Difference between revisions of "Part:BBa K1722001"

Revision as of 03:33, 4 September 2015

shTERT is a cancer cell specific promoter with high efficiency.

Telomerase reverse transcriptase(abbreviated to TERT, or hTERT in humans) is a catalytic subunit of the enzyme telomerase, which, together with the telomerase RNA component (TERC), comprises the most important unit of the telomerase complex.

The telomerase is a ribonucleoprotein enzyme to which multiple functions have been attributed, the most important of these is the maintenance of the telomere which is related with cellular immortalization and cancer. 85% of human tumors have telomerase activity, that in normal cells goes undetected. These characteristics make the telomerase an attractive target for chemotherapy. The TERT promoter can specifically identify TERT proteins which are largely produced in human tumor cells, thus being expected to be tumor-specific in human body. TERT promoter mutations were highly frequent in glioblastoma (83.9%), urothelial carcinoma (64.5%), oligodendroglioma (70.0%), medulloblastoma (33.3%) and hepatocellular carcinoma (31.4%). These mutations differentially enhanced the transcriptional activity of the TERT core promoter.TERT promoter mutations are frequent in multiple tumour types and have similar distributions in Chinese cancer patients.

More importantly , we mutated the normal hTERT promoter into a super hTERT promoter(shTERT) with higher promote activity so as to make our system work more efficiently. Four base pairs of the TERT promoter sequence were mutated to form shTERT. Eventually, the shTERT can be activated with the identification of specific RNA polymerase. In our system, we use shTERT to initiate the expression of downstream DNA sequence. Together with bladder-specific promoter hUPII we can achieve the targeted recognization of bladder cancer cells. By using therapeutic genes(such as p21 and Bax) as effectors, targeted gene therapy for bladder cancer can be carried out.  In our experiment, we constructed three plasmids system and two plasmids system before and after to verify the function using high-efficency shTERT.It can be activated inside cancer cells. Similar to our system, alike synthesizing gene circuits are also expected to be one of the promising approaches to the treatment of other cancer.

Design Notes

We designed the following primers and amplified hTERT promoter from the vector psi-Check2:Up: CCGGAATTCGGCACCTCCCTCGGGTTAG Down: TGCACTGCAGACTAGTCGCGTGGGTGGCCG. By incorporating these primers into hTERT promoter, the promoter is flanked by the iGEM prefix and suffix after amplification.

Source

The telomerase reverse transcriptase promoter can be found in human cancer cells. In our experiment, we got the part from Shenzhen Second People's Hospital. Additionally, the verification of our system's function was also carried out in Shenzhen Second People's Hospital.

References

[1]Castillo Ureta H, Barrera Saldaña HA, Martínez Rodríguez HG. Telomerase: an enzyme with multiple applications in cancer research. Rev. Invest. Clin. 54 (4): 342–8. 

[2]Huang DS, Wang ZH, He XJ, et al. Recurrent TERT promoter mutations identified in a large-scale study of multiple tumour types are associated with increased TERT expression and telomerase activation, European Journal of Cancer, 51: 969-976.