Mechanism for IL-15-Driven B Cell Chronic Lymphocytic Leukemia Cycling: Roles for AKT and STAT5 in Modulating Cyclin D2 and DNA Damage Response Proteins.

Clonal expansion of B cell chronic lymphocytic leukemia (B-CLL) occurs within lymphoid tissue pseudofollicles. IL-15, a stromal cell-associated cytokine found within spleens and lymph nodes of B-CLL patients, significantly boosts in vitro cycling of blood-derived B-CLL cells following CpG DNA priming. Both IL-15 and CpG DNA are elevated in microbe-draining lymphatic tissues, and unraveling the basis for IL-15-driven B-CLL growth could illuminate new therapeutic targets. Using CpG DNA-primed human B-CLL clones and approaches involving both immunofluorescent staining and pharmacologic inhibitors, we show that both PI3K/AKT and JAK/STAT5 pathways are activated and functionally important for IL-15→CD122/ɣc signaling in ODN-primed cells expressing activated pSTAT3. Furthermore, STAT5 activity must be sustained for continued cycling of CFSE-labeled B-CLL cells. Quantitative RT-PCR experiments with inhibitors of PI3K and STAT5 show that both contribute to IL-15-driven upregulation of mRNA for cyclin D2 and suppression of mRNA for DNA damage response mediators ATM, 53BP1, and MDC1. Furthermore, protein levels of these DNA damage response molecules are reduced by IL-15, as indicated by Western blotting and immunofluorescent staining. Bioinformatics analysis of ENCODE chromatin immunoprecipitation sequencing data from cell lines provides insight into possible mechanisms for STAT5-mediated repression. Finally, pharmacologic inhibitors of JAKs and STAT5 significantly curtailed B-CLL cycling when added either early or late in a growth response. We discuss how the IL-15-induced changes in gene expression lead to rapid cycling and possibly enhanced mutagenesis. STAT5 inhibitors might be an effective modality for blocking B-CLL growth in patients.

Mechanistic Insights into CpG DNA and IL-15 Synergy in Promoting B Cell Chronic Lymphocytic Leukemia Clonal Expansion.

Malignant cell growth within patients with B cell chronic lymphocytic leukemia (B-CLL) is largely restricted to lymphoid tissues, particularly lymph nodes. The recent in vitro finding that TLR-9 ligand (oligodeoxynucleotide [ODN]) and IL-15 exhibit strong synergy in promoting B-CLL growth may be particularly relevant to growth in these sites. This study shows IL-15-producing cells are prevalent within B-CLL-infiltrated lymph nodes and, using purified B-CLL cells from blood, investigates the mechanism for ODN and IL-15 synergy in driving B-CLL growth. ODN boosts baseline levels of phospho-RelA(S529) in B-CLL and promotes NF-κB-driven increases in IL15RA and IL2RB mRNA, followed by elevated IL-15Rα and IL-2/IL-15Rß (CD122) protein. IL-15→CD122 signaling during a critical interval, 20 to 36-48 h following initial ODN exposure, is required for optimal induction of the cycling process. Furthermore, experiments with neutralizing anti-IL-15 and anti-CD122 mAbs indicate that clonal expansion requires continued IL-15/CD122 signaling during cycling. The latter is consistent with evidence of heightened IL2RB mRNA in the fraction of recently proliferated B-CLL cells within patient peripheral blood. Compromised ODN+IL-15 growth with limited cell density is consistent with a role for upregulated IL-15Rα in facilitating homotypic trans IL-15 signaling, although there may be other explanations. Together, the findings show that ODN and IL-15 elicit temporally distinct signals that function in a coordinated manner to drive B-CLL clonal expansion.

Autologus dendritic cell vaccine for chronic hepatitis B carriers: a pilot, open label, clinical trial in human volunteers.

Antigen-presenting autologous dendritic cells (ADCs), primed with antigen, have been used in immunotherapy. We evaluated ADCs for treatment of chronic hepatitis B (CHB). ADCs were administered to 380 CHB patients. Virological, biochemical, and serological responses were evaluated in each patient over the course of 48 weeks. Undetectable levels of HBV DNA were reported in 46.36% of patients negative for the hepatitis B "e" antigen (HBeAg) and 3.13% HBeAg-positive patients. Normalization of alanine aminotransferase levels occurred in both HBeAg-positive (P=0.007) and HBeAg-negative (P=0.003) patients. It appears that ADC vaccination effectively reconstructed the immunity and elicited virological, serological, and biochemical improvements in some patients with chronic HBV. No side effects were observed.

Lifespan is prolonged in autoimmune-prone (NZB/NZW) F1 mice fed a diet supplemented with indole-3-carbinol.

Dietary modulation has the potential to prevent or ameliorate systemic lupus erythematosus (SLE). Indole-3-carbinol (I3C), which is abundant in cruciferous vegetables, was evaluated in a murine model of SLE because of its antiestrogenic activities. Female (NZB x NZW) F1 mice, which develop SLE, were fed an AIN76A diet without or with 0.2 g/kg I3C, starting soon after weaning or at 5 mo of age. At 12 mo of age, 80% of mice fed the I3C-supplemented diet soon after weaning were alive compared with only 10% of controls. When experimental diets were initiated at 5 mo of age, 100% of I3C fed mice and 30% of controls were alive at 12 mo of age. Anti-double-stranded DNA (dsDNA) antibodies developed in all groups, although at several time points, the levels produced in I3C-fed mice were significantly lower. Renal disease (proteinuria, histologic changes, IgG immune complex deposition, subepithelial deposits and diffuse epithelial cell foot process effacement) was more severe in controls with both protocols. The estrogen urinary metabolite ratio of 2- to 16alpha-hydroxyestrone was increased in I3C-fed mice. These findings demonstrate a profound effect of dietary I3C in experimental SLE.