Molecular sequelae of histone deacetylase inhibition in human retinoblastoma cell lines: clinical implications.
OBJECTIVE
To characterize the molecular sequelae induced in retinoblastoma (Rb) cells by histone deacetylase inhibitors (HDACIs). Hydroxamic acid-based HDACIs such as vorinostat (suberoylanilide hydroxamic acid) induce the differentiation and apoptosis of transformed cells. Vorinostat has demonstrated significant anticancer activity against hematologic and solid tumors at doses well tolerated by patients and has been approved for the treatment of patients with cutaneous T-cell lymphoma.


METHODS
The authors evaluated the effects of the HDACIs vorinostat and m-carboxycinnamic acid bis-hydroxamide on the Rb cell lines Y79 and WERI-Rb1 with the use of the MTT assay, BrdU incorporation assay, flow cytometry, immunoblotting, gene-expression profiling, quantitative RT-PCR, and NF-kappaB DNA-binding assay.


RESULTS
Both HDACIs were effective against both Rb cell lines, inducing growth arrest and apoptosis in vitro. Vorinostat increased p53 expression and activated caspases -8, -9 and -3, whereas caspase inhibition abrogated vorinostat-induced apoptosis. Vorinostat downregulated baseline NF-kappaB activity and potentiated the activity of the DNA-damaging chemotherapeutic doxorubicin. Gene expression profiling and qRT-PCR demonstrated that vorinostat modulated the mRNA levels of genes important for signal transduction, cell cycle, cellular metabolism, stress response, apoptosis, extracellular matrix synthesis, and cell differentiation. Notably, several transcripts involved in the ephrin and Notch signaling pathways were upregulated.


CONCLUSIONS
HDACIs, such as vorinostat, induce caspase-dependent apoptosis in Rb cells, downregulate baseline NF-kappaB activity, and potentiate the effectiveness of conventional chemotherapy. The finding that vorinostat augments the effectiveness of doxorubicin provides a rationale for future clinical studies looking at the use of vorinostat in combination with conventional chemotherapy in Rb.
Poulaki
Vassiliki
V
Department of Ophthalmology, Angiogenesis Laboratory, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Massachusetts, USA
Mitsiades
Constantine S
CS
Kotoula
Vassiliki
V
Negri
Joseph
J
McMullan
Ciaran
C
Miller
Joan W
JW
Marks
Paul A
PA
Mitsiades
Nicholas
N
D000107:Acetylation; D017209:Apoptosis; D001973:Bromodeoxyuridine / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002934:Cinnamates / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D056572:Histone Deacetylase Inhibitors; D006657:Histones / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D015151:Immunoblotting; D016328:NF-kappa B / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics; D019572:Retinal Neoplasms / Q000201:enzymology / Q000473:pathology*; D012175:Retinoblastoma / Q000201:enzymology / Q000473:pathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D000077337:Vorinostat
D002934:Cinnamates; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D006877:Hydroxamic Acids; D016328:NF-kappa B; D009363:Neoplasm Proteins; D016159:Tumor Suppressor Protein p53; C111238:carboxycinnamic acid bishydroxamide; D000077337:Vorinostat; D004317:Doxorubicin; D020169:Caspases; D001973:Bromodeoxyuridine