• Vol. 38 No. 7, 621–629
  • 15 July 2009

Use of Phage Display to Isolate Specific Human Monoclonal Antibody Fragments Against a Potential Target for Multiple Myeloma

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ABSTRACT

Introduction: Multiple myeloma (MM), a malignancy of plasma cells, accounts for 10% of all haematological malignancies and is currently incurable. Although it can be treated, the disease tends to relapse after several years and becomes increasingly resistant to conventional therapy. Investigations into using humoral therapy for MM are now underway with a view that novel therapeutic agents may provide a more targeted therapy for MM. Materials and Methods: Here, phage display, a faster and more efficient method compared to classical hybridoma fusion technology, was used as a proof-of-concept to isolate several single-chain Fragment variables (scFv) against Ku86. Results: Anti-Ku86 polyclonal scFvs biopanning was successful where third round scFvs (A450~1.1) showed a 1/3 increase in binding as compared to the first round scFvs (A450~0.4) with 100ug/mL of antigen (purified human Ku86). Subsequent selection and verification of monoclonal antibodies using third round biopanning revealed 4 good affinity binding clones ranging from A450~0.1 to A450~0.15 on 12.5ug/mL of antigen as compared to low binders (A450~0.07) and these antibodies bind to Ku86 in a specific and dose-dependent manner. Comparative studies were also performed with commercially available murine antibodies and results suggest that 2 of the clones may bind close to the following epitopes aa506-541 and aa1- 374. Conclusions: These studies using phage display provide an alternative and viable method to screen for antibodies quickly and results show that good affinity antibodies against Ku86 have been successfully isolated and they can be used for further studies on MM and form the basis for further development as anti-cancer therapeutic agents.


Multiple myeloma (MM) is a malignant plasma-cell neoplasm that is characterised by an excess of mature B-cells in the bone marrow (plasmacytosis), in association with secretion of monoclonal immunoglobulins (Ig) in serum and/ or urine, decreased normal Ig and lytic bone disease.1,2 MM accounts for 10% to 20% of all malignant haematological cancers and myeloma patients often have extensive skeletal destruction, which are a main cause of morbidity and mortality.2-4 Although MM can be treated, the disease ultimately relapses in most cases and becomes increasingly resistant to therapy until conventional options are exhausted, prompting an urgent need for novel treatment.5 Major advances have been made in the understanding of plasma cell biology and the importance of the microenvironment in MM development6-9 in the past decade, leading to the development of newer agents such as thalidomide, its analog lenalidomide10 and the proteasome inhibitor bortezomib.11 These agents have been used in combination with existing agents against MM, but prospects of a cure of MM with these agents is still uncertain and major limitations exist, such as patient resistance and toxicities relating to the use of these drugs.

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