Cloning of the Kynureninase gene into theAd۵ human Adenovirus vector

Background: In cancer cells, One of tryptophan cycle metaboliteswhich is called Kynurenine cause suppression of the immunesystem including apoptosis of effective immune cells, Consequently,one of the solutions is to disrupt the tryptophan cycleinside the tumor. the tumors produce a metabolite called kynurenineusing tryptophan, which has been observed that the amountof kynurenine inside the tumors increasing the tumor’s ability toescape from the immune system, as a result, one of the basic solutionsis to inhibit and eliminate Kynurenine with Kynureninase.Materials and Methods: In this project, the kynureninase genewas first synthesized, then it was cloned inside the PAd-Track-CMV shuttle vector the accuracy of the cloning was confirmedby confirmatory tests, then complete the Ad۵ human adenovirusvector and prepare it for packaging and virus production. Homologousrecombination between linearized PAd-Track-CMVshuttle vector and PAd-Easy-۱ backbone vector was performedby BJ۵۱۸۳ bacteria.Results: It has been observed that as the cancer progressesand the cancer enters more advanced stages, the expression ofkynurenine increases and instead the expression of kynureninasedecreases, and it is also observed that the expression ofkynureninase is correlated with the expression of other cancerbiomarkers. It was also observed that by increase the expressionof kynureninase in the culture medium, the rate of growthand division of cancer cells decreased drastically, and this issuehas provened by repeating the experiment in mice. It has alsobeen observed that preventing the production of kynurenine bytargeting its upstream pathways, such as the two enzymes IDOand TDO, does not have a good result because their expressionlevels may be different in different cancer cells, so the bestway is to directly target kynurenine with kynureninase. due tothe advantages of the Ad۵ adenovirus vector system, the Ad۵adenovirus vector method was used to produce this enzyme sothat we can have a high expression level of kynureninase in atargeted manner.

کلیدواژه ها:

Tryptophan cycle ، Kynurenine ، Kynureninase ، Ad۵human adenovirus vector