Exosome-mediated genetic reprogramming of tumor-associated macrophages by exoASO-STAT6 leads to potent monotherapy antitumor activity.

Effectiveness of checkpoint immunotherapy in cancer can be undermined by immunosuppressive tumor-associated macrophages (TAMs) with an M2 phenotype. Reprogramming TAMs toward a proinflammatory M1 phenotype is a novel approach to induce antitumor immunity. The M2 phenotype is controlled by key transcription factors such as signal transducer and activator of transcription 6 (STAT6), which have been "undruggable" selectively in TAMs. We describe an engineered exosome therapeutic candidate delivering an antisense oligonucleotide (ASO) targeting STAT6 (exoASO-STAT6), which selectively silences STAT6 expression in TAMs. In syngeneic models of colorectal cancer and hepatocellular carcinoma, exoASO-STAT6 monotherapy results in >90% tumor growth inhibition and 50 to 80% complete remissions. Administration of exoASO-STAT6 leads to induction of nitric oxide synthase 2 (NOS2), an M1 macrophage marker, resulting in remodeling of the tumor microenvironment and generation of a CD8 T cell-mediated adaptive immune response. Collectively, exoASO-STAT6 represents the first platform targeting transcription factors in TAMs in a highly selective manner.

A versatile platform for generating engineered extracellular vesicles with defined therapeutic properties.

Extracellular vesicles (EVs) are an important intercellular communication system facilitating the transfer of macromolecules between cells. Delivery of exogenous cargo tethered to the EV surface or packaged inside the lumen are key strategies for generating therapeutic EVs. We identified two "scaffold" proteins, PTGFRN and BASP1, that are preferentially sorted into EVs and enable high-density surface display and luminal loading of a wide range of molecules, including cytokines, antibody fragments, RNA binding proteins, vaccine antigens, Cas9, and members of the TNF superfamily. Molecules were loaded into EVs at high density and exhibited potent in vitro activity when fused to full-length or truncated forms of PTGFRN or BASP1. Furthermore, these engineered EVs retained pharmacodynamic activity in a variety of animal models. This engineering platform provides a simple approach to functionalize EVs with topologically diverse macromolecules and represents a significant advance toward unlocking the therapeutic potential of EVs.

In vitro characterization of an acetylcholine receptor-transferrin fusion protein for the treatment of myasthenia gravis.

SHG2210, a fusion protein containing the N-terminus of human nicotinic acetylcholine receptor α (AchR-α; aa1-210) and human transferrin (TF), was characterized as a potential therapeutic for myasthenia gravis (MG) caused predominately by α subunit autoantibodies. SHG2210 was shown to be able to bind to α subunit autoantibodies and the TF receptor (TFR). SHG2210 and SHG2210-anti-AchR antibody complex are internalized through TFR-mediated endocytosis. The SHG2210 and SHG2210-anti-AchR antibody complex is present in Lamp1-positive lysosomal compartments after internalization; however, neither SHG2210 nor SHG2210-antibody complex is present in Rab11-positive recycling endosomes. SHG2210 bound to α subunit of AChR autoantibodies may be cleared by the lysosome, resulting in short cellular half-life relative to SHG2210. SHG2210 is shown to have a protective effect on antigenic modulation of the AChR induced by serum from select patients with MG, suggesting that a fusion protein approach may be an effective therapeutic for treating MG.