Combination strategy targeting the hypoxia inducible factor-1 alpha with mammalian target of rapamycin and histone deacetylase inhibitors.
OBJECTIVE
The hypoxia-inducible factor-1 alpha (HIF-alpha) is a key regulator of tumor angiogenesis. Mammalian target of rapamycin (mTOR) and histone deacetylase (HDAC) inhibitors suppress tumor-induced angiogenesis by reducing tumor HIF-1 alpha protein expression. Thus, we hypothesized that combination treatment of rapamycin and the HDAC inhibitor LBH589 has greater antiangiogenic and antitumor activity compared with single agents.


METHODS
To evaluate the effect of LBH589 and rapamycin on HIF-1 alpha in human prostate PC3, renal C2 carcinoma cell lines, and endothelial cells (human umbilical vein endothelial cells), we did Western blot analysis. To determine the antitumor activity of LBH589 and rapamycin, cell proliferation assays and xenograft experiments were conducted.


RESULTS
Western blotting showed that combination treatment of human umbilical vein endothelial cells, C2 and PC3, significantly reduced HIF-1 alpha protein expression compared with single agents. Treatment with rapamycin resulted in inhibition of the downstream signals of the mTOR pathway and increased phosphorylation of Akt in C2 cells, whereas the constitutively activated Akt in PC3 cells was not modulated. LBH589 decreased both constitutively expressed and rapamycin-induced phosphorylated Akt levels in PC3 and C2 cell lines. In clonogenic assays, the combination treatment had a greater inhibitory effect in PC3 cells (93 +/- 1.4%) compared with single agents (66 +/- 9% rapamycin and 43 +/- 4% LBH589). Combination of rapamycin and LBH589 significantly inhibited PC3 and C2 in vivo tumor growth and angiogenesis as measured by tumor weight and microvessel density.


CONCLUSIONS
Combination treatment of mTOR and HDAC inhibitors represents a rational therapeutic strategy targeting HIF-1 alpha that warrants clinical testing.
Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, 1650 Orleans Street, Cancer Research Building I, 1M52, Baltimore, MD 21231, USA.
Henk M W
HM
Verheul
Brenda
B
Salumbides
Karen
K
Van Erp
Hans
H
Hammers
David Z
DZ
Qian
Tolib
T
Sanni
Peter
P
Atadja
Roberto
R
Pili
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000008:administration & dosage*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D042783:Endothelial Cells / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000008:administration & dosage; D006655:Histone Deacetylases / Q000187:drug effects; D006801:Humans; D006877:Hydroxamic Acids / Q000008:administration & dosage*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors / Q000187:drug effects; D007150:Immunohistochemistry; D007211:Indoles; D009374:Neoplasms, Experimental / Q000188:drug therapy*; D009389:Neovascularization, Pathologic / Q000188:drug therapy; D051058:Oncogene Protein v-akt / Q000187:drug effects / Q000378:metabolism; D000077767:Panobinostat; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000008:administration & dosage*; D058570:TOR Serine-Threonine Kinases; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
D004791:Enzyme Inhibitors; D006877:Hydroxamic Acids; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007211:Indoles; D042461:Vascular Endothelial Growth Factor A; D000077767:Panobinostat; D011494:Protein Kinases; C546842:MTOR protein, human; D051058:Oncogene Protein v-akt; D058570:TOR Serine-Threonine Kinases; D006655:Histone Deacetylases; D020123:Sirolimus