Effects of a soluble CD4 and CD4-Pseudomonas exotoxin A chimeric protein on human peripheral blood lymphocytes: lymphocyte activation and anti-HIV activity in vitro.
Recombinant sCD4-based proteins were evaluated for their effects on antigen-stimulated proliferation of human peripheral blood mononuclear cells (PBMC) and for antiviral activity against PBMC infected with human immunodeficiency virus (HIVD34). Two sCD4-based proteins were solubilized, refolded, and purified to homogeneity from recombinant E. coli and consisted of the 178 amino-terminal residues of CD4 fused with the translocating and catalytic domains of Pseudomonas exotoxin A (sCD4-PE40) or 183 amino-terminal residues of CD4 (sCD4-183); a third sCD4 consisting of 369 amino acids of CD4 was purified from recombinant mammalian cells for comparative purposes (sCD4-369). Increasing molar concentrations of these sCD4s were evaluated for inhibition of PBMC proliferation induced by alloantigen (MLR), by tetanus toxoid (TTOX), or in response to crosslinking with antibody to CD3 (OKT3). In addition, the concentrations of each protein required to inhibit replication of the HIVD34 isolate in primary PBMC was determined by quantitation of HIV p24 antigen released into supernatant fluids by infected cells. By comparing antiviral activity with anti-proliferative activity a relative estimate of the selectivity index for each recombinant sCD4 was determined. Proliferation of PBMC in response to alloantigen or OKT3 was less sensitive to inhibition than proliferation induced by TTOX, and the selectivity indices estimated for sCD4-PE40 were 170, 170 and 17, respectively. The selectivity index for sCD4-183 was greater than 350 under all assay conditions. Comparative evaluation of alloantigen-stimulated proliferation with antiviral activity of sCD4-183 versus sCD4-369 suggested that the E. coli-derived sCD4-183 may have a higher selectivity index under these conditions than its mammalian cell-derived counterpart.
Rubino
K L
KL
Cancer and Infectious Diseases Research, Upjohn Laboratories, Kalamazoo, Michigan 49007.
Tarpley
W G
WG
Nicholas
J A
JA
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D015704:CD4 Antigens; D005098:Exotoxins / Q000494:pharmacology*; D006678:HIV / Q000502:physiology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000187:drug effects* / Q000382:microbiology; D008213:Lymphocyte Activation / Q000187:drug effects*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology*; D037521:Virulence Factors; D014779:Virus Replication / Q000187:drug effects*; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D015704:CD4 Antigens; D005098:Exotoxins; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A