Targeting the Over-Expressed Transcription Factor Yin-Yang 1 (YY1) Sensitizes Resistant Multiple Myeloma (MM) Cell Lines to Apoptosis by Bortezomib or Melphalan,

Abstract 3991

The transcription factor Yin-Yang 1 (YY1) is a multi-functional DNA-binding protein which can activate, repress, or initiate transcription depending on the context in which it binds. We have previously reported that YY1 is a repressor for the death receptors Fas and DR5 as well as it regulates both chemo- and immuno- resistance in many tumors such as prostate carcinoma and lymphoma. YY1 has also been reported to be a prognostic factor for some cancers. Due to the inherent resistance of multiple myeloma (MM) cell lines to various cytotoxic drugs, we examined the expression levels of YY1 in both MM cell lines and bone marrow (BM)-derived MM tissues. Analysis for YY1 was performed by immunohistochemistry (IHC) and western blot. We found that, compared to normal peripheral blood mononuclear cells and normal bone marrow, the expression of YY1 was significantly upregulated in both the cell lines and patient-derived MM tissues. The expression of YY1 was distributed in both the cytoplasm and the nucleus. Further, the intensity and frequency of cells expressing YY1 in the nuclei of MM tissues were significantly higher among patients with progressive disease as compared to patients with stable disease. In order to ascertain the role of YY1 in the maintenance of drug resistance in MM, we hypothesized that, due to the overexpression of YY1 in MM, inhibition of YY1 expression and activity should lower the threshold of resistance and render the MM cells more sensitive to drug-induced apoptosis. This hypothesis was tested using the MM U266 cell line and the drugs bortezomib and melphalan as models. The inhibition of YY1 was done by knocking down YY1 following transfection with YY1 siRNA. Analysis of the transfected cells with YY1 siRNA, but not with control siRNA, inhibited YY1 expression as analyzed by IHC and western blot. The U266 cells were first transfected with YY1 siRNA or control siRNA for 48 hours and then treated with various concentrations of bortezomib (2.5 and 5.0 nM) or melphalan (10 and 20 μM) that were not toxic to the cells for 48 hours. The cells were analyzed for apoptosis by activation of the effector caspase 3. The findings demonstrate that YY1/siRNA-treated cells were sensitized to apoptosis by bortezomib or melphalan. Analysis of the underlying mechanism by which the inhibition of YY1 sensitizes the MM cells via regulation of the apoptotic pathways will be presented. We are also currently testing specific chemical inhibitors for YY1 that are being tested both in vitro and pre-clinically in mice.

The present findings demonstrate that YY1 is a potential target for therapeutic intervention and the combination of YY1 inhibitors with non-toxic doses of bortezomib, melphalan or other cytotoxic drugs may be beneficial for the treatment of MM patients who are no longer responding to current treatments. Further, we propose that, based on our findings that the level and nuclear localization of YY1 dictates disease progression in a subset of patients, inhibition of YY1 with treatment may reduce the frequency of patients with progressive disease. *Contributed equally