Safety Answers to Questions

1. Would any of your project ideas raise safety issues in terms of : • researcher safety, • public safety, or • environmental safety?

• Actually, it may raise partially safety issues since our project, Wound Dressing, actually raise safety issues about cancer and microbial infection. However, when we deeply search about the cancer and its relation with our project, we find out that cancer doesn’t depend on EGF amount. It depends on the EGF receptor amount . Since we are just increasing the EGF amount, as our most society knows the opposite, there is no cancer risk. In addition to this, for microbial infection risk, we have taken many precautions. Our embedded switch mechanism, granulysin protein and the polyurethane layers provide enough safety and these precautions are approved by authorities of university. The other two project ideas, bacteriaflow and bactoplus also don’t raise any safety issue since they are both require working only with bacteria.

2. Is there a local biosafety group, committee, or review board at your institution?

• Actually, since the meetings of our biosafety groups do not meet in very short term periods, our project ideas are not discussed at a meeting. However, we have planned meetings with all the academic personal who may be responsible from safety in his/her field. We have arranged meetings with most of these personnel and they applied that our ideas and works apply the safety rules.

3. What does your local biosafety group think about your project?

• Every single member of biosafety member has approached suspicious to this issue;since, we interfere with the actual healing mechanism. However, when we explain the procedures and the precautions, they apply what we have done so far.

4.Do any of the new BioBrick parts that you made this year raise any safety issues? If yes, did you document these issues in the Registry?

• The BBa_K258010 of our biobrick part has raised suspicion because it is known to induce cancer in most society. However, when we talked about this part with academic staff who is making research about cancer, they have guaranteed that in most cancer cases, the problem is about the receptors; not the EGF protein . Therefore, because we mention in this page, we do not need to mention about this issue about the part’s page since it may be counted as unnecessary information. The other parts of us do not raise any safety issue.

OUR SAFETY PRECAUTIONS

1. The on/off switch mechanism of our bacteria(via Quaroum Sensing): Since we do not want to produce excess amount of proteins and these proteins being accumulated in the second layer (thin polyurethane, we added a switch mechanism to our product. When not used our bacteria will not synthesize EGF proteins. The mechanism is opened with the activation of EGF synthesizing E. Coli with oxygen presence and lysis of both type of bacteria with the quorum sensing mechanism of KGF synthesizing bacteria.

2. Lyophilized bacteria: Owing to the fact that metabolism of bacteria needs to be stopped, we lyophilized our bacteria. Therefore, when not used, our bacteria will not release any protein to outside until it is used.

3. Granulysin embedded gelatin sponge: The granulysin protein is an antimicrobial protein affecting both gram positive and gram negative bacteria. Therefore, by using this protein in the last layer, we provided both the invasion of our bacteria to the wound and sterilize the wounded area.

4. Thin polyurethane layer: This layer will function as a filter which inhibits the transport of bacteria to gelatin layer. By this layer, invasion of bacteria to the wound will also be prevented.

5. Thick Polyurethane layer: This layer will only allow the small molecules such as gases to enter the wound dressing. Since we don’t want any contamination of the the area which bacteria have been cultivated, we added this layer. Moreover, this layer has many advantegous properties. One of them is its protective character. This layer will protect the wounded area from shocks and other interference. The other property is elastic properties. Since wounds have flexible environment, if something which is not flexible is placed, they will not stay at the place which it was placed. Therefore, its plasticity help us to establish wound dressing on wounds.

6. Parafilm covering: For our mechanism to function properly, oxygen should not enter the wound dressing until utilization. This layer will help us to prevent gas entrance to our wound dressing.

Biosafety of Used Proteins-EGF and KGF- in Wound Dressing

There are some medical concerning about the utilization of wound dressing. Actually not all the people is capable of using the Wound Dressing because of the risks. You can find out the risks of our wound dressing and possible solutions to those problems are as follows.

1. Cancer Risk: When it comes to fast proliferation, mostly cancer comes to mind. However, cancer is not that risky for our mechanism. Since cancer occurs due to enormous expression of EGF receptors, our addition of EGF will not induce cancer because normal cells only have limited amount of EGF receptor. Therefore, increasing HEGF amount does not cause cancer. However, if a patient has cancer risk, s/he shouldn’t use wound dressing since tumor cells have many amount of EGF receptors on their cell membrane. So the people who sholdn’t use wound dressing are:

 Having history about having the skin cancer : Since after successful cancer therapies, some tumors may remain. Adding EGF to these patients will induce cancer again.

 Having skin cancer in family pedigree: Since cancer is accounted to be genetic disease, familly pedigree sustain great importance. If people have lots of EGF receptors on their skin cells congenitally, the risk of having skin cancer is also increased.

 Sunbathing a lot: Skin cancer is caused by overexposure to the sun’s harmful UV rays. A suntan isn’t healthy; in fact it’s a sign of skin damage. The results of sunbathing with combined with our wound dressing may induce skin cancer.

 Heavy Metal exposure: Heavy metal exposure is also risk for having cancer. Therefore, people who have connected with heavy metal, should use wound dressing only if the wounds become unbearable for patient. The metals wich have to be avoided are as follows;

2. Microbial Infection: Since our wound dressing depends on E.coli microbial cell, any invasion from our wound dressing to wound may cause microbial infection. However, we lowered the risk of infection which is based on our E. Coli bacteria cells by embedding the antimicrobial granulysin protein into the gelatin sponge. The gelatin sponge will prevent any invasion coming from the upper layers. Moreover, the gelatin sponge will also kill gram positive and gram negative bacteria on wound by forming pores on bacterial cell membrane. Fastening the inflammatroy phase is other effect of the granulysin protein.

3. Allergic Reactions and Mad-Cow Disease: There is some concern about the safety of gelatin because it comes from animal sources. Some people are worried that unsafe manufacturing practices might lead to contamination of gelatin products with diseased animal tissues including those that might transmit mad cow disease (bovine spongiform encephalopathy). Although this risk seems to be low, many experts advise against using animal-derived supplements like gelatin. Moreover, owing to the fact that our gelatin is biomedical grade, we avoid that risk. However, before using the Wound Dressing, people shold know whether they are allergic to gelatin or not.

4. Metabolic disorders: Since we have used gelatin, a protease inhibitor, we could come up with some metabolic disorders because of delayed activity proteases. Since we do not pass the mammalian experiments, we do not know whether its metabolic effects are. However, our literature search shows that in short –term, the delayed activity of proteases is not harmful.