title: Morphoproteomic profile of mTOR, Ras/Raf kinase/ERK, and NF-kappaB pathways in human gastric adenocarcinoma.
abstract: Preclinical studies using human gastric adenocarcinoma (GAC) cell lines have shown that the mammalian target of rapamycin (mTOR) inhibitor, rapamycin, can inhibit tumor growth and that the extracellular signal-regulated kinase (ERK) of the Ras/Raf kinase/ERK pathway is related to chemoresistance and apoptosis. We examined the state of activation of components of mTOR, Ras/Raf kinase/ERK, and nuclear factor (NF)-kappaB signal transduction pathways, as well as cell cycle protein analyte correlates in GAC cases. Formalin-fixed paraffin-embedded tissue microarray blocks containing samples from 210 cases of GAC were examined. Immunohistochemistry was utilized to detect the following antigens: S100P, upstream stimulator of ERK, and NF-kappaB pathways; phosphorylated (p)-mTOR (Ser 2448), p-ERK-1/2 (Thr 202/Tyr 204), and one of their common down-stream effectors, p-p70S6K(Thr 389); p-NF-kappaBp65(Ser 536); and cell cycle associated proteins, Ki-67, and S phase kinase-associated protein (Skp)2. Immunoreactivity (0 to 4+) of protein expression and compartmentalization were assessed by bright-field microscopy. The majority of cases showed positive (1+ to 4+) cytoplasmic/plasmalemmal p-mTOR (88%), and moderate-strong (2+ to 4+) nuclear p-p70S6K (93%) and nuclear S100P (81%) expression. A subset of cases exhibited moderate-strong nuclear p-ERK-1/2 (15%) and p-NF-kappaBp65 (36%) expression. The majority of cases showed concomitant moderate-strong (2+ to 4+) nuclear Ki-67 (71%) and Skp2 (68%). Nuclear expression levels of p-ERK-1/2 and p-NF-kappaBp65, of p-p70S6K and p-NF-kappaB, and of Ki-67 and Skp2, respectively, showed significant linear correlations in GAC (p <0.001). Additionally, there were statistically significant differences in the mean expression levels of p-ERK-1/2 and p-NF-kappaBp65 in diffuse vs intestinal types of GAC, with higher levels of both in the diffuse type ( p = 0.001 and p <0.0001, respectively). In summary, morphoproteomic analysis reveals constitutive activation of mTOR and to some extent, Ras/Raf kinase and NF-kappaB pathways in GAC, as evidenced by increased cytoplasmic p-mTOR, nuclear translocation of p-p70S6K and p-ERK-1/2 phosphorylated at putative sites of activation (Ser 2448, Thr 389, and Thr 202/Tyr 204, respectively), as well as correlative expression of cell cycle analytes, Ki-67, and Skp2. These results suggest that a prospective study is warranted to evaluate the use of morphoproteomic profiling of individual patients with GAC in order to design combinatorial treatment strategies that target the mTOR, Ras/Raf kinase/ERK, and/or NF-kappaB pathways.
authors: Department of Pathology, MD Anderson Cancer Center, Houston, TX 77030, USA.
authors: Wei
authors: W
authors: Feng
authors: Robert E
authors: RE
authors: Brown
authors: Camtu D
authors: CD
authors: Trung
authors: Wei
authors: W
authors: Li
authors: Liwei
authors: L
authors: Wang
authors: Thaer
authors: T
authors: Khoury
authors: Sadir
authors: S
authors: Alrawi
authors: James
authors: J
authors: Yao
authors: Keping
authors: K
authors: Xia
authors: Dongfeng
authors: D
authors: Tan
mesh_terms: D000230:Adenocarcinoma / Q000201:enzymology / Q000473:pathology; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002135:Calcium-Binding Proteins / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005260:Female; D005753:Gastric Mucosa / Q000201:enzymology / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D019394:Ki-67 Antigen / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D016328:NF-kappa B / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D040901:Proteomics; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D044786:S-Phase Kinase-Associated Proteins / Q000378:metabolism; D013274:Stomach Neoplasms / Q000201:enzymology* / Q000473:pathology; D058570:TOR Serine-Threonine Kinases; D051996:Transcription Factor RelA / Q000378:metabolism; D048490:raf Kinases / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism*
chemical_list: D002135:Calcium-Binding Proteins; D019394:Ki-67 Antigen; D016328:NF-kappa B; D009363:Neoplasm Proteins; D010750:Phosphoproteins; D044786:S-Phase Kinase-Associated Proteins; C103003:S100P protein, human; D051996:Transcription Factor RelA; D011494:Protein Kinases; C546842:MTOR protein, human; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D048490:raf Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D018631:ras Proteins