Investigation of atovaquone-induced spatial changes in tumour hypoxia assessed by hypoxia PET/CT in non-small cell lung cancer patients

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

Improved Immunotherapy Efficacy by Vascular Modulation

Several strategies have been developed to modulate the tumour vasculature for cancer therapy including anti-angiogenesis and vascular normalisation. Vasculature modulation results in changes to the tumour microenvironment including oxygenation and immune cell infiltration, therefore lending itself to combination with cancer therapy. The development of immunotherapies has led to significant improvements in cancer treatment. Particularly promising are immune checkpoint blockade and CAR T cell therapies, which use antibodies against negative regulators of T cell activation and T cells reprogrammed to better target tumour antigens, respectively. However, while immunotherapy is successful in some patients, including those with advanced or metastatic cancers, only a subset of patients respond. Therefore, better predictors of patient response and methods to overcome resistance warrant investigation. Poor, or periphery-limited, T cell infiltration in the tumour is associated with poor responses to immunotherapy. Given that (1) lymphocyte recruitment requires leucocyte–endothelial cell adhesion and (2) the vasculature controls tumour oxygenation and plays a pivotal role in T cell infiltration and activation, vessel targeting strategies including anti-angiogenesis and vascular normalisation in combination with immunotherapy are providing possible new strategies to enhance therapy. Here, we review the progress of vessel modulation in enhancing immunotherapy efficacy.

Vessel compression biases red blood cell partitioning at bifurcations in a haematocrit-dependent manner: implications for tumour blood flow

The tumour microenvironment is abnormal and associated with tumour tissue hypoxia, immunosuppression, and poor response to treatment. One important abnormality present in tumours is vessel compression. Vessel decompression has been shown to increase survival rates in animal models via enhanced and more homogeneous oxygenation. However, our knowledge of the biophysical mechanisms linking tumour decompression to improved tumour oxygenation is limited. In this study, we propose a computational model to investigate the impact of vessel compression on red blood cell (RBC) dynamics in tumour vascular networks. Our results demonstrate that vessel compression can alter RBC partitioning at bifurcations in a haematocrit-dependent and flowrate-independent manner. We identify RBC focussing due to cross-streamline migration as the mechanism responsible and characterise the spatiotemporal recovery dynamics controlling downstream partitioning. Based on this knowledge, we formulate a reduced-order model that will help future research to elucidate how these effects propagate at a whole vascular network level. These findings contribute to the mechanistic understanding of haemodilution in tumour vascular networks and oxygen homogenisation following pharmacological solid tumour decompression.

Effects of hypoxic preconditioning on neuroblastoma tumour oxygenation and metabolic signature in a chick embryo model

Hypoxia episodes and areas in tumours have been associated with metastatic dissemination and poor prognosis. Given the link between tumour tissue oxygen levels and cellular metabolic activity, we hypothesised that the metabolic profile between metastatic and non-metastatic tumours would reveal potential new biomarkers and signalling cues. We have used a previously established chick embryo model for neuroblastoma growth and metastasis, where the metastatic phenotype can be controlled by neuroblastoma cell hypoxic preconditioning (3 days at 1% O2). We measured, with fibre-optic oxygen sensors, the effects of the hypoxic preconditioning on the tumour oxygenation, within tumours formed by SK-N-AS cells on the chorioallantoic membrane (CAM) of chick embryos. We found that the difference between the metastatic and non-metastatic intratumoural oxygen levels was small (0.35% O2), with a mean below 1.5% O2 for most tumours. The metabolomic profiling, using NMR spectroscopy, of neuroblastoma cells cultured in normoxia or hypoxia for 3 days, and of the tumours formed by these cells showed that the effects of hypoxia in vitro did not compare with in vivo tumours. One notable difference was the high levels of the glycolytic end-products triggered by hypoxia in vitro, but not by hypoxia preconditioning in tumours, likely due to the very high basal levels of these metabolites in tumours compared with cells. In conclusion, we have identified high levels of ketones (3-hydroxybutyrate), lactate and phosphocholine in hypoxic preconditioned tumours, all known to fuel tumour growth, and we herein point to the poor relevance of in vitro metabolomic experiments for cancer research.