Phosphatidylinositol 3-kinase/Akt/mammalian Target Of Rapamycin Research Articles

Thymoquinone Inhibits JAK/STAT and PI3K/Akt/ mTOR Signaling Pathways in MV4-11 and K562 Myeloid Leukemia Cells.

Constitutive activation of Janus tyrosine kinase-signal transducer and activator of transcription (JAK/STAT) and Phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathways plays a crucial role in the development of acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). Thymoquinone (TQ), one of the main constituents of Nigella sativa, has shown anti-cancer activities in several cancers. However, the inhibitory effect mechanism of TQ on leukemia has not been fully understood. Therefore, this study aimed to investigate the effect of TQ on JAK/STAT and PI3K/Akt/mTOR pathways in MV4-11 AML cells and K562 CML cells. FLT3-ITD positive MV4-11 cells and BCR-ABL positive K562 cells were treated with TQ. Cytotoxicity assay was assessed using WSTs-8 kit. The expression of the target genes was evaluated using RT-qPCR. The phosphorylation status and the levels of proteins involved in JAK/STAT and PI3K/Akt/mTOR pathways were investigated using Jess western analysis. TQ induced a dose and time dependent inhibition of K562 cells proliferation. TQ significantly downregulated PI3K, Akt, and mTOR and upregulated PTEN expression with a significant inhibition of JAK/STAT and PI3K/Akt/mTOR signaling. In conclusion, TQ reduces the expression of PI3K, Akt, and mTOR genes and enhances the expression of PTEN gene at the mRNA and protein levels. TQ also inhibits JAK/STAT and PI3K/Akt/mTOR pathways, and consequently inhibits proliferation of myeloid leukemia cells, suggesting that TQ has potential anti-leukemic effects on both AML and CML cells.

Fucoxanthin prevents cell growth and induces apoptosis in endometrial cancer HEC-1A cells by the inhibition of the PI3K/Akt/mTOR pathway.

Endometrial cancer is the major type of gynecological cancer and ranks as the sixth most common cancer in women. Endometrial cancer usually is diagnosed in an advanced stage, complicating the treatments in many cases. The present research was focused on unveiling the in vitro anticancer role of fucoxanthin against the endometrial cancer HEC-1A cells by inhibiting the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling axis. The cytotoxicity of fucoxanthin against the endometrial cancer HEC-1A cells was studied using the MTT test. The level of reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) status, and apoptotic cell death in the 7.5 and 10 µM administered HEC-1A cells were assayed using fluorescent staining techniques. The messenger RNA expression was analyzed using RT-PCR for PI3K/Akt/mTOR signaling molecules, proapoptotic (Bax and caspase-3) antiapoptotic (cyclin D1 and Bcl-2) genes, and inflammatory markers like tumour necrosis factor α (TNFα), nuclear factor kappa B (NF-κB), Cox-2, and interleukin (IL)-6. The cell viability assay proved that fucoxanthin effectively prevented HEC-1A cell viability, where the IC50 was 7.5 µM. Fucoxanthin at 7.5 and 10 µM remarkably improved ROS production and apoptosis and decreased the MMP in HEC-1A cells. The fucoxanthin effectively inhibited the PI3K/Akt/mTOR cascade along with the expression of TNF-α, NF-κB, Cox-2, and IL-6 and antiapoptotic genes cyclin D1 and Bcl-2 in the HEC-1A cells. Fucoxanthin treatment also enhanced the Bax and caspase-3 expressions in the HEC-1A cells. Our results from this work unveiled that fucoxanthin triggered growth inhibition and apoptosis in endometrial cancer HEC-1A cells. Besides, fucoxanthin inhibited the PI3K/Akt/mTOR cascade and improved apoptotic marker expressions in the HEC-1A cells.

MicroRNA-31 inhibits traumatic brain injury-triggered neuronal cell apoptosis by regulating hypoxia-inducible factor-1A/vascular endothelial growth factor A axis.

MicroRNAs are dysregulated in traumatic brain injury and are involved in neuronal cell behaviors. Previous studies identified miR-31 as a spinal cord injury-related microRNA, while its role in traumatic brain injury remains indistinct. Herein, we explored the participation of miR-31 in traumatic brain injury. Traumatic brain injury model was established after traumatic neuron injury. Neurocytes were transfected with miR-31 mimic or inhibitor. Cell counting kit-8, lactate dehydrogenase assay, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling, and western blot were applied to examine cell viability, lactate dehydrogenase releasing, apoptosis, and apoptosis-related protein. The binding between miR-31 and hypoxia-inducible factor-1A was verified by luciferase assay. Quantitative reverse transcription-PCR was used to detect the regulation of traumatic neuron injury or hypoxia-inducible factor-1A overexpression on vascular endothelial growth factor A level. The effects of hypoxia-inducible factor-1A or vascular endothelial growth factor A on neuronal cell injury were examined. Additionally, phosphatidylinositol 3kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway was also examined using western blot. Downregulation of miR-31 promoted traumatic neuron injury-induced neuronal cell injury, and its overexpression did the opposite. Hypoxia-inducible factor-1A acted as a downstream mRNA of miR-31 and its downregulation was involved in miR-31-regulated neuronal cell injury. Vascular endothelial growth factor A level was elevated by traumatic neuron injury or hypoxia-inducible factor-1A overexpression. Hypoxia-inducible factor-1A enhanced neuronal cell injury via promoting vascular endothelial growth factor A expression. Furthermore, miR-31/hypoxia-inducible factor-1A/vascular endothelial growth factor A regulated PI3K/AKT/mTOR pathway in neuronal cells. Our study demonstrated miR-31 inhibited neuronal cell apoptosis via regulating hypoxia-inducible factor-1A/vascular endothelial growth factor A axis.

Persistent activation of the mammalian target of rapamycin signalling pathway in cutaneous squamous cell carcinomas in cats.

Cutaneous squamous cell carcinoma (CSCC) represents the most common malignant tumour of the feline skin. Emerging evidence suggests that the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway may represent a potential target for pharmacological intervention in human and canine CSCC. The present study aimed to explore the expression pattern and status of activation of relevant signalling proteins of the PI3K/Akt/mTOR signalling pathway in feline CSCC. The expression of pEGFRTyr1068 , pAktSer473 , pS6Ser235/236 combined with Ki-67, and the tumour suppressor protein PTEN was evaluated by immunohistochemical analysis in 45 samples of feline CSCC, using a tissue microarray. The immunodetection using phosphospecific antibodies to detect the activated forms of signalling proteins showed that the PI3K/Akt/mTOR signalling pathway is frequently activated in feline CSCCs, and may be independent of the activation of EGFR. The results also showed that PTEN expression is not significantly altered in feline CSCCs. Our study shows that the persistent activation of the PI3K/Akt/mTOR signalling pathway represents a key event in feline CSCC, pointing to this signalling pathway being a potential therapeutic target in feline patients with CSCC.

Heregulin controls ERα and HER2 signaling in mammospheres of ERα-positive breast cancer cells and interferes with the efficacy of molecular targeted therapy

Estrogen receptor (ER)α and the human epidermal growth factor receptor (HER) family are inversely expressed in ERα-positive cancer in association with resistance to hormonal therapy, but the mechanism underlying their relationship remains unknown. We analyzed the effect of HER family ligands on the expression of ER and the HER family in ERα-positive MCF-7 and T47D breast cancer cell lines in 3D spheroid culture. Here, we demonstrated for the first time that heregulin-1β (HRG), a HER3 and HER4 ligand, most effectively regulated ER/HER family expression by decreasing ERα mRNA expression and increasing HER family mRNA expression. HRG treatment attenuated fulvestrant-mediated growth inhibition, and promoted the migration of MCF-7 cells. Moreover, HRG increased the CD44+/CD24− cell fraction and side population cells, both of which are recognized as prospective breast cancer stem cell markers. HRG activated both phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) and mitogen-activated protein kinase (MAPK) pathways. Inhibitors of these pathways reduced the growth of MCF-7 cells, but the addition of HRG has different effects on these pathways. HRG blocked the inhibitory effect of mTOR inhibitors, such as rapamycin and everolimus, on cell growth but not that of a PI3K inhibitor. Furthermore, HRG slightly decreased the inhibitory effect of an AKT inhibitor on cell growth. In contrast, HRG enhanced the MEK inhibitor-induced inhibition of cell growth. These findings suggest that HRG-stimulated signaling pathways allow ERα-positive breast cancer cells to escape from growth inhibition caused by everolimus, via MAPK signaling and/or other signaling pathways. Everolimus improves progression-free survival in combination with exemestane as second-line therapy for metastatic hormone receptor-positive breast cancer. Our study suggests that HRG is a novel target for ERα-positive breast cancer therapy.

Mangiferin ameliorates placental oxidative stress and activates PI3K/Akt/mTOR pathway in mouse model of preeclampsia

Preeclampsia is an inflammatory disease which can induce oxidative stress in placenta. Oxidative stress in preeclampsia is regulated by the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway. Mangiferin, an anti-oxidative molecule, is reported to ameliorate oxidative stress in the kidney and brain through activating the PI3K/Akt/mTOR pathway. We aimed to investigate the effects of mangiferin in a mouse model of preeclampsia, which was induced by phosphatidylserine/dioleoyl-phosphatidycholine (PS/PC) from day 5 to 17 of pregnancy. The female pregnant mice were divided into five groups according to drug treatment. Animals received mangiferin orally at doses of 10, 20, 40 mg/kg/day from day 0.5 to 17. In preeclampsia mouse model, elevated systolic blood pressure and proteinuria were ameliorated by mangiferin treatment. Mangiferin attenuated fms-like tyrosine kinase-1 and placental growth factor expression and oxidative stress in both blood and placenta of preeclampsia mice. The suppressed PI3K/Akt/mTOR pathway in placenta was activated by mangiferin treatment. This study demonstrates that mangiferin ameliorates placental oxidative stress and activates PI3K/Akt/mTOR pathway in a mouse model of preeclampsia.

Advance in research of microRNA-21-5p regulate autophagy by targeting gene

Autophagy is a dynamic process that degrades intracellular proteins and damaged organelles, and maintains environmental stability within the cell and provides good conditions for cell survival. Hyperoxic acute lung injury (HALI) is one of the serious complications of clinical oxygen therapy. The pathogenesis of HALI is still unclear. There are studies having shown that autophagy is involved in the pathogenesis of HALI. There are many pathway mechanisms that regulate autophagy, including phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway, mitogen-activated protein kinase/extracellular signaling-regulated protein kinase (MAPK/ERK) signaling pathway, adenosine 5'-monophosphate-activated protein kinase/unc-51 like autophagy activating kinase 1 (AMPK/ULK1) signaling pathway, transforming growth factor β (TGF-β) and forkhead box O1 (FoxO1) and Ras guanosine triphosphatease superfamily member Rab11a, each of which is referred to as microRNA-21-5p (miR-21-5p) target gene having a role in regulating autophagy activity in many diseases. In this paper, the above-mentioned signaling pathways of miRNA-21-5p target genes regulating autophagy were reviewed in order to find clues about the mechanism of miRNA-21-5p regulating autophagy in HALI and provide a theoretical basis for subsequent basic research.

Dysregulated Expression of Phosphorylated Epidermal Growth Factor Receptor and Phosphatase and Tensin Homologue in Canine Cutaneous Papillomas and Squamous Cell Carcinomas

The molecular mechanisms contributing to the development of cutaneous papillomas (CPs) and cutaneous squamous cell carcinomas (CSCCs) are still poorly understood, limiting the ability to identify molecular suitable targets for the development of novel therapies. Persistent activation of the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway is a component of epidermal carcinogenesis in dogs. The present study describes the immunohistochemical expression pattern of two key regulatory molecules involved in the PI3K/Akt/mTOR signalling pathway, phosphorylated epidermal growth factor receptor (pEGFR)Tyr1068 and phosphatase and tensin homologue (PTEN), in samples of normal canine epidermis, CP, preneoplastic epidermis and CSCC using tissue microarrays to determine whether the deregulated activity of these molecules is involved in the pathogenesis of these relevant epidermal tumours of dogs. Expression of pEGFR and PTEN was dysregulated in most samples of CP, preneoplastic epidermis and CSCC. Overexpression of pEGFR, together with decreased expression of PTEN, may facilitate the progression of some canine CPs and CSCCs by deregulation of the key cellular functions in which the PI3K/Akt/mTOR signalling pathway is involved. These findings suggest that the PI3K/Akt/mTOR signalling molecules may be potential therapeutic targets for canine patients with CP and CSCC.

DPEP1 is a direct target of miR-193a-5p and promotes hepatoblastoma progression by PI3K/Akt/mTOR pathway

Hepatoblastoma (HB) is the most common hepatic neoplasm in childhood and the therapeutic outcomes remain undesirable due to its recurrence and metastasis. Increasing evidence shows that dipeptidase 1 (DPEP1) has pivotal function in tumorigenesis in multiple tumors. However, the expression pattern, biological function, and underlying mechanism of DPEP1 in HB have not been reported. Here we showed that DPEP1 was significantly upregulated and was associated with poor prognosis in HB patients. In vitro and in vivo assays indicated that silencing DPEP1 significantly suppressed HB cell proliferation, migration, and invasion, while DPEP1 overexpression exhibited the opposite effect. In addition, we identified that DPEP1 was a direct target of microRNA-193a-5p (miR-193a-5p). Functional experiments demonstrated that overexpression of miR-193a-5p significantly inhibited cell proliferation and invasion of HB cells, while the inhibitory effect could be reversed by DPEP1 overexpression. Moreover, miR-193a-5p was decreased in HB tumor tissues and associated with a poor clinical prognosis. Mechanistically, our results indicated that the miR-193a-5p/DPEP1 axis participated to the progression of HB via regulating the PI3K/Akt/mTOR (phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin) signaling. In conclusion, our findings suggest that the miR-193a-5p /DPEP1 axis might be a good prognostic predictor and therapeutic target in HB.

Dysregulated expression of the key effectors of the mammalian target of rapamycin signalling pathway in cutaneous papillomas of dogs.

Cutaneous papillomas (CP) are one of the most common skin neoplasms in dogs. Different murine models have shown that persistent activation of the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway has a central role in the development and progression of CP. The purpose of this study were to evaluate the immunohistochemical expression pattern of two key molecules involved in the PI3K/Akt/mTOR signalling pathway, pAktSer473 , and pS6Ser235/236 , on 36 canine specimens of CP using a tissue microarray. The results show that the PI3K/Akt/mTOR signalling pathway is persistently activated in CP of dogs, pointing to this pathway as a potential therapeutic target.

Neuroprotective effect of ginkgetin in experimental cerebral ischemia/reperfusion via apoptosis inhibition and PI3K/Akt/mTOR signaling pathway activation

Ginkgetin, extracted from Ginkgo biloba L leaves, has been demonstrated to have potential anti-inflammatory and immune-suppressive properties. But the neuroprotective effect and potential mechanisms of ginkgetin on cerebral ischemia/reperfusion (IR) injury remain unclear. In this research, we studied the neuroprotective effect of ginkgetin in the middle part of the middle cerebral artery occlusion/reperfusion rat model, by analyzing the apoptosis of brain tissues harvested from treatment groups and control groups using the terminal deoxynucleotidyl transferase dUTP nick-end labeling and apoptosis assays. In addition, we detected the association of the neuroprotective effect of ginkgetin with apoptosis inhibition via the activation of the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway using Western blot analysis. Our results showed that administration of ginkgetin remarkably reduced brain infarction volumes and neurologic deficits; in addition, reducing apoptotic cell numbers, downregulating the levels of cleaved caspase-3 and Bax, and upregulating the level of Bcl-2 in rats subjected to IR injury in a dose-dependent manner. Moreover, high-dose ginkgetin treatment (100 mg/kg) significantly increased the phosphorylations of Akt and mTOR. Blocking of PI3K by LY294002 clearly decreased its antiapoptotic effect and reduced both Akt and mTOR phosphorylation levels. Taken together, these results for the first time suggest that ginkgetin antagonizes cerebral IR-induced injury by inhibiting apoptosis in rats, and this effect was attenuated by the activation of PI3K/Akt/mTOR signaling pathway.

A Comparison of DNA Mutation and Copy Number Profiles of Primary Breast Cancers and Paired Brain Metastases for Identifying Clinically Relevant Genetic Alterations in Brain Metastases.

Improving the systemic treatment of brain metastases (BM) in primary breast cancer (PBC) is impaired by the lack of genomic characterization of BM. To estimate the concordance of DNA copy-number-alterations (CNAs), mutations, and actionable genetic alterations (AGAs) between paired samples, we performed whole-genome array-comparative-genomic-hybridization, and targeted-next-generation-sequencing on 14 clinical PBC–BM pairs. We found more CNAs, more mutations, and higher tumor mutational burden, and more AGAs in BM than in PBC; 92% of the pairs harbored at least one AGA in the BM not observed in the paired PBC. This concerned various therapeutic classes, including tyrosine-kinase-receptor-inhibitors, phosphatidylinositol 3-kinase/AKT/ mammalian Target of Rapamycin (PI3K/AKT/MTOR)-inhibitors, poly ADP ribose polymerase (PARP)-inhibitors, or cyclin-dependent kinase (CDK)-inhibitors. With regards to the PARP-inhibitors, the homologous recombination defect score was positive in 79% of BM, compared to 43% of PBC, discordant in 7 out of 14 pairs, and positive in the BM in 5 out of 14 cases. CDK-inhibitors were associated with the largest percentage of discordant AGA appearing in the BM. When considering the AGA with the highest clinical-evidence level, for each sample, 50% of the pairs harbored an AGA in the BM not detected or not retained from the analysis of the paired PBC. Thus, the profiling of BM provided a more reliable opportunity, than that of PBC, for diagnostic decision-making based on genomic analysis. Patients with BM deserve an investigation of several targeted therapies.

The Esophageal Cancer and the PI3K/AKT/mTOR Signaling Regulatory microRNAs: a Novel Marker for Prognosis, and a Possible Target for Immunotherapy.

The Phosphatidylinositol 3-kinase/AKT/Mammalian Target of Rapamycin (PI3K/AKT/mTOR) pathway has a critical regulatory role in cell biology including translation, transcription, and autophagy. Dysregulation of this pathway is involved in the pathogenesis, development, and prognosis of esophageal cancer that has been assessed in the recent years and its potential as a target in therapy. This report summarizes the current knowledge about PI3K/AKT/mTOR pathway and its cross-talk with a focus on the value of targeting this pathway as a potential therapeutic target in the treatment of esophageal cancer.

Immunohistochemical expression of selected phosphoproteins of the mTOR signalling pathway in canine cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) represents one of the most common malignant skin tumors in dogs. Research aimed at clarifying how the deregulated activity of signalling pathways contributes to cSCC progression can help to identify molecular suitable targets for the development of novel therapies. The present study describes the immunohistochemical expression pattern of two proteins (pAktSer473 and pS6Ser235/236, the latter combined with Ki-67) involved in the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway in canine specimens of normal epidermis, pre-neoplastic epidermis, and cSCC using tissue microarrays.The results suggest that the PI3K/Akt/mTOR signalling pathway has a low expression in the normal canine epidermis, and that selected molecules involved in this signalling pathway are dysregulated during the canine epidermal carcinogenesis process. These findings provide important evidence that the persistent activation of the PI3K/Akt/mTOR signalling pathway represents one of the key events during cSCC progression in canine patients, pointing to the PI3K/Akt/mTOR pathway as a potential therapeutic target.

Mangiferin Potentiates Neuroprotection by Isoflurane in Neonatal Hypoxic Brain Injury by Reducing Oxidative Stress and Activation of Phosphatidylinositol-3-Kinase/Akt/Mammalian Target of Rapamycin (PI3K/Akt/mTOR) Signaling.

BackgroundHypoxic-ischemic brain injury in the perinatal period is a main cause of perinatal mortality and neurologic complications in neonates and children. Recent studies have focused on the neuroprotective effect of anesthetic drugs. The volatile anesthetic isoflurane has been shown to exert neuroprotective effects in cerebral ischemia. Mangiferin is a natural polyphenol with various pharmacological properties, including antioxidant and ant-tumor effects. This study aimed to determine whether mangiferin potentiates the neuroprotective effects of isoflurane and also if mangiferin when administered alone exerts neuroprotective effects following hypoxic-ischemic brain injury.Material/MethodsSprague-Dawley rats were subjected to cerebral hypoxic ischemia on postnatal day 10 (P10). Mangiferin (50, 100, or 200 mg/kg b.w.) was intragastrically administered from P3 to P12 and 1 h prior to insult on the day of ischemic induction. At 3 h after hypoxia-ischemia (HI) insult, separate groups of rat pups were exposed to isoflurane (1.5%) for 6 h. Following 48 h of HI, the rats were sacrificed and brain tissues were used for analysis.ResultsMangiferin treatment attenuated neuronal apoptosis and reduced cerebral infarct volume. The expression of cleaved caspase-3 and apoptotic cascade proteins were regulated. The levels of reactive oxygen species (ROS) and malondialdehyde were reduced by mangiferin and/or isoflurane exposure. The levels of antioxidant glutathione were considerably raised under HI injury, which was modulated by mangiferin and isoflurane exposure. The PI3K/Akt signaling pathway, which was downregulated following HI insult, was activated by mangiferin and/or isoflurane.ConclusionsThis study reveals the potent neuroprotective efficacy of mangiferin against HI-induced brain injury via effectively modulating apoptotic pathways, ROS levels, and PI3K/Akt cascades while potentiating protective effects of isoflurane.

Blocking of YY1 reduce neutrophil infiltration by inhibiting IL-8 production via the PI3K-Akt-mTOR signaling pathway in rheumatoid arthritis

Our previous study revealed that Yin Yang 1(YY1) played an important part in promoting interleukin (IL)-6 production in rheumatoid arthritis (RA). However, whether YY1 has any role in regulation of IL-8 in RA remains unclear. YY1 and IL-8 expression in RA patients were analyzed by real-time polymerase chain reaction (PCR). Ingenuity pathway analysis (IPA) was used to analyze the signaling pathway involved in YY1-induced IL-8 production. The expression of YY1 and proteins involved in the pathway were detected by Western blot and enzyme-linked immunosorbent assay (ELISA). Migration of neutrophils was performed by chemotaxis assay. In this study, we found that high expression of IL-8 was positively associated with YY1 expression in RA. Blocking YY1 expression by YY1-short hairpin (sh)RNA lentivirus reduced IL-8 production. Mechanistically, we showed YY1 activated IL-8 production via the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway. Further, using a co-culture system consisting of peripheral blood mononuclear cells (PBMC) and neutrophils, we found that migration of neutrophils would be inhibited by YY1 RNA interference. Finally, using the collagen-induced arthritis animal model, we showed that treatment with the YY1-shRNA lentivirus led to reduction of IL-8 levels and attenuation of inflammation and neutrophil infiltration in vivo. Our results reveal a role of YY1 involved in neutrophil infiltration in RA via the PI3K/Akt/mTOR/IL-8 signaling pathway. YY1 may be a new therapeutic target for treatment of RA.

Two acute myeloid leukemia patient subsets are identified based on the constitutive PI3K-Akt-mTOR signaling of their leukemic cells; a functional, proteomic, and transcriptomic comparison

ABSTRACTObjectives: Constitutive signaling through the phosphatidylinositol-3-kinase-Akt-mammalian target of rapamycin (PI3K-Akt-mTOR) pathway is present in acute myeloid leukemia (AML) cells. The aim of the study was to compare constitutive PI3K-Akt-mTOR activation of primary AML cells for a large group of unselected patients.Methods: We investigated expression and phosphorylation of 18 mediators in the PI3K-Akt-mTOR main track by flow cytometry for AML cells derived from 77 patients, and compared this with global gene expression profiles, proteomic, and transcriptomic profiles, and susceptibility to antileukemic agents.Results: Patients were divided into two main subsets showing generally high or low constitutive pathway activation. The high activation subset was characterized by decreased frequency of cells showing monocytic differentiation, increased frequency of adverse karyotypes, decreased constitutive cytokine release, and increased expression of certain integrins. Finally, the two groups differed in their expression of genes encoding regulators of protein phosphorylation, whereas phosphoproteomic analyses showed differences especially with regard to transcriptional regulation. Antiproliferative effects of pathway inhibition were generally stronger for the low phosphorylation subset.Conclusion: The constitutive PI3K-Akt-mTOR activation differed between patients; this difference appears to be a part of complex phenotypic differences including cell communication, intracellular signaling through other pathways, and transcriptional regulation.

Ursolic Acid Attenuates High Glucose-Mediated Mesangial Cell Injury by Inhibiting the Phosphatidylinositol 3-Kinase/Akt/Mammalian Target of Rapamycin (PI3K/Akt/mTOR) Signaling Pathway.

BackgroundTo investigate the protective effect of ursolic acid (UA) on high glucose (HG)-induced human glomerular mesangial cell injury and to determine whether UA inhibits cell proliferation and reactive oxygen species (ROS) production by suppressing PI3K/Akt/mTOR pathway activation.Material/MethodsHuman mesangial cells were cultured with normal glucose (NG group), high glucose (HG group), mannitol (mannitol hypertonic control group), or high glucose with different concentrations (0.5, 1.0, and 2.0 mmol/L) of UA (HG+UA groups). Cell proliferation and intracellular ROS levels were assessed by methyl thiazolyl tetrazolium (MTT) and dichloro-dihydro-fluorescein diacetate (DCFH-DA) flow cytometry assays, respectively. Western blotting was used to detect mesangial cell expression of PI3K/Akt/mTOR pathway components, including Akt, p-Akt, mTOR, and p-mTOR, and proteins related to cell injury, including TGF-β1 and fibronectin (FN). mRNA expression of TGF-β1 and FN were evaluated using real-time quantitative polymerase chain reaction (PCR).ResultsAbnormal proliferation was observed in human glomerular mesangial cells at 48 h after treatment with HG, and UA suppressed the HG-induced proliferation of mesangial cells in a dose-dependent manner. UA inhibited ROS generation and oxidative stress in mesangial cells and mitigated mesangial cell injury. Treatment with UA reduced Akt and mTOR phosphorylation levels in mesangial cells exposed to HG (p<0.05 vs. HG) and downregulated protein and mRNA expression of TGF-β1 and FN in these cells (p<0.05 vs. HG).ConclusionsUA attenuated mesangial cell proliferation and ROS generation by inhibiting HG-mediated PI3K/Akt/mTOR pathway activation, thereby ameliorating mesangial cell damage.

Design, Synthesis, and Preclinical Evaluation of Fused Pyrimidine-Based Hydroxamates for the Treatment of Hepatocellular Carcinoma.

Class I histone deacetylases (HDACs) are highly expressed and/or upregulated in hepatocellular carcinoma (HCC) and are associated with aggressiveness, spread, and increased mortality of HCC. Activation of phosphatidylinositol 3-kinase-Akt-mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway was involved in the development of HCC and acquired resistance to sorafenib. A series of purine or 5H-pyrrolo[3,2-d]pyrimidine based hydroxamates were designed and developed as multitarget drugs to modulate both HDACs and the PI3K/Akt/mTOR pathway. Among 39 cell lines screened, the molecules (e.g., 20e, 20f, and 20q) were the most selective against leukemia, lymphoma, and HCC cells; they also demonstrated target modulation in cancer cell lines and in mice bearing MV4-11 and HepG2 tumors. Compound 20f in particular showed significant single agent oral efficacy in hypervascular liver cancer models (e.g., HepG2, HuH-7, and Hep3B) and was well-tolerated. These encouraging results, along with its favorable target profile and tissue distribution, warrant further development of 20f.

Novel Molecular and Phenotypic Insights into Congenital Lung Malformations.

Disruption of normal pulmonary development is a leading cause of morbidity and mortality in infants. Congenital lung malformations are a unique model to study the molecular pathogenesis of isolated structural birth defects, as they are often surgically resected. To provide insight into the molecular pathogenesis of congenital lung malformations through analysis of cell-type and gene expression changes in these lesions. Clinical data, and lung tissue for DNA, RNA, and histology, were obtained from 58 infants undergoing surgical resection of a congenital lung lesion. Transcriptome-wide gene expression analysis was performed on paired affected and unaffected samples from a subset of infants (n = 14). A three-dimensional organoid culture model was used to assess isolated congenital lung malformation epithelium (n = 3). Congenital lung lesions express higher levels of airway epithelial related genes, and dysregulated expression of genes related to the Ras and PI3K-AKT-mTOR (phosphatidylinositol 3-kinase-AKT-mammalian target of rapamycin) signaling pathways. Immunofluorescence confirmed differentiated airway epithelial cell types throughout all major subtypes of congenital lung lesions, and three-dimensional cell culture demonstrated a cell-autonomous defect in the epithelium of these lesions. This study provides the first comprehensive analysis of the congenital lung malformation transcriptome and suggests that disruptions in Ras or PI3K-AKT-mTOR signaling may contribute to the pathology through an epithelial cell-autonomous defect.