Synthesis and preliminary evaluation of ^99mTc-Hynic-fragments [F(ab')₂ and F(ab')] of Rituximab as radioimmunoscintigraphic agents for patients with Non-Hodgkin's lymphoma

Highlights: • F(ab')₂-Rituximab and F(ab')-Rituximab prepared and characterized by SDS-PAGE and SE-HPLC. • Fragments of Rituximab conjugated with Hynic and radiolabeled with ^99mTc. • ^99mTc-Hynic-F(ab')₂/F(ab')-Rituximab exhibited good in vitro stability up to 24 h. • In vitro cell binding studies confirmed specificity of radioimmunoconjugates to CD20 antigen. • ^99mTc labeled Rituximab fragments cleared predominantly through the renal route. - Abstract: This study was to evaluate the potential of ^99mTc-Hynic-fragments of Rituximab as radioimmunoscintigraphic agents for diagnosis of patients with Non-Hodgkin's Lymphoma (NHL). Rituximab was digested with immobilized pepsin and papain to yield F(ab')₂-Rituximab and F(ab')-Rituximab fragments respectively. Purified fragments were characterized by SE-HPLC and SDS-PAGE and subsequently radiolabeled with technetium-99m using Hynic as bifunctional chelator. The ^99mTc-Hynic-F(ab')₂-Rituximab and ^99mTc-Hynic-F(ab')-Rituximab exhibited good in-vitro stability and specificity to Raji cells. Biodistribution studies demonstrated rapid pharmacokinetics and clearance predominantly through renal route.