Acquired epilepsy (i.e., after an insult to the brain) is often considered to be a progressive disorder, and the nature of this hypothetical progression remains controversial. Antiepileptic drug treatment necessarily confounds analyses of progressive changes in human patients with acquired epilepsy. Here, we describe experiments testing the hypothesis that development of acquired epilepsy begins as a continuous process of increased seizure frequency (i.e., proportional to probability of a spontaneous seizure) that ultimately plateaus. Using nearly continuous surface cortical and bilateral hippocampal recordings with radiotelemetry and semiautomated seizure detection, the frequency of electrographically recorded seizures (both convulsive and nonconvulsive) was analyzed quantitatively for ϳ100 d after kainate-induced status epilepticus in adult rats. The frequency of spontaneous recurrent seizures was not a step function of time (as implied by the "latent period"); rather, seizure frequency increased as a sigmoid function of time. The distribution of interseizure intervals was nonrandom, suggesting that seizure clusters (i.e., short interseizure intervals) obscured the early stages of progression, and may have contributed to the increase in seizure frequency. These data suggest that (1) the latent period is the first of many long interseizure intervals and a poor measure of the time frame of epileptogenesis, (2) epileptogenesis is a continuous process that extends much beyond the first spontaneous recurrent seizure, (3) uneven seizure clustering contributes to the variability in occurrence of epileptic seizures, and (4) the window for antiepileptogenic therapies aimed at suppressing acquired epilepsy probably extends well past the first clinical seizure.

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.

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Purpose Subregional hypoxia is a common feature of tumors and is recognized as a limiting factor for the success of radiotherapy and chemotherapy. TH-302, a hypoxia-activated prodrug selectively targeting hypoxic regions of solid tumors, delivers a cytotoxic warhead to the tumor, while maintaining relatively low systemic toxicity. The antitumor activity, different dosing sequences, and dosing regimens of TH-302 in combination with commonly used conventional chemotherapeutics were investigated in human tumor xenograft models. Methods Seven chemotherapeutic drugs (docetaxel, cisplatin, pemetrexed, irinotecan, doxorubicin, gemcitabine, and temozolomide) were tested in combination with TH-302 in eleven human xenograft models, including non-small cell lung cancer (NSCLC), colon cancer, prostate cancer, fibrosarcoma, melanoma, and pancreatic cancer. Results The antitumor activity of docetaxel, cisplatin, pemetrexed, irinotecan, doxorubicin, gemcitabine, and temozolomide was increased when combined with TH-302 in nine out of eleven models tested. Administration of TH-302 2–8 h prior to the other chemotherapeutics yielded superior efficacy versus other sequences tested. Simultaneous administration of TH-302 and chemotherapeutics increased toxicity versus schedules with dosing separations. In a dosing optimization study, TH-302 administered daily at 50 mg/kg intraperitoneally for 5 days per week in the H460 NSCLC model showed the optimal response with minimal toxicity. Conclusions TH-302 enhances the activity of a wide range of conventional anti-neoplastic agents in a broad panel of in vivo xenograft models. These data highlight in vivo effects of schedule and order of drug administration in regimen efficacy and toxicity and have relevance to the design of human regimens incorporating TH-302.

These data suggest that animal models with spontaneous seizures, such as kainate- and pilocarpine-treated rats, can be used efficiently for rapid testing of AEDs with a repeated-measures, crossover protocol. Furthermore, the results indicate that this design allows both dose-effect and time-course-of-recovery studies.