Effects of HIV protease inhibitor ritonavir on Akt-regulated cell proliferation in breast cancer.
OBJECTIVE
These studies were designed to determine whether ritonavir inhibits breast cancer in vitro and in vivo and, if so, how.


METHODS
Ritonavir effects on breast cancer cell growth were studied in the estrogen receptor (ER)-positive lines MCF7 and T47D and in the ER-negative lines MDA-MB-436 and MDA-MB-231. Effects of ritonavir on Rb-regulated and Akt-mediated cell proliferation were studied. Ritonavir was tested for inhibition of a mammary carcinoma xenograft.


RESULTS
ER-positive estradiol-dependent lines (IC50, 12-24 micromol/L) and ER-negative (IC50, 45 micromol/L) lines exhibit ritonavir sensitivity. Ritonavir depletes ER-alpha levels notably in ER-positive lines. Ritonavir causes G1 arrest, depletes cyclin-dependent kinases 2, 4, and 6 and cyclin D1 but not cyclin E, and depletes phosphorylated Rb and Ser473 Akt. Ritonavir induces apoptosis independent of G1 arrest, inhibiting growth of cells that have passed the G1 checkpoint. Myristoyl-Akt, but not activated K-Ras, rescues ritonavir inhibition. Ritonavir inhibited a MDA-MB-231 xenograft and intratumoral Akt activity at a clinically attainable serum Cmax of 22 +/- 8 micromol/L. Because heat shock protein 90 (Hsp90) substrates are depleted by ritonavir, ritonavir effects on Hsp90 were tested. Ritonavir binds Hsp90 (K(D), 7.8 micromol/L) and partially inhibits its chaperone function. Ritonavir blocks association of Hsp90 with Akt and, with sustained exposure, notably depletes Hsp90. Stably expressed Hsp90alpha short hairpin RNA also depletes Hsp90, inhibiting proliferation and sensitizing breast cancer cells to low ritonavir concentrations.


CONCLUSIONS
Ritonavir inhibits breast cancer growth in part by inhibiting Hsp90 substrates, including Akt. Ritonavir may be of interest for breast cancer therapeutics and its efficacy may be increased by sustained exposure or Hsp90 RNA interference.
Department of Medicine, Indiana University, Indianapolis, Indiana 46202, USA.
Anjaiah
A
Srirangam
Ranjana
R
Mitra
Mu
M
Wang
J Christopher
JC
Gorski
Sunil
S
Badve
LeeAnn
L
Baldridge
Justin
J
Hamilton
Hiromitsu
H
Kishimoto
John
J
Hawes
Lang
L
Li
Christie M
CM
Orschell
Edward F
EF
Srour
Janice S
JS
Blum
David
D
Donner
George W
GW
Sledge
Harikrishna
H
Nakshatri
David A
DA
Potter
D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D019938:Cyclin D1 / Q000378:metabolism; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005260:Female; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D008325:Mammary Neoplasms, Experimental / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D019438:Ritonavir / Q000097:blood / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
D017320:HIV Protease Inhibitors; D018841:HSP90 Heat-Shock Proteins; D050505:Mutant Proteins; D016159:Tumor Suppressor Protein p53; D019938:Cyclin D1; D051057:Proto-Oncogene Proteins c-akt; D018844:Cyclin-Dependent Kinases; D019438:Ritonavir