12 / 15
Page 34
allied
academies
August 23-24, 2018 | London, UK
Hematology and Oncology
2
nd
International Conference on
Journal of Hematology and Blood Disorder | Volume 2
FBXO11 is a frequently mutated oncosuppressor in Burkitt Lymphoma
Roberto Chiarle
Harvard Medical School, USA
Introduction: FBXO11 is a ubiquitin ligase involved in the
degradationofBCL6,akeyoncogeneinlymphomapathogenesis.
We previously described inactivating mutations of the FBXO11
gene in Diffuse Large B Cell Lymphoma (DLBCL) (Duan et al,
Nature 2012). Thus, FBXO11 acts as an oncosuppressor inDLBCL
by promoting the accumulation of BCL6. In the present work we
searched for FBXO11 mutations in BCL6-positive lymphomas
and we investigated its role in lymphoma development in vivo.
Methods: We sequenced the FBXO11 coding sequence in
100 cases of Follicular Lymphoma (FL), 36 cases of Burkitt
Lymphoma (BL), 8 BL cell lines and 8 Anaplastic Large cell
lymphoma cell lines, all BCL6-positive lymphomas, and 50
cases of Marginal Zone B Cell Lymphoma (MZL), with variable
expression of BCL6. We functionally validated the FBXO11
mutations by testing their ability to induce BCL6 degradation.
We then applied the CRISPR/Cas9 system to disrupt the
endogenous FBXO11 gene in BL cells and evaluated its effect
on BCL6 stability. We tested the sensitivity of FBXO11 knock-out
(KO) BL cells to a BCL6 inhibitor (FX1) alone or in combination
with chemotherapy (doxorubicin). To dissect the in vivo role
of FBXO11 in lymphomagenesis we generated conditional
FBXO11 KO mice (CD19/Cre-FBXO11fl/fl) and we crossed them
with Eμ-myc transgenic mice to investigate whether FBXO11
inactivation cooperates with c-myc in lymphomagenesis.
Results: We identified FBXO11 mutations in BL cases and cell
lines (10/44, 22.7%), one case of FL (1/100) and one case of
MZL (1/50). Recurrent FBXO11 mutations in BL were further
identified in publicly available sequencing databases of 66
BL cases (13/66, 19.7%) (Love et al Nat Genet 2012, Grobner
et al Nature 2018). BL mutations found in our series were
mostly missense and splice-site mutations located in the
functional domains and all of them impaired FBXO11 ability
to induce BCL6 degradation. CRISPR/Cas9 mediated KO of
FBXO11 in BL cells resulted in an almost complete stabilization
of BCL6, thus suggesting that FBXO11 is the main ubiquitin
ligase that controls BCL6 stability in BL. FBXO11-KO BL cells
showed increased resistance to standard chemotherapy as
well as increased sensitivity to BCL6 inhibition compared to
the FBXO11 WT BL cells. The simultaneous combination of
FX1 with doxorubicin restored the sensitivity of FBXO11-KO
BL cells to standard chemotherapy. Finally, we observed an
acceleration of lymphoma development in the CD19/Cre-
FBXO11fl/fl mice crossed with Eμ-myc transgenic mice. The
lymphomas showed histologic features of high-grade disease
with a more mature B-cell phenotype, stabilization of BCL6 and
reduced apoptotic fraction compared to Eμ-myc only tumors.
Conclusion: Our results demonstrate that FBXO11 is frequently
mutated in BL with a mutation frequency of about 20% of
cases. Thus, FBXO11 is one of the top five most frequently
mutated genes in BL. Biological experiments in vitro and
in vivo show that FBXO11 deletion cooperates with c-myc
in accelerating lymphomagenesis. Remarkably, FBXO11
deletion in the context of c-myc overexpression generates
more mature lymphomas that closely resemble human BL,
providing a novel tool for potential preclinical testing of
therapies with BCL6 inhibitors. Indeed, the combination of
BCL6-targeted therapy restored the sensitivity of FBXO11-
KO BL cells to standard chemotherapy suggesting potential
combinational strategies for the treatment of BL patients
e:
roberto.chiarle@childrens.harvard.edu