Human Antibodies

The prevalence of antibodies to human platelet antigens (HPA) and human leukocyte antigens (HLA) class 1 antigens among Nigerian pregnant women has not been reported in our country. This study was therefore aimed at screening the obstetric population for evidence of alloimmunization due to human platelet and HLA class 1 antigens. One hundred and forty four (144) pregnant women attending the obstetric clinic of Military Hospital, Port Harcourt, participated in the study. Their sera were tested for antibodies to HPA and HLA class 1 antigens using GTI PakPlus solid phase ELISA Kit. The total prevalence rate of antibody production was 60.5% (87 out of 144). Among the positive samples, 60 had platelet glycoprotein specific antibodies (41.7%) and 27 had HLA class 1 antibodies (18.8%). In 39.6% of the pregnant women, both platelet specific antibodies and HLA class 1 antibodies appeared. The prevalence of platelet specific glycoprotein antibodies were obtained as follows: GP 11b/111a 12 (8.3%), GP 1a/11a 35 (20.8%), GP Ib/IX 18 (12.5%) and GP IV 9 (6.3%). The prevalence of each platelet antibody subgroup was obtained as follows: anti-HPA-1a,-3a,-4a (4.2%), anti-HPA-1b,-3b,-4a (4.2%), anti-HPA-30 5a and anti-GP Ib/IX (12.5% each), anti-HPA-5b (8.3%) and anti-GP IV (6.3%). A high prevalence rate of human platelet arid cytotoxic antibodies has been observed in our obstetric population. There is need to establish platelet serology laboratory for the proper antenatal and postnatal management of pregnant mothers in this region.

Laboratory screening for the diagnosis of hepatitis B virus (HBV) infection in blood donors currently consists of testing for hepatitis B surface (HBsAg) antigen alone. The prevalence of isolated anti-HBc-IgM is not yet known in the semi arid region of Nigeria. The major objective of this study was to determine the sero-prevalence of antibody to hepatitis B core antigen (anti-HBc-IgM) and other infectious agent markers; HBsAg, HCV, HIV and Syphilis among blood donors in the North Eastern region of Nigeria. In a cross sectional study from October 2010 to January 2011, 266 blood donors were tested for the infectious disease markers using standard ELISA procedures as contained in the manufacturer's standard operating procedures. The prevalence of various infectious markers obtained were as follows: HBsAg (8.6%); anti-HCV (1.5%); HIV (2.6%) and anti-HBc-IgM(18.4%). There was a zero percent prevalence of Syphilis in this donor population. The proportion of isolated anti- HBc-IgM antibody obtained was 18.1%. Performance indices for HBsAg were as follows: Sensitivity (10.2%), specificity (91.7%), positive predictive value (PPV) (21.7%), Negative predictive value (81.9%), and efficiency (76.7%). The prevalence of anti-HBc- IgM antibody was higher among first time blood donors (21.4%), and in some ethnic groups. There is a high prevalence of isolated anti-HBc-IgM antibody among blood donors in Maiduguri. The sensitivity of HBsAg was found to be very low and as such many recent HBV infections may be missed during pre-transfusion screening. The use of anti-HBc-IgM screening as a mandatory pre-transfusion screening test is hereby advocated.

PAR-2 belongs to a family of G-protein coupled Protease-Activated Receptors (PAR) which are activated by specific proteolytic cleavage in the extracellular N-terminal region. PAR-2 is activated by proteases such as trypsin, tryptase, proteinase 3, factor VIIa, factor Xa and is thought to be a mediator of inflammation and tissue injury, where elevated levels of proteases are found. Utilizing the HuCAL GOLD® phage display library we generated fully human antibodies specifically blocking the protease cleavage site in the N-terminal domain. In vitro affinity optimization resulted in antibodies with up to 1000-fold improved affinities relative to the original parental antibodies with dissociation constants as low as 100 pM. Corresponding increases in potency were observed in a mechanistic protease cleavage assay. The antibodies effectively inhibited PAR-2 mediated intracellular calcium release and cytokine secretion in various cell types stimulated with trypsin. In addition, the antibodies demonstrated potent inhibition of trypsin induced relaxation of isolated rat aortic rings ex vivo. In a short term mouse model of inflammation, the trans vivo DTH model, anti-PAR-2 antibodies showed inhibition of the inflammatory swelling response. In summary, potent inhibitors of PAR-2 were generated which allow further assessment of the role of this receptor in inflammation and evaluation of their potential as therapeutic agents.

Background: Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular dysplasia and severe, recurrent epistaxis is a common clinical phenotype associated with HHT. An intranasal treatment regime of diluted Avastin™ (Bevacizumab; recombinant humanized anti-vascular epithelial growth factor immunoglobin G1) using apulsatile nasal irrigator has proven efficacious in clinical practice. However, concerns regarding the stability of Avastin™ following dilution and prolonged storage in standard containers used for drug delivery, such as polyethylene bottles, have so far prevented a more widespread clinical use. Compatibility with the preservative benzalkonium chloride was also unknown.

Objective: This study aimed at determining, whether dilution, prolonged refrigerated storage and the presence of the preservative benzalkonium chloride – as required for novel Avastin™ formulations – affected the biochemical and electrochemical properties of the drug.

Methods: We performed a detailed biochemical and electrochemical analysis of Avastin™, including native and sodium dodecyl sulfate polyacrylamide gel electrophoresis, enzyme-linked immunosorbent assay and isoelectric focusing.

Results: We did not detect any evidence of degeneration or aggregation following dilution and prolonged, refrigerated storage or from the presence of benzalkonium chloride. All biochemical and electrochemical properties of Avastin™ after dilution and prolonged, refrigerated storage were undistinguishable from control.

Conclusions: Our data provide important insight into the stability of Avastin™ and allow the consideration of novel Avastin™ formulations, including its use in a metered-dose nasal spray for the treatment of HHT and other applications.