ALBERT

Description: To compare the clinical and serological outcomes of patients receiving donors' marrow positive or negative for hepatitis B surface antigen (HBsAg), we studied 18 patients of allogeneic hematopoietic cell transplantation receiving HBsAg-positive marrow (group 1) and 18 receiving HBsAg-negative marrow (group 2). The recipients of the 2 groups were matched for hepatitis B virus (HBV) serology, sex, age, underlying hematological diseases, conditioning regimen, and prophylaxis against graft-versus-host diseases. Eight (44.4%) recipients in group 1 and 2 (11.1%) in group 2 suffered from HBV-related hepatitis posttransplant (P = .03). Furthermore, HBV-related hepatic failure was seen in 6 group 1 patients, but in none of the group 2 patients (P = .007). Five of the 9 (55.5%) HBsAg-negative recipients in group 1 became positive after receiving HBsAg-positive marrow. Serum HBV DNA was positive in all 5 donors of these patients, but in none of the donors of recipients who remained HBsAg negative (P = .008). Group 1 patients developing HBV-related hepatitis posttransplant were more likely to have a donor carrying a precore A1896 and/or core promoter T1762/A1764 HBV variant (62.5% versus 0%, P = .007). This study has demonstrated that a high incidence of HBV-related hepatitis was associated with the use of HBsAg-positive marrow for transplant, and a high viral load in the donor appeared to predispose recipients to the development of HBV-related hepatitis posttransplant. Further clinical trials will be necessary to determine the optimal management approach to this problem, including the use of the antiviral agents in the donors and the recipients.

Description: To compare the clinical and serological outcomes of patients receiving donors' marrow positive or negative for hepatitis B surface antigen (HBsAg), we studied 18 patients of allogeneic hematopoietic cell transplantation receiving HBsAg-positive marrow (group 1) and 18 receiving HBsAg-negative marrow (group 2). The recipients of the 2 groups were matched for hepatitis B virus (HBV) serology, sex, age, underlying hematological diseases, conditioning regimen, and prophylaxis against graft-versus-host diseases. Eight (44.4%) recipients in group 1 and 2 (11.1%) in group 2 suffered from HBV-related hepatitis posttransplant (P = .03). Furthermore, HBV-related hepatic failure was seen in 6 group 1 patients, but in none of the group 2 patients (P = .007). Five of the 9 (55.5%) HBsAg-negative recipients in group 1 became positive after receiving HBsAg-positive marrow. Serum HBV DNA was positive in all 5 donors of these patients, but in none of the donors of recipients who remained HBsAg negative (P = .008). Group 1 patients developing HBV-related hepatitis posttransplant were more likely to have a donor carrying a precore A1896 and/or core promoter T1762/A1764 HBV variant (62.5% versus 0%, P = .007). This study has demonstrated that a high incidence of HBV-related hepatitis was associated with the use of HBsAg-positive marrow for transplant, and a high viral load in the donor appeared to predispose recipients to the development of HBV-related hepatitis posttransplant. Further clinical trials will be necessary to determine the optimal management approach to this problem, including the use of the antiviral agents in the donors and the recipients.

Description: The dose of 400mg per day of imatinib is currently considered standard therapy for patients with newly diagnosed chronic myeloid leukemia (CML) in chronic phase (CP). The IRIS trial demonstrated superior response rates in imatinib treated patients compared to interferon alfa, and evaluated molecular response up to 24 months of imatinib therapy in patients who achieved a complete cytogenetic response (CCR). Those patients with a major molecular response (MMR, ≥3 log reduction of BCR-ABL) by 12 months had 100% progression free survival at 24 months. However, very few patients had undetectable BCR-ABL levels. In the current study, we monitored the molecular response for a median of 42 months (25th to 75th percentile 39–45 months) in all patients enrolled in the IRIS trial in Australia and New Zealand who commenced imatinib as their first-line therapy (n=28 patients). We aimed to determine if the BCR-ABL levels continued to decrease and whether additional patients achieve a MMR with a longer follow up. BCR-ABL transcript levels were monitored by real-time quantitative PCR at 3 to 6 month intervals. A CCR (approximately equivalent to a greater than 2-log reduction of BCR-ABL) was achieved in 24 of the 28 patients (85%). The table demonstrates that while the frequency of achieving a MMR increased between 12 and 42 months, most of the improvement occurred between 12 and 24 months. Conversely, from 24 to 42 months the number of patients achieving a ≥4-log reduction increased significantly (P

Description: HLA incompatibility between the donor and recipient is the most critical factor governing the incidence of rejection and GVHD after conventional allogeneic stem cell transplantation. But the impact of HLA disparity on GVHD and graft rejection after RICT remains to be elucidated. We retrospectively analyzed the outcomes of 437 patients who underwent bone marrow (n=95) or peripheral blood stem cell RICT (n=342). The numbers of patients who received a graft from a HLA-matched (275 from siblings, 11 from family members, and 54 from unrelated donors), one-locus-mismatched, 2- or 3-loci-mismatched donor were 340, 65, and 32, respectively. The HLA-matched group included significantly higher population of patients who received cyclosporine alone for GVHD prophylaxis. The overall cumulative incidence of grade II-IV acute GVHD was 40% for all subjects. It was 38% (95% CI; 33%–43%) in recipients of HLA-matched donors, 43% (95% CI; 31%–54%) in those of one-locus-mismatched donors, and 54% (95% CI; 37%–68%) in those of 2–3-loci-mismatched donors. A Cox regression model adjusted for potential confounders including GVHD prophylaxis demonstrated that 2-3 loci-mismatch was identified as an independent risk factor of grade II-IV acute GVHD (Table). Use of antithymocyte globulin was identified as an independent better protective factor for GVHD (HR;0.66, p=.003). Cumulative incidence of rejection was significantly higher after one-locus mismatch RICT (Table) and the risk tended to increase in relation to an increase of HLA disparity. Malignant disease was identified as an independent prognostic factor for rejection. In patients with hematologic malignancies, overall survival (OS) of recipients of 2–3-loci-mismatched RICT at 1 year (38%, 95%CI; 21%–54%) was significantly worse than that after HLA-matched RICT (65%, 95%CI; 59%–70%). By contrast, there was no statistical difference in the incidence of grade II-IV acute GVHD and OS between HLA-matched RICT and one-locus-mismatched RICT. Multivariate analysis demonstrated 2–3-loci-mismatch (Table) and high-risk disease (HR; 2.3, p=.001) as independent risk factors for OS. Thus, HLA incompatibility between the donor and recipient is an important risk factor for rejection, acute GVHD and overall survival after RICT. Therefore RICT from a one-locus-mismatched donor may represent an effective alternative approach in patients lacking HLA-matched sibling donors. multivariate analysis n acute GVHD Rejection OS HR (95%CI) p HR (95%CI) p HR (95%CI) p match 340 1.0 1.0 1.0 1-mismatch 65 1.4 (0.9–2.2) 0.20 4.5 (1.1–17.9) 0.03 1.0 (0.6–1.6) 0.88 2–3-mismatch 32 2.2 (1.2–4.1) 0.02 7.0 (0.8–64.8) 0.08 3.3 (1.8–6.2)

Description: Monocytes from patients with sickle cell disease (SCD) are in an activated state. However, the mechanism of activation of monocytes in SCD is not known. Our studies showed that placenta growth factor (PlGF) activated monocytes and increased mRNA levels of cytokines (tumor necrosis factor-α [TNF-α] and interleukin-1β [IL-1β]) and chemokines (monocyte chemotactic protein-1 [MCP-1], IL-8, and macrophage inflammatory protein-1β [MIP-1β]) in both normal monocytes and in the THP-1 monocytic cell line. This increase in mRNA expression of cytochemokines was also reflected in monocytes derived from subjects with SCD. We studied the PlGF-mediated downstream cellular signaling events that caused increased transcription of inflammatory cytochemokines and chemotaxis of THP-1 monocytes. PlGF-mediated cytochemokine mRNA and protein expression was inhibited by PD98059 and wortmannin, inhibitors of mitogen-activated protein kinase kinase (MAPK/MEK) kinase and phosphatidylinositol-3 (PI3) kinase, respectively, but not by SB203580, a p38 kinase inhibitor. PlGF caused a time-dependent transient increase in phosphorylation of extracellular signal–regulated kinase-1/2 (ERK-1/2), which was completely inhibited by wortmannin, indicating that activation of PI3 kinase preceded MEK activation. PlGF also induced transient phosphorylation of AKT. MEK and PI3 kinase inhibitors and antibody to Flt-1 abrogated PlGF-induced chemotaxis of THP-1 monocytes. Overexpression of a dominant-negative AKT or a dominant-negative PI3 kinase p85 subunit in THP-1 monocytes attenuated the PlGF-mediated phosphorylation of ERK-1/2, cytochemokine secretion, and chemotaxis. Taken together, these data show that activation of monocytes by PlGF occurs via activation of Flt-1, which results in activation of PI3 kinase/AKT and ERK-1/2 pathways. Therefore, we propose that increased levels of PlGF in circulation play an important role in the inflammation observed in SCD via its effects on monocytes.

Description: Sickle cell disease (SCD) results in chronic hypoxia and secondarily increased erythropoietin concentrations. Leukocytosis and activated monocytes are also observed in SCD in absence of infection or vaso-occlusion (steady state), the reasons for which are unknown. We found that erythroid cells produced placenta growth factor (PlGF), an angiogenic growth factor belonging to the vascular endothelial growth factor (VEGF) family, and its expression was induced in bone marrow CD34+ progenitor cells in the presence of erythropoietin. Furthermore, the steady state circulating PlGF levels in subjects with severe SCD (at least 3 vaso-occlusive crises [VOCs] per year) were 18.5 ± 1.2 pg/mL (n = 9) compared with 15.5 ± 1.2 pg/mL (n = 13) in those with mild SCD (fewer than 3 VOCs per year) and 11.3 ± 0.7 pg/mL (n = 9) in healthy controls (P 〈 .05), suggesting a correlation between PlGF levels and SCD severity. In addition, PlGF significantly increased mRNA levels of the proinflammatory cytochemokines interleukin-1β, interleukin-8, monocyte chemoattractant protein-1, and VEGF in peripheral blood mononuclear cells (MNCs) of healthy subjects (n = 4; P 〈 .05). Expression of these same cytochemokines was significantly increased in MNCs from subjects with SCD at steady state (n = 14), compared with healthy controls. Of the leukocyte subfractions, PlGF stimulated monocyte chemotaxis (P 〈 .05, n = 3). Taken together, these data show for the first time that erythroid cells intrinsically release a factor that can directly activate monocytes to increase inflammation. The baseline inflammation seen in SCD has always been attributed to sequelae secondary to the sickling phenomenon. We show that PlGF contributes to the inflammation observed in SCD and increases the incidence of vaso-occlusive events.

Description: The immune dysfunction and cell destruction that occur in the human immunodeficiency virus (HIV)-infected host appear to result from the direct cytopathic effects of viral infection and the effects of viral proteins on uninfected bystander cells. Recently, the α-chemokine receptor CXCR4 has been reported to mediate apoptosis in neuronal cells and in CD4+ and CD8+ T cells after its binding to HIV-1 envelope proteins. In the current study, it was observed that human umbilical vein endothelial cells (HUVEC) undergo apoptosis after their treatment with the HIV-1 envelope proteins gp120/160. Anti-CXCR4 monoclonal antibody decreased HIV-1 gp120/160-induced apoptosis, suggesting that the CXCR4 chemokine receptor mediates the apoptotic effects of these HIV envelope glycoproteins. Further studies revealed that caspases play an important role in this process because the pretreatment of cells with a general caspase enzyme inhibitor decreased the extent of HUVEC apoptosis induced by gp120/160. In addition, it was found that caspase-3 was activated on HIV-1 gp120/160 treatment of these cells. It was also observed that gp120/160 treatment slightly increased the expression of the pro-apoptotic molecule Bax. These results suggest that HIV-1 envelope glycoproteins can disrupt endothelial integrity through the interaction with CXCR4, thereby facilitating virus transit out of the bloodstream and contributing to the vascular injury syndromes seen in acquired immunodeficiency syndrome.

Description: The IRIS trial demonstrated that imatinib at 400 mg/day was superior to alpha interferon as first line therapy for chronic phase CML. To assess the tolerability and efficacy of higher doses of imatinib in this setting we are conducting a Phase II trial (TIDEL) using imatinib 600 mg initially, increasing to 800 mg if specified response criteria are not met: complete hematologic response (CHR) at 3 Mo; major cytogenetic response (MCR) at 6 Mo; complete cytogenetic response (CCR) at 9 Mo, and 〉4 log reduction in BCR-ABL at 12 Mo. Filgrastim was used in cases of neutropenia to maintain dose intensity. Of 103 patients enrolled, 8 came off study in the first 12 Mo (2 unrelated deaths, 3 blast crisis, 3 poor response). 80 patients are currently assessable at 12 Mo (median age 47 years, range 21–75). Protocol mandated dose increases to 800 mg for failure to achieve MCR or CCR targets were activated in 7 patients before 12 months. We made a historical comparison of the best response by 12 Mo to responses in the IRIS trial (95% confidence intervals). Response rates at 12 Mo MCR (0–35% Ph+) CCR (0% Ph+) MMR (≥3 log reduction in BCR-ABL) 400mg - imatinib arm of IRIS n=556 84.1% (81.0 – 87.2%) 69.3% (65.3 – 73.2%) 40% (NA) 600mg - imatinib in TIDEL trial n=80 94.2% (87.3 – 97.5%) 88.5% (80.3 – 93.5%) 47.4% (37.5 – 57.6%) P-value for z-test 0.0004

Description: Preclinical models have shown that transplantation of marrow mesenchymal cells has the potential to correct inherited disorders of bone, cartilage, and muscle. The report describes clinical responses of the first children to undergo allogeneic bone marrow transplantation (BMT) for severe osteogenesis imperfecta (OI), a genetic disorder characterized by defective type I collagen, osteopenia, bone fragility, severe bony deformities, and growth retardation. Five children with severe OI were enrolled in a study of BMT from human leukocyte antigen (HLA)–compatible sibling donors. Linear growth, bone mineralization, and fracture rate were taken as measures of treatment response. The 3 children with documented donor osteoblast engraftment had a median 7.5-cm increase in body length (range, 6.5-8.0 cm) 6 months after transplantation compared with 1.25 cm (range, 1.0-1.5 cm) for age-matched control patients. These patients gained 21.0 to 65.3 g total body bone mineral content by 3 months after treatment or 45% to 77% of their baseline values. With extended follow-up, the patients' growth rates either slowed or reached a plateau phase. Bone mineral content continued to increase at a rate similar to that for weight-matched healthy children, even as growth rates declined. These results suggest that BMT from HLA-compatible donors may benefit children with severe OI. Further studies are needed to determine the full potential of this strategy.

Description: We recently reported that chronic lymphocytic leukemia (CLL) cells synthesize and release vascular endothelial growth factor (VEGF) under normoxic and hypoxic conditions. CLL B cells also express VEGF membrane receptors (VEGF-R1 and VEGF-R2), suggesting that they use VEGF as a survival factor. To assess the mechanism of apoptosis resistance related to VEGF, we determined the impact of VEGF on CLL B cells, and we studied the impact of epigallocatechin-3-gallate (EGCG), a known receptor tyrosine kinase (RTK) inhibitor, on VEGF receptor status and viability of CLL B cells. VEGF165 significantly increased apoptotic resistance of CLL B cells, and immunoblotting revealed that VEGF-R1 and VEGF-R2 are spontaneously phosphorylated on CLL B cells. EGCG significantly increased apoptosis/cell death in 8 of 10 CLL samples measured by annexin V/propidium iodide (PI) staining. The increase in annexin V/PI staining was accompanied by caspase-3 activation and poly–adenosine diphosphate ribose polymerase (PARP) cleavage at low concentrations of EGCG (3 μg/mL). Moreover, EGCG suppressed the proteins B-cell leukemia/lymphoma-2 protein (Bcl-2), X-linked inhibitor of apoptosis protein (XIAP), and myeloid cell leukemia-1 (Mcl-1) in CLL B cells. Finally, EGCG (3-25 μg/mL) suppressed VEGF-R1 and VEGF-R2 phosphorylation, albeit incompletely. Thus, these results suggest that VEGF signaling regulates survival signals in CLL cells and that interruption of this autocrine pathway results in caspase activation and subsequent leukemic cell death.