Broadening and Enhancing Functions of Antibodies by Self-Assembling Multimerization at Cell Surface.

Monoclonal antibody therapy has offered treatment benefits. Nonetheless, a lack of efficacy still exists, partially because monovalent binding of antibodies to specific receptors fails to translate into an active response. Here, we report a pretargeting-postassembly approach that exploits the selective Watson-Crick base pairing properties of oligonucleotides and multivalently tethers receptor-prebound antibodies to albumin at the cell surface. We demonstrate that this two-step self-assembling strategy allows sequential actions of receptor binding and clustering that broadens and strengthens the functions of antibodies. We show that anti-CD20 obinutuzumab (OBN) modified with one morpholino oligonucleotide (OBN-MORF1) maintains the feature of naked OBN antibody upon CD20 binding, and results in actin redistribution, homotypic adhesion, and lysosome-mediated cell death. Consecutive treatment with albumin grafted with multiple copies of a complementary morpholino oligonucleotide (HSA-(MORF2)x) hybridizes with surface-attached OBN-MORF1, manipulates CD20 clustering, and engages additional signals to induce calcium influx and caspase-related apoptosis. With the two types of different mechanisms collaborating in one system, the simple design exerted a notable survival extension of mice bearing disseminated B-cell lymphomas.

Human Serum Albumin-Based Drug-Free Macromolecular Therapeutics: Apoptosis Induction by Coiled-Coil-Mediated Cross-Linking of CD20 Antigens on Lymphoma B Cell Surface.

A therapeutic platform-drug-free macromolecular therapeutics (DFMT)-that induces apoptosis in B cells by cross-linking of CD20 receptors, without the need for low molecular weight cytotoxic drug, is developed. In this report, a DFMT system is synthesized and evaluated based on human serum albumin (HSA) and two complementary coiled-coil forming peptides, CCE and CCK. Fab' fragment of anti-CD20 monoclonal antibody rituximab is attached to CCE (Fab'-CCE); multiple grafts of CCK are conjugated to HSA (HSA-(CCK)7 ). The colocalization of both nanoconjugates at the surface of non-Hodgkin's lymphoma (NHL) Raji cells is demonstrated by confocal fluorescence microscopy. The colocalization leads to coiled-coil formation, CD20 cross-linking, and apoptosis induction. The apoptotic levels are evaluated by Annexin V, Caspase 3, and terminal deoxynucleotidyl transferase dUTP nick end labeling assays. Selective surface binding of DFMT to CD20+ cells is validated in experiments on a coculture of CD20+ (Raji) and CD20-(DG-75) cells. It is found that DFMT can trigger calcium influx only in Raji cells, but not in DG-75 cells. A highly specific treatment for NHL and other B cell malignancies with considerable translational potential is presented by HSA-based DFMT system.

Rational design of "heat seeking" drug loaded polypeptide nanoparticles that thermally target solid tumors.

This paper demonstrates the first example of targeting a solid tumor that is externally heated to 42 °C by "heat seeking" drug-loaded polypeptide nanoparticles. These nanoparticles consist of a thermally responsive elastin-like polypeptide (ELP) conjugated to multiple copies of a hydrophobic cancer drug. To rationally design drug-loaded nanoparticles that exhibit thermal responsiveness in the narrow temperature range between 37 and 42 °C, an analytical model was developed that relates ELP composition and chain length to the nanoparticle phase transition temperature. Suitable candidates were designed based on the predictions of the model and tested in vivo by intravital confocal fluorescence microscopy of solid tumors, which revealed that the nanoparticles aggregate in the vasculature of tumors heated to 42 °C and that the aggregation is reversible as the temperature reverts to 37 °C. Biodistribution studies showed that the most effective strategy to target the nanoparticles to tumors is to thermally cycle the tumors between 37 and 42 °C. These nanoparticles set the stage for the targeted delivery of a range of cancer chemotherapeutics by externally applied mild hyperthermia of solid tumors.