ABCG2 is a direct transcriptional target of hedgehog signaling and involved in stroma-induced drug tolerance in diffuse large B-cell lymphoma

Abstract

Successful treatment of diffuse large B-cell lymphoma (DLBCL) is frequently hindered by development of resistance to conventional chemotherapy resulting in disease relapse and high mortality. High expression of anti-apoptotic and/or drug transporter proteins induced by oncogenic signaling pathways has been implicated in the development of chemoresistance in cancer. Previously, our studies showed high expression of ATP-binding cassette drug transporter ABCG2 in DLBCL correlated inversely with disease-free and failure-free survival. In this study, we have implicated activated hedgehog (Hh) signaling pathway as a key factor behind high ABCG2 expression in DLBCL through direct upregulation of ABCG2 gene transcription. We have identified a single binding site for GLI transcription factors in the ABCG2 promoter and established its functionality using luciferase reporter, site-directed mutagenesis and chromatin-immunoprecipitation assays. Furthermore, in DLBCL tumor samples, significantly high ABCG2 and GLI1 levels were found in DLBCL tumors with lymph node involvement in comparison to DLBCL tumor cells collected from pleural and/or peritoneal effusions. This suggests a role for the stromal microenvironment in maintaining high levels of ABCG2 and GLI1. Accordingly, in vitro co-culture of DLBCL cells with HS-5 stromal cells increased ABCG2 mRNA and protein levels by paracrine activation of Hh signaling. In addition to ABCG2, co-culture of DLBCL cells with HS-5 cells also resulted in increase expression of the antiapoptotic proteins BCL2, BCL-xL and BCL2A1 and in induced chemotolerance to doxorubicin and methotrexate, drugs routinely used for the treatment of DLBCL. Similarly, activation of Hh signaling in DLBCL cell lines with recombinant Shh N-terminal peptide resulted in increased expression of BCL2 and ABCG2 associated with increased chemotolerance. Finally, functional inhibition of ABCG2 drug efflux activity with fumitremorgin (FTC) or inhibition of Hh signaling with cyclopamine-KAAD abrogated the stroma-induced chemotolerance suggesting that targeting ABCG2 and Hh signaling may have therapeutic value in overcoming chemoresistance in DLBCL.

Transcriptional Regulation of Serine/Threonine Protein Kinase (AKT) Genes by Glioma-associated Oncogene Homolog 1

“…This activation of GLI1 is not due to amplifications of GLI1 (22) or mutations of Hhrelated genes (23,24). Our data support that the Hh pathway in DLBCL is aberrantly activated as its activation is in part mediated by external Hh ligands (autocrine and paracrine Hh signaling loops) (5,16) but also intrinsically by cross-talks with other oncogenic pathways (22).…”

Hedgehog signaling pathway affects the sensitivity of hepatoma cells to drug therapy through the ABCC1 transporter

“…26 These findings are further supported by the observation that ABCB1 and ABCG2 promoter regions have the consensus sequence of Gli-binding site, and Gli-1 directly bound to the consensus GACCACCCA-like motif located in the promoter regions of ABCB1 and ABCG2 to regulate their transcription in B-cell lymphoma and ovarian cancer cells. 27,28 However, our results demonstrate that ABCC1 was highly expressed in Huh-7-DN and Huh-7-trans cells; in Figure 7 The effects of LDE225 and itraconazole on expression of Gli-1, Gli-2, and ABCC1 in Huh-7-DN subpopulation. Huh-7-DN cells were exposed to LDE225 or itraconazole at various concentrations as indicated for 24 h. mRNA levels of Gli-1, Gli-2, and ABCC1 in Huh-7-DN subpopulation were determined by quantitative RT-PCR, using untreated Huh-7 cells as a control.…”

Active IKKβ promotes the stability of GLI1 oncogene in diffuse large B-cell lymphoma

“…We further demonstrated that the canonical Hh ligand-PTCH1-SMO-GLI1 axis plays important roles in cell proliferation, survival, and chemotolerance in this lymphoma subtype. [12][13][14][15][16] Regulators of GLI1's activities include SNF5, a core subunit of the adenosine trisphophate (ATP)-dependent SWItch/Sucrose NonFermentable chromatin remodeling complex, which modulates its transcriptional activities. 17 Hh signaling, meanwhile, promotes the transcriptional activity of both GLI1 and GLI2 proteins by promoting their deacetylation via HDAC1 upregulation.…”