Mechanism Of Proteolysis Inducing Factor In The Cancer Cachexia

Cancer cachexia is one of the most important characteristics of malignant tumor. The common manifestations of cancer cachexia includes anoxia, anemia, accelerated proteolysis, loss of lean body mass, continuing weight loss, fatigue and organ function failure. Among these, continuing muscle proteolysis and the break down of LBM are the most distinguished symptoms with subsequent poor life quality and finally lead to patients’ death.Recent study showed that a kind of sulfated glycoprotein can be detected in the urine of cancer cachexia patients, while it was negative in those without of with mild weight loss patients. In vitro experiments demonstrated that this protein can induce mice cultured muscle cell proteolysis and it was called proteolysis induce factor, PIF. PIF is directly connected with muscle proteolysis caused by cancer cachexia. So it is very important to better understand the working mechanism of PIF. Our research here is to unveil the signal transduce pathway of PIF so we can find targeted reagents precisely interfering with PIF pathway.In the first part of research, we examined urine PIF expression in clinical cancer cachexia patients and compared the urine PIF positive rate in cancer cachexia patients, patients with benign diseases, and cancer patients without cachexia as well. Through this examination, we had a better understanding of urine PIF expression pattern in different GI cancer patients. In the second part, we inspected the possible target of PIF in its signal tranduce pathway through intervention and rescue experiment in mice muscle culture.We found that in a portion of GI cancer cachexia patients, urine PIF expression was positive while it was negative in none cachexia patients. PIF can induce muscle proteolysis through PI3K-Akt pathway, and this can be rescued by5’-dFUrd and EPA. Our experiments demonstrated therapeutic potent towards cancer cachexia of these two reagent through there proteolysis attenuation ability.Part I Examination of urine PIF in cancer cachexia patients[Objective] To examine urine PIF expression pattern and to demonstrate specific expression of PIF in cancer cachexia patients.[Method] We collected urine samples from28GI cancer cachexia patients,13GI cancer patients without cachexia and12weight loss benign patients. PIF examination was conducted through western blot.[Results] The positive rate in28cancer cachexia patients were66.67%,57.14%, and37.50%in gastric cancer, colon cancer, and pancreatic cancer respectively with a total positive rate of53.57%. In other two control groups, no positive result was detected.[Conclusin] PIF is specifically expressed in cancer cachexia patients and it can be detected from patients’urine with no expression in none cachexia and benign cachexia patients.Part II in vitro muscle cell experiment of PIF mechanism[Objective] To inspect the mechanism of PIF during proteolysic process and its possible target in PI3K-Akt pathway. Find out the possible anti-cachexia mechanism of two clinical prescribed reagents through drug intervention strategy.[Method] First, we found out the optimum concentration and duration for Akt phosphorylation in mice cultured C2C12muscle cell. Then we used LY294002,5’-dFUrd and EPA to treat this cell at the time with concentration described above to examine the morphological and Akt phosphorylation change after drug intervention.[Results] PIF can successfully induce Akt phosphorylation. The peak of this phosphorylation can be detected after half an hour of the initiation of PIF at a concentration of4nM. The phosphorylation can be blocked by LY294002(a specific inhibitor of PI3K) and EPA but not5’-dFUrd.[Conclusion] PIF can activate Akt phosphorylation and induce downstream proteolysis. PI3K is in the upperstream of Akt to make this activation possible. EPA exhibits anti-proteolysis ability through the blockage of this pathway.5’-dFUrd can attenuate cancer cachexia without any effect on this pathway. There maybe other mechanism at large.