Volumetric growth rates of sessile serrated adenomas/polyps observed in situ at longitudinal CT colonography.

OBJECTIVE: Sessile serrated adenomas/polyps (SSA/Ps) are now recognized as potential cancer precursors, but little is known about their natural history. We assessed the in vivo growth rates of histologically proven SSA/Ps at longitudinal CT colonography (CTC) and compared results with non-advanced tubular adenomas (TAs). METHODS: We identified a cohort of 53 patients (mean age, 54.8 ± 5.5 years; M:F, 26:27) from one center with a total of 58 SSA/Ps followed longitudinally at CTC (mean follow-up interval, 5.3 ± 1.9 years). Initial and final size measurements were determined using dedicated CTC software. Findings were compared with 141 non-advanced TAs followed at CTC (mean, 4.1 ± 2.3 years) in 113 patients (mean age, 56.8 ± 6.9 years). RESULTS: SSA/Ps were more often flat (62% [36/58] vs. 14% [20/141], p < 0.0001) and right-sided (98% [57/58] vs. 46% [65/141], p < 0.0001) compared with TAs. Initial average diameter was greater for SSA/Ps (9.3 mm vs. 6.3 mm; p < 0.0001). Mean annual volumetric growth was + 12.7%/year for SSA/Ps vs. + 36.4%/year for TAs (p = 0.028). Using a previously defined threshold of + 20% increase in volume/year to define progression, 22% (13/58) of SSA/Ps and 41% (58/141) of TAs progressed (p = 0.014). None of the SSA/Ps had dysplasia or invasive cancer at histopathology. CONCLUSIONS: Sessile serrated adenoma/polyps demonstrate slower growth compared with conventional non-advanced tubular adenomas, despite larger initial linear size. This less aggressive behavior may help explain the more advanced patient age for serrated pathway cancers. Furthermore, these findings could help inform future colonoscopic surveillance strategies, as current guidelines are largely restricted to expert opinion related to the absence of natural history data. KEY POINTS: • Sessile serrated adenoma/polyps (SSA/Ps) tend to be flat, right-sided, and demonstrate slower growth compared with conventional non-advanced tubular adenomas. • This less aggressive behavior of SSA/Ps may help explain the more advanced patient age for serrated pathway cancers.

PIK3CA mutations can initiate pancreatic tumorigenesis and are targetable with PI3K inhibitors.

Aberrations in the phosphoinositide 3-kinase (PI3K) signaling pathway have a key role in the pathogenesis of numerous cancers by altering cell growth, metabolism, proliferation and apoptosis. Interest in targeting the PI3K signaling cascade continues, as new agents are being clinically evaluated. PIK3CA mutations result in a constitutively active PI3K and are present in a subset of pancreatic cancers. Here we examine mutant PIK3CA-mediated pancreatic tumorigenesis and the response of PIK3CA mutant pancreatic cancers to dual PI3K/mammalian target of rapamycin (mTOR) inhibition. Two murine models were generated expressing a constitutively active PI3K within the pancreas. An increase in acinar-to-ductal metaplasia and pancreatic intraepithelial neoplasms (PanINs) was identified. In one model these lesions were detected as early as 10 days of age. Invasive pancreatic ductal adenocarcinoma developed in these mice as early as 20 days of age. These cancers were highly sensitive to treatment with dual PI3K/mTOR inhibition. In the second model, PanINs and invasive cancer develop with a greater latency owing to a lesser degree of PI3K pathway activation in this murine model. In addition to PI3K pathway activation, increased ERK1/2 signaling is common in human pancreatic cancers. Phosphorylation of ERK1/2 was also investigated in these models. Phosphorylation of ERK1/2 is demonstrated in the pre-neoplastic lesions and invasive cancers. This activation of ERK1/2 is diminished with dual PI3K/mTOR inhibition. In summary, PIK3CA mutations can initiate pancreatic tumorigenesis and these cancers are particularly sensitive to dual PI3K/mTOR inhibition. Future studies of PI3K pathway inhibitors for patients with PIK3CA mutant pancreatic cancers are warranted.

Gastrin-releasing peptide is a mitogen and a morphogen in murine colon cancer.

Little is known about the factors involved in regulating the appearance, or differentiation, of solid tumors including those arising from the colon. We herein demonstrate that the mitogen gastrin-releasing peptide (GRP) is a morphogen, critically important in regulating the differentiation of murine colon cancer. Although epithelial cells lining the mouse colon do not normally express GRP and its receptor (GRP-R), both are aberrantly expressed by all better differentiated cancers in wild-type C57BL/6J mice treated with the carcinogen azoxymethane. Whereas small tumors in both wild-type and GRP-R-deficient (i.e., GRP-R-/-) mice are histologically similar, larger tumors become better differentiated in the former but degenerate into more poorly differentiated mucinous adenocarcinomas in the latter. This alteration in phenotype is attributable to GRP increasing focal adhesion kinase expression in GRP-R-expressing tumors. Consistent with GRP acting as a mitogen, GRP/GRP-R coexpressing tumors in wild-type animals also contain more proliferating cells than those occurring in GRP-R-/- mice. Yet tumors are similarly sized in animals of either genotype receiving azoxymethane for identical times, a finding attributable to the significantly higher number of apoptotic cells detected in GRP/GRP-R coexpressing cancers. Thus, these findings indicate that although GRP is a mitogen, aberrant expression does not result in increased tumor growth. Rather, the mitogenic properties of GRP are subordinate to it acting as a morphogen, where it and its receptor are critically involved in regulating colon cancer histological progression by promoting a well-differentiated phenotype.

Quantitative immunohistochemistry by measuring cumulative signal strength using commercially available software photoshop and matlab.

Currently available techniques for performing quantitative immunohistochemistry (Q-IHC) rely upon pixel-counting algorithms and therefore cannot provide information as to the absolute amount of chromogen present. We describe a novel algorithm for true Q-IHC based on calculating the cumulative signal strength, or energy, of the digital file representing any portion of an image. This algorithm involves subtracting the energy of the digital file encoding the control image (i.e., not exposed to antibody) from that of the experimental image (i.e., antibody-treated). In this manner, the absolute amount of antibody-specific chromogen per pixel can be determined for any cellular region or structure. (J Histochem Cytochem 48:303-311, 2000)

Pathogenic Escherichia coli increase Cl- secretion from intestinal epithelia by upregulating galanin-1 receptor expression.

Galanin is widely distributed in enteric nerve terminals lining the human gastrointestinal (GI) tract. We have shown previously that galanin-1 receptors (Gal1-R) are expressed by epithelial cells lining the human GI tract, and upon activation cause Cl- secretion. Because expression of this receptor is transcriptionally regulated by nuclear factor-kappa B (NF-kappa B), which is activated by enteric pathogens as a part of the host epithelial response to infection, we investigated whether such bacterial pathogens could directly increase Gal1-R expression in the T84-cell model system. Pathogenic Escherichia coli, but not nonpathogenic E. coli, activate a p50/p65 NF-kappa B complex that binds to oligonucleotides corresponding to a recognition site located within the 5' flanking region of the human GAL1R gene. Pathogenic E. coli, but not normal commensal organisms, increase Gal1-R mRNA synthesis and [(125)I]galanin binding sites. Whereas galanin increases short-circuit current (Isc) approximately 5-fold in uninfected T84 cells, exposure to pathogenic, but not nonpathogenic, E. coli results in galanin increasing Isc approximately 20-fold. To confirm the validity of these in vitro observations, we also studied C57BL/6J mice infected with enterohemorrhagic E. coli (EHEC) by gavage. Infection caused a progressive increase in both NF-kappa B activation and Gal1-R expression, with maximal levels of both observed 3 days after gavage. Ussing chamber studies revealed that colons infected with EHEC, but not those exposed to normal colonic flora, markedly increased Isc in response to galanin. These data indicate that pathogen-induced increases in Gal1-R expression by epithelial cells lining the colon may represent a novel unifying pathway responsible for at least a portion of the excessive fluid secretion observed during infectious diarrhea.

Aberrant expression of gastrin-releasing peptide and its receptor by well-differentiated colon cancers in humans.

Epithelial cells lining the adult human colon do not normally express gastrin-releasing peptide (GRP) or its receptor (GRPR). In contrast, approximately one-third of human colon cancers and cancer cell lines have been shown to express GRP-binding sites. Because GRPR activation causes the proliferation of many cancer cell lines, GRP has been presumed to act as a clinically significant growth factor. Yet GRP has not been shown to be expressed by colon cancers in humans nor has the effect of GRP and/or GRPR coexpression on tumor behavior been investigated. We therefore determined GRP and GRPR expression by immunohistochemistry in 50 randomly selected colon cancers resected between 1980 and 1997, all 37 associated lymph node and liver metastases, and 20 polyps. Tumor sections studied were those that contained the margin and adjacent nonmalignant epithelium. Overall, 84% of cancers aberrantly expressed GRP or GRPR, with 62% expressing both ligand and receptor, whereas expression was not observed in adjacent normal epithelium. Consistent with the previously established mitogenic capabilities of GRP, tissues coexpressing GRP and GRPR were more likely to express proliferating cell nuclear antigen than tissues not expressing both ligand and receptor. Yet GRP/GRPR coexpression was seen with equal frequency in stage A as in stage D cancers and was only detected in 1 in 37 metastases. Furthermore, Kaplan-Meier analysis did not reveal any difference in patient survival between those whose tumors did or did not express GRP/GRPR. In contrast, GRP/GRPR coexpression was found in all well-differentiated tumor regions, whereas poorly differentiated tissues never coexpressed GRP/GRPR. Overall, these data indicate that, although GRP is a mitogen, it is not a clinically significant growth factor in human colon cancers. Rather, the strong association of GRP/GRPR coexpression with tumor differentiation raises the possibility that these proteins primarily act in vivo as morphogens.

Electrophysiological characterization of human distal colon epithelium isolated using a novel technique.

Electrophysiological studies of human colonic epithelia traditionally have been hampered by the lack of tissue availability and by poor tissue quality. Human colonic epithelium is usually obtained surgically from individuals with underlying disease, while surgery itself can injure or alter the resected tissue. As a result, a wide range in electrophysiological parameters is reported in previous studies of human colonic epithelium. Such factors may also account for differences in measurements between humans and the few other species studied. We therefore devised a novel and rapid endoscopic technique, endoscopic mucosal resection (EMR), that allows for the removal and study of intestinal mucosal epithelium from normal volunteers. Using EMR we rapidly (7.2+/-2.4 min) isolated surgical-sized epithelial sheets from the distal colon (1.4+/-0.4 by 1.3+/-0.4 cm) that were readily mounted in a 0.64-cm2 Ussing chamber. We observed stable resistance (289+/-30 omega cm2), potential difference (1.6+/-0.6 mV), and I(SC)(24+/-9 microA/cm2) for at least 90 min, after which all experiments were terminated. Exposure to carbachol increased I(SC)2.2+/-0.5-fold, while forskolin increased I(SC) 4.4+/-0.5-fold. These data show that the electrophysiological characteristics of the human distal colon removed by EMR more closely approximate values reported for other mammals than when removed using other techniques. Thus EMR represents a significant advance over traditional techniques for isolating human tissues and will increase the availability of this tissue for future studies.