The Antitumor Activity Of A Novel Dual PI3Kα/mTOR Inhibitor PI-103 In Cancer Therapy

Lung cancer is the leading cause of morbility and mortality of malignant tumors which seriously threat our health and lives. Colon cancer and primary hepatic carcinoma are also two common malignant tumors, which are characterized with high morbility and mortality. The major therapeutics of malignant tumors is surgery, traditional chemotherapy and radiotherapy. Traditional tumor therapies show high side-effects due to theirs non-specificity, which not only kill tumor cells but do harm to normal tissue. With the development of tumor molecular biology, tumor targeted therapy become increasingly important for cancer therapy, targeted therapy and compound therapy will provide several promising strategies for cancer therapy. In the recent years, a series of studies have shown that PI3K/Akt/mTOR and Ras/Erk signaling pathways play a key role in growth, proliferation, invasion and metastasis of cancer cells. Blockade of the two pathways will induce growth inhibition and apoptosis. The novel PI3Ka/mTOR dual inhibitor was synthesized by scientists in University of California in San Francisco, U0126 is the MEK kinase inhibitor. These two inhibitors are being used in preclinical experiments. PI-103 was purchased from Merck company, and U0126 from Sigma company. In the present study, the antitumor activity of PI-103 alone or in combination with U0126, paclitaxel and DHA was observed in non-small lung cancer cell line A549 and H460, colon caner cell line CaCO2 and hepatic carcinoma cell line HepG2. The results were presented as follows:1. PI-103, the novel PI3Kα/mTOR dual inhibitor showed high antitumor effects in A549 and H460 cells. H460 cells with activating mutations of PIK3CA, the gene encoding the p110αcatalytic subunit of PI3K, were more sensitive to low-dose PI-103. The CI value of PI-103 in combination with U0126 was less than one, suggesting that the combination of PI-103 and U0126 acted synergistically to induce growth inhibition. The synergistic antitumor effects were derived from concomitant inhibition of the PI3K/Akt/mTOR and Ras/Erk signaling pathways. Additionally, Akt phosphorylation induced by mTOR inhibition with rapamycin was inhibited by PI-103. We also found that mTOR functions downstream of Akt and Erk. The phosphorylation of ribosomal protein s6 and the eukaryotic translation initiation factor 4B (eIF4B) was concomitantly inhibited by PI-103 and U0126, which may cooperatively inhibited translation initiation complex assembly and translation initiation. This data explained why blockade of the PI3K/Akt/mTOR and Ras/Erk shows synergistic antitumor effects.2. G0/G1 arrest was induced by PI-103 in A549 and H460 cells, and H460 cells were more sensitive to PI-103. PI-103 synergized with U0126 to induce G0/G1 arrest in A549 and H460 cells. Our results showed that PI-103 or U0126 exerts its effects on cell cycle progression via the upregulation of p21 and p27, and the simultaneous downregulation of cyclin D1 and cyclin E1. The synergistic antiproliferative effects of PI-103 in combination with U0126 were derived from markedly up-regulating p21 and p27 protein levels, and significantly inhibiting the expression of cyclin D1 and cyclin E1.3. PI-103 not only potentiated paclitaxel-induced growth inhibition but also enhanced paclitaxel-induced apoptosis in A549 and H460 cells. Apoptosis induced by paclitaxel and PI-103 was related to alterations of apoptosis proteins. Compared with treatment of paclitaxel alone, the combination of PI-103 and paclitaxel significantly up-regulated tumor suppressor P53 and pro-apoptosis protein Bad, and simultaneously down-regulated anti-apoptosis protein Bcl-2. Alterations of these apoptosis-related proteins finally triggered mitochondrial-related apoptosis pathway and led to cell apoptosis. Our study showed that compound therapy is the most effective tumor therapeutics, and provide a potential therapy strategy for gefitinib-resistant non-small cell lung cancer.4. High antitumor activity of PI-103 was also showed in CaCO2 and HepG2 cells. PI-103 in combination with U0126 also showed synergistic antiproliferative effects with induction of G0/G1 arrest in CaCO2 and HepG2 cells. We also observed that Paclitaxel-induced apoptosis was enhanced by PI-103 in HepG2 cells. Our study suggested that PI-103 alone or in combination with other kinase inhibitors, or chemotherapy agents is a promising therapeutics for treating colon cancer and hepatic carcinoma.5. The antitumor effects of Docosahexaenic Acid (DHA) were associated with lipid peroxidation. Lipid peroxidation may decrease DHA levels in cell membrane and lead to reduced phosphatidylserine in intracellular membrane, which blocks the PI3K/Akt signaling pathway and leads to cell growth inhibition. Antitumor effects of DHA were significantly antagonisted by serum. High-concentration DHA promoted cell growth in A549 cells in culture with 10% FBS. It is possible that growth factors or BSA impedes the lipid peroxidation of DHA enriched in plasma membrane and promotes the signaling of the PI3K/Akt pathway, which is attenuated by PI-103. Our study further elucidated the antitumor mechanism of DHA and provided meaningful directions for development of DHA as an adjuvant antitumor agent.