LAUSR

Simple Summary Today, many incurable blood malignancies, such as multiple myeloma (MM), acute myeloid leukemia, and myelodysplastic syndrome, lack effective immunotherapy options. In this study, we used phage display technology to develop candidate therapeutic antibodies targeting cell surface epitopes on MM cells. Antibody derivatives against surface glycoforms were developed and optimized for the induction of antibody-dependent cellular cytotoxicity against MM cells. Additionally, an antibody-mimetic peptide was developed, and it was used to deliver a pro-apoptotic peptide that killed MM cells. The engineered antibody derivatives show promise for the further development of cancer therapeutics. Abstract Despite therapeutic advances in recent years, there are still unmet medical needs for patients with multiple myeloma (MM). Hence, new therapeutic strategies are needed. Using phage display for screening a large repertoire of single chain variable fragments (scFvs), we isolated several candidates that recognize a heavily sulfated MM-specific glycoform of the surface antigen syndecan-1 (CD138). One of the engineered scFv-Fc antibodies, named MM1, activated NK cells and induced antibody-dependent cellular cytotoxicity against MM cells. Analysis of the binding specificity by competitive binding assays with various glycan ligands identified N-sulfation of glucosamine units as essential for binding. Additionally, site-directed mutagenesis revealed that the amino acids arginine and histidine in the complementarily determining regions (CDRs) 2 and 3 of the heavy chain are important for binding. Based on this observation, a heavy-chain antibody, known as a nanobody, and a peptide mimicking the CDR loop sequences were designed. Both variants exhibited high affinity and specificity to MM cells as compared to blood lymphocytes. Specific killing of MM cells was achieved by conjugating the CDR2/3 mimic peptide to a pro-apoptotic peptide (KLAKLAK)2. In a co-culture model, the fusion peptide killed MM cells, while leaving normal peripheral blood mononuclear cells unaffected. Collectively, the development of antibodies and peptides that detect tumor-specific glycoforms of therapeutic targets holds promise for improving targeted therapies and tumor imaging.