Rapid PCR construction of a gene containing Lym-1 antibody variable regions.

Abstract

1Department of Internal Medicine, University of California, Davis, Medical Center, Sacramento, California 95817; 2Department of Chemistry, University of California, Davis, California 95616 Clinical applications of monoclonal antibodies (mAbs) to tumor-associated antigens provide potential for targeted cancer therapy. Radiolabeled mAb Lym-1 has been used successfully to treat >50 patients with B-cell lymphoma. ~-3) The uptake of Lym-1 in lymphomas is rapid and efficient compared with solid tu m o r s . However, the amount of the radioactive dose injected on these large molecules reaching each gram of tumor is usually <0.1%. When Lym-1 is labeled with radiometal chelates to enhance its tumoricidal effects, there is undesirable accumulat ion and slow clearance of radioactivity in the liver. Evidence suggests that this is related to receptors for components of the mAb Fc region. (4's) Recent studies (6'7) have demonstrated that two recombinant radio-directed single-chain ant igen-binding (SCA) proteins derived from monoclonal antibodies have in vivo tumor-targeting capacity. Although the percent injected dose per gram of tumor is low, the therapeutic index (tumor to normal organ radiation dose) is several times that of the intact mAb, F(ab')2 , or Fab' fragments. (7) This improvement in biokinetics properties of the radiolabeled SCA protein make it a promising agent for alternative therapeutic strategies. Furthermore, SCA proteins facilitate development of single or bispecific radioimmunoconjugates , humanized antibodies, or new versions of tumor-reactive molecules for immunotherapy. This may be particularly rewarding for a mAb such as Lym-1 that has already been demonstrated to be successful in clinical radioimmunoconjugate cancer therapy. Gene engineering techniques have been used to produce the SCA protein, which consists of the heavy-chain variable region (VH) and the light-chain variable region (VL) of an antibody synthesized as a single polypeptide. (8) In this engineered protein, the carboxyl terminus of one variable region is linked by a linker peptide to the amino terminus of the other. Some are VH--V L and some are