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438 BCL6 Is Critical to Overcome Oncogene-Induced Senescence in RAS-Mediated B Cell Transformation

Disordered Gene Expression in Hematologic Malignancy, including Disordered Epigenetic Regulation
Program: Oral and Poster Abstracts
Type: Oral
Session: 602. Disordered Gene Expression in Hematologic Malignancy, including Disordered Epigenetic Regulation: Mouse Models of Epigenetic Regulation
Sunday, December 4, 2016: 5:45 PM
Marriott Grand 5-6 (Marriott Marquis San Diego Marina)

Lai N Chan, PhD1*, Christian Hurtz, PhD2*, Gang Xiao, PhD1*, Seyedmehdi Shojaee, PhD1*, Rebecca Caeser3*, Huimin Geng, PhD1*, Ari Melnick, MD4 and Markus Müschen, MD, PhD1

1Department of Laboratory Medicine, University of California San Francisco, San Francisco, CA
2Division of Hematology and Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA
3University of California San Francisco, San Francisco, CA
4Division of Hematology/Oncology, Weill Cornell Medicine, New York, NY

Background and Hypothesis: The transcriptional repressor and proto-oncogene BCL6 is a therapeutic target in subtypes of diffuse large B cell lymphoma (DLBCL) and modulates drug-resistance in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL; Duy et al., Nature 2011). BCL6 was shown to be a critical factor that bypasses p53-dependent senescence and thereby enables RAS-driven transformation of mouse embryonic fibroblasts (Shvarts et al., Genes Dev. 2002). Given that ~50% of pediatric ALL cases carry genetic lesions that lead to hyperactivated RAS-ERK signaling (Zhang et la., Blood 2012), we examined the role of BCL6 in RAS-driven pre-B ALL and identified a novel mechanism by which RAS-ERK signaling can mediate BCL6 expression.

Results: Using a doxycycline-inducible TetOn- NRASG12D vector system, we found that inducible activation of RAS-ERK signaling strongly upregulated BCL6 expression at both the mRNA (~350-fold) and protein (~50-fold) levels in murine pre-B cells. Increases in BCL6 expression were abrogated upon treatment with a MEK inhibitor (PD325901). In addition, Cre-mediated deletion of Mapk1 suppressed upregulation of BCL6 expression upon imatinib treatment in BCR-ABL1-driven pre-B ALL cells. These findings suggested that elevated expression of BCL6 is a consequence of ERK activation. Previously, we demonstrated that BCL6 expression is negatively regulated by STAT5 in BCR-ABL1 pre-B ALL (Duy et al., Nature 2011). Interestingly, oncogenic NRASG12D inhibited phosphorylation of STAT5-Y694 by activating the inhibitory protein tyrosine phosphatase Ptpn6. Cre-mediated deletion of Ptpn6 induced STAT5 activity. Furthermore, loss of Ptpn6 function abrogated upregulation of BCL6 expression induced by imatinib in BCR-ABL1 pre-B ALL. Taken together, RAS-ERK signaling induces BCL6 expression by suppressing STAT5 activity. To directly test the role of BCL6 in RAS-transformed pre-B ALL, we generated a novel mouse model for inducible Cre-mediated deletion of Bcl6 exons 5-10, flanked by loxP sites. Inducible deletion of Bcl6 in NRASG12D-transformed pre-B ALL cells led to rapid depletion from the cell culture and reduced colony forming ability in vitro. These findings suggested that BCL6 is required for maintenance of fully established RAS-transformed ALL. Notably, we found that initiation of NRASG12D-driven leukemia in vivo depends on BCL6 as NRASG12D ALL failed to give rise to leukemia in the absence of Bcl6 in transplant recipient mice. Studying a diagnostic (KRAS wild-type) and a relapsed (KRASG12V) sample from one pre-B ALL patient revealed increased BCL6 expression in KRASG12V relapsed ALL cells. In addition, selective sensitivity to PD325901 and a retro inverso BCL6 peptide inhibitor (RI-BPI) was observed in KRASG12V relapsed ALL cells. Finally, RI-BPI prolonged overall survival of recipient mice transplanted with KRASG12V relapsed ALL cells in vivo. 

Conclusions: In summary, we demonstrated a novel mechanism by which oncogenic RAS signaling induces expression of BCL6, and showed that BCL6 is critical for RAS-driven transformation in pre-B ALL. Importantly, ALL clones often acquire drug resistance and activating mutations in the RAS pathway (Bhojwani and Pui, Lancet Oncol. 2013). Our findings suggest that pharmacological inhibition of BCL6 may provide a novel therapeutic avenue to overcome drug-resistance and prevent leukemia relapse after initial remission in RAS-driven ALL.

Disclosures: Hurtz: Incyte: Research Funding. Melnick: Roche: Consultancy, Research Funding; GSK: Research Funding; Eli Lilly: Consultancy, Research Funding; Epizyme: Consultancy; Boehringer-Ingelheim: Consultancy; Janssen: Research Funding.

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