Tumor acidity, ion trapping and chemotherapeutics: II. pH-dependent partition coefficients predict importance of ion trapping on pharmacokinetics of weakly basic chemotherapeutic agents
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)
- et al.
Acute metabolic alkalosis enhances response of C3H mouse mammary tumors to the weak base mitoxantrone
Neoplasia (New York)
(2001) Mitoxantrone: an active, new antitumor agent with an improved therapeutic index
Cancer Treat. Rev.
(1987)- et al.
Carbonic anhydrase inhibitors: sulfonamides as anti-tumor agents
Bioorg. Med. Chemother.
(2001) - et al.
pH and drug resistance in tumors
Drug Resist. Updates
(2000) - et al.
The relevance of tumour pH to the treatment of malignant disease
Radiother. Oncol.
(1984) - Mahoney BP, Raghunand N, Baggett B, Gillies RJ. Tumor acidity, ion trapping and chemotherapeutics. I. Acid pH affects...
- et al.
Enhancement of chemotherapy by manipulation of tumour pH
Br. J. Cancer
(1999) - et al.
Activity of adriamycin (NSC-123127) and daunomycin (NSC-82151) against mouse mammary carcinoma
Cancer Chemother. Rep.—Part 1
(1972) - et al.
Topoisomerase II activities in AML and their correlation with cellular sensitivity to anthracyclines and epipodophyllotoxins
Leukemia
(1996) - et al.
Effects of DNA intercalating agents on topoisomerase II induced DNA strand cleavage in isolated mammalian cell nuclei
Biochemistry
(1985)
Further studies on the generation of reactive oxygen species from activated anthracyclines and the relationship to cytotoxic action and cardiotoxic effects
Biochem. Pharmacol.
Mitoxantrone: a review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential in the chemotherapy of cancer
Drugs
The cardiotoxicity of anticancer agents
Semin. Oncol.
Toxicity and anticancer activity of a new triazine antifolate
Cancer Res.
Circadian changes in mitoxantrone toxicity in mice: relationship with plasma pharmacokinetics
Int. J. Cancer
Influence of low pH on cytotoxicity of paclitaxel, mitoxantrone and topotecan
Br. J. Cancer
Effects of acid-base status on acute hypoxic pulmonary vasoconstriction and gas exchange
J. Appl. Physiol.
The pulmonary consequences of aspiration of gastric contents at pH values greater than 2.5
Am. Rev. Respir. Dis.
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These authors contributed equally to this work.