- Email: [email protected]
The effect of first-dose doxorubicin on the cyclic nucleotide levels of the human myocardium
TOXICOLOGY
AND
APPLIED
PHARMACOLOGY
SHORT
60,
151-154
(1981)
COMMUNICATION
The Effect of First-Dose Doxorubicin on the Cyclic Levels of the Human Myocardiuml
Nucleotide
The Effect of First-Dose Doxorubicin on the Cyclic Nucleotide Levels of the Human Myocardium. UNVERFERTH, D. V., FERTEL, R. H., TALLEY, R. L. MAGORIEN, R. D.. AND BALCERZAK, S. P. (1981). Toxicol. Appl. Pharmacol. 60, 151-1.54. This study investigated changes in cyclic nucleotide concentration in human myocardium during the first 24 hr after the initial injection of doxorubicin. Ten patients with a normal cardiac status and no previous radiation or chemotherapy received 44.5 t 3.5 mg/m2 (8 f SEM) of doxorubicin. Cyclic nucleotide analysis of endomyocardial biopsy tissue removed at baseline and 4 and 24 hr after injection of doxorubicin demonstrated no change of cyclic AMP, but did show a fall of cyclic GMP at both 4 (p < 0.01) and 24 hr (p < 0.02) and a consequent rise of the cyclic AMP:cyclic GMP ratio. These changes in cyclic GMP levels in the myocardium of patients receiving doxorubicin may reflect injury induced by free radicals.
Doxorubicin hydrochloride (adriamycin) is an effective cancer chemotherapeutic agent for the treatment of acute leukemias, lymphomas and certain solid tumors (O’Bryan et al., 1973; Gottlieb er al., 1974). The clinical application of long-term doxorubicin therapy is limited by the development of a dose-related cardiomyopathy; this complication has led to the recommendation that the total cumulative dose not exceed 550 mg per square meter of body surface area (Lefrak et al., 1973). This study was designed to investigate the changes of myocardial cyclic nucleotides in the 24-hr period immediately following the initial doxorubicin dose in human subjects. The purpose was to document early changes and to determine if these changes might supply a clue to the etiology and pathogenesis of doxorubicin cardiotoxicity. METHODS Porient characteristics. The 10 patients studied ranged in age from 34 to 58 years with an average age
I This work Comprehensive the NCI, Grant Chapter of the
was supported in part by the OSU Cancer Center, Grant ACS-IN-16Q: CA-15147; and the Central Ohio Heart American Heart Association.
of 48. There were eight females and two males. Each patient had a tumor which was felt to be responsive to doxorubicin chemotherapy (breast cancer-6, lung cancer-2, sarcoma- 1, lymphoma-1). The pretreatment cardiac history and cardiovascular examination were unremarkable. None of the patients had received prior chemotherapy or radiation therapy. Each patient gave written informed consent prior to study. Doxorubicin was given in a single dose of 30 to 60 mg/m’ (k = 44.5) as an intravenous injection over a 5-min period. The electrocardiogram was monitored during the injection. Endomyocurdial biopsy procedure. An endomyocardial biopsy procedure was performed immediately prior to and at 4 and 24 hr after the intravenous injection of doxorubicin. No premeditation was administered and local anesthesia was used for the procedure. An internal jugular venous catheter was left in place between biopsy procedures so that only one internal jugular puncture was required. At least three pieces of tissue were removed from the right side of the interventricular septum during each procedure. The specimens were submerged in liquid nitrogen immediately upon removal. The time delay from the closure of the jaws of the bioptome on the septal tissue until submersion was less than eight seconds. Each tissue sample measured 2 mm” in size and weighed 2 to 5 mg. Determinations of’ cyclic nucleoride levels. The biopsies from each procedure were stored in liquid nitrogen. The frozen biopsies were placed in I ml of 570 trichloracetic acid (TCA) and homogenized. The tissue was sonicated for approximately 30 set and then centrifuged at 3OOg for 15 min. Cyclic nucleotides were assayed by a radioimmunoassay procedure (Steiner et ul., 1972) modified to pro151
0041-008X/81/100151-04$02.00/O Copyright All rights
0 1981 by Academic Press. Inc. of reproduction in any form rewrved.
152
SHORT COMMUNICATION TABLE
1
CHANGEOF CYCLIN NUCLEOTIDES MYOCARDIUMAFTERDOXORUBICIN
IN HUMAN
Time after doxorubicin Pre-doxolubicin (Baseline) CAMP’
cGMP" cAMP/cGMP
2.6 z 0.4 0.24 + 0.05 11 r2
4 hr
24 hr
2.5 + 0.3 0.18 + 0.04’ 14 2 2
2.9 + 0.5 0.16 c 0.09 18 + 30
a Cyclic AMP (CAMP) and cyclic GMP (cGMP) fmollpg protein (X 2 SE). b Significantly different from baseline (p < 0.05).
expressed
as
duce increased sensitivity (Delaage ef al., 1978). Crossreactivity of the antibody to cyclic AMP was less than 2% with cyclic GMP and less than 0.005% with all other nucleotides tested. Cross-reactivity of the antibody to cyclic GMP was less than 0.001% with cyclic AMP and ATP and less than 0.001% with all other nucleotides tested. Essentially all of the material recorded as cyclic nucleotide in the assay was destroyed by phosphodiesterase. Protein content was determined by the method of Lowry et al. (1951). The adenosine 3’5’~cyclic monophosphate (CAMP) and guanosine 3’,5’-cyclic monophosphate (cGMP) values were expressed as femtomoles cyclic nucleotide per microgram of protein. Each sample was run in duplicate for both CAMP and cGMP and the tubes were averaged for each sample. Two tissue specimens from each time period for each patient were analyzed. Reproducibility of the cyclic nucleotide analysis on endomyocardial biopsies has been previously reported by our laboratory (Unverferth et al., 1981). Statistical analysis. The cyclic nucleotide levels were compared to the baseline measurements for each test by Student’s t test for paired data. A p value of 0.05 or less was considered to be statistically significant.
RESULTS
AND DISCUSSION
The results of the cyclic nucleotide determinations are shown in Table 1. The CAMP did not vary from baseline but the cGMP fell from a baseline of 0.24 & 0.05 fmol/pg protein (x ? SE) to 0.18 t 0.04 fmol/pg protein at 4 hr and to 0.16 f. 0.05 fmol/pg protein at 24 hr. The ratio CAMP/ cGMP rose significantly from baseline (11 ? 2) to 14 f 2 at 4 hr and 18 ? 3 at 24 hr after doxorubicin.
Doxorubicin has a wide spectrum of biologic activity. It intercalates with DNA (Pigram et al., 1972)) generates free radical compounds (Bachur et al., 1979; Sato et al., 1977), interferes with oxidative phosphorylation (Iwamoto et al., 1974) and alters the electrolyte composition and movement in cells (Moore et al., 1977). Intercalation with DNA and inhibition of DNA polymerase may be most important in doxorubicin’s antineoplastic activity (Myers et al., 1977). The inhibition of nucleic acid synthesis is less important in cardiac toxicity because human myocardial cells do not proliferate after 2 months of age (Zak, 1973). Although protein synthesis is necessary for cellular repair, doxorubicin does not inhibit RNA or protein synthesis to the same degree as it inhibits DNA synthesis (Fialkoff et al., 1979; Momparier et al., 1976). A proposed cause of doxorubicin cardiotoxicity is free radical formation (Myers et al., 1977). Electron spin resonance studies have shown that the anthraquinone nucleus of doxorubicin is reversibly converted to a free radical semiquinone (Bachur et al., 1977) with the resultant formation of superoxide and hydrogen peroxide (Sato el al., 1977). These reactive compounds may damage biologic membranes by lipid peroxidation. The heart is susceptible to free radical injury because it has low levels of superoxide dismutase (Doroshow et al., 1980) and catalase and because its glutathione peroxidase is easily depleted (Revis and Marusic, 1978). These are key enzymes for detoxification of free radicals (Goodman and Hochstein, 1977). Studies on the effect of doxorubicin on the metabolism of heart slices showed that cardiac tissue does not degrade hydrogen peroxide adequately (Burton et al., 1979). Doxorubicin inhibits cardiac guanylate cyclase. Lehotay et al. (1979) have demonstrated in rats that a single injection of doxorubicin (IO-20 mg/kg) resulted in a 3050% decrease in cardiac guanylate cyclase
153
SHORT COMMUNICATION
within 72 hr. In vitro studies using tissue homogenates show doxorubicin induced inhibition of cardiac guanylate cyclase, but not spleen, liver, lung, or kidney guanylate cyclase. The same authors have also demonstrated in vitro inhibition of human cardiac guanylate cyclase (Levey et al., 1979). The importance of our study is that we have demonstrated that pharmacologic doses of doxorubicin, when administered to patients with normal hearts, can lower cGMP presumably by inhibition of cardiac guanylate cyclase. The inhibition of guanylate cyclase by free radicals has been suggested. Hydrogen peroxide mediates the inhibition of guanylate cyclase (White et al., 1976) and can be prevented by cysteine or glutathione. Thus, it is reasonable to postulate that free radical formation associated with doxorubicin administration may be responsible for the inhibition of cardiac guanylate cyclase and that antioxidants such as a-tocopherol and N-acetylcysteine may prevent doxorubicininduced guanylate cyclase inhibition in cardiac tissue. Cardiac guanylate cyclase activity and cGMP levels may be a biochemical marker of doxorubicin cardiotoxicity. ACKNOWLEDGMENTS The authors would like to thank Marlene Griffin for her assistance in performing the biopsies, Kathryn Tight for the performance of the cyclic nucleotide assays, and Tami West for the preparation of the manuscript.
REFERENCES BACHUR. N. R., GORDON, S. L., AND GEE, M. V. (1977). Anthracycline antibiotic augmentation of microsomal electron transport and free radical formation. Mol. Pharmacol. 13, 901-910. BACHUR, N. R., GORDON, S. L., GEE, M. V., AND KON, M. (1979). NADPH cytochrome P-450 reductase activation of quinone anticancer agents to free radicals. Proc. Nat. Acad. Sci. USA 76, 9.54957. BURTON, G. M., HENDERSON, C. A., BALCERZAK,
S. P., AND SAGONE, A. L. (1979). Effect of adriamycin on the metabolism of heart slices. Inr. J. Radiat.
Oncol.
Biol.
Phys.
5, 1287-1289.
DELAAGE, M. R., Roux, D., AND CAILLA, H. L. (1978). Recent Advances in cyclic nucleotide radioimmunoassay. In Molecular Biology and Pharmocology of Cyclic Nucleotides (G. Folco and R. Paoletti, eds.), Vol. 1, pp. 155-171. Elsevier/NorthHolland, New York. DOROSHOW, J. H., LOCKER, G. Y., AND MYERS. C. E. (1980). Enzymatic defenses of the mouse heart against reactive oxygen metabolites. J. C/in. In~.cst. 65, 128-135. FIALKOFF, H., GOODMAN, M. F., AND SERAYDARI~N, M. W. (1979). Differential effect of adriamycin on DNA replicative and repair synthesis in cultured neonatal rat cardiac cells. Cancer Re.c. 39, 1321-1327. GOODMAN. J., AND HOCHSTEIN, P. (1977). Generation of free radicals and lipid peroxidation by redox cycling of adriamycin and daunomycin. Biochrm Biophys.
Res. Commun.
77, 797-803.
GOTTLIEB. J. A., BAKER, L. H., O’BRYAN. R. M., LUCE, J. K., SINKOVICS, J. G., AND QUAGLIANA, J. M. (1974). Chemotherapy of metastatic sarcomas using combinations with adriamycin. Biorlrrm. Pharmacol. Suppl. 2, 183-192. IWAMOTO, Y., HANSEN, I. L., PORTER, T. H., h~;o FOLKERS, K. (1974). Inhibition of coenzyme Q,(,enzymes, succinoxidase and NADH-oxidase. by adriamycin and other quinones having antitumor activity. Biochem. Biophys. Res. Commute. 58, 633-638. LEFRAK. E. A., PITHA, J., ROSENHEIM, S.. .\NU GOTTLIEB. J. A. (1973). A clinicopathologic analysis of adriamycin cardiotoxicity. Cancer 32, 302-314. LEHOTAY. D. C., LEVEY, B. A., ROGERSON, B., ANI) LEVEY, G. S. (1979). Inhibition of rat cardiac guanylate cyclase by doxorubicin (adriamycin) in vivo. Cl&. Res. 27, 616A. LEVEY, G. S., LEVEY, B. A., Rutz, E., AND LEHO I ~1, D. C. (1979). Selective inhibition of rat and human cardiac guanylate cyclase in vitro by doxorubicin (adriamycin): Possible link to anthracycline cardiotoxicity. J. Mol. Cell. Cardiol. 11, 591-599. LOWRY, 0. H., ROSEBOROUGH. N. J., FARR. A. L.. AND RANDALL, R. J. (1951). Protein measurement with a Folin phenol reagent. J. Biol. Chcm. 193, 265-275. MOMPARIER, R. L., KARON, M., SIEGEL, S. E.. .,\NI) AVILA. F. (1976). Effect of adriamycin on DNA, RNA. and protein synthesis in cell-free systems and intact cells. Cancer Res. 36, 2891-2895. MOORE, L., LANDON, E. J., AND COONEY. D. A. (1977). Inhibition of the cardiac mitochondrial calcium pump by adriamycin in vitro. Biochem Zled. 18, 131-138.
154
SHORT COMMUNICATION
MYERS, C. E., MCGUIRE, W. P., LISS, R. H., GROTZINGER, K., AND YOUNG, R. C. (1977). Adriamycin:
The role of lipid peroxidation in cardiac toxicity and tumor response. Science 197, 165- 167. O’BRYAN, R. M., LUCE, J. K., TALLEY, R. W., GOTTLIEB,
J. A., BAKER,
L. H., AND BONADONNA,
G. (1973). Phase II evaluation of adriamycin in human neoplasia. Cancer 32, l-8. PIGRAM, W. J., FULLER, W., AND HAMILTON, L. D. (1972). Stereochemistry of intercalation: Interaction of daunomycin with DNA. Nature New Biol. 235, 17-19. REVIS, N. W., AND MARUSIC, N. (1978). Glutathione peroxidase activity and selenium concentration in the hearts of doxorubicin-treated rabbits. J. Mol. Cell. Cardiol. 10, 945-951. SATO, A., IWAIZUMI, M., HANDA, K., AND TAMURA, Y. (1977). Electron spin resonance study on the mode of generation of free radicals of daunomytin, adriamycin, and carboquone in NAD (P) Hmicrosome system. Gann 58,603-608. STEINER,
A. L.,
PARKER,
C. W., AND KIPNIS,
D. M.
(1972). A radioimmunoassay for measurement of cyclic nucleotides. J. Biol. Chem. 247, 1106- 1113. UNVERFERTH, MAGORIEN,
D. V., FERTEL, R. H., ALTSCHULD, R., R. D., LEWIS, R. P., AND LEIER, C. V.
(1981). Biochemical measurements of endomyocardial biopsies. Catheter Cardiovasc. Diag. 7, 55-64. WHITE,
A. A., CRAWFORD,
P. J. (1976). Activation of soluble guanylate cyclase from rat lung by incubation or by hydrogen peroxide. J. Biol. Chem. 251, 7304-7312. ZAK, R. (1973). Cell proliferation during cardiac growth. Amer. J. Cardiol. 31, 211-219. LAD,
K. M.,
PATT,
C. S., AND
V. UNVERFERTH’
DONALD Division of Cardiology Department of Medicine
RICHARD
H. FERTEL
Department of Pharmacology ROBERT
L. TALLEY
Division of Hematology and Oncology Department of Medicine RAYMOND D. MAGORIEN Division of Cardiology Department of Medicine STANLEY Division of Department Ohio State Columbus, Received
P. BALCERZAK
Hematology and Oncology of Medicine University Hospitals Ohio 43210 December 9. 1980
* To whom reprint requests should be sent to: Donald V. Unverferth, M.D., 657 Means Hall, 466 West Tenth Avenue, Columbus, Ohio 43210.
AND
APPLIED
PHARMACOLOGY
SHORT
60,
151-154
(1981)
COMMUNICATION
The Effect of First-Dose Doxorubicin on the Cyclic Levels of the Human Myocardiuml
Nucleotide
The Effect of First-Dose Doxorubicin on the Cyclic Nucleotide Levels of the Human Myocardium. UNVERFERTH, D. V., FERTEL, R. H., TALLEY, R. L. MAGORIEN, R. D.. AND BALCERZAK, S. P. (1981). Toxicol. Appl. Pharmacol. 60, 151-1.54. This study investigated changes in cyclic nucleotide concentration in human myocardium during the first 24 hr after the initial injection of doxorubicin. Ten patients with a normal cardiac status and no previous radiation or chemotherapy received 44.5 t 3.5 mg/m2 (8 f SEM) of doxorubicin. Cyclic nucleotide analysis of endomyocardial biopsy tissue removed at baseline and 4 and 24 hr after injection of doxorubicin demonstrated no change of cyclic AMP, but did show a fall of cyclic GMP at both 4 (p < 0.01) and 24 hr (p < 0.02) and a consequent rise of the cyclic AMP:cyclic GMP ratio. These changes in cyclic GMP levels in the myocardium of patients receiving doxorubicin may reflect injury induced by free radicals.
Doxorubicin hydrochloride (adriamycin) is an effective cancer chemotherapeutic agent for the treatment of acute leukemias, lymphomas and certain solid tumors (O’Bryan et al., 1973; Gottlieb er al., 1974). The clinical application of long-term doxorubicin therapy is limited by the development of a dose-related cardiomyopathy; this complication has led to the recommendation that the total cumulative dose not exceed 550 mg per square meter of body surface area (Lefrak et al., 1973). This study was designed to investigate the changes of myocardial cyclic nucleotides in the 24-hr period immediately following the initial doxorubicin dose in human subjects. The purpose was to document early changes and to determine if these changes might supply a clue to the etiology and pathogenesis of doxorubicin cardiotoxicity. METHODS Porient characteristics. The 10 patients studied ranged in age from 34 to 58 years with an average age
I This work Comprehensive the NCI, Grant Chapter of the
was supported in part by the OSU Cancer Center, Grant ACS-IN-16Q: CA-15147; and the Central Ohio Heart American Heart Association.
of 48. There were eight females and two males. Each patient had a tumor which was felt to be responsive to doxorubicin chemotherapy (breast cancer-6, lung cancer-2, sarcoma- 1, lymphoma-1). The pretreatment cardiac history and cardiovascular examination were unremarkable. None of the patients had received prior chemotherapy or radiation therapy. Each patient gave written informed consent prior to study. Doxorubicin was given in a single dose of 30 to 60 mg/m’ (k = 44.5) as an intravenous injection over a 5-min period. The electrocardiogram was monitored during the injection. Endomyocurdial biopsy procedure. An endomyocardial biopsy procedure was performed immediately prior to and at 4 and 24 hr after the intravenous injection of doxorubicin. No premeditation was administered and local anesthesia was used for the procedure. An internal jugular venous catheter was left in place between biopsy procedures so that only one internal jugular puncture was required. At least three pieces of tissue were removed from the right side of the interventricular septum during each procedure. The specimens were submerged in liquid nitrogen immediately upon removal. The time delay from the closure of the jaws of the bioptome on the septal tissue until submersion was less than eight seconds. Each tissue sample measured 2 mm” in size and weighed 2 to 5 mg. Determinations of’ cyclic nucleoride levels. The biopsies from each procedure were stored in liquid nitrogen. The frozen biopsies were placed in I ml of 570 trichloracetic acid (TCA) and homogenized. The tissue was sonicated for approximately 30 set and then centrifuged at 3OOg for 15 min. Cyclic nucleotides were assayed by a radioimmunoassay procedure (Steiner et ul., 1972) modified to pro151
0041-008X/81/100151-04$02.00/O Copyright All rights
0 1981 by Academic Press. Inc. of reproduction in any form rewrved.
152
SHORT COMMUNICATION TABLE
1
CHANGEOF CYCLIN NUCLEOTIDES MYOCARDIUMAFTERDOXORUBICIN
IN HUMAN
Time after doxorubicin Pre-doxolubicin (Baseline) CAMP’
cGMP" cAMP/cGMP
2.6 z 0.4 0.24 + 0.05 11 r2
4 hr
24 hr
2.5 + 0.3 0.18 + 0.04’ 14 2 2
2.9 + 0.5 0.16 c 0.09 18 + 30
a Cyclic AMP (CAMP) and cyclic GMP (cGMP) fmollpg protein (X 2 SE). b Significantly different from baseline (p < 0.05).
expressed
as
duce increased sensitivity (Delaage ef al., 1978). Crossreactivity of the antibody to cyclic AMP was less than 2% with cyclic GMP and less than 0.005% with all other nucleotides tested. Cross-reactivity of the antibody to cyclic GMP was less than 0.001% with cyclic AMP and ATP and less than 0.001% with all other nucleotides tested. Essentially all of the material recorded as cyclic nucleotide in the assay was destroyed by phosphodiesterase. Protein content was determined by the method of Lowry et al. (1951). The adenosine 3’5’~cyclic monophosphate (CAMP) and guanosine 3’,5’-cyclic monophosphate (cGMP) values were expressed as femtomoles cyclic nucleotide per microgram of protein. Each sample was run in duplicate for both CAMP and cGMP and the tubes were averaged for each sample. Two tissue specimens from each time period for each patient were analyzed. Reproducibility of the cyclic nucleotide analysis on endomyocardial biopsies has been previously reported by our laboratory (Unverferth et al., 1981). Statistical analysis. The cyclic nucleotide levels were compared to the baseline measurements for each test by Student’s t test for paired data. A p value of 0.05 or less was considered to be statistically significant.
RESULTS
AND DISCUSSION
The results of the cyclic nucleotide determinations are shown in Table 1. The CAMP did not vary from baseline but the cGMP fell from a baseline of 0.24 & 0.05 fmol/pg protein (x ? SE) to 0.18 t 0.04 fmol/pg protein at 4 hr and to 0.16 f. 0.05 fmol/pg protein at 24 hr. The ratio CAMP/ cGMP rose significantly from baseline (11 ? 2) to 14 f 2 at 4 hr and 18 ? 3 at 24 hr after doxorubicin.
Doxorubicin has a wide spectrum of biologic activity. It intercalates with DNA (Pigram et al., 1972)) generates free radical compounds (Bachur et al., 1979; Sato et al., 1977), interferes with oxidative phosphorylation (Iwamoto et al., 1974) and alters the electrolyte composition and movement in cells (Moore et al., 1977). Intercalation with DNA and inhibition of DNA polymerase may be most important in doxorubicin’s antineoplastic activity (Myers et al., 1977). The inhibition of nucleic acid synthesis is less important in cardiac toxicity because human myocardial cells do not proliferate after 2 months of age (Zak, 1973). Although protein synthesis is necessary for cellular repair, doxorubicin does not inhibit RNA or protein synthesis to the same degree as it inhibits DNA synthesis (Fialkoff et al., 1979; Momparier et al., 1976). A proposed cause of doxorubicin cardiotoxicity is free radical formation (Myers et al., 1977). Electron spin resonance studies have shown that the anthraquinone nucleus of doxorubicin is reversibly converted to a free radical semiquinone (Bachur et al., 1977) with the resultant formation of superoxide and hydrogen peroxide (Sato el al., 1977). These reactive compounds may damage biologic membranes by lipid peroxidation. The heart is susceptible to free radical injury because it has low levels of superoxide dismutase (Doroshow et al., 1980) and catalase and because its glutathione peroxidase is easily depleted (Revis and Marusic, 1978). These are key enzymes for detoxification of free radicals (Goodman and Hochstein, 1977). Studies on the effect of doxorubicin on the metabolism of heart slices showed that cardiac tissue does not degrade hydrogen peroxide adequately (Burton et al., 1979). Doxorubicin inhibits cardiac guanylate cyclase. Lehotay et al. (1979) have demonstrated in rats that a single injection of doxorubicin (IO-20 mg/kg) resulted in a 3050% decrease in cardiac guanylate cyclase
153
SHORT COMMUNICATION
within 72 hr. In vitro studies using tissue homogenates show doxorubicin induced inhibition of cardiac guanylate cyclase, but not spleen, liver, lung, or kidney guanylate cyclase. The same authors have also demonstrated in vitro inhibition of human cardiac guanylate cyclase (Levey et al., 1979). The importance of our study is that we have demonstrated that pharmacologic doses of doxorubicin, when administered to patients with normal hearts, can lower cGMP presumably by inhibition of cardiac guanylate cyclase. The inhibition of guanylate cyclase by free radicals has been suggested. Hydrogen peroxide mediates the inhibition of guanylate cyclase (White et al., 1976) and can be prevented by cysteine or glutathione. Thus, it is reasonable to postulate that free radical formation associated with doxorubicin administration may be responsible for the inhibition of cardiac guanylate cyclase and that antioxidants such as a-tocopherol and N-acetylcysteine may prevent doxorubicininduced guanylate cyclase inhibition in cardiac tissue. Cardiac guanylate cyclase activity and cGMP levels may be a biochemical marker of doxorubicin cardiotoxicity. ACKNOWLEDGMENTS The authors would like to thank Marlene Griffin for her assistance in performing the biopsies, Kathryn Tight for the performance of the cyclic nucleotide assays, and Tami West for the preparation of the manuscript.
REFERENCES BACHUR. N. R., GORDON, S. L., AND GEE, M. V. (1977). Anthracycline antibiotic augmentation of microsomal electron transport and free radical formation. Mol. Pharmacol. 13, 901-910. BACHUR, N. R., GORDON, S. L., GEE, M. V., AND KON, M. (1979). NADPH cytochrome P-450 reductase activation of quinone anticancer agents to free radicals. Proc. Nat. Acad. Sci. USA 76, 9.54957. BURTON, G. M., HENDERSON, C. A., BALCERZAK,
S. P., AND SAGONE, A. L. (1979). Effect of adriamycin on the metabolism of heart slices. Inr. J. Radiat.
Oncol.
Biol.
Phys.
5, 1287-1289.
DELAAGE, M. R., Roux, D., AND CAILLA, H. L. (1978). Recent Advances in cyclic nucleotide radioimmunoassay. In Molecular Biology and Pharmocology of Cyclic Nucleotides (G. Folco and R. Paoletti, eds.), Vol. 1, pp. 155-171. Elsevier/NorthHolland, New York. DOROSHOW, J. H., LOCKER, G. Y., AND MYERS. C. E. (1980). Enzymatic defenses of the mouse heart against reactive oxygen metabolites. J. C/in. In~.cst. 65, 128-135. FIALKOFF, H., GOODMAN, M. F., AND SERAYDARI~N, M. W. (1979). Differential effect of adriamycin on DNA replicative and repair synthesis in cultured neonatal rat cardiac cells. Cancer Re.c. 39, 1321-1327. GOODMAN. J., AND HOCHSTEIN, P. (1977). Generation of free radicals and lipid peroxidation by redox cycling of adriamycin and daunomycin. Biochrm Biophys.
Res. Commun.
77, 797-803.
GOTTLIEB. J. A., BAKER, L. H., O’BRYAN. R. M., LUCE, J. K., SINKOVICS, J. G., AND QUAGLIANA, J. M. (1974). Chemotherapy of metastatic sarcomas using combinations with adriamycin. Biorlrrm. Pharmacol. Suppl. 2, 183-192. IWAMOTO, Y., HANSEN, I. L., PORTER, T. H., h~;o FOLKERS, K. (1974). Inhibition of coenzyme Q,(,enzymes, succinoxidase and NADH-oxidase. by adriamycin and other quinones having antitumor activity. Biochem. Biophys. Res. Commute. 58, 633-638. LEFRAK. E. A., PITHA, J., ROSENHEIM, S.. .\NU GOTTLIEB. J. A. (1973). A clinicopathologic analysis of adriamycin cardiotoxicity. Cancer 32, 302-314. LEHOTAY. D. C., LEVEY, B. A., ROGERSON, B., ANI) LEVEY, G. S. (1979). Inhibition of rat cardiac guanylate cyclase by doxorubicin (adriamycin) in vivo. Cl&. Res. 27, 616A. LEVEY, G. S., LEVEY, B. A., Rutz, E., AND LEHO I ~1, D. C. (1979). Selective inhibition of rat and human cardiac guanylate cyclase in vitro by doxorubicin (adriamycin): Possible link to anthracycline cardiotoxicity. J. Mol. Cell. Cardiol. 11, 591-599. LOWRY, 0. H., ROSEBOROUGH. N. J., FARR. A. L.. AND RANDALL, R. J. (1951). Protein measurement with a Folin phenol reagent. J. Biol. Chcm. 193, 265-275. MOMPARIER, R. L., KARON, M., SIEGEL, S. E.. .,\NI) AVILA. F. (1976). Effect of adriamycin on DNA, RNA. and protein synthesis in cell-free systems and intact cells. Cancer Res. 36, 2891-2895. MOORE, L., LANDON, E. J., AND COONEY. D. A. (1977). Inhibition of the cardiac mitochondrial calcium pump by adriamycin in vitro. Biochem Zled. 18, 131-138.
154
SHORT COMMUNICATION
MYERS, C. E., MCGUIRE, W. P., LISS, R. H., GROTZINGER, K., AND YOUNG, R. C. (1977). Adriamycin:
The role of lipid peroxidation in cardiac toxicity and tumor response. Science 197, 165- 167. O’BRYAN, R. M., LUCE, J. K., TALLEY, R. W., GOTTLIEB,
J. A., BAKER,
L. H., AND BONADONNA,
G. (1973). Phase II evaluation of adriamycin in human neoplasia. Cancer 32, l-8. PIGRAM, W. J., FULLER, W., AND HAMILTON, L. D. (1972). Stereochemistry of intercalation: Interaction of daunomycin with DNA. Nature New Biol. 235, 17-19. REVIS, N. W., AND MARUSIC, N. (1978). Glutathione peroxidase activity and selenium concentration in the hearts of doxorubicin-treated rabbits. J. Mol. Cell. Cardiol. 10, 945-951. SATO, A., IWAIZUMI, M., HANDA, K., AND TAMURA, Y. (1977). Electron spin resonance study on the mode of generation of free radicals of daunomytin, adriamycin, and carboquone in NAD (P) Hmicrosome system. Gann 58,603-608. STEINER,
A. L.,
PARKER,
C. W., AND KIPNIS,
D. M.
(1972). A radioimmunoassay for measurement of cyclic nucleotides. J. Biol. Chem. 247, 1106- 1113. UNVERFERTH, MAGORIEN,
D. V., FERTEL, R. H., ALTSCHULD, R., R. D., LEWIS, R. P., AND LEIER, C. V.
(1981). Biochemical measurements of endomyocardial biopsies. Catheter Cardiovasc. Diag. 7, 55-64. WHITE,
A. A., CRAWFORD,
P. J. (1976). Activation of soluble guanylate cyclase from rat lung by incubation or by hydrogen peroxide. J. Biol. Chem. 251, 7304-7312. ZAK, R. (1973). Cell proliferation during cardiac growth. Amer. J. Cardiol. 31, 211-219. LAD,
K. M.,
PATT,
C. S., AND
V. UNVERFERTH’
DONALD Division of Cardiology Department of Medicine
RICHARD
H. FERTEL
Department of Pharmacology ROBERT
L. TALLEY
Division of Hematology and Oncology Department of Medicine RAYMOND D. MAGORIEN Division of Cardiology Department of Medicine STANLEY Division of Department Ohio State Columbus, Received
P. BALCERZAK
Hematology and Oncology of Medicine University Hospitals Ohio 43210 December 9. 1980
* To whom reprint requests should be sent to: Donald V. Unverferth, M.D., 657 Means Hall, 466 West Tenth Avenue, Columbus, Ohio 43210.