- We have chosen hTert-p53 as our favorite part because it can help solve the off-target effect of traditional photothermal therapy. We tend to describe the part as a gunsight. HTert is human telomerase reverse transcriptase, which is constitutively activated only in immortalized cells including tumor cells.
P35 is a tumor suppressor which can increase the thermosensitivity of cancer cells. Theoretically, combined with hTert promoter, p53 will be overexpressed in tumor cells specifically, so the thermosensitivity of tumor cells would be increased rather than of normal cells. After equipping our gun with such a gunsight, the accuracy rate will increase. Namely, majority of our enemies, the tumor cells will be ablated while the innocent citizens, the normal cells can stay safe and sound with rational gold nanorods concentration and light intensity. In addition, the part can become a gun itself because it will induce apoptosis of cancer cells specifically. Cooperated with this part, photothermal therapy will achieve an astonishing result which demonstrates that one plus one is greater than two.
|BBa_K1922004||regulatory||(hsp70 promoter) pro-moter||477|
|BBa_K1922008||reporter||Green fluorescent protein||717|
|BBa_K1922010||reporter||Green fluorescent protein||993|
|BBa_K1922003||coding||coding for tumor supressor, wild type p53||993|
- Approximately 80% of cancers are immortalized by constitutive activation of telomerase to maintain telomeres throughout rapid cellular proliferation. HTert promoter is the promoter we cloned from human cell, which can drive the transcription of telomerase in human. We hoped that it can promote the transcription of the certain gene following it in tumor cells specifically, instead of in normal cells.
- The cells of all tissues react to various stresses through the quick synthesization of HSPs. As the most important HSPs family, HSP70 has the highest temperature sensitivity and conservation. Having synthesized human hsp70 promoter, we hope it can drive the transcription of selected gene after heat shock at certain temperature. From the literature we finally set the preform condition as 42℃ for 1h
- The cells of all tissues react to various stresses through the quick synthesization of HSPs.As the most important HSPs family. HSP70 has the highest temperature sensitivity and conservation. Having synthesized human hsp70 promoter, we hope it can drive the transcription of selected gene after heat shock at certain tem-perature. From the literature we finally set the preform condition as 42℃ for 1h
- To test if it works, we also constructed a plasmid linking a GFP as a reporter gene linked with it. So when we transfected it into cancer cells like hcc 1937 or hela, and telomerase negative cells, we could observe GFP fluorescent in cancer cells instead of normal cells.
- We hope we can optimize the conventional theotherapy into a responsive system, which means if the laser is working, we want to know it. It will be more convenient and efficient for researchers to set appropriate condition of the equipment, such as the power of laser. So a gene of luciferase was linked after p53 as a reporter.
- With the combination of hTert promoter and p53, if we deliver the plasmid into mouse by virus packaging, p53 will be overexpressed in tumor cells specifically, so the thermosensitivity of tumor cells would be increased rather than of normal cells. When tumor is treated by laser with certain-length wave, with the heat induced by our golden nano rods(GNRs), the apoptosis of tumor cells whose thermosensitivity has been increased would start more efficiently. Thus, because of the tumor-targeted hTert promoter and the tumor suppressor following it, we improved the targeting of conventional thermotherapy.
- As we mentioned above, heat shock protein 70(hsp
70) will be synthesized rapidly after heat shock. While convention-al thermotherapy has already got great efficiency in clinical application, we wondered if we can make the whole system responsive, which means there can be a reporter system telling people that the heat treatment of laser is working. Hsp70 promoter can drive the transcription of p53 gene after heat shock which is done by laser and GNRs in thermotherapy.
- GFP gene was linked after hTert promoter,after transfecting it into different cell lines, we can examine if the promoter can drive the transcription of the gene after it in hTert positive cells but not negative ones.
- After inserting the hsp70-p53 fraction into pGL3 vector which con-tains a luciferase gene, we could observe if the tumor cells had been heat treated in in vivo bioluminescence imaging machine with the injection of luciferase substance. If the result can be re-flected from mouse direcly, it will be easier to set experimental condition and save the use of model animals. At the same time, p53 can work to increase the thermosesitivity of tumor cells. We hope hsp70-p53-luciferase can be a novel reporter system to make conventional thermotherapy a responsive method for us to treat cancer in a more efficient way.