The promise of effective treatments that can be produced cheaply, delivered orally, and with minimal systemic side effects continues to fuel interest in using engineered bacteria as delivery systems to bring about a new wave of therapeutics. In the field of synthetic biology, a wide array of genetic tools have been developed for several microbial host organisms, or chassis, that can be engineered to sense and respond to environmental signals within the body. Different bacterial chassis have been designed to treat diseases in gastrointestinal tract or in the microenvironment of solid tumors (1, 2). The use of recombinant bacteria producing and delivering in situ the molecule of interest, has been extensively described (3-5) (6, 7) (8-13) (14). As an example, Escherichia coli has been engineered to eliminate and prevent Pseudomonas aeruginosa gut infection (15). Engineered Lactobacillus reuteri as a probiotic has been shown to decrease high levels of phenylalanine in blood in homozygous PAHenu2 mice (Phenylketonuria model) (16). Despite most bacterial vectors have been designed to treat gut diseases there are some examples of using engineered bacteria to treat diseases in other organs. As an example, both Lactobacillus and Saccharomyces cerevisiae, have been engineered to prevent HIV infection in women (17-19).

Although these recombinant bacteria have shown high efficacy in animal models, they have yet to yield clinical successes. This is due in part due to the challenge of achieving and maintaining sufficiently high concentrations of the therapeutic molecule at the site of disease, without causing an adverse body reaction (20). Selecting the bacteria to be used to deliver therapeutic molecules depending on the site of action, could be a strategy to ensure survival of the bacteria without causing an adverse reaction and local delivery of the therapeutic agents. Despite the problems encountered there is a consensus that the engineering of bacteria as delivery systems to treat diseases could be the new wave of therapeutics.

Recently, considerations for the design and development of engineered live biotherapeutics to achieve regulatory and patient acceptance have been reported (21), noting both the opportunities and challenges associated to applying synthetic biology tools to the development of therapeutics for human disease. Important aspects that need to be covered include: i) regulatory considerations, ii) the strategies for how these therapeutics can be evaluated for their pharmacokinetic and pharmacodynamic properties and iii) manufacturability of engineered microbes to enable production at scale, as well as achieving formulations and presentations that support the needs of patients. In order to deal successfully with these challenges, proper standardization of tools and processes is required to ensure the maximum reproducibility and efficacy of the developed product.

In line with this, BIOROBOOST aims to integrate the expertise of a scientific community that does rational engineering of different biological systems (Pseudomonas, E. coli, Mycoplasma pneumoniae, Bacillus subtilis…) to develop a wide range of applications not only for human health but also to solve environmental problems. Independently of the system and the application, a number of molecular and bioinformatics tools are used by members of the synthetic biology field, and it is critical to standardize that ensure the reproducibility of these methods across different groups.

References

  1. Riglar DT, Giessen TW, Baym M, Kerns SJ, Niederhuber MJ, Bronson RT, et al. Engineered bacteria can function in the mammalian gut long-term as live diagnostics of inflammation. Nat Biotechnol. 2017;35(7):653-8.
  2. Stritzker J, Weibel S, Hill PJ, Oelschlaeger TA, Goebel W, Szalay AA. Tumor-specific colonization, tissue distribution, and gene induction by probiotic Escherichia coli Nissle 1917 in live mice. Int J Med Microbiol. 2007;297(3):151-62.
  3. Bermudez-Humaran LG, Aubry C, Motta JP, Deraison C, Steidler L, Vergnolle N, et al. Engineering lactococci and lactobacilli for human health. Curr Opin Microbiol. 2013;16(3):278-83.
  4. Kuehn MJ. Genetically engineered probiotic competition. Gastroenterology. 2006;130(6):1915-6.
  5. Wu MR, Jusiak B, Lu TK. Engineering advanced cancer therapies with synthetic biology. Nat Rev Cancer. 2019;19(4):187-95.
  6. Pouwels PH, Vriesema A, Martinez B, Tielen FJ, Seegers JF, Leer RJ, et al. Lactobacilli as vehicles for targeting antigens to mucosal tissues by surface exposition of foreign antigens. Methods Enzymol. 2001;336:369-89.
  7. Lalsiamthara J, Kim JH, Lee JH. Engineering of a rough auxotrophic mutant Salmonella Typhimurium for effective delivery. Oncotarget. 2018;9(39):25441-57.
  8. Martin R, Chain F, Miquel S, Natividad JM, Sokol H, Verdu EF, et al. Effects in the use of a genetically engineered strain of Lactococcus lactis delivering in situ IL-10 as a therapy to treat low-grade colon inflammation. Hum Vaccin Immunother. 2014;10(6):1611-21.
  9. Steidler L, Neirynck S, Huyghebaert N, Snoeck V, Vermeire A, Goddeeris B, et al. Biological containment of genetically modified Lactococcus lactis for intestinal delivery of human interleukin 10. Nat Biotechnol. 2003;21(7):785-9.
  10. Steidler L, Hans W, Schotte L, Neirynck S, Obermeier F, Falk W, et al. Treatment of murine colitis by Lactococcus lactis secreting interleukin-10. Science. 2000;289(5483):1352-5.
  11. Schotte L, Steidler L, Vandekerckhove J, Remaut E. Secretion of biologically active murine interleukin-10 by Lactococcus lactis. Enzyme Microb Technol. 2000;27(10):761-5.
  12. Vandenbroucke K, de Haard H, Beirnaert E, Dreier T, Lauwereys M, Huyck L, et al. Orally administered L. lactis secreting an anti-TNF Nanobody demonstrate efficacy in chronic colitis. Mucosal Immunol. 2010;3(1):49-56.
  13. Vandenbroucke K, Hans W, Van Huysse J, Neirynck S, Demetter P, Remaut E, et al. Active delivery of trefoil factors by genetically modified Lactococcus lactis prevents and heals acute colitis in mice. Gastroenterology. 2004;127(2):502-13.
  14. Praveschotinunt P, Duraj-Thatte AM, Gelfat I, Bahl F, Chou DB, Joshi NS. Engineered E. coli Nissle 1917 for the delivery of matrix-tethered therapeutic domains to the gut. Nat Commun. 2019;10(1):5580.
  15. Hwang IY, Koh E, Wong A, March JC, Bentley WE, Lee YS, et al. Engineered probiotic Escherichia coli can eliminate and prevent Pseudomonas aeruginosa gut infection in animal models. Nat Commun. 2017;8:15028.
  16. Durrer KE, Allen MS, Hunt von Herbing I. Genetically engineered probiotic for the treatment of phenylketonuria (PKU); assessment of a novel treatment in vitro and in the PAHenu2 mouse model of PKU. PLoS One. 2017;12(5):e0176286.
  17. Liu X, Lagenaur LA, Simpson DA, Essenmacher KP, Frazier-Parker CL, Liu Y, et al. Engineered vaginal lactobacillus strain for mucosal delivery of the human immunodeficiency virus inhibitor cyanovirin-N. Antimicrob Agents Chemother. 2006;50(10):3250-9.
  18. Liu X, Lagenaur LA, Lee PP, Xu Q. Engineering of a human vaginal Lactobacillus strain for surface expression of two-domain CD4 molecules. Appl Environ Microbiol. 2008;74(15):4626-35.
  19. Palma ML, Garcia-Bates TM, Martins FS, Douradinha B. Genetically engineered probiotic Saccharomyces cerevisiae strains mature human dendritic cells and stimulate Gag-specific memory CD8(+) T cells ex vivo. Appl Microbiol Biotechnol. 2019;103(13):5183-92.
  20. Smart AL, Gaisford S, Basit AW. Oral peptide and protein delivery: intestinal obstacles and commercial prospects. Expert Opin Drug Deliv. 2014;11(8):1323-35.
  21. Charbonneau MR, Isabella VM, Li N, Kurtz CB. Developing a new class of engineered live bacterial therapeutics to treat human diseases. Nat Commun. 2020;11(1):1738.

 

Text by: Maria Lluch (Pulmobiotics), April 2020

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