Immunotherapy of B-Cell Lymphoma With CD3x19 Bispecific Antibodies: Costimulation via CD28 Prevents "Veto" Apoptosis of Antibody-Targeted Cytotoxic T Cells

Bispecific antibodies (CD3x19) against the CD3-chain of theT-cell-receptor/CD3 complex and the CD19 antigen on B cells can targetpolyclonal, nontumor-specific T cells to B lymphoma cells. This inducesT-cell activation, and generation of cytotoxic T cells (CTLs). Thesepolyclonal CTLs, targeted by the CD3x19 bispecific antibodies, can lyseCD19⁺ B-lymphoma cells. In a xenotransplant model insevere combined immunodeficiency deficient (SCID) mice, weand others observed that CD28 triggering is required for efficientelimination of B-lymphoma cells and cure from the tumor in addition toCD3x19 administration. We also showed that the activation and targeting of CTLs to the target cell by signal one alone, ie, the CD3x19 mab,induces T-cell death by apoptosis. In blocking experiments we showedthat this "veto" apoptosis is mediated by the CD95/Fas ligand.Addition of anti-CD28 (signal 2) renders the T cells resistant for vetoapoptosis both in vitro and in vivo. We therefore conclude that therole of costimulation in immunotherapy with bispecific antibodies orother T-cell-based immune strategies is not only to facilitate T-cellactivation but also to prevent T-cell deletion by apoptosis.     

Abnormal development of secondary lymphoid tissues in lymphotoxin β-deficient mice

The tumor necrosis factor (TNF) family cytokines lymphotoxin (LT) α and LTβ form heterotrimers that are expressed on the surface of activated lymphocytes and natural killer cells; LTα homotrimers can be secreted as well. Mice with a disrupted LTα gene lack lymph nodes (LN), Peyer’s patches (PP), and follicular dendritic cell (FDC) networks and reveal profound defects of the splenic architecture. However, it is unclear which of these abnormalities is the result of the absence in LTα homotrimers or LTαβ heterotrimers. To distinguish between these two possibilities, a mouse strain deficient in LTβ was created employing Cre/loxP-mediated gene targeting. Mice deficient in LTβ reveal severe defects in organogenesis of the lymphoid system similar to those of LTα^−/− mice, except that mesenteric and cervical LN are present in most LTβ-deficient mice. Both LTβ- and LTα-deficient mice show significant lymphocytosis in the circulation and peritoneal cavity and lymphocytic infiltrations in lungs and liver. After immunization, PNA-positive B cell clusters were detected in the splenic white pulp of LTβ-deficient mice, but FDC networks were severely underdeveloped. Collectively, these results indicate that LTα can signal independently from LTβ in the formation of PNA-positive foci in the spleen, and especially in the development of mesenteric and cervical LN.     

Definition of MHC and T cell receptor contacts in the HLA-DR4restricted immunodominant epitope in type II collagen and characterization of collagen-induced arthritis in HLA-DR4 and human CD4 transgenic mice

Rheumatoid arthritis (RA) is an autoimmune disease associated with the HLA-DR4 and DR1 alleles. The target autoantigen(s) in RA is unknown, but type II collagen (CII) is a candidate, and the DR4- and DR1-restricted immunodominant T cell epitope in this protein corresponds to amino acids 261–273 (CII 261–273). We have defined MHC and T cell receptor contacts in CII 261–273 and provide strong evidence that this peptide corresponds to the peptide binding specificity previously found for RA-associated DR molecules. Moreover, we demonstrate that HLA-DR4 and human CD4 transgenic mice homozygous for the I-A^b_β⁰ mutation are highly susceptible to collagen-induced arthritis and describe the clinical course and histopathological changes in the affected joints.     

Interleukin-2 induces β2-integrin-dependent signal transduction involving the focal adhesion kinase-related protein B (fakB)

β₂ integrin molecules are involved in a multitude of cellular events, including adhesion, migration, and cellular activation. Here, we studied the influence of β₂ integrins on interleukin-2 (IL-2)-mediated signal transduction in human CD4⁺ T cell lines obtained from healthy donors and a leukocyte adhesion deficiency (LAD) patient. We show that IL-2 induces tyrosine phosphorylation of a 125-kDa protein and homotypic adhesion in β₂ integrin (CD18)-positive but not in β₂-integrin-negative T cells. EDTA, an inhibitor of integrin adhesion, blocks IL-2-induced tyrosine phosphorylation of the 125-kDa protein but not other proteins in β₂-integrin-positive T cells. Likewise, a β₂ integrin (CD18) antibody selectively inhibits induction of the 125-kDa phosphotyrosine protein, whereas cytokine-mediated tyrosine phosphorylation of other proteins is largely unaffected. Immunoprecipitation experiments indicate that the IL-2-induced 125-kDa phosphotyrosine protein is the focal adhesion kinase-related protein B (fakB). Thus, IL-2 induces strong tyrosine phosphorylation of fakB in β₂-integrin-positive but not in β₂-integrin-negative T cells, and CD18 mAb selectively blocks IL-2-induced fakB-tyrosine phosphorylation in β₂-integrin-positive T cells. In parallel experiments, IL-2 does not induce or augment tyrosine phosphorylation of p125^FAK. In conclusion, our data indicate that IL-2 induces β₂-integrin-dependent signal transduction events involving the tyrosine kinase substrate fakB.     

Low antibody levels against cell wall-attached proteins of Streptococcus pyogenes predispose for severe invasive disease

Acute-phase serum samples from 70 patients with group A streptococcal (GAS) invasive disease were analyzed for IgG antibodies against 6 recently characterized GAS virulence factors (SclA, SclB, GRAB, MtsA, EndoS, and IdeS) and SpeB. Antibody levels against the cell wall-attached GAS antigens SclA, SclB, and GRAB were significantly lower in patients with severe invasive disease (streptococcal toxic shock syndrome [STSS] and/or necrotizing fasciitis [NF]; n=35), compared with levels in patients with nonsevere GAS bacteremia (n=35). Among patients with severe invasive disease, significantly lower antibody levels against GRAB were found in patients with STSS (n=10) than in patients with NF (n=17). Antibody levels against SpeB in patients with severe bacteremia were similar to those in patients with nonsevere bacteremia, and levels in patients with STSS were similar to those in patients with NF. The data indicate that immunity to cell wall-attached proteins may play a role in the protection against severe invasive disease and that antibodies against GRAB may be of importance in the pathogenesis of STSS.     

Fecal calprotectin concentration in patients with colorectal carcinoma

PURPOSE: The study contained herein was undertaken to investigate fecal calprotectin excretion in a series of patients with colorectal carcinoma and to determine whether the excretion was influenced by localization or stage of the tumor. Furthermore, the effect of surgical treatment on the concentrations was studied. Fecal calprotectin was also compared with plasma concentrations of calprotectin, carcinoembryonic antigen, and C-reactive protein. METHODS: Fecal calprotectin was measured in 119 consecutive patients admitted for treatment of colorectal carcinoma. In 116 (97.5 percent) patients, resectional surgery was performed. Plasma calprotectin was measured in 90 (76 percent) patients, carcinoembryonic antigen in 88 (74 percent) patients, and C-reactive protein in 82 (69 percent) patients. RESULTS: Median fecal calprotectin concentration in the 119 patients was 50 (range, 2-950) mg/l, which was significantly (P<0.0001) higher than in 125 control patients (median, 5.2 mg/l). In 23 patients studied also after resection, the excretion fell greatly. There were no significant differences in fecal calprotectin concentration among patients with different tumor stages. Elevated plasma calprotectin concentrations were found in 67 of 90 (73.3 percent) patients with colorectal carcinoma, compared with elevated fecal calprotectin in 111 of 119 (93.3 percent) patients, and there was no significant correlation between plasma and fecal calprotectin concentrations. Plasma calprotectin concentrations were significantly lower in patients with T1 or T2 tumors than in those with more advanced stages (P=0.0025). CONCLUSION: Measurement of fecal calprotectin may become a diagnostic tool in detecting colorectal carcinoma. The specificity in relation to colorectal carcinoma has not, however, been completely investigated. Both neoplastic and inflammatory conditions may be associated with elevated values; therefore, it is unlikely that calprotectin can predict specific colonic disorders.