National Institute on Aging's 50th anniversary: Advancing aging research and the health and well-being of older adults.

The National Institute on Aging (NIA), part of the National Institutes of Health (NIH), was founded in 1974 to support and conduct research on aging and the health and well-being of older adults. Fifty years ago, the concept of studying aging generated much skepticism. Early NIA-funded research findings helped establish the great value of aging research and provided the foundation for significant science advances that have improved our understanding of the aging process, diseases and conditions associated with aging, and the effects of health inequities, as well as the need to promote healthy aging lifestyles. Today, we celebrate the many important contributions to aging research made possible by NIA, as well as opportunities to continue to make meaningful progress. NIA emphasizes that the broad aging research community must continue to increase and expand our collective efforts to recruit and train a diverse next generation of aging researchers.

Immunoglobulin superfamily receptor translocation associated 2 protein on lymphoma cell lines and hairy cell leukemia cells detected by novel monoclonal antibodies.

PURPOSE: The immunoglobulin superfamily receptor translocation associated 2 (IRTA2) gene encodes a cell surface receptor homologous to the family of Fc receptors. Because of the restricted expression of mRNA in B cell-lineage cells, IRTA2 is a new potential target for the immunotherapy of B cell malignancies. To study the expression of the IRTA2 gene product, we produced monoclonal antibodies (MAbs) specific to IRTA2. EXPERIMENTAL DESIGN: A mouse used for cell fusion was DNA-immunized with an expression plasmid encoding the IRTA2 cDNA. The reactivity of MAbs secreted from the hybridomas were characterized with recombinant proteins of IRTA family members in an enzyme immunoassay and a fluorescence-activated cell sorter (FACS). Nineteen human lymphoma cell lines and blood cells from five patients with hairy cell leukemia (HCL) were analyzed with IRTA2 expression using FACS. RESULTS: Three MAbs (F25, F56, and F119) were selected based on their specific reactivity with recombinant IRTA2 and lack of cross-reactivity with other IRTA family members. In a FACS analysis, MAbs F56 and F119 detected IRTA2 expression in six of seven B cell non-Hodgkin's lymphoma and one of six Burkitt's lymphoma cell lines. Reverse transcriptase-PCR experiments and Western blotting using MAb F25 confirmed the expression profile. We also found that HCL cells from five patients expressed IRTA2. CONCLUSIONS: Our results provide the first evidence that IRTA2 is expressed on the surface of human lymphoma cell lines and HCL cells. IRTA2 could be useful as a new target for immunotherapy.

Increased affinity and stability of an anti-HIV-1 envelope immunotoxin by structure-based mutagenesis.

HIV-infected cells are selectively killed by an immunotoxin in which a truncated form of Pseudomonas exotoxin A is joined to the variable region of a broadly neutralizing antibody (3B3) that recognizes the viral envelope glycoprotein (Env). To improve the efficacy of this molecule, we used three-dimensional structural information and phage selection data to design 23 single and multiple point mutations in the antibody variable region sequences that contact Env. Substituting an aromatic residue for an aspartate in the third complementarity-determining region of V(H) increased the potency of the immunotoxin by approximately 10-fold in a cell-killing assay. Detailed analysis of one such mutant, N31H/Q100eY, revealed both a higher affinity for monomeric and cell surface Env and an increased stability against aggregation compared with the starting immunotoxin. Conversion to a disulfide-linked two-chain format further stabilized the protein. N31H/Q100eY retained the ability to bind to Env from multiple viral isolates, to inhibit Env-mediated cell fusion, and to limit spreading viral infection in peripheral blood mononuclear cells. Such site-directed mutants may increase the utility of immunotoxins for reducing or eradicating persistent HIV-1 infection in humans.

Novel anti-CD30 recombinant immunotoxins containing disulfide-stabilized Fv fragments.

PURPOSE: To develop a novel targeting reagent to CD30 expressed on Hodgkin'sdisease and anaplastic large cell lymphoma, we made a panel of recombinant immunotoxins specific for CD30 using Fvs of newly produced anti-CD30 monoclonal antibodies (MAbs) and a M(r) 38,000 truncated mutant of Pseudomonas exotoxin. EXPERIMENTAL DESIGN: A group of MAbs against CD30 was produced and characterized for their reactivity and epitopes. Recombinant immunotoxins were made using the Fv genes cloned from the hybridomas. Their cytotoxic activities were examined on various CD30-positive cell lines. RESULTS: Six MAbs were produced. All reacted with recombinant soluble CD30 and to a CD30-Fc fusion protein, and bound to native CD30 expressed on Hodgkin's lymphoma-derived cell lines. The epitopes of the six MAbs were classified into two groups by a mutual competition assay for the binding to CD30 on cells. Sequencing the cDNAs revealed that all of the variable chains are unique except one valiable light that is shared by two MAbs. We made four disulfide stabilized Fv-based recombinant immunotoxins, in which the valiable heavy, which is genetically fused with truncated mutant of Pseudomonas exotoxin, forms a disulfide bond with the valiable light. The purified immunotoxins bound to recombinant soluble CD30 immobilized on a biosensor chip with K(d)s of 4-400 nM. Fluorescence-activated cell sorter analysis confirmed their specific binding. In vitro cytotoxicity tests showed that the immunotoxins specifically kill a variety of CD30-positive lymphoma cell lines as well as CD30-transfected A431 cells. The IC(50) ranged from 0.3 to 100 ng/ml. CONCLUSIONS: Four anti-CD30 disulfide stabilized Fv immunotoxins were successfully produced. Two of these showed good cytotoxic activity to various CD30-positive cell lines. These newly produced immunotoxins should be additionally evaluated for the treatment of CD30-positive lymphomas.