Helicobacter pylori Status May Differentiate Two Distinct Pathways of Gastric Adenocarcinoma Carcinogenesis

Abstract

Background: We evaluated the phenotype of sporadic gastric cancer based on HP status and binding of a tumor risk marker monoclonal, Adnab-9.

Methods: We compared a familial GC kindred with an extremely aggressive phenotype to HP-positive (HP+) and -negative (HP-) sporadic gastric adenocarcinoma (GC) patients in the same community to determine if similar phenotypes exist. This might facilitate gene discovery to understand the pathogenesis of aggressive GC phenotypes, particularly with publications implicating immune-related gene-based signatures, and the development of techniques to gauge the stance of the innate immune system (InImS), such as the FERAD ratio (blood ferritin:fecal Adnab-9 binding OD-background binding). Resection specimens for the sporadic and familial group were stained for HP and examined for intestinal metaplasia (IM) and immunostaining for Adnab-9. Familial kindred specimens were also tested for the E-cadherin mutation and APC (adenomatous polyposis coli). Survival was evaluated.

Results: Of 40 GC patients, 25% were HP+ with a greater proportion of intestinal metaplasia (IM) and gastric atrophy than the HP- group. The proband of the familial GC kindred, a 32-year-old mother with fatal GC, was survived by 13-year-old identical twins. Twin #1 was HP- with IM and Twin #2 was HP+. Both twins subsequently died of GC within two years. The twins did not have APC or E-cadherin mutations. The mean overall survival in the HP+ sporadic GC group was 2.47 ± 2.58 years and was 0.57 ± 0.60 years in the HP- group (p = 0.01). Survival in the kindred was 0.22 ± 0.24 years. Adnab-9 labeling was positive in fixed tissues of 50% of non-familial GC patients and in gastric tissue extract from Twin #2. The FERAD ratio was determined separately in six prospectively followed patient groups (n = 458) and was significantly lower in the gastric cancer patients (n = 10) and patients with stomach conditions predisposing them to GC (n = 214), compared to controls (n = 234 patients at increased risk for colorectal cancer but without cancer), suggesting a failure of the InImS.

Conclusion: The HP+ sporadic GC group appears to proceed through a sequence of HP infection, IM and atrophy before cancer supervenes, and the HP- phenotype appear to omit this sequence. The familial cases may represent a subset with both features, but the natural history strongly resembles that of the HP- group. Two different paths of carcinogenesis may exist locally for sporadic GC. The InImS may also be implicated in prognosis. Identifying these patients will allow for treatment stratification and early diagnosis to improve GC survival.

Keywords: Adnab-9; FERAD ratio; H. pylori; familial gastric cancer; monoclonal antibody.

Figure 1

Adnab-9 binding via ELISA A. (A) bar diagram shows the Adnab-9 tissues binding in Twin #2 and APC patients. Results expressed in terms of percent of positive control in Adnab-9 binding. The Adnab-9 level standardized for protein was 2 times higher that of the APC patient and 2.7 times higher that of the control. (B) A scattergram shows the Adnab-9 stool ELISA in Twin #2 and Chinese gastric cancer patients. Results are expressed in terms of optical density. When the code was broken for the stool samples, the binding in specimens from cancer patients was greater than the controls; however, the level in Twin #2 was five times greater than the mean level of stools from the five patients with gastric cancer and much greater than five patients in the negative control group.

Figure 2

Kaplan–Meier curve depicts survival cases. (A) Overall survival was significantly longer in the HP+ group than in the HP− group (p < 0.005; CI 1.4–6.17); all above parameters analyzed via univariate testing using the log rank test). (B) Atrophy (p = 0.04); (C) metaplasia (0.039); (D) race (p = 0.029). (A–D) is depicted below showing Kaplan-Meier mortality curves for Hp status (A); gastric atrophy (B); (IM) intestinal metaplasia (C); and African Americans (AA—D).

Figure 3

A silver stain of the gastric mucosa taken at surgery from Twin #1. Despite being HP-negative, there is marked intestinal metaplasia present. Magnification was 100×.

Figure 4

Adnab-9 labeling of sporadic gastric cancer. (A) This 70-year-old patient had HP + chronic atrophic gastritis and intestinal metaplasia with stage 4 intestinal-type gastric adenocarcinoma. Adnab-9 staining was 4+ in the deep glands in the vicinity of the cancer. The substrate was EAC, imparting a reddish-brown color. Magnification was 100×. (B) This 40-year-old patient had HP− chronic atrophic gastritis and stage 4 diffuse-type gastric adenocarcinoma. Adnab-9 staining was 3+ in the cytoplasm of very large cancer cells (arrows). The substrate was EAC, imparting a reddish-brown color. Magnification was 400×.

Figure 5

E-cadherin labeling in Twin #1 showed intense labeling in both the cancerous and corresponding benign tissue. (A) Diffuse cytoplasmic staining in low power of cancer tissue. The substrate was diaminobenzidine (DAB), imparting a golden-brown color. Magnification was ×75. (B) Membrane staining in hyperplastic glands in high power. This suggests that there was no mutation of E-cadherin in the kindred. The substrate was diaminobenzidine (DAB), imparting a golden-brown color. Magnification was ×100.

Figure 6

A bar diagram depicting FERAD ratios in each patient group. The premorbid FERAD ratios are the lowest for GC and tend to be less than either HP+ or HP− groups (p = 0.08; Mann–Whitney) as summarized in Table 3.