Expression, Role in Transformation and Prognostic Significance of the Transcription Factor Yin Yang 1 (YY1) in Non-Hodgkin’s Lymphoma: Analyses in NHL Tissues by Experimental and Bioinformatic Approaches
Authors: Libra Massimo , Castellano Giancarlo , Baritaki Stavroula , Huerta-Yepez Sara , Ligresti Giovanni , Militello Loredana , Malaponte Grazia , Stivala Franca , Canevari Silvana , Bonavida Benjamin 
Published: April 2020

Abstract
Objective and Rationale Non-Hodgkin lymphomas (NHL) are a heterogeneous group of lymphoproliferative malignancies with variable patterns of behavior and responses to treatment. Although at present a significant number of high grade lymphoma patients can be cured with intensive regimens of cytotoxic and immunotherapeutic drugs, still these patients may relapse or become resistant to treatment. Therefore, there is a need to identify novel factors involved in lymphoma development and progression and hopefully to be used as targeted therapies. Thus, the objective of the present study was to explore the potential involvement of Yin Yang 1 (YY1) in the development as well as in the prognosis of NHL.
Hypothesis Our previous studies showed that YY1 is overexpressed in several human cancer cell lines and its expression correlates with resistance to chemo- immune-mediated apoptosis. Prognostic and diagnostic significance of YY1 has been recently shown in prostate cancer. Thus, we hypothesized that YY1 overexpression in NHL may be involved in transformation and act as a prognostic biomarker.
Methods and Designs Overexpression of YY1 has been shown by western blot and RT-PCR in NHL cell lines compared to normal B-cells. Validation of in vitro results has been performed by immunohistochemistry (IHC) and bioinformatics in NHL tissues. IHC analyses were performed in two commercial NHL tissue arrays containing 104 samples.
Results Gene expression analysis and association with clinical features of the transcription factor YY1 were evaluated using three different datasets of publicly available microarray data from lymphoma tumor-biopsy specimens. By analyzing the data set (221 NHL and 25 normal B-cells) by Basso et al., (Nat Genet 37:382, 2005) higher YY1 transcript levels were found in lymphoma tissues (BL and DLBCL) in comparison to normal B-cells. No significant differences were found with the other lymphoma histotypes. This class comparison analysis revealed that YY1 could be involved in the transformation of B cells giving rise to high-grade lymphomas. Expression levels of YY1 were analyzed in association with the available main clinical and biological parameters using the expression data of two different high grade lymphoma studies with a total of approximately 400 samples (Rosenwald et al., N Engl J Med.346:1937, 2002; Hummel et al., N Engl J Med.354:2419, 2006). Upregulated expression of YY1 was found in the subgroups of patients with positive proliferation signature scores with respect to those with negative proliferation signature scores. A significant correlation was found between BCL6, post-germinal center (GC) marker and YY1 expression. The presence of BCL6 protein in the tumors is associated with high levels of YY1 gene transcription. These data confirm that YY1 is mostly overexpressed among high-grade lymphomas that usually are post-GC transit. Survival analysis in both datasets reveal that higher levels of YY1 gene transcription are associated with poor outcome. Finally, biological network analysis, using the Proprietary Ingenuity Pathway analysis software, was performed to better understand the biological significance of these findings. Among all genes present in the data set by Hummel et al. (2006) a significant positive correlation with YY1 expression was found in 374 genes. In this data set, Top Networks Functions associated with YY1 expression levels were cellular movement, cell morphology, cell cycle, and cell-to-cell adhesion. These analyses indicate the potential direct involvement of YY1 in the regulation of cell cycle pathways and in the regulation of cellular motility.
Conclusions and Implications Overall, the findings reveal that YY1 is involved in lymphoma development and may be useful as a biomarker of NHL transformation as well as a potential target for therapeutic interventions.