High TRAIL-R3 expression on leukemic blasts is associated with poor outcome and induces apoptosis-resistance which can be overcome by targeting TRAIL-R2.

Activation of the TNF-related apoptosis-inducing ligand (TRAIL) pathway can induce apoptosis in a broad range of human cancer cells. Four membrane-bound receptors have been identified. TRAIL-R1 and TRAIL-R2 contain a functional death domain; TRAIL-R3 and TRAIL-R4 lack a functional death domain and function as decoy receptors. Flow-cytometric analysis revealed that acute myeloid leukemic (AML) blasts expressed significantly more pro-apoptotic receptors compared to normal blasts. However, about 20% of AML patients highly expressed decoy receptor TRAIL-R3, which was strongly correlated to a shortened overall survival. TRAIL-R3 expression was also high on CD34+/CD38- cells, the compartment that harbors the leukemia initiating stem cell. Expression levels of pro-apoptotic TRAIL receptors were not correlated to the susceptibility for soluble TRAIL, which was generally low (mean level of cell death induction 14%). Cell death could be enhanced by down-modulation of TRAIL-R3, confirming its decoy function on AML blasts. Bypassing of TRAIL-R3 by treatment with antibodies directly targeting TRAIL-R2 resulted in higher rates of induced cell death (max. 80%). In conclusion, AML blasts do express pro-apoptotic TRAIL receptors. However, co-expression of decoy receptor TRAIL-R3 results in significant shortened overall survival. AML blasts could be targeted by anti-TRAIL-R2 antibodies, yielding a new therapeutic option for AML patients.

Involvement of caspase-8 in chemotherapy-induced apoptosis of patient derived leukemia cell lines independent of the death receptor pathway and downstream from mitochondria.

Resistance of leukemic cells to chemotherapy frequently occurs in patients with acute leukemia, which may be caused by alterations in common apoptotic pathways. Controversy exists whether cytostatic agents induce the mitochondrial or death receptor pathway of apoptosis. In the mitochondrial pathway cytochrome C release and caspase-9 activation play a central role in the induction of apoptosis, while formation of a Death Inducing Signaling Complex (DISC) and caspase-8 activation have been reported to be essential in death receptor-induced apoptosis. Here, we show in human derived myeloid and lymphoblastic leukemia cell lines that caspase-8 plays a more important role than previously expected in apoptosis mediated via the mitochondrial pathway. We demonstrated in these malignant cells chemotherapy-induced apoptosis independent of the death receptor pathway, since blocking this pathway using a retroviral construct encoding Flice inhibitory protein (FLIP) did not inhibit drug-induced apoptosis or caspase-8 activation, while overexpression of Bcl-2 completely inhibited both events. Furthermore, we showed that activation of caspase-8 by cytostatic agents occurred downstream from mitochondria. Since caspase-8 plays a central role in both death receptor- and chemotherapy-induced apoptosis of malignant cells from patients with acute leukemia, therapeutic strategies focusing at modulation and activation of caspase-8 may be successful in the treatment of drug-resistant malignancies.

Outcome of intensive immunosuppression and autologous stem cell transplantation in patients with severe rheumatoid arthritis is associated with the composition of synovial T cell infiltration.

OBJECTIVE: To determine clinical and immunological correlates of high dose chemotherapy (HDC) + autologous stem cell transplantation (ASCT) in patients with severe rheumatoid arthritis (RA), refractory to conventional treatment. METHODS: Serial samples of peripheral blood and synovial tissue were obtained from seven patients with RA treated with HDC and autologous peripheral blood grafts enriched for CD34+ cells. Disease activity was assessed with the Disease Activity Score (DAS), serum concentrations of C reactive protein (CRP), and human immunoglobulin (HIg) scans, and the extent of immunoablation was determined by immunophenotyping of peripheral blood mononuclear cells, and immunohistochemistry and double immunofluorescence of synovium. RESULTS: Clinical responders (n = 5) had a larger number of cells at baseline expressing CD3, CD4, CD27, CD45RA, CD45RB, and CD45RO in synovium (p < 0.05), higher activity on HIg scans (p = 0.08), and a trend towards higher concentrations of CRP in serum than non-responders (n = 2). Subsequent remissions and relapses in responders paralleled reduction and re-expression, respectively, of T cell markers. A relatively increased expression of CD45RB and CD45RO on synovial CD3+ T cells was seen after HDC + ASCT. No correlations were found between DAS and changes in B cells or macrophage infiltration or synoviocytes. CONCLUSIONS: HDC + ASCT results in profound but incomplete immunoablation of both the memory and naïve T cell compartment, which is associated with longlasting clinical responses in most patients. The findings provide strong circumstantial evidence for a role of T cells in established RA, and demonstrate a role for the synovium in post-transplantation T cell reconstitution.