Elsevier

Biochemical Pharmacology

Volume 66, Issue 7, 1 October 2003, Pages 1219-1229
Biochemical Pharmacology

Tumor acidity, ion trapping and chemotherapeutics: II. pH-dependent partition coefficients predict importance of ion trapping on pharmacokinetics of weakly basic chemotherapeutic agents

https://doi.org/10.1016/S0006-2952(03)00468-4Get rights and content

Abstract

Ion-trapping theory predicts that alkalinization of tumor extracellular pH will enhance the anti-tumor activity of weak-base chemotherapeutics. We have previously demonstrated that chronic and acute treatment of tumor-bearing mice with sodium bicarbonate results in tumor-specific alkalinization of extracellular pH. Furthermore, bicarbonate pretreatment enhances the anti-tumor activity of doxorubicin and mitoxantrone in two different mouse tumor models. Previous work has indicated subtle, yet significant differences between the pH sensitivities of the biodistribution and anti-tumor efficacies of doxorubicin and mitoxantrone in vitro. The present study demonstrates that systemic alkalinization selectively enhances tumor uptake of radiolabeled mitoxantrone, but not doxorubicin. Results using these two drugs are quantitatively and qualitatively very different, and can be explained on the basis of differences in the octanol–water partition coefficients of their charged forms. These results suggest that inducing metabolic alkalosis in patients would have a positive effect on response to mitoxantrone therapy. However, the therapeutic index would not increase if sodium bicarbonate also caused increased retention of mitoxantrone in susceptible normal tissues in the host. The major dose-limiting organ systems for mitoxantrone are heart, liver, bone marrow, spleen and blood cells. Bicarbonate was found to have no significant effect on the distribution of mitoxantrone to any of these tissues except for spleen. However, neither spleen weights nor lymphocyte counts were adversely affected by NaHCO3 pretreatment, indicating that this co-therapy does not enhance myelosuppression due to mitoxantrone therapy. These findings suggest that metabolic alkalosis would produce a net gain in mitoxantrone therapeutic index.

Introduction

Previous work has indicated subtle, yet significant differences between the pH sensitivities of the anti-tumor efficacies of doxorubicin and mitoxantrone in vitro[1]. Ion-trapping theory predicts that alkalinization of tumor extracellular pH (pHe) would enhance the anti-tumor activity of these weakly basic drugs. In agreement with this prediction, more of these drugs are taken up in cells cultured at pH 7.4 than at pH 6.6 in vitro. Furthermore, both of these drugs are more cytotoxic at pH 7.4 than at pH 6.6 [1]. However, in both cytotoxicity and distribution studies, mitoxantrone was consistently more pH-sensitive than doxorubicin. In vivo, chronic and acute treatment of tumor-bearing mice with sodium bicarbonate results in tumor-specific alkalinization of pHe, and bicarbonate pretreatment enhances the anti-tumor activity of doxorubicin and mitoxantrone in two different tumor models [2], [3]. However, similar to the in vitro results, these studies also showed greater pH sensitivity of the anti-tumor activity of mitoxantrone than doxorubicin. The purpose of the current study is to further examine the difference in pH-dependent activities of these weakly basic chemotherapeutics, in order to better understand the contribution of ion trapping to physiological drug resistance.

Both mitoxantrone and doxorubicin are weakly basic chemotherapeutics that intercalate into DNA, inhibit topoisomerase II and generate reactive oxygen species [4], [5], [6], [7]. They are, however, chemically distinct. Doxorubicin consists of an amino sugar, daunosamine, linked via a glycosidic bond to the planar tricyclic, adriamycinone. Mitoxantrone also has a planar polycyclic aromatic ring structure, but lacks a sugar moiety. It has two polar side chains attached to the aromatic rings, which render the molecule water-soluble [8]. Doxorubicin has a single ionizable amine with a pKa of approximately 8.3, and mitoxantrone has two ionizable amines with pKa values of 8.3–8.6. Previous work has shown that the uptake and cytotoxicity of these drugs is enhanced at elevated pHe, with the effects being greater for mitoxantrone compared to doxorubicin [1]. Subtle differences were observed in the kinetics of uptake of the radiolabeled drugs. Although the uptake of both drugs were enhanced at elevated pHe, the differences were not as significant for doxorubicin as might be expected on the basis of published pKa values. In vivo, we have found that tumor alkalinization significantly enhances mitoxantrone efficacy in the C3H tumor/C3H mouse model [3], and also enhances doxorubicin efficacy in the MCF-7 tumor/SCID mouse model, albeit to a lesser extent [2]. The current investigation examines the differences between mitoxantrone and doxorubicin in greater detail by determining the in vivo pharmacokinetic consequences of systemic metabolic alkalosis induced in the host animal by bolus or chronic administration of NaHCO3. These data confirm the previous in vitro observations, i.e. that only modest improvements in doxorubicin efficacy can be expected with tumor alkalinization and that these effects may be independent of ion trapping. On the other hand, mitoxantrone showed a large and robust enhancement in both tumor uptake and response following systemic metabolic alkalosis, suggesting that this approach may improve response to mitoxantrone in the clinic. The differences in the behavior of these two weakly basic drugs could be predicted by differences in their pH-dependent partition coefficients, and such a model may be useful for further drug design.

Gains in therapeutic index would require that the bicarbonate pretreatment protocol not have an adverse effect on drug distribution to normal tissues. In the case of doxorubicin the dose-limiting toxicity is cardiomyopathy [9]. The dose-limiting toxicity associated with mitoxantrone is bone marrow suppression, typically observed 7–14 days after treatment [10]. In studies using radiolabeled doxorubicin, no increase in drug distribution to the heart was observed following bicarbonate pretreatment. In studies with radiolabeled mitoxantrone, there was a slight (14%) increase in the spleen drug accumulation following bicarbonate pretreatment. However, follow-up studies showed no effect of bicarbonate on the effects of mitoxantrone on spleen weights or lymphocyte counts, suggesting that this co-therapy does not introduce additional myelosuppression. Furthermore, bicarbonate pretreatment had no effect on the ld50 values for normal mice. These control studies indicate that alkalinization via bicarbonate pretreatment will effectively increase the therapeutic index for mitoxantrone.

Section snippets

Cells and tumor

MCF-7 cells were cultured in RPMI-1640 or Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) (HyClone). For in vivo culturing, a suspension of 5×106 MCF-7 cells in 0.05 mL of Matrigel were implanted in the mammary fat pads of 6- to 7-week-old female severe combined immunodeficient (SCID) mice. Since MCF-7 cells are estrogen-dependent, 17β-estradiol pellets (0.72 mg, 60-day release; Innovative Research of America) were subcutaneously implanted in the shoulder

NaHCO3 enhances anti-tumor effects of anthracyclines but not taxol

Sodium bicarbonate has been previously shown to increase the pHe of tumors, whether delivered ad libitum chronically [2] or acutely by gavage or i.p. injection [3]. The increase in pHe relative to the corresponding tissues in untreated mice is greater in tumors than in normal tissues. This is possibly due to the low pHe in tumors in control mice which reduces the buffering power of endogenous bicarbonate. These treatments would be expected to increase the chemotherapeutic efficacy of weakly

Discussion

In a companion manuscript, the effect of ion trapping on a series of ionizable chemotherapeutic drugs was examined [1]. These results identified a subtle, yet significant, difference between the pH sensitivities of the anti-tumor activities of doxorubicin and mitoxantrone in vitro[1]. The current work was designed to further explore the potential mechanisms underlying these differences in vivo.

Sodium bicarbonate causes alkalinization of tumors in SCID and C3H/Hen model systems [2], [3], [29].

Acknowledgements

National Institutes of Health grant R01 CA77575.

References (37)

  • J.W. Lown et al.

    Further studies on the generation of reactive oxygen species from activated anthracyclines and the relationship to cytotoxic action and cardiotoxic effects

    Biochem. Pharmacol.

    (1982)
  • D. Faulds et al.

    Mitoxantrone: a review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential in the chemotherapy of cancer

    Drugs

    (1991)
  • D.D. Von Hoff et al.

    The cardiotoxicity of anticancer agents

    Semin. Oncol.

    (1982)
  • T.H. Corbett et al.

    Toxicity and anticancer activity of a new triazine antifolate

    Cancer Res.

    (1982)
  • F. Levi et al.

    Circadian changes in mitoxantrone toxicity in mice: relationship with plasma pharmacokinetics

    Int. J. Cancer

    (1994)
  • V. Vukovic et al.

    Influence of low pH on cytotoxicity of paclitaxel, mitoxantrone and topotecan

    Br. J. Cancer

    (1997)
  • J.A. Loeppky et al.

    Effects of acid-base status on acute hypoxic pulmonary vasoconstriction and gas exchange

    J. Appl. Physiol.

    (1992)
  • D.J. Schwartz et al.

    The pulmonary consequences of aspiration of gastric contents at pH values greater than 2.5

    Am. Rev. Respir. Dis.

    (1980)
  • Cited by (165)

    • Tumor acidic environment directs nanoparticle impacts on cancer cells

      2023, Journal of Colloid and Interface Science
    View all citing articles on Scopus
    1

    These authors contributed equally to this work.

    View full text