The DLEU2/Mir-15a/Mir-16-1 Locus, Commonly Deleted in B-Cell Chronic Lymphocytic Leukemia (CLL), Controls B-Cell Compartment Expansion and Its Deletion Leads to CLL in Mice

B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) is derived by the malignant transformation of mature B cells. Approximately 50% of CLL cases have monoallelic or biallelic deletions of the chromosomal region 13q14, suggesting the presence of a tumor suppressor involved in CLL pathogenesis. Similar deletions are associated with lymphoproliferative disorders, such as CD5⁺ monoclonal B-cell lymphocytosis (MBL), that may predispose to CLL, B-cell non-Hodgkin lymphoma (NHL) and multiple myeloma. We and others previously identified a minimal deleted region (MDR) at 13q14 which contains:

1. the DLEU2 gene, encoding a long (1.2 Kb) non-coding RNA that is necessary for miR-15a/miR-16-1 production and forms a processed mRNA that is stable in the cytoplasm suggestive of an independent function; and

2. the microRNAs miR-15a/miR-16-1, located within the intronic region of DLEU2.

Although miR-15a/miR-16-1 have been proposed as the culprit of the deletion, no conclusive proof of the respective roles of the DLEU2 or miR-15a/miR-16-1 genes in tumor suppression has been obtained. We therefore investigated the consequences of eliminating these genetic elements in vivo by establishing two transgenic mouse lines that carry deletions of either the ~120 kb large MDR, including dLeu2 and miR-15a/miR-16-1, or of miR-15a/miR-16-1 alone.

Young MDR^−/− and miR-15a/miR-16-1^−/− mice did not display phenotypic abnormalities, and showed normal lymphoid development. However, between 12–18 months, both MDR^−/− and miR-15a/miR-16-1^−/− mice developed MBL (~15% of cases), CLL/SLL with multiorgan involvement (~20%), or CD5⁻ NHL subtypes (~5%). Analogous lymphoproliferations were observed, albeit at lower frequencies in heterozygous mice, suggesting a haploinsufficient role for the 13q14 locus. Both MDR^−/− and miR-15a/miR-16-1^−/− mice succumbed to their disease earlier than their heterozygous or wild-type littermates. To ascertain whether the observed lymphoproliferative disorders were B-cell autonomous, we generated B-cell conditional deletions by crossing “floxed” MDR and miR-15a/miR-16-1 alleles with Cd19-Cre. These mice developed pathologies comparable in frequency and type to the constitutional knock-out mice, indicating that the B-cell proliferative disorders were not dependent on other cell types.

In conclusion, these results indicate that the 13q14 region contains a locus controlling the expansion of the B-cell compartment that includes both CD5⁺ and CD5⁻ B cells. Deletion of the locus leads to B cell-autonomous lymphoproliferative disorders, indicating a tumor suppressor function for which the miR-15a/miR-16-1 cluster but not the mature DLEU2 RNA, is required. The development of indolent lymphoproliferative disorders in mice with 13q14 deletions, such as MBL, CLL, and other NHL recapitulates the spectrum of CLL-associated phenotypes, suggesting that these mice are representative of the natural history of this disease. As such, these mice represent genotypically and phenotypically faithful models of the human disease uniquely suited for studies of mechanisms underlying disease progression and for testing novel anti-CLL therapeutic modalities.