The effect of 3-aminobenzamide, inhibitor of poly(ADP-ribose) polymerase, on human osteosarcoma cells.
This study demonstrates that in human osteosarcoma cells treatment with 3-aminobenzamide (3-AB), a potent inhibitor of poly(ADP-ribose) polymerase (PARP), induces morphological and biochemical features of differentiation, the duration of which depends on whether or not the normal RB gene is expressed. In Saos-2 cells expressing a non-functional Rb protein, 3-AB treatment induced the formation of transient, short dendritic-like protrusions. In RB-transfected-Saos-2 cells (a clone previously generated in our laboratory that shows stable expression of wild-type Rb protein), 3-AB induced marked and prolonged changes with the formation of long dendritic-like protrusions and the appearance of stellate (osteocyte-like) cells. In MG-63 cells producing a wild-type Rb protein, 3-AB treatment had more marked effects, with a larger number of cells assuming the stellate appearance of osteocytes, which were connected to each other via junctions resembling small channels. Regardless of cell type, at some point after 3-AB treatment the differentiative attempt failed and the cells died. Death was apoptotic, as demonstrated by chromatin condensation and fragmentation, specific cleavage of PARP and Lamin-B, processing of caspase-3 and the appearance of Bax immunoreactive species. Enzymatic assay and RT-PCR of alkaline phosphatase (ALP) - an enzyme whose levels markedly decrease when osteoblasts undergo terminal differentiation into osteocytes - showed that 3-AB treatment markedly lowered ALP expression. Simultaneously, 3-AB treatment markedly increased the expression of CD44, a transmembrane multifunctional adhesion molecule and sensitive marker of osteocytic differentiation. This study hypothesizes a cross-talk between pRb and PARP and suggests that PARP may be a useful target for anticancer drugs.
De Blasio
Anna
A
Department of Cell and Development Biology, Section of Biochemistry, University of Palermo, Policlinico, I-90127 Palermo, Italy.
Musmeci
Maria T
MT
Giuliano
Michela
M
Lauricella
Marianna
M
Emanuele
Sonia
S
D'Anneo
Antonella
A
Vassallo
Barbara
B
Tesoriere
Giovanni
G
Vento
Renza
R
D000469:Alkaline Phosphatase / Q000378:metabolism; D017209:Apoptosis; D001549:Benzamides / Q000494:pharmacology*; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D002470:Cell Survival; D002843:Chromatin / Q000378:metabolism; D053938:DNA Fragmentation; D003712:Dendrites / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis / Q000378:metabolism; D034921:Lamin Type B / Q000378:metabolism; D008858:Microscopy, Phase-Contrast; D012516:Osteosarcoma / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D001549:Benzamides; D002843:Chromatin; D004791:Enzyme Inhibitors; D018960:Hyaluronan Receptors; D034921:Lamin Type B; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D016160:Retinoblastoma Protein; D051028:bcl-2-Associated X Protein; C025160:3-aminobenzamide; D000469:Alkaline Phosphatase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases