Positron emission tomography (PET)/computed tomography (CT) using the radiotracer 18F-Fluoromisonidazole (FMISO) has been widely employed to image tumour hypoxia and is of interest to help develop novel hypoxia modifiers and guide radiation treatment planning. Yet, the optimal post-injection (p.i.) timing of hypoxic imaging remains questionable. Therefore, we investigated the correlation between hypoxia-related quantitative values in FMISO-PET acquired at 2 and 4 h p.i. in patients with non-small cell lung cancer (NSCLC). Patients with resectable NSCLC participated in the ATOM clinical trial (NCT02628080) which investigated the hypoxia modifying effects of atovaquone. Two-hour and four-hour FMISO PET/CT images acquired at baseline and pre-surgery visits (n = 58) were compared. Cohort 1 (n = 14) received atovaquone treatment, while cohort 2 (n = 15) did not. Spearman’s rank correlation coefficients (ρ) assessed the relationship between hypoxia-related metrics, including standardised uptake value (SUV), tumour-to-blood ratio (TBR), and tumour hypoxic volume (HV) defined by voxels with TBR ≥ 1.4. As the primary imaging-related trial endpoint used to evaluate the action of atovaquone on tumour hypoxia in patients with NSCLC was change in tumour HV from baseline, this was also assessed in patients (n = 20) with sufficient baseline 2- and 4-h scan HV to reliably measure change (predefined as ≥ 1.5 mL). Tumours were divided into four subregions or distance categories: edge, outer, inner, and centre, using MATLAB. In tumours overall, strong correlation (P < 0.001) was observed for SUV max ρ = 0.87, SUV mean ρ = 0.91, TBR max ρ = 0.83 and TBR mean ρ = 0.81 between 2- and 4-h scans. Tumour HV was moderately correlated (P < 0.001) with ρ = 0.69 between 2- and 4-h scans. Yet, in tumour subregions, the correlation of HV decreased from the centre ρ = 0.71 to the edge ρ = 0.45 (P < 0.001). SUV, TBR, and HV values were consistently higher on 4-h scans than on 2-h scans, indicating better tracer-to-background contrast. For instance, for TBR max , the mean, median, and interquartile range were 1.9, 1.7, and 1.6–2.0 2-h p.i., and 2.6, 2.4, and 2.0–3.0 4-h p.i., respectively. Our results support that FMISO-PET scans should be performed at 4 h p.i. to evaluate tumour hypoxia in NSCLC. Trial registration: ClinicalTrials.gov, NCT02628080. Registered 11/12/2015, https://clinicaltrials.gov/ct2/show/NCT02628080 .

Background Tumour hypoxia promotes an aggressive tumour phenotype and enhances resistance to anticancer treatments. Following the recent observation that the mitochondrial inhibitor atovaquone increases tumour oxygenation in NSCLC, we sought to assess whether atovaquone affects tumour subregions differently depending on their level of hypoxia. Methods Patients with resectable NSCLC participated in the ATOM trial (NCT02628080). Cohort 1 ( n = 15) received atovaquone treatment, whilst cohort 2 ( n = 15) did not. Hypoxia-related metrics, including change in mean tumour-to-blood ratio, tumour hypoxic volume, and fraction of hypoxic voxels, were assessed using hypoxia PET imaging. Tumours were divided into four subregions or distance categories: edge, outer, inner, and centre, using MATLAB. Results Atovaquone-induced reduction in tumour hypoxia mostly occurred in the inner and outer tumour subregions, and to a lesser extent in the centre subregion. Atovaquone did not seem to act in the edge subregion, which was the only tumour subregion that was non-hypoxic at baseline. Notably, the most intensely hypoxic tumour voxels, and therefore the most radiobiologically resistant areas, were subject to the most pronounced decrease in hypoxia in the different subregions. Conclusions This study provides insights into the action of atovaquone in tumour subregions that help to better understand its role as a novel tumour radiosensitiser. Trial registration : ClinicalTrials.gov, NCT0262808. Registered 11th December 2015, https://clinicaltrials.gov/ct2/show/NCT02628080