PB2103 TRANSCRIPTOME ANALYSIS AFTER DEXAMETHASONE EXPOSURE DIFFERENTIATES RESPONSE TO GLUCOCORTICOIDS IN MULTIPLE MYELOMA
Authors: P. Krzeminski, L. Corchete, R. García-Sanz, N. Gutiérrez
Journal: HemaSphere (2019)

Background: Multiple myeloma (MM) is a plasma cell disorder, which remains incurable despite the remarkable progress made in therapeutic strategies. Glucocorticoids (GC), mainly dexamethasone, have always been part of the standard of care of MM patients, used in combination with proteasome inhibitors and immunomodulatory drugs. Surprisingly, little attention has been devoted to elucidating the mechanism of resistance to GC compared to the research made for investigating those mechanisms for other drugs. Dexamethasone is a well-known synthetic GC that acts through NR3C1 glucocorticoid receptor inducing changes in the expression of hundreds of genes. However, consequence of GC treatment can reach far beyond gene expression affecting transcripts and splicing balance. Aims: To investigate the effect of dexamethasone treatment on gene, transcript and splice event expression in MM cell lines with different sensitivity to GC. Methods Gene expression, transcriptome and splice event balance were investigated using Affymetrix Clariom™ D Arrays, which include probes that match not only to gene exons, but also to linking regions between exons, introns and UTRs. Thus, any exon change, intron retention or UTR sequence difference between experimental conditions (control vs dexamethasone 50 nM 24 h) can be detected. Five MM cell lines, MM1s, MM1r, OPM2, KMS12PE and U266 were investigated. All experiments were performed in triplicate. Raw data were quantile normalized, background corrected and log2 transformed. Differential expression analyses were conducted using the samr R package. Genes and transcripts with q-value <0.05 were considered statistically significant and were included in the KEGG pathway analysis. Results: Dexamethasone induced significant gene expression changes only in sensitive (MM1s and OPM2) cell lines compared to resistant ones (MM1r, KMS12PE and U266). Interestingly, the number of deregulated genes in MM1s was dramatically higher than in OPM2 (1832 vs 20 genes, respectively) despite the fact that both cell lines are equally sensitive to dexamethasone. Analysis of both cell lines identified six overlapping genes: FAR2, FKBP5, DDIT4, INSR, GBR14 and ZC3H12. Although expression of the majority of genes decreased upon treatment with dexamethasone in MM1s cell line, the six deregulated genes shared with OPM2 showed increased expression. Gene pathway analysis in MM1s cell line revealed that dexamethasone treatment affected mainly biological processes such as, cell cycle, DNA replication and proteasome pathways. The analysis of transcript expression showed a similar pattern to that found in gene expression profiling, with 21,484 transcripts deregulated in MM1s cell line, and only 477 changed in OPM2. Expression of transcripts was almost unaffected in resistant cell lines. Interestingly, the deregulated transcripts in MM1s were predominantly decreased, while in OPM2 the number of transcripts displaying increased expression was higher. Transcript set enrichment analysis also identified “cell cycle” and “DNA replication” as the most overrepresented pathways. The analysis of splice variants showed that the number of splice events was significantly higher in MM1s compared to OPM2, consistent with the results obtained in the gene and transcript differential expression analysis. Summary/Conclusion: Dexamethasone induces transcriptome and splice event changes that characterize glucocorticoid sensitivity, but are mainly cell line specific.