Gene expression-based prediction of myeloma cell sensitivity to histone deacetylase inhibitors

Abstract

Background: Multiple myeloma (MM) is still a fatal plasma cell cancer. Novel compounds are currently clinically tested as a single agent in relapsing patients, but in best cases with partial response of a fraction of patients, emphasising the need to design tools predicting drug efficacy. Histone deacetylase inhibitors (HDACi) are anticancer agents targeting epigenetic regulation of gene expression and are in clinical development in MM.

Methods: To create a score predicting HDACi efficacy, five MM cell lines were treated with trichostatin A (TSA) and gene expression profiles were determined.

Results: The expression of 95 genes was found to be upregulated by TSA, using paired supervised analysis with Significance Analysis of Microarrays software. Thirty-seven of these 95 genes had prognostic value for overall survival in a cohort of 206 newly diagnosed MM patients and their prognostic information was summed up in a histone acetylation score (HA Score); patients with the highest HA Score had the shorter overall survival. It is worth noting that MM cell lines or patients' primary MM cells with a high HA Score had a significant higher sensitivity to TSA, valproic acid, panobinostat or vorinostat.

Conclusion: In conclusion, the HA Score allows identification of MM patients with poor survival, who could benefit from HDACi treatment.

Figure 1

Histone acetylation score in normal and malignant PCs. Histone acetylation score in normal BMPCs (N=7), in premalignant PCs of patients with monoclonal gammopathy of undetermined significance (MGUS, N=5), in MM cells of patients with intramedullary MM (N=206) and in HMCLs (N=40). ** Indicate that the score value is significantly different with a P-value <0.01.

Figure 2

Prognostic value of HA Score in MM. (A) Patients of the HM cohort were ranked according to increased HA Score and a maximum difference in OS was obtained with HA Score=−11.3 splitting patients into high-risk (42.7%) and low-risk (57.3%) groups. Histone acetylation score also had a prognostic value of an independent cohort of 345 patients from University of Arkansas for Medical Science (UAMS) treated with TT2 therapy (UAMS-TT2 cohort). The parameters to compute the HA Score of patients of UAMS-TT2 cohort and the proportions delineating the two prognostic groups were those defined with HM cohort. (B) Clustergram of HA Score genes ordered from best to worst prognosis. The level of the probe set signal is displayed from low (deep blue) to high (deep red) expression. MM patients (N=206) were ordered by increasing GE-based risk score.

Figure 3

Histone acetylation score in MM molecular subgroups. The HA Score was computed for MMCs of patients belonging to the eight groups of the University of Arkansas for Medical Science (UAMS) molecular classification of multiple myeloma, using UAMS-TT2 cohort. *Score value is significantly higher in the group compared with all the patients of the cohort (P<0.05). **Score value is significantly lower in the group compared with all the patients of the cohort (P<0.05). Abbreviations: CD1, cyclin D1; CD2, cyclin D2; HY, hyperdiploid; LB, low bone disease; MF, MAF; MS, MMSET; MY, myeloid; PR, proliferation.

Figure 4

Histone acetylation score predicts for sensitivity of HMCLs to TSA. HMCLs with high HA Score (N=5) exhibit significant higher TSA sensitivity compared with HMCLs with low HA Score (N=5). Human myeloma cell lines were cultured for 4 days in 96-well flat-bottom microtitre plates in RPMI 1640 medium, 10% fetal calf serum, 2 ng ml⁻¹ IL-6 culture medium (control) and graded TSA concentrations. Data are mean values ±standard deviation (s.d.) of five experiments determined on sextuplet culture wells.

Figure 5

Histone acetylation score predicts for TSA sensitivity of primary myeloma cells of patients. Mononuclear cells from tumour samples of 13 patients with MM were cultured for 4 days in the presence of IL-6 (2 ng ml⁻¹) with or without graded TSA concentrations. At day 4 of culture, the count of viable MMCs was determined using CD138 staining by flow cytometry. Black color represents patients with high HA Score (N=8) and white represents patients with low HA Score values (N=5).

Figure 6

Histone acetylation score predicts for sensitivity of HMCLs to panobinostat, valproic acid (VPA) or SAHA HDACi. Human myeloma cell lines with a high HA Score (N=5) exhibit significant higher sensitivity to panobinostat, VPA or SAHA compared with HMCLs with a low HA Score (N=5). Human myeloma cell lines were cultured for 4 days in 96-well flat-bottom microtitre plates in RPMI 1640 medium, 10% fetal calf serum, 2 ng ml⁻¹ IL-6 culture medium (control) and graded concentration of panobinostat (A), VPA (B) or SAHA (vorinostat) (C). Data are mean values± standard deviation (s.d.) of five experiments determined on sextuplet culture wells.