Constitutive Baff Signalling Plays a Key Role in CLL Development by Promoting Tumor Cell Survival

Abstract

The B cell-activating factor of the tumor necrosis factor family (BAFF) is a potent B-cell survival factor. We recently found that nurselike cells, which presumably reside in the leukemia-microenvironment, express BAFF, which promotes survival of leukemia cells of patients with chronic lymphocytic leukemia (CLL) cells through activation of the classical NF-kB pathway. To study the influence of BAFF on leukemogenesis, we crossed BAFF transgenic (Tg) mice with either Eμ-TCL-Tg mice, which develop a lymphoproliferative disease resembling human CLL at about 12 months of age, or Ea-MYC-Tg mice, which experience higher rates of apoptosis in their mature B cells, but do not develop overt lymphoproliferative disease per se. We found that BAFF/TCL1-Tg mice had a shorter mean survival than either TCL1-Tg or BAFF-Tg mice due to the early development of a CD5+CD3−CD45Rlo leukemia B-cell population resembling human CLL at the age of about 3–4 months as compared to 7–9 months in TCL1-Tg mice. In contrast none of the BAFF-Tg or wt mice developed lymphoproliferative disease over the 26-month period of observation. The CD5+CD3−CD45Rlo cell population increased in BAFF/TCL1-Tg mice relatively rapidly, coming to represent >99% of the total blood mononuclear cells of 9-month-old double Tg animals. At this age, these mice also developed massive splenomegaly and their spleens were heavily infiltrated with leukemia B cells. Southern blot analyses of splenocytes harvested at various ages detected oligoclonal/monoclonal splenic B cell populations at approximately 4 months of age in the BAFF/TCL1-Tg mice compared to 9–12 months in the TCL1-Tg mice. Similarly, Ea-MYC-Tg mice, generated by inserting a single copy of MYC into the mouse Ig heavy-chain Ca locus, were crossed with the BAFF-Tg mice in order to obtain BAFF/MYC-Tg mice. In contrast to Ea-MYC-Tg mice, the BAFF/MYC-Tg mice developed expansions of CD5+CD3−CD45Rlo CLL-like leukemia B cells in the blood and spleen starting at 4 months of age, resulting in splenomegaly. These cells could be adoptively transferred into syngeneic BAFF-Tg mice, allowing for continued proliferation of such CD5+CD3−CD45Rlo leukemia B-cells and development lymphoproliferative disease. Staining of splenic sections for proliferating-cell nuclear antigen (PCNA) demonstrated similar high-rates of cell-proliferation in Ea-MYC-Tg mice and BAFF/MYC-Tg mice, which were twice greater than those of BAFF-Tg mice or wt mice. In contrast, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) of splenic sections revealed that Ea-MYC-Tg mice had 4 times greater proportions of apoptotic splenocytes than did BAFF/MYC-Tg mice, suggesting that BAFF promoted B cell expansion by inhibiting B cell turnover. Consistent with this, we found that BAFF induced in TCL1-Tg, as well as MYC-Tg mice, high-level B cell expression of anti-apoptotic proteins such as Bcl-XL, Bcl-2, A1/Bfl1, and Pim-2 in leukemia B cells. Also, these leukemia B cells transferred more efficiently into syngeneic BAFF-Tg mice than in mice lacking expression of BAFF. We treated BAFF-Tg mice with intraperitoneal (i.p.) injections of BAFF-neutralizing BR3-Fc or control protein prior to adoptive transfer of leukemia cells of Eμ-TCL1-Tg mice. We found that i.p. injection of 200 mg BR3-Fc into the recipient animals reduced the numbers of circulating CLL cells relative to that found in control-treated mice by nearly 20% (18.2%±5.3%; n=3) within 6 days. These findings indicate that BAFF accelerates development of leukemia in Eμ-TCL1-Tg and Ea-MYC-Tg mice primarily by enhancing leukemia B-cell survival, but not leukemia-cell proliferation, resulting in accelerated rates of tumor development and apparent enhancement of leukemia B cell proliferation in vivo. These findings suggest that targeting BAFF or BAFF-expressing cells within the leukemia microenvironment might be an effective strategy for treatment of CLL and related B-cell malignancies.