Difference between revisions of "Team:MIT"

Genetic Circuit to Sense Endometriosis

This year, the MIT iGEM team is creating a circuit constructed of five plasmids that will be able to sense the presence of endometriosis in the endometrial biopsies of affected women. Endometriosis is a disease caused by cells from the endometrial lining of the uterus growing elsewhere in the body, usually on the ovaries, which causes significnt and chronic pain as well as infertility. Our tool could expedite the diagnosis process, something sorely needed when currently there is an average 7-year wait between the onset of disease symptoms and an accurate diagnosis. The circuit will function through a double-check, two "latch" system, checking the cells' miRNA profiles in both the estrogen-high proliferative phase and the estrogen-low secretory phase of the menstrual cycle. The circuit will also sense for progesterone-resistance which is characteristic of the disease. While the team is still unsure if the circuit will function as a diagnostic tool, a treatment, or some combination of the two, the design of the circuit upstream of the output has been completed.

Current Design

The circuit will first rely on endogenous estrogen receptors (ERa) to report the presence of estrogen to the circuit through an ERa binding region called ERE in a synthetic promoter.

In the estrogen high phase-reactive half of the circuit, this interaction, combined with the correct miRNA conditions, will activate the expression of a recombinase that will flip the first latch before the output. The wrong miRNA conditions will result in the degradation of the recombinase genes's mRNA, and the lack of a small molecule drug will result in the degredation of the protein if by chance the recombinase mRNA reaches a ribosome before it is degraded by the miRNA complex. This is because the recombinase will be synthetically combined with a degradation tag, DDd, that is stabilized by that small molecule drug.

In the estrogen low phase-reactive half of the circuit, the binding of ERa to the synthetic promoter region activates expression of a repressor protein that prevents the expression of the second recombinase. This way, when there is very little estrogen, the repressor is not expressed in significant amounts and the constitutive promoter before the recombinase can activate its expression. As before, the miRNA profile of the cell in the estrogen low phase is used to degrade mRNA of the repressor at that point, and degrades the mRNA of the recombinase during the estrogen high phase. The second recombinase is also controlled by a degradation tag and small molecule drug. Through this system of repressors, the team hopes that the second recombinase will only be expressed in the estrogen low phase.

The first possible output would be the apoptosis of cells identified by the circuit as endometriotic. This is a risky option as there is a chance for leaky expression of the recombinases despite the number of repressors. The second possible output would be a fluorescence gene, so the disease cells can be more easily identified during surgery, allowing for better complete removal of the lesions. This is a less ideal option because through the circuit, the team hopes to decrease the need for women with endometriosis to undergo surgery, and while this output may decrease the number of surgeries required, it would not provide an alternative to surgery. The third possible output would be the production of a biomarker that can be sensed through blood or urine testing as a diagnostic, an alternative to laparoscopic surgery that is the sole diagnostic tool at this time. This option would remove the need for one surgery, but would not be able to actually provide a treatment. It's also possible to do a combination of these outputs.

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