MR imaging and spectroscopy reveals metabolic changes in glioblastoma driven by treatment with temozolomide and lonidamine

Resistance to temozolomide (TMZ) limits the efficacy of treatment in glioblastoma (GBM) which is fuelled by a metabolic reprogramming of the cancer cells and alterations in the tumour microenvironment. Lonidamine (LND), a glycolytic and mitochondrial inhibitor, modulates tumour metabolism and can alter tumour response to treatment. This study investigated the effects of LND in combination with TMZ and used MR imaging and spectroscopy to evaluate metabolic and microstructural changes in a syngeneic GBM model.

C57BL/6 mice bearing intracranial GL261 tumours (n = 66) were treated with saline, LND, TMZ, or LND + TMZ for five consecutive days. Longitudinal imaging (days 0, 3, 6) included T2-weighted MRI, diffusion weighted imaging (DWI), and conventional and lactate-edited ¹H-MRS. Tumour volume, survival, DWI parameters, MCT4 expression, and mitotic indices were assessed.

In vitro, acute and chronic bioenergetic reprogramming of GL261 cells was assessed using the Seahorse extracellular flux analysis. Lactate is a not simply an end product of high glycolytic cancer cells, but it has also emerged as a molecule capable to shaping the tumour microenvironment. A significant decrease in lactate observed in the LND and LND + TMZ groups indicated a reduction in the steady-state lactate pool suggestive of reduced lactate production, utilization or altered glycolytic flux.

However, lactate remained high in the control and TMZ-only groups. This metabolic shift in the LND treatment group was consistent with a marked reduction in MCT4 protein, the lactate transporter, in LND-treated tumours. In vitro bioenergetic profiling of GL261 cells revealed an acute decrease in mitochondrial respiration in response to LND, with partial recovery at later time points.

Finally, even though not significant, an enhanced survival benefit of the treatment combination was observed, with no significant differences observed between the groups for tumour volume, DWI-derived parameters, or mitotic index after 6 days. LND induces early metabolic changes detectable by MRS. Although a limited therapeutic benefit of addition LND to TMZ was observed in the early stages of tumour development, its tumour-selective activity and effect on tumour microenvironment is promising and its long-term effects on tumour development should be investigated as part of an optimized combination strategy for TMZ treatment.

The authors acknowledge the use of the Centre for Preclinical Imaging (CPI) and Biobank facilities provided by Liverpool Shared Research Facilities (LIVSRF), University of Liverpool. We also acknowledge the Histology Laboratory, Leahurst Campus, University of Liverpool. Tareq Alrashidi has been funded by a PhD studentship grant from the Ministry of Education, Saudi Arabia.