Connie: My dearest friends, thank you for joining us today on this beautiful Saturday evening. Today, we are glad to have invited our old friend, Dr. Hofstadter, to introduce applications of new probiotics. Hello, Dr. Hofstadter. Thank you for being here.
Dr. Hofstadter: Thanks for inviting me, Connie. Good evening, everyone. It is quite nice to see you again. I’m very excited to be here.
Connie: In the last program, we introduced the application of recombinant DNA technology in lactic acid bacteria metabolic engineering. We learned that metabolic engineering lactic acid bacteria can be roughly divided into food fermentation initiators and cell factories that produce chemicals, vitamins, and secondary metabolites. By using metabolic engineering to modify a gene of lactic acid bacteria, it is possible to solve the problem of acidosis in children. In addition, recombinant lactic acid bacteria can make food taste better. In today’s episode, we will explore the application of lactic acid bacteria modified by metabolic engineering in the field of disease treatment. Where should we start, Dr. Hofstadter?
Dr. Hofstadter: Let’s first review the role of metabolically engineered lactic acid bacteria in food. Engineered lactic acid bacteria are not only used in the dairy industry but also play an additional role in protein production and the food fermentation industry. These industries produce vitamin B, diacetyl, acetaldehyde, and folic acid. Recombinant metabolites and proteins produced with the help of Engineering organisms do not need food quality carriers in the production process. However, they have the greatest usage when co-administered to humans or animals.
Connie: Do you mean some carriers need to be taken orally?
Dr. Hofstadter: That’s right. The concept of oral carrier evolved from lactic acid bacteria and bifidobacteria. Because they are acid resistant and still adhere to the mucosal epithelium. In addition, these bacteria also cultivate genes that express therapeutic molecules, such as oral interleukin.
Connie: In these years of research, have any strains with therapeutic effects been obtained?
Dr. Hofstadter: Yes, and I will give you an example using a potential probiotic strain named Lactobacillus acidophilus NCK56. This strain was integrated by two-hybrid and excision after phosphoglyceryl-transferase deletion, and ended up with eliminated antibiotic resistance. Here is another example. Eosinophils have been reported to help regulate inflammatory response and protect colonic polyposis in a mouse model.
Connie: I have learned that integrating food quality vectors into chromosomes can produce more stable vectors. Have you seen any specific cases?
Dr. Hofstadter: Food quality carriers are becoming more widely used. Let me share with you a practical application. Some research institutes have clarified the integrated chromosome expression system in paralactobacillus. The system is based on the aggregation promoter gene of Lactobacillus curlicus. Which can be expressed stably.
Connie: Today, there is a large number of patients with metabolic disorders. What benefits can metabolically engineered probiotics bring to these patients?
Dr. Hofstadter: Although patients with metabolic disorders are not uncommon, we know that the treatment of these diseases is quite expensive. And the treatment drugs and means for patients need to be strictly observed, because they will affect the patient’s lifestyle and survival rate. But engineered microorganisms can help patients with various diseases and provide long-term benefits.
Connie: It is estimated that by 2025, more than 1 billion people in the world will suffer from hypertension. Therefore, it is urgent to solve the problem of hypertension. Do engineered microorganisms help with hypertension as well?
Dr. Hofstadter: Yeah. Angiotensin-converting enzyme inhibitory peptide is highly associated with hypertension. It may help relax blood vessels, reduce the reabsorption of water by the kidney, and ultimately help reduce blood pressure. Angiotensin-converting enzyme inhibitory peptide can be carried out with the help of Lactobacillus Plantarum engineering strain nc8. The coding sequences of tuna structural protein and albacore structural protein are integrated by arginine ligands. Animal experiments show that arginine ligand plays a role in the reduction of systolic blood pressure, endothelin, and angiotensin II in spontaneously hypertensive rats.
Connie: What are the advantages of this engineered strain of Lactobacillus Plantarum over traditional inhibitors?
Dr. Hofstadter: Compared with traditional angiotensin-converting enzyme inhibitors, some studies have found that the modified Lactobacillus Plantarum showed an additional antihypertensive effect in vivo for 10 days, and no side effects were observed.
Connie: We have mentioned several times in previous episodes about the beneficial effects of probiotics on diabetes treatment. So, what is the contribution of modified probiotics in this regard?
Dr. Hofstadter: Again, I will refer to some results from published studies. In an animal experiment, the production of Lactococcus lactis expressing a fusion protein can prevent type 1 diabetes in nonobese diabetic mice, and significantly improve glucose tolerance and reduce insulin resistance. There are also data that suggest that engineered bacteria can help type 2 diabetes. Glucagon-like peptide 1 is an insulin-like hormone. It can promote pancreatic function, but its half-life is short. However, when symbiotic bacteria such as Bifidobacterium longum are designed to directly express and secrete bioactive glucagon-like peptide 1 as the penetrating fusion protein of glucagon-like peptide 1 into the colon, their potential therapeutic applications can be realized.
Connie: So in other words, using engineered bacteria to express fusion protein that promotes blood glucose levels can achieve the goal of treating diabetes. Some studies have reported that the number of multi-drug-resistant pathogens is increasing, while the choice of antibiotics is decreasing. So we really need to find alternative methods to fight deadly microorganisms. Is engineering probiotics a new method?
Dr. Hofstadter: Yes, this is a promising alternative method because of its several advantages, including stronger specificity, the controllable release of antimicrobial agents, and a narrow risk of antibiotic resistance.
Connie: How can we use engineered probiotics to solve the problem of multidrug resistance? And what probiotics are we looking at here?
Dr. Hofstadter: Genetically engineered Streptococcus lactis has been used as a target for multidrug-resistant enterococci. The main function of Lactobacillus is to identify multi-drug resistant Enterococcus faecalis. It is designed to produce bacteriocin under the control of the chlorine-induced promoter. This promoter is triggered in the gut to attack and sense Enterococcus faecium pheromones. Clostridium difficile is a pathogen causing antibiotic-associated diarrhea that is often seen in developed countries. Clostridium difficile infection can be treated by fecal microbiota transplantation. The virulence factor of engineered Lactobacillus expresses the non-toxic TcdA and TcdB fragments of Clostridium difficile and produces immunity to this pathogen in vivo.
Connie: Wasn’t there some news that engineering probiotics are also used to develop vaccines. Can you give us some examples?
Dr. Hofstadter: Sure. So mutant Bacillus subtilis spores were outlined in a study to express Helicobacter pylori urease B protein on the surface. It was observed that the recombinant spores produced a humoral reaction in mice, which significantly reduced the burden of Helicobacter pylori. Bacillus subtilis spores are also designed to express Mycobacterium tuberculosis antigen for the development of tuberculosis vaccine. In addition, the designed probiotics have been used in vaccines against HIV and other viruses. HIV gag antigen was expressed on the surface of a lactic acid bacterium. The antigen showed the ability to initiate an immune response, which established its potential use as a vaccine.
Connie: That’s interesting. The World Health Organization launched a project on the global cancer Observatory in 2012. The project estimates that the number of new cancer cases will increase by 14 million in the next 20 years, which is roughly a 70 percent increase. The use of bacteria to treat cancer was developed in the 19th century. Can you tell us more about it?
Dr. Hofstadter: Yeah. Over the years, microbiologists have proposed several diverse anaerobic bacteria. They preferentially proliferate in solid tumors and encourage the use of microorganisms as transmitters of anticancer agents. They have analyzed various anaerobes, including Clostridium, Bifidobacterium, Salmonelle, and Escherichia coli. Studies have shown that in tumor model mice, combining oral L-arginine with anti-PD-L1 immunotherapy can synergistically enhance the antitumor efficacy. This is due to the construction of engineered probiotics based on E. coli, which can specifically colonize tumors and continuously convert ammonia to L-arginine. Ammonia is one of the metabolic wastes accumulated in tumors. Intratumoral or systemic administration of the bacterium to mice can increase the concentration of L-arginine in the tumor, increase the number of tumors infiltrating T cells and promote the function of effector T cells. So we know that engineered bacteria can synergistically enhance the effect of anti-PD-L1 therapy and form long-term anti-tumor immunity.
Connie: Interesting. A recent study reported that engineering long bacterium expressing herpes simplex virus thymidine kinase successfully fused with ganciclovir. This has been proved to be a well-studied cancer gene therapy. Why is that?
Dr. Hofstadter: I think it’s because that the transgenic microorganisms used in combination with ganciclovir can shield the xenotransplantation of colon, liver, stomach, and breast tumors in mice. In addition to the ones we just mentioned, another facultative anaerobe commonly used in cancer treatment is Salmonella.
Connie: That’s it for our episode today. In fact, we have just finished our series on probiotics. I hope you enjoyed all of our episodes. Thanks, Dr. Hofstadter for being here with us throughout the series. And thanks everyone for listening. We will continue to come up with the most interesting topic in the biomedical industry. Please stay tuned to our next series of episodes. Have a wonderful evening.