We investigated the genotype-dependent therapeutic potential of targeting the PI3K/Akt pathway

We investigated the genotype-dependent therapeutic potential of targeting the PI3K/Akt pathway for thyroid cancers. and OCUT1 cells that harbored genetic alterations in the PI3K/Akt pathway but had no effect on SW1736 and KAT18 cells that did not. Transfection with PIK3CA mutants greatly sensitized SW1736 cells to perifosine and temsirolimus. Growth of xenograft tumors derived from FTC133 cells but not SW1736 cells in nude mice was dramatically inhibited by perifosine. Thus this work for the first time demonstrates that genetic alterations in the PI3K/Akt pathway confer thyroid cancer cells addiction to this pathway and their sensitivity to inhibition by targeting Akt and mTOR. This genotype-based targeting of the PI3K/Akt pathway using Akt and mTOR inhibitors may offer an effective therapeutic strategy for thyroid cancer and warrants further studies. product PTEN which is a phosphatase that terminates the signaling of this pathway by dephosphorylating PI(3 4 5 (8). Driven by genetic alterations the PI3K/Akt pathway is frequently over-activated in human cancers including thyroid cancer (1 9 10 Follicular thyroid cell-derived thyroid cancer is the most common endocrine malignancy. This cancer is classified into differentiated papillary thyroid cancer (PTC) and follicular thyroid cancer (FTC) and the undifferentiated anaplastic thyroid cancer (ATC) (11). PTC and FTC may progress into poorly differentiated thyroid cancer (PDTC). Genetic alterations in the PI3K/Akt pathway are common in thyroid cancer including the amplification and mutations mutations mutations and amplifications of some key genes in this pathway (12-18). These genetic alterations are particularly common and important in aggressive thyroid cancers such as PDTC and ATC (13 14 17 18 which account for most of the incurable and fetal cases of EGT1442 thyroid cancer. Therefore the PI3K/Akt pathway is a potentially important and effective therapeutic target in thyroid caner. We propose that the activating genetic alterations in Rabbit Polyclonal to CD3EAP. the PI3K/Akt pathway may confer special sensitivity of thyroid cancer cells to inhibition by targeting the pathway which may form a basis for the development of novel genetic-based therapeutic strategies for this cancer. In the present study we tested this hypothesis using two clinically applicable inhibitors perifosine and temsirolimus as well as the shRNA approach in a large panel of thyroid cancer cell lines for which we characterized the genotypes of EGT1442 the PI3K/Akt pathway. Materials and Methods Thyroid cancer cell lines The thyroid tumor cell lines C643 Hth7 Hth74 and SW1736 had been originally from Dr. N.E. Heldin (College or university of Uppsala Uppsala Sweden); KAT18 from Dr. Kenneth B. Ain (College or university of Kentucky INFIRMARY Lexington KY); OCUT1 from Dr. Naoyoshi Onoda (Osaka Town College or university Graduate College of Medication Osaka Japan); BCPAP from Dr. Massimo Santoro (College or university of Federico II Naples Italy); K1 from Dr. David Wynford-Thomas (College or university of Wales University of Medication Cardiff UK); WRO-82-1 from Dr. G. J. F. Juillard (College or university of California-Los Angeles College of Medicine LA CA); and FTC133 from Dr. Georg Brabant (College or university of Manchester Manchester UK). The standard thyroid cell-derived cell range TAD2 was from Dr. Mario Vitale (Università Federico II Naples Italy). The TPC1 cell range was supplied by Dr. Alan P Dackiw (Johns Hopkins College or university EGT1442 Maryland). These tumor cells have already been lately characterized to become distinct thyroid tumor cell EGT1442 lines (19). These were all expanded at 37°C in RPMI 1640 moderate with 10% fetal bovine serum (FBS) aside from FTC133 that was cultured with DMEM/HAM’S F-12 moderate. For some tests cells had been treated with perifosine or temsirolimus using the indicated concentrations and period and the moderate and agents had been replenished every 24 h. Perifosine and temsirolimus had been from Cayman Chemical substance (Ann Arbor MI USA) dissolved in DMSO and ethanol respectively having a share concentration of 10 mM and stored at -20°C. Analysis of genetic alterations in the PI3K/Akt pathway in thyroid cancer cell lines We analyzed the major genetic EGT1442 alterations in the PI3K/Akt pathway in all the thyroid cancer cell lines in the present study. (exons 1 and 2) (exons 1 and 2) (exons 1 and 2) (exons 9 and 20) and (exons 5-7) were analyzed for mutations using our previously designed primers (14 18 For genomic DNA amplification of all the genes by PCR after 4 min initial.