Selective Tumor Hypoxia Targeting by Hypoxia-Activated Prodrug TH-302 Inhibits Tumor Growth in Preclinical Models of Cancer

Abstract

Purpose: Tumor hypoxia underlies treatment failure and yields a more aggressive, invasive, and metastatic cancer phenotype. TH-302 is a 2-nitroimidazole triggered hypoxia-activated prodrug of the cytotoxin bromo-isophosphoramide mustard (Br-IPM). The purpose of this study is to characterize the antitumor activity of TH-302 and investigate its selective targeting of the hypoxic cells in human tumor xenograft models.Experimental Design: Antitumor efficacy was assessed by tumor growth kinetics or by clonogenic survival of isolated cells after tumor excision. Hypoxic fractions (HF) were determined by immunohistochemistry and morphometrics of pimonidazole staining. Tumor hypoxia levels were manipulated by exposing animals to different oxygen concentration breathing conditions. The localization and kinetics of TH-302 induced DNA damage was determined by gH2AX immunohistochemistry.Results: TH-302 antitumor activity was dose-dependent and correlated with total drug exposure. Correlation was found between antitumor activity and tumor HF across 11 xenograft models. Tumorbearing animals breathing 95% O 2 exhibited attenuated TH-302 efficacy, with whereas those breathing 10% O 2 exhibited enhanced TH-302 efficacy, both compared with air (21% O 2 ) breathing. TH-302 treatment resulted in a reduction in the volume of the HF 48 hours after dosing and a corresponding increase in the necrotic fraction. TH-302 induced DNA damage as measured by gH2AX was initially only present in the hypoxic regions and then radiated to the entire tumor in a time-dependent manner, consistent with TH-302 having a "bystander effect."Conclusions: The results show that TH-302 has broad antitumor activity and selectively targets hypoxic tumor tissues. Clin Cancer Res; 18(3); 758-70. Ó2011 AACR.

Activity of the hypoxia‐activated pro‐drug TH‐302 in hypoxic and perivascular regions of solid tumors and its potential to enhance therapeutic effects of chemotherapy

“…These authors showed that induction of gH2aX was located preferentially near pimonidazole positive cells. Our results support the findings of Sun et al 11 with respect to gH2aX expression near hypoxic regions (here identified by EF5); however, our evaluation of gH2aX and other biomarkers (cleaved caspase -3 or -6 and reduction of Ki-67) in regions close to functional blood vessels identified by DiOC7 revealed that TH-302 can also change the expression of these biomarkers in well-oxygenated regions of tumors. We have evaluated previously biomarker distributions following treatment with another nitrogen mustard melphalan, and found increased gH2aX, cleaved caspase and reduced Ki-67 expression close to blood vessels.…”

“…The ability of TH-302 to induce expression of gH2aX has also been reported by Sun et al, 11 who administered TH-302 to mice as well as pimonidazole (to detect hypoxia) given an hour before animal death. These authors showed that induction of gH2aX was located preferentially near pimonidazole positive cells.…”

“…15,23 TH-302 is a pro-drug that is activated specifically in hypoxic cells, and has demonstrated promising results both in preclinical studies and in early clinical trials. [11][12][13]25,26 However, since TH-302 is non-fluorescent, its direct visualization in tumor tissue is not possible. Therefore, we extended our techniques of assessing biomarker distribution following drug treatment to evaluate the effects of TH-302 in hypoxic and vascular regions of solid tumors.…”

“…11,[16][17][18] In most experiments evaluating drug combinations, TH-302 was administered 4 hr before doxorubicin or docetaxel, but we also evaluated gH2aX expression following concurrent delivery of TH-302 and doxorubicin in MCF-7 tumors; this was done to test observations of a previous study that TH-302 administration 4 hr before chemotherapy was superior to concurrent administration. 19 Mice were killed and tumors excised from saline (control) and treated animals at 10 min and 24 hr following drug injection; EF5 and DiOC7 were administered 2 hr and 1 min, respectively, before animal death.…”

“…TH-302 is a 2-nitroimidazole containing a toxic alkylating moiety that is released only when its attached nitro-heterocyclic trigger molecule fragments in hypoxic cells to release the DNA crosslinking agent bromo-isophosphoramide mustard. 11 Traditional chemotherapy combined with TH-302 can lead to greater cell kill and growth delay of various rodent tumors and human xenografts, 12 and promising results have been reported from early clinical trials evaluating TH-302 combined with gemcitabine for treatment of pancreatic carcinoma. 13 However, there have been few studies evaluating the microenvironmental distribution of activity of TH-302 within solid tumors and the mechanisms which lead to antitumor effects remain uncertain.…”