Antitumor effects of doxorubicin followed by zoledronic acid in a mouse model of breast cancer.
BACKGROUND
The potent antiresorptive drug zoledronic acid (Zol) enhances the antitumor effects of chemotherapy agents in vitro. We investigated the effects of clinically achievable doses of doxorubicin (Dox) and Zol, given alone, in sequence, and in combination, on the growth of established breast tumors in vivo.


METHODS
Female MF1 nude mice were inoculated subcutaneously with 5 x 10(5) human breast cancer MDA-MB-436 cells that stably expressed green fluorescent protein (ie, MDA-G8 cells). Beginning on day 7 after tumor cell injection, the mice were injected weekly for 6 weeks with saline, Dox (2 mg/kg body weight via intravenous injection), Zol (100 microg/kg body weight via intraperitoneal injection), Dox plus Zol, Zol followed 24 hours later by Dox, or Dox followed 24 hours later by Zol (n = 8-9 mice per group). The effects of treatment on tumor growth were determined by measuring tumor volume; on tumor cell apoptosis and proliferation by immunohistochemistry using antibodies for caspase-3 and Ki-67, respectively; and on bone by microcomputed tomography and bone histomorphometry. All P values are two-sided.


RESULTS
Treatment with Dox or Zol alone or Zol followed 24 hours later by Dox did not statistically significantly decrease final tumor volume compared with saline. Mice treated with Dox plus Zol had statistically significantly smaller final tumor volumes than those treated with Dox alone (mean = 122 mm(3) vs 328 mm(3), difference = 206 mm(3), 95% confidence interval [CI] = 78 to 335 mm(3), P < .001), with Zol alone (122 mm(3) vs 447 mm(3), difference = 325 mm(3), 95% CI = 197 to 454 mm(3), P < .001), or with Zol followed 24 hours later by Dox (122 mm(3) vs 418 mm(3), difference = 296 mm(3), 95% CI = 168 to 426 mm(3), P < .001). Treatment with Dox followed 24 hours later by Zol almost completely abolished tumor growth. Tumors from mice that were treated with Dox followed by Zol had more caspase-3-positive cells than tumors from mice treated with saline (mean number of caspase-3-positive cells per square millimeter: 605.0 vs 82.19, difference = 522.8, 95% CI = 488.2 to 557.4, P < .001), with Zol alone (605.0 vs 98.44, difference = 506.6, 95% CI = 472.0 to 541.2, P < .001), or with Zol followed by Dox (605.0 vs 103.1, difference = 501.9, 95% CI = 467.3 to 536.5, P < .001). The treatment-induced increase in the number of caspase-3-positive cells was mirrored by a decrease in the number of tumor cells positive for the proliferation marker Ki-67. No evidence of bone disease was detected in any of the treatment groups following microcomputed tomography and histological analysis of bone.


CONCLUSIONS
Sequential treatment with Dox followed by Zol elicited substantial antitumor effects in subcutaneous breast tumors in vivo, in the absence of bone disease.
Ottewell
Penelope D
PD
Academic Unit of Clinical Oncology, School of Medicine and Biomedical Sciences, University of Sheffield, Beech Hill Road, Sheffield, S10 2RX, UK. i.holen@sheffield.ac.uk.
Mönkkönen
Hannu
H
Jones
Mark
M
Lefley
Diane V
DV
Coleman
Robert E
RE
Holen
Ingunn
I
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000008:administration & dosage / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000008:administration & dosage / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D050071:Bone Density Conservation Agents / Q000494:pharmacology; D053148:Caspase 3 / Q000032:analysis; D049109:Cell Proliferation / Q000187:drug effects; D004164:Diphosphonates / Q000008:administration & dosage / Q000494:pharmacology*; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000008:administration & dosage / Q000494:pharmacology*; D004334:Drug Administration Schedule; D004357:Drug Synergism; D005260:Female; D006801:Humans; D007093:Imidazoles / Q000008:administration & dosage / Q000494:pharmacology*; D007150:Immunohistochemistry; D019394:Ki-67 Antigen / Q000032:analysis; D008325:Mammary Neoplasms, Experimental / Q000188:drug therapy* / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D012107:Research Design; D014057:Tomography, X-Ray Computed; D000077211:Zoledronic Acid
D000903:Antibiotics, Antineoplastic; D050071:Bone Density Conservation Agents; D004164:Diphosphonates; D007093:Imidazoles; D019394:Ki-67 Antigen; D000077211:Zoledronic Acid; D004317:Doxorubicin; D053148:Caspase 3