Abstract
Trichostatin A produces predominantly G(1) cell-cycle blockade and differentiation of the cisplatinum-sensitive A2780 ovarian cancer cell line. Given the propensity of ovarian tumors to become resistant to cisplatinum, often leading to cross-resistance to other agents, we have extended these observations by examining how the emergence of resistant phenotypes in A2780 cells affects the actions of histone deacetylase (HDAC) inhibitors. Trichostatin A exposure (100 ng/mL, 24 hours) induced ultrastructural differentiation of the "intrinsically" cisplatinum-resistant A2780-9M subline, with the reappearance of intercellular junctions and lumina containing primitive microvilli. Similar trichostatin A exposure in the acquired resistance A2780CP cells produced minimal differentiation consisting of occasional weak intercellular junctions. Independent of the differences in trichostatin A-induced differentiation, in both resistant sublines trichostatin A produced a similar reduction in cell viability, by >90%, within 5 days of treatment. Diminished viability in both A2780-9M and CP cells was associated with the absence of cell cycle arrest in G1, resulting in predominant G2-checkpoint arrest accompanied by a 10- to 20-fold increase in Annexin V binding and the reemergence of apoptosis. Similar cell cycle arrests and apoptosis were also observed using other HDAC inhibitors and in other resistant ovarian cancer cell lines (OVCAR-3 and SK-OV-3). Trichostatin A-induced apoptosis in resistant cells is in sharp contrast to its effects on the parental cisplatinum-sensitive A2780 and normal MRC-5 fibroblast cell lines (predominant cycle arrest in G1 with no detectable apoptosis). Western immunoblot analysis indicated trichostatin A triggers apoptosis in resistant ovarian cancer cells via p53-independent activation of the intrinsic "mitochondrial" pathway, commensurate with induction of the Bcl-2-related protein Bad. These results suggest cisplatinum resistance alters the effects of HDAC inhibition through a shift in cell cycle arrest from the G1 to the G2 checkpoint and reactivation of the intrinsic mitochondrial apoptotic cascade.
Highlights
The use of platinum-based combination chemotherapy remains the standard treatment for ovarian epithelial cancers, with an initial clinical response rate of f70% [1]
Our results indicate that both ‘‘intrinsic’’ and ‘‘acquired’’ resistance in A2780 sublines alter the actions of histone deacetylase (HDAC) inhibitors, resulting in predominant G2-checkpoint cell cycle arrest and subsequent loss of cell viability due to reactivation of apoptotic cascades
To examine what effects cisplatinum-resistant phenotypes have on the action of HDAC inhibitors, we used the previously described cisplatinum-sensitive A2780 epithelial derived ovarian cancer cell line and its acquired resistance A2780CP subline [23]
Summary
The use of platinum-based combination chemotherapy remains the standard treatment for ovarian epithelial cancers, with an initial clinical response rate of f70% [1]. Multiple mechanisms have been described that contribute to the ability of cells to resist the actions of chemotherapeutic agents [4], including increased DNA damage tolerance, due to increased DNA damage repair, induction of survival factors, and alterations in apoptotic signaling. Cisplatinum resistance is often associated with diminished apoptotic signaling, involving the silencing of genes for cell cycle regulation Studies have shown that gene silencing is often associated with hypoacetylation of histone proteins through the aberrant actions of the histone deacetylase (HDAC) enzymes. Given the close association between histone acetylation and the transcriptional silencing of genes associated with oncogenesis, inhibitors of HDAC enzymes are emerging as a potentially important new class of anticancer agents [9].
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