5 min were 27.3 μg/ml and 5.3 μg/ml, respectively. Plasma Auc0-t for Link-N3 and Link-F3 were 1759.6 μg·min/ml and 137.9 μg·min/ml, respectively. Plasma T1/2 of Link-N3 and Link-F3 were 394 and 7 min, respectively. The volume of distribution was approximately 3204 ml/kg and 35 ml/kg for Link-N3 and Link-F3, respectively. Clearances for Link-N3 and Link-F3 were 5.6 ml/min/kg and 3.6 ml/min/kg, respectively. Peak tumor concentration of Link-N3 at 30 min was 0.77 μg/g and Link-F3 at 5 min was 0.49 μg/g, both much lower than peak plasma concentrations.
Conclusion: The lack of significant antitumor activity of Link-N3 or Link-F3 in tumor-bearing mice is related to the low concentrations that reached the xenografts. Modification will be required in these bivalent compounds to make them more drug-like.
Support: 1R01CA142580 and P30-CA47904
Citation Format: Jianxia Guo, Dana M. Clausen, Edward V. Prochownik, Steven J. Metallo, Robert A. Parise, Jan H. Beumer, Julie L. Eiseman. Cytotoxicity in vitro, pharmacokinetics and tissue distribution of Link-N3 and Link-F3, two bivalent small molecules inhibitors of c-Myc/Max interactions in C.B-17 SCID mice bearing Daudi Burkitt's lymphoma xenografts. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1019. doi:10.1158/1538-7445.AM2013-1019