Team:Fudan/HP Supplement
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HP Supplement
Limits of Existing Treatments
Lactose intolerance is a common disease in China, prevalence of Primary Lactase intolerance has reached up to 95%-100%[1], but some adults with lactose intolerance don't care about it because they can just live a life without milk. When it comes to babies ,things are different.
This year, in January, when Yichen Gong (One of our team members) was doing voluntary work at Obstetrics and Gynecology Hospital of Fudan University, she found that lots of babies were suffering from lactose intolerance. Milk is very important to them, so they can't avoid drinking milk to treat this disease. At hospital, she realized that there are two main ways in treating lactose intolerance, one is to use lactase and another is to use probiotics, but according to the babies' parents, both ways are inconvenient for babies. Lactase might be inactivated by gastric acid, and the process of using lactase is complicated. Babies have to take lactase around 15 minutes before they drink milk, and the water temperature for brewing lactase has to be controlled.
Besides lactase, probiotics are not genetic modified so they are not able to compete with the local microflora, so babies have to take probiotic preparation over and over again.
After reading related papers, we found out that among infants less than six months old in China, 10.91% of the babies are suffering from primary lactose intolerance, and babies suffering from secondary lactose intolerance has reached up to 64.70%[2]. Diseases like Intestinal mucosal injury caused by diarrhea may lead to lactase intolerance. So lactose intolerance in babies is really a local problem in China. When babies are treating lactose intolerance, whatever using probiotics or lactase, they have to stop drinking breast milk, and stop drinking breast milk for a long time is bad for baby's immune system because they can get antibodies from their mom through drinking breast milk.
Because of the problems about lactase and probiotics, we want to create a medicine which can both survive in baby's gut and didn't have to be taken constantly.
[1]:Swagerty J D, Walling A D, Klein R M. Lactose intolerance.[J]. American Family Physician, 2002, 65(9):1845-50.
[2]: Yuan Yun. Clinical observation of lactase in the treatment of lactose intolerance in children[J]. Chinese Community Doctors, 2017, 33(22); Discussion, Clinical Forum.
Survey
Before we asked the subjects whether they would accept the treatment of putting Nissle into babies' gut, we asked them which existing method of treating lactose intolerance had they known. As is showing below, 31.34% of subjects have heard about taking Shuhua Milk and 19.40% of subjects have heard about taking lactase before drinking milk. It means these two ways of treating lactose intolerance are quite common.
Taking ShuHua Milk is not good for babies, because the galactose which is the decomposition of lactose is used form cerebroside and mucopolysaccharide, so taking ShuHua Milk for a long time is bad for babies' nervous system. ShuHua Milk is better use in adults than in babies. Except taking ShuHua Milk, taking lactase before drinking milk also has its shortcomings. Lactase might be inactivated by gastric acid, and the process of using lactase is complicated. Babies have to take lactase around 15 minutes before they drink milk, and the water temperature for brewing lactase has to be controlled. The chart below is why subjects feel it inconvinent to use lactase.
Since the existing treatment is inconvenient, we asked whether the subjects would accept our treatment: putting probiotics into babies' gut. 88% of the subjects indicated that they can accept employing transgenic probiotics to treat lactose intolerance, but 55.22% of them are not sure enough, they still have some concerns.
As to their concerns, 89.55% subjects shows that they have concerns about safety, and 68.66% shows that they concerned about the therapeutic effect of the method. Since most people really care about safety problems, we found one of the members of China biosafety Committee, Baorong Lu, for advice.
Safety
Few of Chinese people have tried genetic modified products, since they are genetic modified, people who are not familiar with them quite concerns about the safety problems. Some thought GMOs might cause unpredictable damages, and some thought GMOs might be bad for human's health. According to our survey, 89.55% of babies' parents have these kind of concerns.
Their concerns are reasonable, and we want to decrease these concerns by focus on ensuring our product's safety. We found one of the members of China Biosafety Committee, Baorong Lu, for advice. China Biosafety Committee is in charge of examining and evaluating risk of the biological factors and research protocols in China. They determined whether a genetic modified product can be released into market.
According to Mr.Lu, China haven't passed any kind of genetic modified drugs (neither in American). It's mainly because genetic modified drugs are a leading edge technique, and it have to take years before a drug be approved, so it doesn't mean genetic modified drugs is harder than other drugs to be approved. On the opposite, unlike genetic modified food, genetic modified drugs for specific diseases are not replaceable, so its easier to be approved.
The e.coli we used, Nissle 1917's biosafety level is Ⅰ, which means the safest. Since we decided to transfer acid-resistance and antibacterial peptide into Nissle, we asked whether this kind of treatment will increase its security risk. Mr.Lu answered that as long as we only increase its survivability in the pressure of human's gut, not in the environment, its acceptable.
Besides, to ensure Nissle won't grow in the environment when it has leak into the environment through excreta, we collaborated with team UCAS. Their project focused on a killer system which can kill e.coli after it leaves human body. Click to see our collaboration.
Adhesion VS Competition
According to the literature we have read and the personal experience of our lactose intolerant classmates, probiotics (such as bifidobacterium) have very limited clinical effects in treating lactose intolerance.
Unable to find an explanation through the literature, we decided to contact Ms. Lihui Feng, an expert specializing in the development mechanism of intestinal flora and its relationship with infant health and disease. We asked her "why oral bifidobacteria, as a probiotic that colonized the gut, is not clinically effective in treating lactose intolerance".
In her experience, the colonization of foreign species depends on survival in competition with the original flora. 1. First, the majority of oral bifidobacteria will die due to gastric acid; 2. second, the original flora in the intestinal tract occupies most of the space, so it is difficult for oral flora to colonize; 3. finally, even if it is colonized and the original flora in the intestinal tract, it will be eliminated, if it has no competitive advantage. Aiming at these problems, we designed three modules of "acid resistance", and "antimicrobial peptide".
In addition, we have the honor to interview professor Yufeng Zhou, who is an experienced clinician. According to his experience, common probiotics have been abused, and doctors will prescribe probiotics whenever there is intestinal dysfunction, and the real effect is no different from placebo.(screenshots of the New England article can be found here.) but because of the probiotics' media hype, the family's acceptance of probiotics was high, which inspired us to use Nissle instead of other e. coli as the modified bacteria.
Market Preparation
The current stage of our project is stopped at laboratory process. If we want to marketize it, we need to ask for professional advice, so we found Hong Lv, a professor who was expert in microbial product marketing. Hong Lv was researching about genetically engineered bacteria producing high-yield yeast for feed enzymes.
After we described our project and our marketization vision to her, she gave us some advices. In the current stage we are using plasmids to perform particular functions, but if we want to make it industrialized, we need to integrate those genes into the genome, otherwise the plasmid might lose or transfer into other bacteria.
Because of that we want to control Nissle into a certain amount, our circuit contains quorum sensing system, but actual production needs a large flora size. Quorum sensing will control Nissle's growth before it reaches the certain size. We can add an inhibitor before the quorum sensing gene. The inhibitor is induced by some chemicals which doesn't exist in human body, such as arabinose. If the inducer is added in actual production, the inhibitor can inhibit quorum sensing to make Nissle reach a certain amount.
Though we have ensured our project's safety, since we are producing a drug, we need further safety monitoring experiments. We need to test whether the secretions of the genetic modified Nissle is toxin to human body, also we have to do drug toxicology and kinetics experiments.
Our target group is babies, so we want to make it freeze-dried to let babies swallow. To expand target groups, we can let teenagers who are quickly growing up and the aged who are more likely to fall osteoporosis. They need to drink more milk than other people, but lactose intolerence limits them. To let them use our product, we need to make Nissle more tasty. We can wrap nissle with gel sugar to make it chewy and sweet.
After the interview with Hong Lv, we learned more about marketizing and were more prepared to industrialize our product.