Upregulated expression of the PSMB5 gene may contribute to drug resistance in patient with multiple myeloma when treated with bortezomib-based regimen. Exp Hematol.2011;39(12):1117-8.

Multiple myeloma (MM) is a hematological malignancy of which complete response (CR) is rarely achieved by routine chemotherapy. Proteasome inhibitors have been recognized as a new generation of chemotherapeutic agents. Bortezomib is a reversible inhibitor that targets primarily the chymotrypsin-like activity at proteasome β5 subunit (PSMB5). Bortezomib not only promotes apoptosis of myeloma cells, but also sensitizes these cells to other chemotherapeutic drugs [1]. High CR and overall response rates have been reported when bortezomib alone or combined with other drugs used to treat refractory or newly diagnosed MM [2-3]. However, resistance to bortezomib develops quickly in many clinical patients [4]. Our previous study [5-7] and some other investigators’s study [8-9] have confirmed mutation and/or overexpression of the PSMB5 gene were important mechanisms associated with bortezomib resistance in leukemia and MM cell lines. Moreover, study of Yang et al showed the anticancer effect of bortezomib can be enhanced through PSMB5 downregulation in cancer cell lines [10]. In this study we explored the possible mechanism of bortezomib resistance in clinical MM patients.上海长海医院血液内科吕书晴

We collected the MM cells from bone marrow of MM patients before treatment with a 4-week bortezomib-based regimen (BADT: intravenous bortezomib 1.0 mg/m² and epirubicin 12 mg/m² on days 1, 4, 8, and 11, dexamethasone 20 mg on days 1, 2, 4, 5, 8, 9, 11, and 12; oral thalidomide 100 mg/m² daily). MM cells were collected again if CR not achieved after 6 cycles BADT treatment. mRNA expression levels of the PSMB5 gene in MM cells were detected by quantitation real-time reverse transcription polymerase chain reaction (QT-PCR). Mutation of the PSMB5 gene was also detected by DNA sequencing. Normal bone marrow mononuclear cells collected from a healthy volunteer were used as control. The detail of QT-PCR and DNA sequencing were similar as our previous report [5-7].

Three refractory MM patients entered this study. Case 1 was a male patient of 56 years old with MM of IgG λ subtype, Durie?Salmon (D-S) stageⅠA, international staging system (ISS) stage Ⅲ. Case 2 was a female patient of 57 years old with MM of IgG λ subtype, D-S stage ⅢA, ISS stage Ⅲ. Case 3 was a male patient of 51 years old with MM of light chain λ subtype, D-S stage ⅢA, ISS stage Ⅲ. Case 1 achieved CR after one cycle BADT treatment, following 3 cycles consolidation treatment of BADT, then he received thalidomide for maintenance therapy and have maintained CR for 40 months. Case 2 achieved CR after one cycle BADT treatment, then she received one cycle consolidation treatment of BADT, but she died of thoracic low grade fibromyxoid sarcomas which may be result from 5-year chemotherapy containing high dose alkylating agents and local radiation 5 years ago before entering this study. Case 3 didn’t achieved CR after 8 cycles BADT treatment, then he stopped bortezomib-based therapy and turned to other rescue regimens.

   The result of QT-PCR showed there were no obvious differences of the mRNA expression levels of PSMB5 between MM cells from these three patients before BADT treatment and control bone marrow mononuclear cells from a healthy volunteer. But the mRNA expression level of PSMB5 in MM cells from Case 3 after 6 cycles BADT treatment was upregulated to 5.03-fold relative to control and 5.19-fold relative to before BADT treatment (Figure 1). We also detected the sequence of the PSMB5 gene in these MM cells and control cells. No mutation was found in the MM cells collected before treatment and control cells. There was also no mutation of the PSMB5 gene found in the MM cells collected from case 3 after 6 cycles BADT treatment.

   Our previous study have confirmed overexpression of the PSMB5 gene and G322A, C323T, or G322A, C326 T conjoined mutation result in substitutions of key Ala49 and/or Ala50 of the PSMB5 protein relative to the inhibitory effect of bortezomib on chymotrypsin-like activity contribute to bortezomib resistance in a T lymphoblastic lymphoma/leukemia cell line JurkatB selected by bortezomib exposure [5-7]. Similar discovery was reported by Oerlemans et al in bortezomib resistant human monocytic/macrophage THP1/BTZ cell line [8]. Recently Ri et al also reported G322A mutation of the PSMB5 gene in two bortezomib-resistant MM cell lines, KMS-11/BTZ and OPM-2/BTZ, but no overexpression of the PSMB5 gene [9]. Our patient of case 3 is the first reported clinical patient who have acquired overexpression of the PSMB5 gene in MM cells resulting in drug resistance during treatment by botezomib-based regimen. To further investigate the mechanism of bortezomib resistance in clinical patients, especially to clarify the role of overexpression and mutations of the PSMB5 gene, a larger number of patients who have acquired bortezomib resistance during or after treatment are required.