Melanoma remains one of the deadliest of human cancers with no effective method for treating the disseminated disease. We propose to develop an effective method for systemic chemotherapy of melanoma by [tumor] selective [intracellular] acidification to sensitize melanomas to N-mustard alkylating agents. Our strategy is to coadminister glucose with the monocarboxylic acid transport (MCT) inhibitor lonidamine LND in order to trap lactic acid produced by glycolysis in the tumor and to inhibit oxidative metabolism by impeding pyruvate trans- port into mitochondria. This approach depends on the MCT being the key pathway for tumor intracellular pH (pHi) homeostasis, which has been confirmed in our melanoma models. Previous treatment with LND under hyperglycemic conditions led to some mortality. We were funded on this 2yr R01 to develop a safe reproducible method for acidification of human DB1 melanoma xenografts with LND. By administering LND 100 mg/kg i.p. without glucose, we have achieved acidification of the tumor to a pHi of 6.40.1 for at least hr during which time the NTP/Pi ratio of the tumor decreased by 50%; there was no mortality, no significant loss of weight and minimal toxicity associated with this treatment. [LND induced minimal changes in extracellular pH (pHe) of normal tissue, but modified the pH gradient across the plasma membrane of the tumor from pretreatment values of pHi 6.9, pHe 7.0 or !pH=~0 to a post-treatment values of pHi=6.4, pHe=6.85 or ?pH=-0.45. Steady state lactate in the tumor increased 3-fold relative to pre-LND levels.] Respiration, EKG, electrolyte levels and blood oxygenation did not change, while blood analysis is still in progress. The pHi and ATP/Pi of skeletal muscle did not change, and there was only a small transient decrease in pHi and ATP/Pi of the liver. Melphalan (LPAM 7.5 mg/kg i.v.)+ LND exhibited a growth delay of 10.50.5 d vs. LND alone, 0.60.7 d. and LPAM alone 1.40.1 d. The [enhanced] response of LPAM to acidification is attributed to an increase in the concentration of the active intermediate (aziridinium ion), a decrease in GSH due to decreased activity of glutathione-S-transferase and a decrease in DNA repair resulting from acid inhibition of O6-alkyl-transferase. Tumor volume decreased by 52.5 12.5%, consistent with the estimated log10 cell kill=0.3012 = log10(2.0). These preliminary studies set the stage for implementation of multidose systemic treatment of xenograft models of human melanoma in preparation for eventual clinical trials.
The Aims of this renewal proposal are: 1) To maximize tumor acidification while still avoiding life threatening toxicity by coadministration of low levels of glucose with LND, and to also extend these studies to the more glycolytic DB8 melanoma human xenograft model. 2) Since other N-mustard agents might exhibit similar enhancements of activity at acidic pHi values but perhaps with lower toxicity, LPAM will be compared with cyclophosphamide, chlorambucil and bendamustine under single and multidose administration. 3) To evaluate potential 1H NMR markers for noninvasive early detection of response--lactate and total choline by MRS and apparent diffusion constant (ADC) and T2 measured by MRI.

Public Health Relevance

Melanoma, the most virulent form of skin cancer, is not responsive to chemotherapy. This project proposes to develop a method to sensitize melanomas to chemotherapy by [selective intracellular acidification of] the cancer cells while minimally affecting normal cells. As a basis for future clinic trials, we will identify the optimal drug and optimal conditions for its administration in multiple doses; we will also develop noninvasive NMR methods for early detection of therapeutic response, which will effectively tailor-fit the therapy to the needs of the individual patient.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Medical Imaging Study Section (MEDI)
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Zhang, Huiming
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University of Pennsylvania
Schools of Medicine
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Xu, He N; Feng, Min; Nath, Kavindra et al. (2018) Optical Redox Imaging of Lonidamine Treatment Response of Melanoma Cells and Xenografts. Mol Imaging Biol :
Nath, Kavindra; Nelson, David S; Roman, Jeffrey et al. (2017) Effect of Lonidamine on Systemic Therapy of DB-1 Human Melanoma Xenografts with Temozolomide. Anticancer Res 37:3413-3421
Nancolas, Bethany; Guo, Lili; Zhou, Rong et al. (2016) The anti-tumour agent lonidamine is a potent inhibitor of the mitochondrial pyruvate carrier and plasma membrane monocarboxylate transporters. Biochem J 473:929-36
Shestov, Alexander A; Mancuso, Anthony; Lee, Seung-Cheol et al. (2016) Bonded Cumomer Analysis of Human Melanoma Metabolism Monitored by 13C NMR Spectroscopy of Perfused Tumor Cells. J Biol Chem 291:5157-71
Nath, Kavindra; Nelson, David S; Putt, Mary E et al. (2016) Comparison of the Lonidamine Potentiated Effect of Nitrogen Mustard Alkylating Agents on the Systemic Treatment of DB-1 Human Melanoma Xenografts in Mice. PLoS One 11:e0157125
Shestov, Alexander A; Lee, Seung-Cheol; Nath, Kavindra et al. (2016) (13)C MRS and LC-MS Flux Analysis of Tumor Intermediary Metabolism. Front Oncol 6:135
Nath, Kavindra; Guo, Lili; Nancolas, Bethany et al. (2016) Mechanism of antineoplastic activity of lonidamine. Biochim Biophys Acta 1866:151-162
Guo, Lili; Worth, Andrew J; Mesaros, Clementina et al. (2016) Diisopropylethylamine/hexafluoroisopropanol-mediated ion-pairing ultra-high-performance liquid chromatography/mass spectrometry for phosphate and carboxylate metabolite analysis: utility for studying cellular metabolism. Rapid Commun Mass Spectrom 30:1835-45
Guo, Lili; Shestov, Alexander A; Worth, Andrew J et al. (2016) Inhibition of Mitochondrial Complex II by the Anticancer Agent Lonidamine. J Biol Chem 291:42-57
Nath, Kavindra; Nelson, David S; Heitjan, Daniel F et al. (2015) Lonidamine induces intracellular tumor acidification and ATP depletion in breast, prostate and ovarian cancer xenografts and potentiates response to doxorubicin. NMR Biomed 28:281-90

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