Induction of Apoptosis in Multiple Myeloma by Ligands of Peroxisome Proliferator-Activated Receptor γ (PPAR-γ).
Authors: Eucker Jan , Baengeroth Katharina , Zavrski Ivana , Krebbel Holger , Zang Chuanbing , Heider Ulrike , Jakob Christian , Elstner Elena , Possinger Kurt , Sezer Orhan 
Published: April 2020

Abstract
Peroxisome proliferator-activated receptor γ (PPAR-g) is a member of a nuclear receptor superfamily, which is expressed in different tumor tissues. Activation of PPAR-γ by its ligands has been shown to reduce tumor growth, interfere with tumor cell differentiation, and induce apoptosis in a variety of human malignancies including solid tumors like colon, breast, lung, liver, prostate cancer, as well as hematological malignancies like myeloid leukemia. Recently, it has been shown that both human B-lymphocytes and B-lymphomas express PPAR-γ and induce apoptosis. 15-deoxy-delta-12,14-prostaglandin J2 (15d-PGJ2) is a natural activator of PPAR-γ. Thiazolidinediones, including troglitazone, rosiglitazone (RGZ), and pioglitazone (PGZ), comprise a group of synthetic PPAR-γ agonists that are currently in use for the treatment of type 2 diabetes mellitus, and have revealed anti-tumor activity in vitro. We investigated in five human multiple myeloma cell lines (LP-1, U-266, RPMI-8226, OPM-2 and IM-9) and sorted human bone marrow myeloma cells whether treatment with PGZ, RGZ or 15d-PGJ2 inhibited tumor cell growth. Expression of PPAR-γ protein was demonstrated by western blot analysis in these cell lines. All 5 cell lines were sensitive to the PPAR-γ agonists. MTT assays revealed growth arrest induced by the natural activator of PPAR-γ 15d-PGJ2 and a lower antiproliferative effect with PGZ and RGZ in a dose dependent manner. At a dose of 50 μM 15d-PGJ2 cell proliferation was reduced to values between 0% and 26% in all multiple myeloma cell lines tested. In most cell lines the anti-proliferative effect was already detectable at 10 μM. At a dose level of 50 μM PGZ cell proliferation was reduced in MTT assay after 48 hours of incubation to 48% in LP-1, 52% in IM-9, 56% in OPM-2, 72% in U-266 and 77% in RPMI-8226. Comparable results were obtained with RGZ. Induction of apoptosis was indicated by annexin V staining. Cell lines were incubated with 50 μM of PPAR-γ agonists, a concentration which had been proven to be effective for growth inhibition in MTT assay before. Again, 15-dPGJ2 was more effective than PGZ and RGZ. All of the 15d-PGJ2 treated cell lines revealed specific apoptosis ranging between 60% and 92%. Apoptosis induced by PGZ in U-266, RPMI-8226-S, IM-9, and OPM-2 cell lines ranged between 17% and 43%, for RGZ it ranged between 20% and 50%. Furthermore, in sorted bone marrow plasma cells from myeloma patients induction of apoptosis was detected. Bone marrow multiple myeloma cells from five different patients were tested. The specific apoptosis rate induced by 15-dPGJ2 lay between 29% and 96%. Apoptosis induced by PGZ showed interindividual differences. In the myeloma cells from four patients the rate of specific apoptosis ranged between 9% and 28%, but in one patient induction of apoptosis was observed neither with PGZ nor with RGZ. For RGZ, the rate of apoptosis induced in the myeloma cells from the other four patients ranged between 7% and 26%. The rate of specific apoptosis induced by 15D-PGJ2 was not statistically different for sorted human bone marrow myeloma cells sensitive versus refractory to conventional chemotherapy with anthracyclines and alkylating agents (p = 0.8). This is one of the first studies evaluating PPAR-γ expression and its therapeutical implications in human multiple myeloma cells. Thiazolidinediones comprise anti-myeloma activity and should be explored further for the treatment of multiple myeloma.