Nerve growth factor may function as a survival factor for human neuroblastoma cells in culture.

Cells from two human neuroblastomas were found to exhibit a marked dependency on nerve growth factor (NGF) for survival in primary cultures. Cells from both tumors also responded to NGF by forming processes, but survival was not necessarily associated with process outgrowth. Lines of replicating cells could not be obtained from either tumor. NGF-dependent survival has not previously been reported as a characteristic of NGF-responsive human neuroblastoma cells in primary cultures or in cultures of established cell lines. Our findings suggest that tumors which require NGF for survival might constitute a biologically distinctive subset of neuroblastomas, or that NGF might function as a survival factor for some human neuroblastomas only in suboptimal or deleterious environments.

HIV infection is blocked in vitro by recombinant soluble CD4.

The T-cell surface glycoprotein, CD4 (T4), acts as the cellular receptor for human immunodeficiency virus, type 1 (HIV-1), the first member of the family of viruses that cause acquired immunodeficiency syndrome. HIV recognition of CD4 is probably mediated through the virus envelope glycoprotein (gp120) as shown by co-immunoprecipitation of CD4 and gp120 (ref.5) and by experiments using recombinant gp120 as a binding probe. Here we demonstrate that recombinant soluble CD4(rsT4) purified from the conditioned medium of a stably transfected Chinese hamster ovary cell line is a potent inhibitor of both virus replication and virus-induced cell fusion (syncytium formation). These results suggest that rsT4 is sufficient to bind HIV, and that it represents a potential anti-viral therapy for HIV infection.

A 113-amino acid fragment of CD4 produced in Escherichia coli blocks human immunodeficiency virus-induced cell fusion.

A gene encoding a 113-amino acid, NH2-terminal fragment of CD4, rsT4.113, was constructed and expressed in Escherichia coli under the control of the tryptophan operon promoter. Following induction, rsT4.113 is produced at 5-10% of total E. coli protein, and it is found in inclusion bodies. The protein is purified in two steps under denaturing and reducing conditions. Solubilized rsT4.113 is first purified on a column of Q-Sepharose to remove low molecular weight contaminants and then purified to greater than 95% homogeneity by gel filtration. Renaturation of rsT4.113 is achieved at approximately 20% yield by dilution and dialysis. High performance liquid chromatography analysis of renatured rsT4.113 reveals a less than 15% contaminant of reduced protein. Purified and renatured rsT4.113 contains epitopes for both OKT4a and Leu3a, anti-CD4 monoclonal antibodies which block CD4-gp 120 association, but lacks measurable affinity toward a nonblocking anti-CD4 monoclonal antibody, OKT4. By comparison to a longer form (375 amino acids) of recombinant soluble T4 produced in mammalian cells that contains the entire extracellular domain, rsT4.113 has a comparable affinity for binding to OKT4a and Leu3a in a radioimmunoassay. Analysis of antiviral activity of rsT4.113 demonstrates that the E. coli-derived protein inhibits human immunodeficiency virus-induced syncytium formation with an IC50 of 5-10 micrograms/ml. These data demonstrate that the human immunodeficiency virus-binding domain of CD4 is localized within the NH2-terminal 113 amino acids of CD4 and is contained within a structure homologous to the kappa variable-like domain of immunoglobulins.