18 F-HX4 Hypoxia PET Holds Promise as a Prognostic and Predictive Imaging Biomarker in a Lung Cancer Xenograft Model Treated with Metformin and Radiotherapy.

Metformin may improve tumor oxygenation and thus radiotherapy response, but imaging biomarkers for selection of suitable patients are still under investigation. First, we assessed the effect of acute metformin administration on non-small cell lung cancer (NSCLC) xenograft tumor hypoxia using positron emission tomography (PET) imaging with the hypoxia tracer 18 F-flortanidazole (18 F-HX4). Second, we verified the effect of a single dose of metformin prior to radiotherapy on long-term treatment outcome. Third, we examined the potential of baseline 18 F-HX4 as a prognostic and/or predictive biomarker for treatment response. Methods: A549 tumor-bearing mice underwent a 18 F-HX4 PET/computed tomography (CT) scan to determine baseline tumor hypoxia. The next day, mice received an intravenous (IV) injection of 100 mg/kg metformin. 18 F-HX4 was administered IV 30 min later and a second PET/CT scan was performed to assess changes in tumor hypoxia. Two days later, mice were divided into three therapy groups: controls (group 1), radiotherapy (group 2), and metformin+radiotherapy (group 3). Animals received saline (groups 1-2) or 100 mg/kg metformin (group 3) IV, followed by a single radiotherapy dose of 10 Gy (groups 2-3) or sham irradiation (group 1) 30 min later. Tumor growth was monitored triweekly by caliper measurement and relative tumor volumes (RTV=Vtime x/Vbaseline) were calculated. The tumor doubling time (TDT), i.e. the time to reach 2× the pre-irradiation tumor volume, was defined as the endpoint. Results: Thirty min post-metformin treatment, 18 F-HX4 demonstrated a significant change in tumor hypoxia with a mean intratumoral reduction in 18 F-HX4 tumor-to-background ratio (TBR) from 3.21±0.13 to 2.87±0.13 ( P = 0.0001). Overall, RTV over time differed across treatment groups (p<0.0001). Similarly, the median TDT was 19, 34 and 52 days in controls, the radiotherapy-group and the metformin+radiotherapy-group, respectively (log-rank p<0.0001). Both baseline 18 F-HX4 TBR (HR 2.0; P = 0.0004) and change from baseline TBR (HR 0.39; P = 0.04) were prognostic biomarkers for TDT irrespective of treatment, and baseline TBR predicted metformin-specific treatment effects which were dependent on baseline tumor hypoxia. Conclusion: Using 18 F-HX4 PET imaging in a NSCLC xenograft model, we showed that metformin may act as radiosensitizer by increasing tumor oxygenation and that baseline 18 F-HX4 shows promise as an imaging biomarker.