Prevalence of high affinity naturally occurring IgG2 antibodies against human chorionic gonadotropin and its subunits in patients with ovarian cyst.

Naturally occurring antibodies to tumour antigens are gaining interest as clinically important cancer biomarkers for early diagnosis, prognosis and for the development of anti-cancer therapeutics. The glycoprotein αß heterodimer hormone human chorionic gonadotropin (hCG) and its ß subunit (hCGß) are produced by various cancers, and their increased serum levels correlate with poor prognosis. We have previously reported that patients with benign ovarian cysts, but not the malignant tumours, were characterized by augmented serum levels of naturally-occurring IgG antibodies to hCG and hCGß. Here we further characterise these antibodies in patients with ovarian cysts. IgG and IgM antibody binding to whole hCG, hCGß, hCGα, hCGß C-terminal peptide (hCGßCTP), and the hCGß core fragment (hCGßCF) were measured in the sera from 36 patients with ovarian cysts and 12 healthy non-pregnant women using a standard ELISA. IgG subclass usage and affinity was also determined together with cross-binding to whole hCG and its subunits of four selected commercial monoclonal antibodies generated against ovarian cyst mucins. Our results showed that 91.7% of the sera tested contained elevated IgG, but not IgM antibodies to one or several antigens, with an overwhelming prevalence of high affinity IgG2 indicating their binding to carbohydrate epitopes and possibly ovarian cyst mucins. Anti-mucin commercial antibody ab212418 (Abcam) produced against Gal1-3GalNAc, exhibited strong cross-binding to hCGαß, hCGß, hCGα and hCGßCTP. The protective anti-cancer potential of these antibodies will be further investigated and could lead to the development of novel treatment strategies for ovarian cancer.

ELEVATED LEVELS OF NATURALLY OCCURRING AUTOANTIBODIES TO HUMAN CHORIONIC GONADOTROPIN Β CORE FRAGMENT IN A FEMALE PATIENT WITH THYROID FOLLICULAR ADENOMA: CASE REPORT.

We report a case of a woman, who had an elevated levels of naturally-occurring autoantibodies to human chorionic gonadotropin (hCG) ß core fragment ( hCGßcf) one year prior to the development of thyroid follicular lesion. The patient underwent surgery and the histology report demonstrated that the lesion was a follicular adenoma. Further investigations of the role of naturally-occurring autoantibodies (NAAbs) to anti-hCGßcf in the pathogenesis of various tumours of thyroid gland might be useful in the development of novel diagnostic methods, using anti-hCGßcf NAAbs as a marker for the detection of unsuspected thyroid tumour.

NATURALLY-OCCURRING AUTOANTIBODIES TO HUMAN CHORIONIC GONADOTROPIN AND ITS SUBUNITS IN OVARIAN CYST PATIENTS.

Growing evidence supports the existence of immune-surveillance mechanisms in ovarian tumour patients, including autoantibodies to tumour associated and tumour specific antigens, tumour growth factors. Glycoprotein hormone human chorionic gonadotropin (hCG) and its hormone-specific hCGß subunit have been associated with epithelial tumours such as bladder, lung, oral/facial, breast, cervical, ovarian, vaginal, prostate, renal and pancreatic carcinomas. It is believed that hCG plays a role of an autocrine growth factor for tumor cells. Here we have demonstrated that sera of patients with ovarian cyst contain naturally-occurring autoantibodies, predominantly of IgG2 isotype, that bind to hCG and its subunits with high affinity. Titration of blood sera from 36 female patients, aged 22-61 after ethical permission and informed consent, diagnosed with ovarian cyst and healthy age-matched controls (n=12) was performed using a classical enzyme-linked immunosorbent assay (ELISA). Binding of the sera to the following antigens was tested: hCGαß, hCGß, hCGα, hCGß C-terminal peptide (hCGßCTP) and hCGß core fragment (hCGßCF). The same type of ELISA (with necessary modifications) was used for further investigation of subclass usage and assessment of binding affinity of the detected autoantibodies. Our data indicates that the sera of the majority of patients with ovarian cyst contain significantly higher levels of the natural IgG antibodies binding to hCGαß, hCGß, hCGα, hCGßCTP and hCGßCF, than those of the healthy controls. Natural IgG antibodies to hCGαß heterodimer were detected in 78% of cases, to hCGß in 61% of cases, to hCGα in 78% of cases, to hCGßCTP in 69% of cases, to hCGßCF in 83% of cases. These autoantibodies predominantly belonged to the IgG2 subclass and were characterized by the high binding affinity. It is plausible that they cross- bind to sugar side chains of hCG and its subunits. Our data demonstrated that sera of patients with the ovarian cyst contains elevated levels of naturally-occurring IgG antibodies, which bind to hCG and/or its subunits. The overwhelming majority of these autoantibodies belong to the IgG2 isotype thus indicating that they might be directed against carbohydrate antigens within highly glycosylated hCG.

Investigation of factors influencing the immunogenicity of hCG as a potential cancer vaccine.

Human chorionic gonadotrophin (hCG) and its ß-subunit (hCGß) are tumour autocrine growth factors whose presence in the serum of cancer patients has been linked to poorer prognosis. Previous studies have shown that vaccines which target these molecules and/or the 37 amino acid C-terminal hCGß peptide (hCGßCTP) induce antibody responses in a majority of human recipients. Here we explored whether the immunogenicity of vaccines containing an hCGß mutant (hCGßR68E, designed to eliminate cross-reactivity with luteinizing hormone) or hCGßCTP could be enhanced by coupling the immunogen to different carriers [keyhole limpet haemocyanin (KLH) or heat shock protein 70 (Hsp70)] using different cross-linkers [1-ethyl-3(3-dimethylaminopropyl)carboiimide (EDC) or glutaraldehyde (GAD)] and formulated with different adjuvants (RIBI or Montanide ISA720). While there was little to choose between KLH and Hsp70 as carriers, their influence on the effectiveness of a vaccine containing the BAChCGßR68E mutant was less marked, presumably because, being a foreign species, this mutant protein itself might provide T helper epitopes. The mutant provided a significantly better vaccine than the hCGßCTP peptide irrespective of the carrier used, how it was cross-linked to the carrier or which adjuvant was used when hCG was the target. Nonetheless, for use in humans where hCG is a tolerated self-protein, the need for a carrier is of fundamental importance. Highest antibody titres were obtained by linking the BAChCGßR68E to Hsp70 as a carrier by GAD and using RIBI as the adjuvant, which also resulted in antibodies with significantly higher affinity than those elicited by hCGßCTP peptide vaccine. This makes this mutant vaccine a promising candidate for therapeutic studies in hCGß-positive cancer patients.

[Alpha-dephenzine concentration in peripheral blood of patients diagnosed with schizophrenia].

Over the past few decades the search for predisposed biological markers has become the course of most priority in scientific psychiatry. Research goal was to reveal the association of alpha-dephenzine concentration in peripheral blood with the disease existence and its progressing (acute and chronic forms) in patients with schizophrenia. In-patients diagnosed with schizophrenia have been investigated (av. age - 45.33±1.86), among them 26 males (av. age - 39.41±1.86) and 13 females (av. age - 45.04±2.44). 27 patients with acute form of schizophrenia (av.age - 44.88±2.42), 12 patients with a chronic form (av. age - 46.33±2.75). 15 healthy volunteers have been investigated as well (av.age - 40.03±2.94), 5 males and 10 females among them. The study of alpha-dephenzine concentration by the immunoenzyme method has been implied in the investigation. Research studies revealed statistically valid increased concentration of alpha-dephenzine in patients diagnosed with schizophrenia comparing with healthy volunteers. The difference was so evident that qualitative difference (marker existence-nonexistence) would rather be considered than quantitative one. Relying on the results of studies we can estimate alpha-dephenzines as possible biomarkers of schizophrenia.