Comparative pharmacology of a new recombinant FSH expressed by a human cell line.

Recombinant FSH proteins are important therapeutic agents for the treatment of infertility, including follitropin alfa expressed in Chinese Hamster Ovary (CHO) cells and, more recently, follitropin delta expressed in the human cell line PER.C6. These recombinant FSH proteins have distinct glycosylation, and have distinct pharmacokinetic and pharmacodynamic profiles in women.

[Pharmacoeconomic model of the use of rifabutin in new cases of pulmonary tuberculosis].

The formalized procedures to commensurate the results and expenditures were used to make a comprehensive evaluation of the clinical efficiency and economical expediency of purposeful application of rifabutin in new cases of disseminated and destructive pulmonary tuberculosis.

Antibodies directed against the MHC-I molecule H-2Dd complexed with an antigenic peptide: similarities to a T cell receptor with the same specificity.

alphabeta TCRs, which use an Ab-like structure to form a combining site, recognize molecular complexes consisting of peptides bound to MHC class I (MHC-I) or class II (MHC-II) molecules. To explore the similarities and differences between Ab and T cell recognition of similar structures, we have isolated two mAbs, KP14 and KP15, that specifically bind H-2D(d) complexed with an HIV envelope gp160-derived peptide, P18-I10. These Abs are MHC and peptide specific.

NK and CTL recognition of a single chain H-2Dd molecule: distinct sites of H-2Dd interact with NK and TCR.

We generated transgenic mice expressing a single-chain beta2-microglobulin (beta2m)-H-2Dd. The cell-surface beta2m-H-2Dd molecule was expressed on a beta2m-deficient background and reacted with appropriate mAbs. It was of the expected m.

An antibody single-domain phage display library of a native heavy chain variable region: isolation of functional single-domain VH molecules with a unique interface.

To develop very small antibody-derived recognition units for experimental, medical, and drug design purposes, a heavy chain variable region (VH) single-domain phage-display library was designed and constructed. The scaffold that was used for library construction was a native sequence of a monoclonal antibody with a unique VH/VL interface. There was no need to modify any residues in the VL interface to avoid non-specific binding of VH domain.

The X-ray crystal structure of a Valpha2.6Jalpha38 mouse T cell receptor domain at 2.5 A resolution: alternate modes of dimerization and crystal packing.

We describe here the structure of a murine T cell receptor (TCR) Valpha2.6Jalpha38 (TCRAV2S6J38) domain, derived from a T cell hybridoma with specificity for the H-2Ddmajor histocompatibility complex class I molecule bound to a decamer peptide, P18-I10, from the HIV envelope glycoprotein gp120, determined by X-ray crystallography at 2.5 A resolution.

Rigidification of the alpha2 helix of an MHC class I molecule by a valine to proline mutation in position 165 does not prevent peptide-specific antigen presentation.

Although classical MHC class I glycoproteins bind peptide Ags for display at the cell surface, some MHC class I-related molecules such as the neonatal Fc receptor (FcRn) execute their function without binding peptide ligands. The three-dimensional structure of the FcRn suggested that a substitution of the conserved valine at position 165 of the alpha2 helix by proline contributed to a kink in the position of this helix relative to the alpha1 helix, and resulted in closing of the potential peptide-binding cleft. To test the contribution of proline 165 to the occlusion of the cleft and the binding of potential antigenic peptides, we introduced this mutation into the classical murine MHC class I molecule, H-2Dd, and characterized the ability of such a mutant to present peptide Ags to either a peptide-specific, H-2Dd-restricted T cell hybridoma (B4.

A three-domain T cell receptor is biologically active and specifically stains cell surface MHC/peptide complexes.

We have expressed in bacteria a single-chain T cell receptor (scTCR) with specificity for an HIV gp120-derived peptide bound to the murine MHC-I molecule, H-2Dd. This scTCR consists of V alpha covalently linked to the VbetaCbeta domains that was solubilized, refolded, and purified in high yield. Specific binding of the scTCR to MHC/peptide complexes was demonstrated by surface plasmon resonance, with a Kd of 2 to 8 x 10(-6) M.

A T cell receptor V alpha domain expressed in bacteria: does it dimerize in solution?

To evaluate the potential for dimerization through a particular T cell receptor (TCR) domain, we have cloned the cDNA encoding a TCR V alpha from a hybridoma with specificity for the human immunodeficiency virus (HIV) envelope glycoprotein 120-derived peptide P18-110 (RGPGRAFVTI) bound to the murine major histocompatibility complex (MHC) class I molecule, H-2Dd. This cDNA was then expressed in a bacterial vector, and protein, as inclusion bodies, was solubilized, refolded, and purified to homogeneity. Yield of the refolded material was from 10 to 50 mg per liter of bacterial culture, the protein was soluble at concentrations as high as 25 mg/ml, and it retained a high level of reactivity with an anti-V alpha 2 monoclonal antibody.

Studying interactions involving the T-cell antigen receptor by surface plasmon resonance.

T-lymphocyte activation is initiated by the interaction of the alpha beta TCR with a complex consisting of a class I or class II MHC-encoded molecule and an antigenic peptide, displayed on the surface of an antigen-presenting cell. Real-time binding measurements using surface plasmon resonance have revealed kinetic and equilibrium parameters for the interactions between purified MHC molecules and peptides, between TCR and MHC-peptide complexes, and between TRC and superantigens. The MHC-peptide interaction is characterized by its high affinity and long half-life, the TCR-MHC/peptide interaction by its low affinity and short half-life, and the TCR-superantigen interaction by its low-to-moderate affinity, which is dependent on the particular superantigen involved.

Highly sensitive immunoassays for detection of barley stripe mosaic virus and beet necrotic yellow vein virus.

Enzyme immunoassays based on the use of monoclonal antibodies (MAbs) were developed for the detection of the barley stripe mosaic virus (BSMV) and the beet necrotic yellow vein virus (BNYVV). Assays employing conjugates of MAbs to horseradish peroxidase (HRP) were compared to systems with biotinylated MAbs and streptavidin conjugated either to monomeric HRP or to HRP homopolymers with different polymerisation degrees including those of 20, 40 and 80. In the ELISA with streptavidin-polymeric HRP conjugates the assay detection limit was about 12-25 times as good as in the monoclonal double antibody sandwich ELISA.

Effective anti-metastatic melanoma vaccination with tumor cells transfected with MHC genes and/or infected with Newcastle disease virus (NDV).

The therapeutic efficacy of active immunization with B16-F10.9 melanoma cells transfected with syngeneic major histocompatibility complex (MHC) class-I genes, modified by infection with Newcastle Disease virus (NDV) or modified by both treatments, was compared. B16-F10.

c-fos and c-jun overexpression in malignant cells reduces their tumorigenic and metastatic potential, and affects their MHC class I gene expression.

Reduced co-expression of the c-fos and c-jun protooncogenes has been correlated with the down regulation of H-2K class I major histocompatibility antigens in high-metastatic cell lines from the Lewis lung carcinoma, B16 melanoma and the K1735 melanoma. Transfection of c-jun and c-fos genes into the high metastatic clones D122 (3LL) and F10.9 (B16 melanoma) resulted in activation of H-2 class I gene expression.

KBF1 (p50 NF-kappa B homodimer) acts as a repressor of H-2Kb gene expression in metastatic tumor cells.

Downregulation of major histocompatibility complex class I expression is causally related to high malignancy and low immunogenicity of certain murine tumors. In this study, we have analyzed the roles of the nuclear factors KBF1/p50 and p65 in regulation of class I expression in high and low metastatic tumor cells. Low class I-expressing cells show at higher levels of KBF1/p50 and NF-kappa B (p50/p65) binding activity than high class I-expressing cells.

H-2Db gene transfer into highly metastatic D122 cells results in tumor rejection in allogeneic recipients, but does not affect metastasis in syngeneic recipients. Implications for mechanisms of allorejection.

Highly metastatic, weakly immunogenic Lewis lung carcinoma clones express very low levels of H-2Kb and moderate levels of H-2Db class-I major histocompatibility complex antigens. These cells metastasize spontaneously in mice with C57BL/6 genetic background possessing the H-2Db locus, and grow as local tumors across allogeneic barriers. Transfection of the H-2Db genes into the highly metastatic clone D122 did not alter the growth or metastatic capacity of these cells in syngeneic mice.

[A new immunochemical method for detecting substance P receptors based on the biotin-streptavidin system].

A novel method for detecting the membrane receptors of Substance P (SP) has been developed. The method does not require radioactive derivatives of SP and is based on quantitation of specifically bound biotinylated SP (Bt-SP), the analysis being performed after destruction of the ligand-receptor complex in acidic conditions and Bt-SP extraction into solution. The acidic extract from the Bt-SP-membrane complex after neutralization is added to immulon microELISA plate coated with affinity purified anti-SP-antibodies.

Abolishment of metastasis formation by murine tumor cells transfected with "foreign" H-2K genes.

High metastatic, low immunogenic Lewis lung carcinoma clones express low levels of H-2Kb major histocompatibility complex antigens. These cells metastasize spontaneously in mice with C57BL/6 genetic background possessing the H-2Db locus. Transfection of different H-2K genes abrogates metastasis in H-2K, H-2D compatible mice and in C57BL/6 recipients.

Reversal of the metastatic phenotype in Lewis lung carcinoma cells after transfection with syngeneic H-2Kb gene.

High metastatic clones of the murine 3LL carcinoma express greatly reduced levels of H-2Kb major histocompatibility complex class I antigens, while low metastatic clones of the same tumor express high levels of H-2Kb. Induced expression of this antigen after transfection with the H-2Kb gene resulted in conversion of a metastatic to a non- or low-metastatic phenotype. Unlike the parental cells, transfected cells are potent inducers of H-2Kb-restricted syngeneic cytotoxic lymphocytes that kill the Kb-positive clones and cross-react with parental nontransfected cells.

The reversal of the metastatic phenotype by gene transfer.

When studying the function of MHC-restricted immune responses in controlling metastatic growth we discovered that highly metastatic clones of mouse tumours express the H-2D but lack expression of the H-2K gene of the MHC system. The de novo expression of the H-2K antigen, after H-2K gene transfection, resulted in the reversal of a metastatic to a non-metastatic phenotype. This reversal was causally related to the acquisition of H-2K-restricted immunogenic properties.

Expression of major histocompatibility class I genes in differentiating leukemic cells is temporally related to activation of c-fos proto-oncogene.

The relationship between the expression of the c-fos proto-oncogene and the expression of the class I major histocompatibility (MHC) antigens during the early stages of induced differentiation in three different leukemic cell lines was examined. In the U937 histiocytic lymphoma line TPA induced an increase in mRNA and cell surface MHC expression which followed induction of c-fos. In contrast, in the murine erythro-leukemia cell line, DMSO induced declining constitutive c-fos levels that were accompanied by declining mRNA and cell surface MHC expression.

[Ultramicromethod of erythrocyte immunoadsorption in screening for somatic antigens in acute myocardial infarct].

Test-systems for the identification of somatic myoglobin antigens and fibrin-fibrinogen degradation products in the sera of patients with acute myocardial infarction have been developed and tested. A significant correlation between erythrocyte immunoadsorption technique and solid-phase immunoenzyme assay was observed, the former technique being simpler and express.