The relation of serum anti-(GalNAc beta) and -para — forssman disaccharide IgG levels to the progression and histological grading of gastrointestinal cancer

Earlier we found two unusual IgG-antibody specificities to GalNAc? and GalNAcβ1-3GalNAcβ (Para — Forssman disaccharide, PFdi) carbohydrate ligands in human serum. The aim of the study was to evaluate whether elevated antibody levels are related to the progression of gastrointestinal cancer and the histopathological grading. Methods: Specific IgG levels were tested in 159 patients with gastric cancer, 88 patients with colorectal cancer and 96 blood donors by the ELISA using synthetic polyacrylamide (PAA) conjugates, GalNAcβ-PAA and PFdi-PAA. Biochemical and haematological analyses were performed using automatic equipment. Results: The anti-PFdi IgG levels were significantly higher in patients with gastric and colorectal cancer than in donors: in stages II–IV, P = 0.0002 – 0.04 (U-test). The elevated anti-PFdi IgG level was associated with the advanced gastric cancer: in stages II, III, IV vs stage I (P = 0.004 – 0.06) and in case of the tumor size T2 + T3 vs T1 (stages I, II; P = 0.03). Differences in anti-GalNAcβ IgG level were insignificant. No relation between antibody levels and the regional and distant metastases of gastric or colorectal cancer was found. The lower anti-GalNAcβ IgG level was associated with lower-differentiated carcinomas (P = 0.01 – 0.04). Prolonged postoperative changes in the levels of both antibodies during the follow-up were established. An elevation of both antibody levels in patients with gastrointestinal cancer was revealed after a surgical removal of G3-tumors (P = 0.003 – 0.01). The anti-PFdi IgG levels correlated with the levels of the C-reactive protein: r = 0.50, P = 0.003. The anti-GalNAcβ IgG levels correlated with the percentage of peripheral blood monocytes: r = 0.42, P = 0.002. Conclusion: The association of the anti-PFdi IgG level with cancer progression suggests the implication of antibodies in the pathogenesis of gastrointestinal cancer. Further studies are required to identify natural targets of antibodies, their relation to other diseases, prognostic significance in cancer.В сыворотке крови человека ранее выявлены IgG-антитела к углеводородным лигандам GalNAcβ и GalNAcβ1-3GalNAcβ (дисахарид Пара — Форсманна, PFdi). Цель исследования — выяснение связи повышенного уровня антител с прогрессией и гистологическими особенностями опухолей желудочно-кишечного тракта. Методы: обследовали 159 больных раком желудка, 88 — раком кишечника и 96 здоровых людей — доноров крови. Уровень специфических IgG-антител определяли методом ИФА с использованием синтетических конъюгатов полиакриламида (ПАА), GalNAcβ-ПАА и PFdi-ПАА. Уделение биохимических и гематологических показателей проводили с использованием автоматических анализаторов. Результаты: уровень анти-PFdi IgG у пациентов со злокачественными новообразованиями в стадиях II–IV (P = 0,0002 – 0,04) достоверно выше, чем у доноров. Повышенный уро вень анти-PFdi IgG ассоциировался с прогрессией рака желудка (в стадиях II–IV по сравнению со стадией I P = 0,004 – 0,06) и увеличением размера опухоли (T2 + T3 по сравнению с T1, стадии I, II; P = 0.03). Различия в уровнях анти- β IgG незначительные. Не выявлено взаимосвязи между уровнем антител и метастазированием опухолей желудка и кишечника. Более низкий уровень анти- β IgG определяли у больных с менее дифференцированными опухолями (P = 0,01 – 0,04). В период послеоперационного наблюдения отмечали изменения уровня обоих типов антител. После хирургического удаления опухолей органов желудочно-кишечного тракта (стадия дифференцировки G3) повышался уровень как анти-PFdi IgG, так и анти- βIgG (P = 0,003 – 0,01). Уровень анти-PFdi IgG коррелировал с уровнем С-реактивного белка (r = 0,50, P = 0,003), а уровень анти-GalNAcβ IgG — с относительным количеством моноцитов в периферической крови (r = 0,42, P = 0,002). Выводы: установлена зависимость уровня анти-PFdi IgG от опухолевой прогрессии, что подтверждает их участие в патогенезе злокачественных новообразований органов желудочно-кишечного тракта. Необходимы дальнейшие исследования по определению природных мишеней для антител, роли в развитии других заболеваний и прогностическом значении при онкологической патологии

The level of anti-(GalNAc beta) and anti-para-Forssman disaccharide igg antibodies in patients with gastrointestinal cancer: relation to survival

Serum anti-(GalNAc beta) and anti-para-Forssman disaccharide (PFdi, GalNAc beta1 - 3GalNAc beta) IgG levels were earlier found to be related to histological grading and progression of gastrointestinal cancer. Aim - to study the relation of serum antibodies level to survival in patients with gastrointestinal cancer. Methods: The level of anti-GalNAc beta, and PFdi IgG was analysed in the serum of patients with gastric (n = 78) and colorectal (n = 48) cancers in the long-term follow-up using ELISA with polyacrylamide glycoconjugates. Survival rate and hazard ratio (HR) were assessed by the Kaplan - Meier method and Cox univariate analysis in different pathomorphological groups. Better survival was observed in patients with an increased preoperative level of GalNAc beta antibodies. These were the gastrointestinal group in stages II, III or tumors T2 - 4 (n = 90 - 104, P = 0,007, HR = 0,48 - 0,49, 95 % CI 0,27 - 0,83, and the group with gastric cancer in stages I, II (n = 49, P = 0,051, HR = 0,39, 95 % CI 0,14 - 1,04). The survival time was significantly longer in the gastrointestinal group in patients whose GalNAc beta antibodies level rose in dynamics (stage III or N1 - 2: P = 0,031 - 0,039, HR = 0,29 - 0,31, 95 % CI 0,09 - 1,00). No significant difference in survival of patients was observed in the evaluation of PFdi antibodies. We suggest that the level of antibodies and its change reflect the enteric microbiota colonization, which may influence cancer progression via different interrelations between microbiota, the tumor and immune system. Key Words: gastrointestinal cancer, survival, GalNAc beta and para-Forssman disaccharide antibodies, enteric bacteria

The characterization of IgG antibodies to GalNAc beta-terminated glycans of gastric cancer survivors

The elevated anti-GalNAcβ IgG level of serum was shown to be associated with the significantly better survival of patients with gastrointestinal cancer. Aim: To characterize the specificity of IgG antibodies to GalNAcβ-terminated glycans of long-term gastric cancer survivors. Methods: Serum antibodies and affinity-isolated antibodies were analysed by the indirect and competitive ELISA using glycan-polyacrylamide (PAA) conjugates as well as by isoelectric focusing and Western blotting. Results: In the serum probes, a partial cross-reactivity of antibodies to GalNAcβ, GalNAcβ1–3Galβ (X2di), GalNAcβ1–3GalNAcβ (PFdi) and GlcNAcβ was observed. The isolated anti-GalNAcβ IgGs demonstrated the cross-reactivity to the X2di glycan mainly. The affinity of the X2di-PAA to anti-GalNAcβ IgGs was 11–21 times lower than that of the GalNAcβ-PAA. Anti-X2di and anti-PFdi IgGs demonstrated monoreactivity to their key glycans-PAA used in isolation. The IC50 values of key glycoconjugates ranged from 1 to 5 · 10⁻⁷ M. No polyreactivity of antibodies to the unrelated antigens (ferritin, casein and DNA) was found. The polyclonal or oligoclonal distribution of IgG bands was established and the monoreactivity of antibodies was not associated with the clonal distribution of bands. Conclusion: The cross-reactivity of anti-GalNAcβ antibodies to X2di and related glycans deserves attention in the clarification of the role of antibodies in cancer progression and enhancement of the prognostic potential in the combined determination of antibody markers. Key Words: gastric cancer, glycan-polyacrylamide, anti-GalNAc-beta, para-Forssman, X2, oligoclonal IgG, microbiota