Synthetic Cells Synthesize Therapeutic Proteins inside Tumors

Abstract

Protocells, artificial cell-like particles, capable of autonomously synthesizing RNA and proteins based on a DNA template, are emerging platforms for studying cellular functions and for revealing the origins-of-life. Here, we show for the first time that artificial lipid-based vesicles, containing the molecular machinery necessary for transcription and translation, can be used to synthesize anti-cancer proteins inside tumors. The particles were engineered as stand-alone systems, sourcing nutrients from their biological microenvironment to trigger protein synthesis. When pre-loaded with template DNA, amino acids and energy-supplying molecules protocells synthesized, up to 2×107 copies of superfolder green fluorescent protein (sfGFP) were synthesized in each liposome. A variety of proteins, having molecular weights reaching 66 kDa and with diagnostic and therapeutic activities, were synthesized inside the particles. Incubating protein producing particles, encoded to secrete Pseudomonas exotoxin A (PE) with 4T1 breast cancer cells in culture, resulted in killing of most of the malignant cells. In mice bearing 4T1 tumors, histological evaluation of the tumor tissue after a local injection of PE-producing particles, indicating robust apoptosis. Protein producing particles are synthetic-biology platforms capable of synthesizing therapeutic proteins on-demand.

Cell-free transcriptional regulation via nucleic-acid-based transcription factors

“…Applications of cell-free synthetic gene circuits are now beginning to emerge, such as portable diagnostics, distributed bio-manufacturing, and therapeutic artificial-cells. 3,30,31 Regulating gene-expression dynamics is desirable in these applications in order to focus a finite amount of energy and resources towards manufacturing the right product at the right time.…”

Bottom-Up Construction of Complex Biomolecular Systems With Cell-Free Synthetic Biology

“…Dwidar et al (2019) engineered a riboswitch for the biologically relevant small molecule histamine into liposomebased artificial cells, which could respond to the presence of histamine in a variety of programmed ways (Figure 5C). Finally, liposome-based artificial cells expressing Pseudomonas exotoxin A were injected in vivo inside mice tumors and an increase in caspase activity was shown (Krinsky et al, 2017), suggesting their potential use in therapeutic or diagnostic applications.…”

Cell‐free protein synthesis: The transition from batch reactions to minimal cells and microfluidic devices

“…PE presence in nanoparticles was approved by experimental results. Also, the percentage of the cells that were killed by PE was above 80% (Krinsky et al, 2018). Furthermore, besides protein synthesis, production of lipids as essential components of the living cellular membrane is achieved.…”