The Mir-17~92 Cluster Enhances Cell Growth and Resistance to Chemotherapy in Mantle Cell Lymphoma by Down-Regulating PTEN, PHLPP2 and BIM
Authors: Rao Enyu , McKeithan Timothy , Jiang Chunsun , Ji Ming , Iqbal Javeed , Wright George , Staudt Louis M. , Zhao Yong , Chan Wing C. , Fu Kai 
Published: April 2020

Abstract
Mantle cell lymphoma (MCL) is an aggressive B-cell malignancy and is characterized by the chromosome translocation t(11;14)(q13;q32), which results in aberrant expression of cyclin D1. The t(11;14)(q13;q32) alone is not sufficient to result in lymphoma, and additional genetic alterations are necessary. Secondary genomic alterations are frequently detected in MCL, of which chromosome 13q31–q32 gain/amplification is one of the most frequent. Amplification at chromosome 13q31–q32 targets a microRNA cluster, miR-17~92, which resides within intron 3 of c13orf25, a non-protein-coding gene at 13q31.3. The miR-17~92 cluster consists of seven miRNAs, and overexpression of the cluster accelerates the development of MYC-induced lymphoma in mice and contributes to carcinogenesis in the lung. Nevertheless, the functional role of miR-17~92 in MCL has not been investigated. Based on gene expression profiling study of 82 primary MCL biopsy specimens using whole genome Affymetrix U133 2.0 plus arrays, we found that overexpression of miR-17~92 was associated with worse overall survival in patients with MCL (p=0.021). We further demonstrated that enforced expression of miR-17~92 reduced chemotherapy-induced apoptosis in retrovirally transduced MCL cell lines. Immunoblot analysis showed that PTEN was down-modulated in MCL cells with overexpression of miR-17~92, and the level of PTEN was restored with reduction of miR-20a levels using an antisense oligonucleotide or a “sponge” plasmid with multiple tandem miR-20a binding sites. The finding was further confirmed by luciferase assays using a reporter plasmid containing the PTEN 3′UTR, and the effect was abrogated by mutating the miR-20 binding site in reporter plasmid, indicating that PTEN was downregulated through the binding of miRNA to the 3′UTR of the transcript. Furthermore, we demonstrated that overexpression of miR-17~92 in MCL cells increased the phosphorylation of AKT and its downstream targets. Interestingly, we found that the protein phosphatase PHLPP2, a negative regulator of the PI3K/Akt pathway, was also a direct target of the miR-17~92 cluster. Moreover, we found that BIM, a BH3-only proapoptotic protein, was a direct target of the miR-17~92 and down-modulation of BIM negatively impacted the intrinsic pathway of apoptosis. In summary, we demonstrated that overexpression of miR-17~92 in MCL cells downmodulates multiple proteins involved in PI3K/Akt signaling and apoptosis, and that downregulation of these targets collaboratively enhances cell growth and chemoresistance in tumor cells. As a consequence, overexpression of miR-17~92 may be associated with poorer survival in MCL patients. Our findings disclose a novel oncogenic pathway in MCL and suggest that targeting the miR-17~92 cluster may provide a novel therapeutic approach for this disease, which is incurable with current chemotherapeutic regimens.