Potentiation of anti-cancer drug activity at low intratumoral pH induced by the mitochondrial inhibitor m-iodobenzylguanidine (MIBG) and its analogue benzylguanidine (BG)

Abstract

Summary Tumour-selective acidification is of potential interest for enhanced therapeutic gain of pH sensitive drugs. In this study, we investigated the feasibility of a tumour-selective reduction of the extracellular and intracellular pH and their effect on the tumour response of selected anti-cancer drugs. In an in vitro L1210 leukaemic cell model, we confirmed enhanced cytotoxicity of chlorambucil at low extracellular pH conditions. In contrast, the alkylating drugs melphalan and cisplatin, and bioreductive agents mitomycin C and its derivative EO9, required low intracellular pH conditions for enhanced activation. Furthermore, a strong and pH-independent synergism was observed between the pHequilibrating drug nigericin and melphalan, of which the mechanism is unclear. In radiation-induced fibrosarcoma (RIF-1) tumour-bearing mice, the extracellular pH was reduced by the mitochondrial inhibitor m-iodobenzylguanidine (MIBG) or its analogue benzylguanidine (BG) plus glucose. To simultaneously reduce the intracellular pH, MIBG plus glucose were combined with the ionophore nigericin or the Na + /H + exchanger inhibitor amiloride and the Na + -dependent HCO 3 -/Cl -exchanger inhibitor 4,4′-diisothiocyanostilbene-2,2′-disulphonic acid (DIDS). Biochemical studies confirmed an effective reduction of the extracellular pH to approximately 6.2, and anti-tumour responses to the interventions indicated a simultaneous reduction of the intracellular pH below 6.6 for at least 3 h. Combined reduction of extra-and intracellular tumour pH with melphalan increased the tumour regrowth time to 200% of the pretreatment volume from 5.7 ± 0.6 days for melphalan alone to 8.1 ± 0.7 days with pH manipulation (P < 0.05). Mitomycin C related tumour growth delay was enhanced by the combined interventions from 3.8 ± 0.5 to 5.2 ± 0.5 days (P < 0.05), but only in tumours of relatively large sizes. The interventions were non-toxic alone or in combination with the anti-cancer drugs and did not affect melphalan biodistribution. In conclusion, we have developed non-toxic interventions for sustained and selective reduction of extra-and intracellular tumour pH which potentiated the tumour responses to selected anti-cancer drugs.

Reduction of Intracellular pH as a Strategy to Enhance the pH-Dependent Cytotoxic Effects of Melphalan for Human Breast Cancer Cells

“…Although lowering the pHe of the environment increases the accumulation of melphalan in cells, this accumulation is not sufficient to account for the increased cell killing (8,9). The results of Kuin et al (12) suggest that a decrease in pHi as a result of the environmental acidity might be the cause of the enhanced cytotoxicity of melphalan. The decrease in pHi might affect the ability of the cell to repair its DNA during and after the treatment.…”

Effects of tumour acidification with glucose+MIBG on the spontaneous metastatic potential of two murine cell lines

“…MIBG also had significant growth inhibitory effects on the KHT-C tumours if the injections were started before a palpable tumour was present. Although cell lines are generally highly capable of maintaining alkaline intracellular pH (pH i ) even when the pH e is as low as 6.2 (Lee and Tannock, 1998;Kozin et al, 2001), there is evidence that suggests that MIBG acidifies tumour cell pH i via an unknown mechanism (Biaglow et al, 1998;Kuin et al, 1999;Zhou et al, 2000). It is therefore possible that MIBG-induced cytotoxic effects may have counteracted the potential metastasis promoting effects of tumour acidification.…”

“…Current evidence suggests that the tumour microenvironmental characteristic, acidic pH e , can be exploited and enhanced to Tumour acidification and metastasis T Kalliomäki and RP Hill improve the efficacy of hyperthermia, certain forms of chemotherapy and photodynamic therapy (Wike-Hooley et al, 1984;Tannock and Rotin, 1989;Jahde et al, 1992;Kuin et al, 1994Kuin et al, , 1999Kozin et al, 2001;Ohtsubo et al, 2001;Piot et al, 2001). However, acidic pH and lactate have also been associated with enhanced metastatic tumour progression both in experimental models (Schlappack et al, 1991;Jang and Hill, 1997) and clinically (Parkins et al, 1997;Walenta et al, 2000;Brizel et al, 2001).…”

“…Several studies have examined ways to further reduce extracellular pH (pH e ) in order to improve the therapeutic index of pH responsive cancer therapies (Wike-Hooley et al, 1984;Thistlethwaite et al, 1987;Tannock and Rotin, 1989;Jahde et al, 1992;Kuin et al, 1994). Acidification of the tumour extracellular space with glucose alone or in combination with meta-iodo-benzylguanidine (MIBG) provides a means for preferential intracellular accumulation and enhanced cytotoxicity of weakly acidic chemotherapeutic drugs, such as chlorambucil (Kuin et al, 1999;Kozin et al, 2001) and increases tumour cell sensitivity to hyperthermia (Ohtsubo et al, 2001). Acidification of human colonic xenografts has also been shown to enhance their sensitivity to aminolevulinic acid-mediated photodynamic therapy (Piot et al, 2001).…”

“…However, low pO 2 levels and acidic pH e are also known to coexist in solid tumours and thus it has been difficult to separate the effects of these two factors on metastatic potential in vivo (Helmlinger et al, 1997;Rofstad, 2000;Hill et al, 2001). To clarify further the relationship between acidic pH and metastatic tumour progression, we injected MIBG in combination with glucose into tumour-bearing mice; this treatment results in selective tumour acidification by inhibiting mitochondrial respiration at complex I of the electron transport chain and by stimulation of lactic acid production through anaerobic glycolysis (Loesberg et al, 1990;Jahde et al, 1992;Kuin et al, 1994Kuin et al, , 1999. We examined the relative contribution of oxygenation and acidity to the spontaneous metastatic potential of the murine KHT-C fibrosarcoma and B16F1 melanoma cells by treating tumourbearing mice daily with glucose þ MIBG alone or in combination with a strategy to simulate acute hypoxia .…”