Prostate cancer, when treated with external beam radiotherapy (RT) in the range of 78 Gy, is frequently associated with gastrointestinal (GI) & genitourinary (GU) toxicities. We hypothesize that tumor sensitization by lonidamine (LND) will enable the use of lower RT doses reducing the risk of side effects. LND effects detected in vivo by 31P and 1H MRS in androgen-independent (PC3) prostate cancer xenografts produced a sustained and tumor-selective decrease in intracellular pH, bioenergetics (βNTP/Pi), oxygen consumption rate and increase in lactate. Selective tumor acidification, deenergization and oxygenation induced by LND potentiated the radiation response in the PC3 prostate cancer model.
PC3, hormone-independent, prostate cancer cells were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum, 2mM L-glutamine, and 1% penicillin-streptomycin. 7×106 PC3 cells were inoculated subcutaneously in each mouse (n=5) as a 0.1 mL suspension.
31P and 1H MRS measurements were performed after positioning the s.c. tumor in a dual-frequency slotted-tube resonator. The intracellular pH (pHi; n=5), extracellular pH (pHe; n=5), bioenergetics (βNTP/Pi; n=5), and lactate (n=3) concentrations were measured after LND (100 mg/kg; i.p.) administration. Procedures for data acquisition, post-processing and parameter estimation were performed as previously described.3
In vitro the oxygen consumption rate and extracellular acidification rate were determined using the Seahorse XF-96 Extracellular Flux Analyzer before and after LND treatment. Glucose and lactate concentrations were measured using a YSI 2300 STAT Plus Glucose & Lactate Analyzer under the same conditions.
The Small Animal Radiation Research Platform (SARRP) was used to irradiate the PC3 xenografts to assess the effect of LND on RT response. Four cohorts of 5 matched animals/tumors; sham irradiated with 0, 2, 4, 8 and 16 Gy single doses were used to determine the dose-response curve measuring tumor growth delay. The 4 Gy dose was selected as providing sufficient dynamic range to demonstrate increased response with LND. The radiation dose was applied 40 min. after LND injection in order to achieve significant acidification and de-energization of the tumor. For the growth-delay determination, the tumor was monitored for 3 doubling times utilizing the log-linear regression phase of growth for each animal. Wilcoxon and t-test analysis were used to determine significant differences between treatment groups.4
The pHi, pHe, bioenergetics and lactate data at various time points following LND administration were compared by One-way ANOVA and t-test analysis.
McCabe Institutional Pilot Grant
1. Nancolas B, Guo LL, Zhou R, et al. The anti-tumour agent lonidamine is a potent inhibitor of the mitochondrial pyruvate carrier and plasma membrane monocarboxylate transporters. Biochemical Journal. 2016;473:929-36. PMCID: PMC4814305
2. Guo LL, Shestov AA, Worth AJ, et al. Inhibition of Mitochondrial Complex II by the Anticancer Agent Lonidamine. Journal of Biological Chemistry. 2016;291(1):42-57. PMCID: PMC4697178
3. Nath K, Nelson DS, Ho AM, et al. 31P and 1H MRS of DB-1 melanoma xenografts: lonidamine selectively decreases tumor intracellular pH and energy status and sensitizes tumors to melphalan. NMR Biomed. 2013;26(1):98-105. PMCID: PMC3465621.
4. Efron TB. Bootstrap Method for Standard Errors, Confidence Intervals, and Other Measures of Statistical Accuracy. Statistical Science. 1986;1(1):54-77.