• HRM test
• PIK3CA
• cetuximab
• colorectal cancer
• mutation spectrum
• nomogram
• the predictive value

• Gastric cancer
• PI3K/Akt/mTOR pathway
• PTEN
• Phosphatidylinositol-3-kinase

Gastric cancer remains a major health concern, and improvement of the therapeutic options is crucial. Treatment with targeted therapeutics such as the EGFR-targeting antibody cetuximab or the HER2-targeting antibody trastuzumab is either ineffective or moderately effective in this disease, respectively. In this study, we analysed the involvement of the HER receptor ligands amphiregulin (AREG), epidermal growth factor (EGF), heparin-binding epidermal growth factor (HB-EGF) and transforming growth factor alpha (TGFα) in the responsiveness of gastric cancer cell lines to cetuximab and trastuzumab.

A panel of 11 gastric cancer cell lines was characterized for cetuximab and trastuzumab sensitivity, ligand secretion and expression and activation of the HER receptors using WST-1 cell proliferation assays, ELISAs and Western blot analyses. We further investigated the effects of an exogenous ligand application on the cetuximab and trastuzumab sensitivity.

We found no correlation between TGFα secretion and the sensitivity to cetuximab or trastuzumab. For AREG, we confirmed previous results indicating that this ligand is a positive predictor of cetuximab sensitivity. Exogenous HB-EGF was effective in rescuing sensitive cell lines from inhibition of cell proliferation by both, cetuximab and trastuzumab.

Our data indicate that HB-EGF may be a useful marker for the prediction of trastuzumab sensitivity in gastric cancer.

The online version of this article (doi:10.1007/s00432-016-2308-z) contains supplementary material, which is available to authorized users.

• Cetuximab
• EGFR
• Gastric cancer
• HER receptors
• Ligand
• Trastuzumab

Many urothelial carcinomas (UC) contain activating PIK3CA mutations. In telomerase-immortalized normal urothelial cells (TERT-NHUC), ectopic expression of mutant PIK3CA induces PI3K pathway activation, cell proliferation and cell migration. However, it is not clear whether advanced UC tumors are PIK3CA-dependent and whether PI3K pathway inhibition is a good therapeutic option in such cases.

We used retrovirus-mediated delivery of shRNA to knock down mutant PIK3CA in UC cell lines and assessed effects on pathway activation, cell proliferation, migration and tumorigenicity. The effect of the class I PI3K inhibitor GDC-0941 was assessed in a panel of UC cell lines with a range of known molecular alterations in the PI3K pathway.

Specific knockdown of PIK3CA inhibited proliferation, migration, anchorage-independent growth and in vivo tumor growth of cells with PIK3CA mutations. Sensitivity to GDC-0941 was dependent on hotspot PIK3CA mutation status. Cells with rare PIK3CA mutations and co-occurring TSC1 or PTEN mutations were less sensitive. Furthermore, downstream PI3K pathway alterations in TSC1 or PTEN or co-occurring AKT1 and RAS gene mutations were associated with GDC-0941 resistance.

Mutant PIK3CA is a potent oncogenic driver in many UC cell lines and may represent a valuable therapeutic target in advanced bladder cancer.

The online version of this article (doi:10.1186/s12885-016-2570-0) contains supplementary material, which is available to authorized users.

• Bladder cancer
• PI3K signaling
• PIK3CA
• Urothelium

• Esophageal squamous cell carcinoma
• Metastasis
• Motility
• PIK3CA
• Prognosis
• Proliferation

• Adult
• Aged
• Apoptosis
• Asian People
• Biomarkers, Tumor/genetics
• Biomarkers, Tumor/metabolism
• Carcinoma, Squamous Cell/enzymology
• Carcinoma, Squamous Cell/ethnology
• Carcinoma, Squamous Cell/genetics
• Carcinoma, Squamous Cell/secondary
• Cell Line, Tumor
• Cell Movement
• Cell Proliferation
• China/epidemiology
• Class I Phosphatidylinositol 3-Kinases
• Esophageal Neoplasms/enzymology
• Esophageal Neoplasms/ethnology
• Esophageal Neoplasms/genetics
• Esophageal Neoplasms/pathology
• Esophageal Squamous Cell Carcinoma
• Female
• Gene Expression Regulation, Neoplastic
• Humans
• Kaplan-Meier Estimate
• Lymphatic Metastasis
• Male
• Middle Aged
• Phosphatidylinositol 3-Kinases/genetics
• Phosphatidylinositol 3-Kinases/metabolism
• RNA Interference
• S Phase Cell Cycle Checkpoints
• Signal Transduction
• Time Factors
• Transfection
• Up-Regulation

PIK3CA is often mutated in a variety of malignancies, including colon, gastric, ovary, breast, and brain tumors. We investigated PIK3CA expression in gastric cancer and explored the relationships between the PIK3CA expression level and clinicopathological features as well as survival of the patients.

We examined PIK3CA expression in a tissue microarray of 178 gastric adenocarcinomas by immunohisto-chemistry and reviewed patients' medical records.

In our study, 112 of the 178 gastric cancer patients displayed positive PIK3CA expression. Overexpression of PIK3CA was correlated with low grade differentiation (P=0.001), frequent lymphatic invasion (P=0.032), and high T stage (P=0.040). Patients with positive PIK3CA staining were more likely to display worse overall survival rate than those with negative PIK3CA staining, as determined by Kaplan-Meier survival analysis with log-rank test (P=0.047) and a univariate analysis using the Cox proportional hazard model (hazard ratio=1.832, P=0.051).

Elevated PIK3CA expression was significantly correlated with tumor invasiveness, tumor phenotypes, and poor patient survival.

• Immunohistochemistry
• PIK3CA
• Stomach neoplasms

Despite advances in clinical therapies and technologies, the prognosis for patients with gastric cancer is still poor. The aim of this study is to investigate new predictive markers for prognosis of gastric cancer.

In this study, we evaluated the expression pattern of PIK3CA in 107 gastric cancer specimens and their adjacent nontumorous tissues. PIK3CA siRNA was synthesized and transfected into gastric cancer cell lines. Colony formation and MTT assays were employed to analyze the cell proliferation. PIK3CA expression was examined by using immunohistochemical analysis and Western blot assay. Transwell invasion assay was used to detect the invasion capability of the cells. Luciferase activity was examined by using 3′-untranslated region luciferase reporter assays.

We observed that PIK3CA was significantly upregulated in gastric cancer tissues. High expression level of PIK3CA was detectable in 48 (44.86%) of the gastric cancer specimens, and correlated with poor prognosis. In addition, our study indicated that miR203 inhibits cell proliferation and invasion via directly targeting and suppressing the PIK3CA expression. MiR203 expression is downregulated in gastric cancer tissues. Moreover, low expression level of miR203 predicted poor prognosis of gastric patients and induced overexpression of PIK3CA. Our further study also reported that overexpression of miR203 inhibited phosphorylation of AKT, while cotransfection of PIK3CA reversed the effect of miR203.

Our study suggested a miR203-PIK3CA-AKT signaling pathway in gastric cancer cells. This signaling pathway might play an important role in gastric cancer genesis and development.

• AKT
• PIK3CA
• gastric cancer
• miR203
• prognosis

• AKT3 and PI3KCA genes
• Apoptosis
• Autophagy
• Glioblastoma multiforme
• siRNA

• Apoptosis/genetics
• Apoptosis Regulatory Proteins/biosynthesis
• Apoptosis Regulatory Proteins/genetics
• Autophagy/genetics
• Cell Line, Tumor
• Gene Expression Regulation
• Gene Knockdown Techniques
• Glioblastoma/genetics
• Glioblastoma/pathology
• Humans
• Microtubule-Associated Proteins/biosynthesis
• Phosphatidylinositol 3-Kinases/deficiency
• Phosphatidylinositol 3-Kinases/metabolism
• Proto-Oncogene Proteins c-akt/deficiency
• Proto-Oncogene Proteins c-akt/metabolism
• RNA Interference

• gastric cancer
• gpr34
• knockdown
• migration
• proliferation

• Apoptosis/genetics
• Apoptosis/physiology
• Blotting, Western
• Cell Line, Tumor
• Cell Proliferation/genetics
• Cell Proliferation/physiology
• Humans
• RNA, Small Interfering/genetics
• Real-Time Polymerase Chain Reaction
• Receptors, Lysophospholipid/genetics
• Receptors, Lysophospholipid/metabolism
• Stomach Neoplasms/genetics
• Stomach Neoplasms/metabolism

The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is one of the key signaling pathways induced by various receptor-tyrosine kinases. Accumulating evidence shows that this pathway is an important promoter of cell growth, metabolism, survival, metastasis, and resistance to chemotherapy. Genetic alterations in the PI3K/Akt/mTOR pathway in gastric carcinoma have often been demonstrated. Many kinds of molecular targeting therapies are currently undergoing clinical testing in patients with solid tumors. However, with the exception of the ErbB2-targeting antibody, targeting agents, including PI3K/Akt/mTOR inhibitors, have not been approved for treatment of patients with gastric carcinoma. This review summarizes the current knowledge on PI3K/Akt/mTOR signaling in the pathogenesis of gastric carcinoma and the possible therapeutic targets for gastric carcinoma. Improved knowledge of the PI3K/Akt/mTOR pathway in gastric carcinoma will be useful in understanding the mechanisms of tumor development and for identifying ideal targets of anticancer therapy for gastric carcinoma.

AIM: To investigate the gene knock-down effect by the phosphoinositide-3-kinase, catalytic, alpha polypeptide (PIK3CA)-targeted double-stranded RNA (dsRNA) and its effect on cell proliferation and cycle distribution in SW948.

METHODS: Two PIK3CA-targeted dsRNAs were constructed and transfected into SW948 cells. Transfections were performed using lipofectamine^TM 2000. The transfection effectiveness was calculated basing on the rate of fluorescence cell of SW948 at 6 h after transfection. Total messenger RNA was extracted from these cells using the RNeasy kit, and semiquantitative reverse transcription polymerase chain reaction was performed to detect the down-regulation of PIK3CA, AKT1, MYC, and CCND1 gene expression. Cells were harvested, proteins were resolved, and western blot was employed to detect the expression levels of PIK3CA, AKT1, MYC, and CCND1 gene. Cell proliferation was assessed by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide assay and the inhibition rate was calculated. Soft agar colony formation assay was performed basing on colonies greater than 60 μm in diameter at ×100 magnification. The effect on cell cycle distribution and apoptosis was assessed by flow cytometry. All experiments were performed in triplicate.

RESULTS: Green fluorescence was observed in SW948 cell transfected with plasmid Pgenesil-1, and the transfection effectiveness was about 65%. Forty-eight hours post-transfection, mRNA expression of PIK3CA in SW948 cells was 0.51 ± 0.04 vs 0.49 ± 0.03 vs 0.92 ± 0.01 vs 0.93 ± 0.03 (P = 0.001 ) in Pgenesil-CA1, Pgenesil-CA2, negative and blank group respectively. mRNA expression of AKT1 was 0.50 ± 0.03 vs 0.48 ± 0.01 vs 0.93 ± 0.04 vs 0.92 ± 0.02 (P = 0.000) in Pgenesil-CA1, Pgenesil-CA2, negative and blank group respectively. mRNA expression of MYC was 0.49 ± 0.01 vs 0.50 ± 0.04 vs 0.90 ± 0.02 vs 0.91 ± 0.03 (P = 0.001) in the four groups respectively. mRNA expression of CCND1 was 0.45 ± 0.02 vs 0.51 ± 0.01 vs 0.96 ± 0.03 vs 0.98 ± 0.01 (P = 0.001) in the four groups respectively. The protein level of PIK3CA was 0.53 ± 0.01 vs 0.54 ± 0.02 vs 0.92 ± 0.03 vs 0.91 ± 0.02 (P = 0.001) in Pgenesil-CA1, Pgenesil-CA2, negative and blank group respectively. The protein level of AKT1 in the four groups was 0.49 ± 0.02 vs 0.55 ± 0.03 vs 0.94 ± 0.03 vs 0.95 ± 0.04, P = 0.000). The protein level of MYC in the four groups was 0.51 ± 0.03 vs 0.52 ± 0.04 vs 0.92 ± 0.02 vs 0.95 ± 0.01 (P = 0.000). The protein level of CCND1 in the four groups was 0.54 ± 0.04 vs 0.56 ± 0.03 vs 0.93 ± 0.01 vs 0.93 ± 0.03 (P = 0.000). Both Pgenesil-CA1 and Pgenesil-CA2 plasmids significantly suppressed the growth of SW948 cells when compared with the negative or blank group at 48 h after transfection (29% vs 25% vs 17% vs 14%, P = 0.001), 60 h after transfection (38% vs 34% vs 19% vs 16%, P = 0.001), and 72 h after transfection (53% vs 48% vs 20% vs 17%, P = 0.000). Numbers of colonies in negative, blank, CA1, and CA2 groups were 42 ± 4, 45 ± 5, 8 ± 2, and 10 ± 3, respectively (P = 0.000). There were more than 4.5 times colonies in the blank and negative control groups as there were in the CA1 and CA2 groups. In addition, the colonies in blank and negative control groups were also larger than those in the CA1 and CA2 groups. The percentage of cells in the CA1 and CA2 groups was significantly higher in G₀/G₁ phase, but lower in S and G₂/M phase when compared with the negative and control groups. Moreover, cell apoptosis rates in the CA1 and CA2 groups were 5.11 ± 0.32 and 4.73 ± 0.32, which were significantly higher than those in negative (0.95 ± 0.11, P = 0.000) and blank groups (0.86 ± 0.13, P = 0.001). No significant difference was found between CA1 and CA2 groups in cell cycle distribution and apoptosis.

CONCLUSION: PIK3CA-targeted short hairpin RNAs can block the phosphoinositide 3-kinase-Akt signaling pathway and inhibit cell growth, increase apoptosis, and induce cell cycle arrest in the PIK3CA-mutant colon cancer SW948 cells.

• Phosphoinositide-3-kinase, catalytic, alpha polypeptide
• RNA interference
• Colon cancer
• Phosphoinositide-3-kinase pathway