PMCCPMCCPMCC

Search tips
Search criteria 

Advanced

 
Logo of molmedLink to Publisher's site
 
Mol Med. 1999 January; 5(1): 35–45.
PMCID: PMC2230376

Two neutralizing human anti-RSV antibodies: cloning, expression, and characterization.

Abstract

BACKGROUND: Respiratory syncytial virus (RSV) infection is a major problem in the newborn and aging populations. Fully human monoclonal antibodies with the ability to neutralize RSV could have a major impact on the immunotherapy of the disease. The generation of human antibodies has been difficult because there exists no general way to activate B cells against an antigen of choice in vitro. MATERIALS AND METHODS: Human spleen cells from individuals exposed to RSV were used to repopulate SCID mice. Hu-SCID mice were boosted with RSV fusion (F)-protein and subsequently developed B cell tumors. The tumors were removed and cultured and subcloned in vitro, using a feeder layer of CD154-expressing T cells. Two of these tumors produced the antibodies designated RF-1 and RF-2. VL genes were isolated by standard PCR techniques, however, it was necessary to use high-temperature reverse transcriptase to clone the VH genes. RESULTS: RF-1 and RF-2 VH genes were both found to be closely related members of the VH2 family. Vk genes originated from the VK III family. RF-1 and RF-2 recombinant antibodies expressed in CHO cells (cRF-1 and cRF-2) were found to have affinities for RSV F-protein of 0.1 nM and 0.07 nM, respectively, and both were able to neutralize several A and B subtypes of RSV. CONCLUSION: The technique of immortalizing human B lymphocytes, by passage in SCID mice and expression as recombinant antibodies in CHO cells, provides a method by which high-affinity human antibodies can be developed for immunotherapy of viral diseases.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.6M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Morrison SL, Johnson MJ, Herzenberg LA, Oi VT. Chimeric human antibody molecules: mouse antigen-binding domains with human constant region domains. Proc Natl Acad Sci U S A. 1984 Nov;81(21):6851–6855. [PubMed]
  • Riechmann L, Clark M, Waldmann H, Winter G. Reshaping human antibodies for therapy. Nature. 1988 Mar 24;332(6162):323–327. [PubMed]
  • Queen C, Schneider WP, Selick HE, Payne PW, Landolfi NF, Duncan JF, Avdalovic NM, Levitt M, Junghans RP, Waldmann TA. A humanized antibody that binds to the interleukin 2 receptor. Proc Natl Acad Sci U S A. 1989 Dec;86(24):10029–10033. [PubMed]
  • Newman R, Alberts J, Anderson D, Carner K, Heard C, Norton F, Raab R, Reff M, Shuey S, Hanna N. "Primatization" of recombinant antibodies for immunotherapy of human diseases: a macaque/human chimeric antibody against human CD4. Biotechnology (N Y) 1992 Nov;10(11):1455–1460. [PubMed]
  • Barbas CF, 3rd, Bain JD, Hoekstra DM, Lerner RA. Semisynthetic combinatorial antibody libraries: a chemical solution to the diversity problem. Proc Natl Acad Sci U S A. 1992 May 15;89(10):4457–4461. [PubMed]
  • Burton DR, Barbas CF., 3rd Human antibodies from combinatorial libraries. Adv Immunol. 1994;57:191–280. [PubMed]
  • Winter G, Griffiths AD, Hawkins RE, Hoogenboom HR. Making antibodies by phage display technology. Annu Rev Immunol. 1994;12:433–455. [PubMed]
  • Griffiths AD, Malmqvist M, Marks JD, Bye JM, Embleton MJ, McCafferty J, Baier M, Holliger KP, Gorick BD, Hughes-Jones NC, et al. Human anti-self antibodies with high specificity from phage display libraries. EMBO J. 1993 Feb;12(2):725–734. [PubMed]
  • Griffiths AD, Williams SC, Hartley O, Tomlinson IM, Waterhouse P, Crosby WL, Kontermann RE, Jones PT, Low NM, Allison TJ, et al. Isolation of high affinity human antibodies directly from large synthetic repertoires. EMBO J. 1994 Jul 15;13(14):3245–3260. [PubMed]
  • Brams P, Nguyen ML, Chamat S, Royston I, Morrow PR. Antigen-specific IgG responses from naive human splenocytes: in vitro priming followed by antigen boost in the SCID mouse. J Immunol. 1998 Mar 1;160(5):2051–2058. [PubMed]
  • Walsh EE, Brandriss MW, Schlesinger JJ. Purification and characterization of the respiratory syncytial virus fusion protein. J Gen Virol. 1985 Mar;66(Pt 3):409–415. [PubMed]
  • Zhang XH, Hauser C, Zubler RH. Soluble factor-independent stimulation of human B cell response by mouse thymoma cells. Cyclosporine A-resistant and -sensitive cell contact signals. J Immunol. 1990 Apr 15;144(8):2955–2960. [PubMed]
  • Tohma S, Lipsky PE. Analysis of the mechanisms of T cell-dependent polyclonal activation of human B cells. Induction of human B cell responses by fixed activated T cells. J Immunol. 1991 Apr 15;146(8):2544–2552. [PubMed]
  • Reff ME, Carner K, Chambers KS, Chinn PC, Leonard JE, Raab R, Newman RA, Hanna N, Anderson DR. Depletion of B cells in vivo by a chimeric mouse human monoclonal antibody to CD20. Blood. 1994 Jan 15;83(2):435–445. [PubMed]
  • Walsh EE, Cote PJ, Fernie BF, Schlesinger JJ, Brandriss MW. Analysis of the respiratory syncytial virus fusion protein using monoclonal and polyclonal antibodies. J Gen Virol. 1986 Mar;67(Pt 3):505–513. [PubMed]
  • Kozbor D, Roder JC. Requirements for the establishment of high-titered human monoclonal antibodies against tetanus toxoid using the Epstein-Barr virus technique. J Immunol. 1981 Oct;127(4):1275–1280. [PubMed]
  • Vandekerckhove BA, Jones D, Punnonen J, Schols D, Lin HC, Duncan B, Bacchetta R, de Vries JE, Roncarolo MG. Human Ig production and isotype switching in severe combined immunodeficient-human mice. J Immunol. 1993 Jul 1;151(1):128–137. [PubMed]
  • Finnern R, Pedrollo E, Fisch I, Wieslander J, Marks JD, Lockwood CM, Ouwehand WH. Human autoimmune anti-proteinase 3 scFv from a phage display library. Clin Exp Immunol. 1997 Feb;107(2):269–281. [PubMed]
  • Kohsaka H, Carson DA, Rassenti LZ, Ollier WE, Chen PP, Kipps TJ, Miyasaka N. The human immunoglobulin V(H) gene repertoire is genetically controlled and unaltered by chronic autoimmune stimulation. J Clin Invest. 1996 Dec 15;98(12):2794–2800. [PMC free article] [PubMed]
  • Sheets MD, Amersdorfer P, Finnern R, Sargent P, Lindquist E, Schier R, Hemingsen G, Wong C, Gerhart JC, Marks JD, et al. Efficient construction of a large nonimmune phage antibody library: the production of high-affinity human single-chain antibodies to protein antigens. Proc Natl Acad Sci U S A. 1998 May 26;95(11):6157–6162. [PubMed]
  • Groothuis JR, Simoes EA, Levin MJ, Hall CB, Long CE, Rodriguez WJ, Arrobio J, Meissner HC, Fulton DR, Welliver RC, et al. Prophylactic administration of respiratory syncytial virus immune globulin to high-risk infants and young children. The Respiratory Syncytial Virus Immune Globulin Study Group. N Engl J Med. 1993 Nov 18;329(21):1524–1530. [PubMed]
  • Wyde PR, Moore DK, Hepburn T, Silverman CL, Porter TG, Gross M, Taylor G, Demuth SG, Dillon SB. Evaluation of the protective efficacy of reshaped human monoclonal antibody RSHZ19 against respiratory syncytial virus in cotton rats. Pediatr Res. 1995 Oct;38(4):543–550. [PubMed]
  • Barbas CF, 3rd, Crowe JE, Jr, Cababa D, Jones TM, Zebedee SL, Murphy BR, Chanock RM, Burton DR. Human monoclonal Fab fragments derived from a combinatorial library bind to respiratory syncytial virus F glycoprotein and neutralize infectivity. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10164–10168. [PubMed]

Articles from Molecular Medicine are provided here courtesy of The Feinstein Institute for Medical Research at North Shore LIJ