A model of sensitivity and resistance to histone deacetylase inhibitors in diffuse large B cell lymphoma

Abstract

Diffuse large B cell lymphoma (DLBCL) is an aggressive form of non-Hodgkin lymphoma. While the initial treatment strategy is highly effective, relapse occurs in 40% of cases. Histone deacetylase inhibitors (HDACi) are a promising class of anti-cancer drugs but their single agent efficacy against relapsed DLBCL has been variable, ranging from few complete/partial responses to some stable disease. However, most patients showed no response to HDACi monotherapy for unknown reasons. Here we show that sensitivity and resistance to the hydroxamate HDACi, PXD101, can be modeled in DLBCL cell lines. Sensitivity is characterized by G2/M arrest and apoptosis and resistance by reversible G1 growth arrest. These responses to PXD101 are independent of several negative prognostic indicators such as DLBCL subtype, BCL2 and MYC co-expression, and p53 mutation, suggesting that HDACi might be used effectively against highly aggressive DLBCL tumors if they are combined with other therapeutics that overcome HDACi resistance. Our investigation of mechanisms underlying HDACi resistance showed that cyclin-dependent kinase inhibitors (CKIs), p21 and p27, are upregulated by PXD101 in a sustained fashion in resistant cell lines concomitant with decreased activity of the cyclin E/cdk2 complex and decreased Rb phosphorylation. PXD101 treatment results in increased association of CKI with the cyclin E/cdk2 complex in resistant cell lines but not in a sensitive line, indicating that the CKIs play a key role in G1 arrest. The results suggest several treatment strategies that might increase the efficacy of HDACi against aggressive DLBCL.

Belinostat and vincristine demonstrate mutually synergistic cytotoxicity associated with mitotic arrest and inhibition of polyploidy in a preclinical model of aggressive diffuse large B cell lymphoma

“…Although mitotic cells appear to accumulate in SUDHL4 and SUDHL8 cells in response to belinostat, these cell lines eventually arrest in G1. 15 It is possible that the increase in mitotic cells is due to a mitotic delay that is eventually resolved. We therefore examined high magnification images of the H3S10Ph-stained cells for evidence of anaphase mitotic figures, which would indicate that cells are progressing beyond prometaphase, when the spindle assembly checkpoint (SAC) is activated.…”

“…15 One is characterized by a reversible arrest in the G1 phase of the cell cycle within 24-48 h of treatment without accompanying apoptosis. In contrast, the second response is characterized by G2/ M arrest within 24 h and subsequent onset of apoptosis by 48 hours.…”

“…5D shows that belinostat treatment for 48 h induced G1 arrest as indicated by increased expression of the cyclin-dependent kinase inhibitor, p27. 15 To measure cell death, we carried out Annexin V/PtdIns assays. As expected, treatment with either drug alone has little effect on PARP cleavage (Fig.…”

“…Previously, we identified 2 major responses to the hydroxamate HDACi, belinostat in our model system, including 1) a reversible, cytostatic arrest in G1 and 2) arrest in G2/M that is followed by apoptosis. 15 We showed that the reversible G1 arrest, which we designated as a form of HDACi resistance, is associated with sustained HDACi-induced expression of the cyclin-dependent kinase inhibitors, p21 and p27, as well as their inhibition of the Cyclin E/cdk2 complex through increased association. 15 A major cause of HDACi-induced tumor cell death has been attributed to perturbed mitotic progression.…”

“…15 We showed that the reversible G1 arrest, which we designated as a form of HDACi resistance, is associated with sustained HDACi-induced expression of the cyclin-dependent kinase inhibitors, p21 and p27, as well as their inhibition of the Cyclin E/cdk2 complex through increased association. 15 A major cause of HDACi-induced tumor cell death has been attributed to perturbed mitotic progression. 16 These drugs have been reported to activate the spindle assembly checkpoint (SAC) resulting in an accumulation of cells in pro-metaphase.…”