MR Imaging Biomarkers to Monitor Early Response to Hypoxia-Activated Prodrug TH-302 in Pancreatic Cancer Xenografts

Abstract

TH-302 is a hypoxia-activated prodrug known to activate selectively under the hypoxic conditions commonly found in solid tumors. It is currently being evaluated in clinical trials, including two trials in Pancreatic Ductal Adenocarcinomas (PDAC). The current study was undertaken to evaluate imaging biomarkers for prediction and response monitoring of TH-302 efficacy in xenograft models of PDAC. Dynamic contrast-enhanced (DCE) and diffusion weighted (DW) magnetic resonance imaging (MRI) were used to monitor acute effects on tumor vasculature and cellularity, respectively. Three human PDAC xenografts with known differential responses to TH-302 were imaged prior to, and at 24 h and 48 hours following a single dose of TH-302 or vehicle to determine if imaging changes presaged changes in tumor volumes. DW-MRI was performed at five b-values to generate apparent diffusion coefficient of water (ADC) maps. For DCE-MRI, a standard clinically available contrast reagent, Gd-DTPA, was used to determine blood flow into the tumor region of interest. TH-302 induced a dramatic decrease in the DCE transfer constant (Ktrans) within 48 hours after treatment in the sensitive tumors, Hs766t and Mia PaCa-2, whereas TH-302 had no effect on the perfusion behavior of resistant SU.86.86 tumors. Tumor cellularity, estimated from ADC, was significantly increased 24 and 48 hours after treatment in Hs766t, but was not observed in the Mia PaCa-2 and SU.86.86 groups. Notably, growth inhibition of Hs766t was observed immediately (day 3) following initiation of treatment, but was not observed in MiaPaCa-2 tumors until 8 days after initiation of treatment. Based on these preclinical findings, DCE-MRI measures of vascular perfusion dynamics and ADC measures of cell density are suggested as potential TH-302 response biomarkers in clinical trials.

Fig 1. Representative K^trans maps for HS766t, Mia PaCa-2 and Su.86.86 tumor type from DCE-MRI (L-R).

As demonstrated in the histogram, K^trans generated from HS766t and Mia-PaCa-2 tumors were dramatically decreased 48 hours after TH-302 treatment compared to pre-treatment values. There were no significant changes observed for a SU.86.86 mouse at the same time point.

Fig 2. Time-course imaging result of DCE-MRI and DW-MRI.

A) Time-courses of normalized mean K^trans values. The mean values of normalized K^trans decreased 69% for TH-302 treated mice in Hs766t tumors, decreased 46% for Mia PaCa-2 tumors and increased 5% in SU.86.86 tumors. B) Changes in normalized mean tumor ADC values over time. A substantial increase in relative mean ADCs was observed for the TH-302 treated group at post- 24 and 48 hours (29% increasing for 24h, p<0.01; 17% increasing for 48h, p<0.01). MIA PaCa-2 is not statistically significant different between conducted groups (8% increasing for 24h, P>0.05; 4% increasing for 48h, p>0.05). For SU.86.86, no significant change was detected by DW-MRI for both TH-302 and control group (3% decreasing for 24h, p>0.05; 0.5% increasing for 48h, p>0.05). The normalization was calculated by the average of the difference of pre- and post-treatment in Th302 group relative to average of the difference of pre- and post-treatment in control group. Error bars stand for standard deviation.

Fig 3. ADC maps and histograms of DW-MRI from representative animals at different time points pre- and post-treatment initiation.

A dramatically increased ADCs observed in HS766T, but not in the other two tumor types.

Fig 4

(A) Histological staining of γ-H2AX, a marker for DNA damage response mechanisms, in PDAC tumors pre- and 48hr post-TH-302 treatment (50 mg/kg). Expression of γ -H2AX was increased in TH-302 treated Hs766t and Mia PaCa-2 tumors when compared to saline control. No detectable change in γ-H2AX staining was observed between Su.86.86 treated and saline tumors. (B) Pimonidazole staining as biomarker of physical tumor hypoxia. Pimonidazole Hydrochloride was injected 2hr prior to tumor removal. The extent of hypoxia was greatest in Hs766t tumors and least in SU.86.86 with Mia PaCa-2 tumors moderately hypoxic. These images are representative images of each tumor type. Scale bars = 200 μM.