RARα2 expression is associated with disease progression and plays a crucial role in efficacy of ATRA treatment in myeloma

Abstract

IntroductionMultiple myeloma (MM) is a plasma cell malignancy. 1,2 Although high-dose therapy and tandem bone marrow autotransplantation can produce higher response rates and longer survival than standard chemotherapy, MM remains largely incurable by current therapeutic strategies. 3,4 Furthermore, clinical outcomes of patients with MM are extremely heterogeneous, with survival ranging from only a few months to more than 15 years. [4][5][6] Increasing evidence suggests that genetic heterogeneity in MM cells largely accounts for the divergent clinical outcomes. 6 We have been pursuing the genetic characteristics of MM for nearly 10 years, which has contributed to the classification of MM diseases. [7][8][9][10][11] However, it remains unknown how to transform MM into a curable disease. Therefore, identification of new targets and genetic alterations, especially those for which there are available drugs, is important.All-trans retinoic acid (ATRA)-based regimens have been used in the therapy of acute promyelocytic leukemia (APL) for more than 20 years and have dramatically raised the 5-year disease-free survival to more than 70%. Furthermore, the toxicity of ATRA has been limited compared with conventional cytotoxic chemotherapy. 12 The high efficacy of retinoic acid (RA)-based therapy in APL and its well-documented safety profile have stimulated considerable interests in the treatment of other malignancies, [13][14][15] including myeloma. 16,17 Despite many efforts, however, clinical benefits of ATRA treatment in other tumors have been minor. RAR␣ has 2 major isoforms, RAR␣1 and RAR␣2, which differ in their expression patterns and N-terminal AF-1 functional domains. 18 Both RAR␣1 and RAR␣2 are considered specific receptors for RA-based reagents. However, the specific cellular roles of RAR␣1 and RAR␣2 are still unclear. In this study, we found that RAR␣2 plays a crucial role in myeloma progression, and more importantly, in mediating RA-based therapy of RAR␣2 ϩ MM diseases. Methods Study subjectsCD138 ϩ myeloma cell samples were obtained from patients who were enrolled on the National Institutes of Health (NIH)-sponsored clinical trials UARK 98-026 (Total Therapy 2 [TT2]) and UARK 03-033 (TT3). The Institutional Review Board of the University of Arkansas for Medical Sciences approved the research studies, and all subjects provided written informed consent in accordance with the Declaration of Helsinki.We have an inventory of more than 30 myeloma cell lines in our laboratory. Cells from these cell lines were cultured in RPMI 1640 containing 10% heat-inactivated fetal calf serum (FCS), 2 mM L-glutamine (Gibco), penicillin (100 U/mL), and streptomycin (100 g/mL) in a humidified incubator at 37°C in 5% CO 2 . Gene expression profilingPlasma cell purifications and gene expression profiling, using the Affymetrix U133Plus2.0 microarray, were performed as previously described. 9,11 Signal intensities were preprocessed and normalized by GCOS1.1 software (Affymetrix). 9,11 Myeloma cell samples from 80 newly diagnosed patien...

RARα2 expression confers myeloma stem cell features

“…We had previously shown that newly diagnosed MM patients with high baseline expression of RARa2 in CD138 selected myeloma cells had an inferior outcome. 9 We now found that RARa2 expression in CD138 selected myeloma cells was positively correlated with Oct4, Sox2, Nanog, Lin28A, TCF1, CCND1, and ABCC3 expression in 23 primary myeloma samples (P , .05), explaining the poor prognosis of these patients. These observations strongly suggest that RARa2 may play a crucial role in maintaining MM stem cell features.…”

“…8 We previously reported that the 30% of newly diagnosed myeloma patients, who expressed the retinoic acid receptor alpha2 (RARa2) in their CD138 selected plasma cells, had a significantly inferior outcome. 9 RARa2 expression was also highly significantly increased in myelomas rapidly relapsing after transplantation compared with paired baseline samples. 9 These findings strongly suggest the existence at diagnosis of a RARa2 expressing drugresistant subclone, which can be CD138…”

“…9 RARa2 expression was also highly significantly increased in myelomas rapidly relapsing after transplantation compared with paired baseline samples. 9 These findings strongly suggest the existence at diagnosis of a RARa2 expressing drugresistant subclone, which can be CD138…”

“…Also, RARa2 expression was significantly increased in CD138 selected myeloma cells at relapse compared with those at diagnosis. 9 We performed quantitative real-time PCR analysis to examine RARa2 expression in MM cell lines and a peripheral blood sample from a MM patient and found higher levels of RARa2 in the CD138 2 fraction of MM cell lines and SP/k 1 fraction in primary sample compared with non-SP/k 1 cells. We had previously shown that newly diagnosed MM patients with high baseline expression of RARa2 in CD138 selected myeloma cells had an inferior outcome.…”

“…9,[24][25][26] The details were described in the supplemental Data (available on the Blood Web site).…”