A potentiometric screen-printed biosensor for analysis of Kynurenineproduction to overcome immune barriers to different cancer cell progression

Analysis of kynurenine (Kyn) is of great importance in cancer biology. Degradation of tryptophan(Trp) to Kyn by indoleamine-2,3-dioxygenases 1 and 2 (IDO1/2) and tryptophan-2, 3-dioxygenase(TDO) in tumors and tumor-draining lymph nodes inhibits antitumor immune responses and isassociated with a poor prognosis in various malignancies. Tumors increase their consumption of theamino acid tryptophan to evade immune control. Many cancers upregulate a liver enzyme,tryptophan dioxygenase, to drive tryptophan consumption. What’s more, the authors find that theprimary product of this process, kynurenine, is an endogenous ligand for the aryl hydrocarbonreceptor, which mediates invasive tumor growth. This second finding links the fields of toxicology,immunology and cancer biology in new ways, and may help to explain how elevated tryptophanconsumption helps tumors to overcome immune barriers to cancer progression. In this work, for thefirst time, we report on the development of biosensor based on constant current potentiometricstripping analysis (cc-PSA) for determination of Kyn in cancer cell culture media samples [1, 2].MethodsTo prepare the potentiometric biosensor, carboxylated multiwall carbon nanotubes (MWCNTs)were uniformly deposited on the gold screen printed electrode surface (MWCNT-AuSPE) and LKynureninemonoclonal antibody (Ab) were uniformly deposited MWCNT-AuSPE (Ab-MWCNTAuSPE)by EDC and NHS agents. The engineered immunosensor was characterised by cyclicvoltammetry (CV), electrochemical impedance spectroscopy (EIS). Moreover, the parameters thataffect the Kyn determination in the biological samples and cell culture were fully investigated.Results and DiscussionIn the optimal condition limit of detection (LOD) was found to be 0.5 nM and two linear detectionrange (LDR) were found to be 0.001 to 1 μM and 1 to 100 μM. Based on these findings, the Kynbiosensor provides a sensitive and reproducible, yet cost-effective, tool with lower LOD forspecific detection of Kyn, even in the presence of different interfering substances, in biological41samples, thus suggesting the utility of this biosensor for the fast and easy determination of Kyn inclinic. Here, we determination of Kyn production in various human tumors cell culture media(HepG2 (hepatocarcinoma), 1321NI (Astrocytoma), Calu-6 (lung carcinoma), NCI-H1299 (lungcarcinoma), HT29 (colorectal carcinoma)). Our results describe a mechanism of tumoral immuneresistance based on Trp consumption and Kyn production. We examined the growth of humantumor cell lines which differ in their metastatic ability, in patients by determination of Kynproduction.ConclusionsHT29 cells with high Kyn production showed a greater ability to migrate to sites which favourtumor growth and to replicate to form micrometastases. These studies point to post-extravasationevents (migration and growth) as being critical in metastasis formation.

کلیدواژه ها:

Immunosensor ، Kynurenine ، Tryptophan ، Antibody ، Constant current potentiometricstripping analysis ، Screen-printed electrode