anti-P Antibodies Research Articles

Anti-Prothrombin Antibodies are Associated with Adverse Pregnancy Outcome

Women with antiphospholipid antibodies (aPL) such as lupus anticoagulant, anticardiolipin antibodies, and anti-β(2) glycoprotein-1 antibodies are at high risk of late pregnancy complications, such as severe pre-eclampsia, placental insufficiency, and fetal loss. It has been observed that aPL consists of a heterogeneous group of antibodies targeting several phospholipid-binding plasma proteins, including also anti-prothrombin (anti-PT), anti-protein S (anti-PS), and anti-protein C (anti-PC) antibodies. Their potential role in late pregnancy complications is not known. The aim of this work was to investigate the association between those autoantibodies and histories for adverse pregnancy outcome. Anti-PT, anti-PS, and anti-PC antibodies were evaluated in 163 patients with previous severe pre-eclampsia, fetal death, and/or placental abruption and in as many women with previous uneventful pregnancies, negative for aPL. The prevalence of anti-PT antibodies was higher in cases than in controls (OR, 95% CI: 10.92, 4.52-26.38). The highest prevalence was observed in subjects with fetal death. Anti-PT antibodies appear to be associated with adverse pregnancy outcome, irrespectively of aPL.

Abstract 621: Targeting phosphatidylserine to improve androgen deprivation therapy of prostate cancer

Abstract Androgen deprivation therapy (ADT) is effective against androgen-dependent prostate cancer, but patients usually relapse from the outgrowth of androgen-independent disease. Here, we tested the hypothesis that relapse can be prevented by combining ADT with treatment with a monoclonal antibody that binds exposed phosphatidylserine (PS). The rationale behind this hypothesis is that ADT will create hypoxia which induces PS exposure on the tumor vascular endothelial cells, leading to immune cell attack on tumor vasculature, collapse of the vessels and death of hormone-resistant and hormone-sensitive tumor cells through deprivation of oxygen and nutrients. In addition, the antibody should block the immunosuppressive effects of PS and induce reactivation of T-cell immunity. To test this hypothesis, we used ADT and anti-PS antibodies (2aG4 or murine chimeric 1N11) to treat mice bearing hormone-sensitive prostate tumors. The tumor models were: i) immunocompromised mice bearing human LNCaP prostate tumors; ii) transgenic (TRAMP) mice which spontaneously develop adenocarcinoma of the mouse prostate. Castration induced exposure of PS on blood vessels and tumor cells in both models. Vascular PS exposure corresponded with hypoxia, as judged by pimonidazole staining. In the LNCaP model, castration alone delayed tumor growth but all animals relapsed with hormone-independent disease, whereas mice treated with castration plus anti-PS antibody did not relapse. In TRAMP mice castrated at 15-weeks of age, only 54% (7/13) of mice treated with castration alone survived beyond 40-weeks of age, as compared with 92% (12/13) of mice treated with castration and 1N11. No toxicity to the mice was caused by the antibody treatment. Tumor blood flow and perfusion were markedly reduced in mice treated with castration and anti-PS antibody. Histological studies revealed extensive disruption of tumor vasculature and abundant infiltration of immune cells, principally M1 macrophages. These results demonstrate that PS becomes exposed on prostate cancer tumor vasculature and tumor cells in response to hypoxia caused by ADT in the tumor microenvironment. The exposed PS could serve as a target to enhance the efficacy of ADT against prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 621. doi:10.1158/1538-7445.AM2011-621

Autoantibodies targeting phosphatidylserine, cardiolipin, β2-glycoprotein I, annexin V and prothrombin in patients with recurrent spontaneous miscarriage (148.18)

Abstract An association between human reproductive failure and the presence of anti-phospholipid autoantibodies (APA) was suggested; both traditional and nontraditional APAs were associated with increased risk of recurrent spontaneous miscarriage (RSM). We measured levels of antibodies to phosphatidylserine (anti-PS), cardiolipin (ACA), β2-glycoprotein-I (anti-β2GPI), annexin V (anti-Anx V) and prothrombin (anti-PT) in 277 women with idiopathic RSM, and 288 control women by ELISA. Anti-PS IgG, ACA IgM/IgG, anti-Anx V IgM/IgG, and anti-PT IgM positivity were significantly associated with higher RSM risk, after controlling for age, BMI, menarche, and gravida. RSM was independently associated with high levels of ACA IgM (OR, 6.5; 95% CI, 2.5-17.1) and IgG (OR, 12.1; 95% CI, 6.6-22.2), anti-PS IgG (OR, 13.3; 95% CI, 6.4-27.4), anti-AnxV IgM (OR, 31.1; 95% CI, 15.7-56.6) and IgG (OR, 23.3; 95% CI, 8.8-51.7), and anti-PT IgM (OR, 2.8; 95% CI, 1.3-5.9). Levels of autoantibodies were categorized into 5 strata (percentiles 1-75, 76-99, 91-95, 96-99, >99 among control patients) and analyzed in logistic regression models. The adjusted OR increased as the percentile value of ACA IgM/IgG ,anti-PS IgG and anti-Anx V IgM/IgG, and anti-PT IgM, increased. In conclusion, the presence of ACA IgM/IgG, anti-Anx IgM/IgG, anti-PS IgG, and anti-PT IgM antibodies are risk factors for RSM. The strongest association was seen for anti-Anx V IgM and IgG, followed by ACA IgG, and anti-PS IgG antibodies.

High serum levels of rheumatoid factor and anti-phosphatidylserine antibody in patients with ischemic heart disease.

Immunopathological and inflammatory processes play important roles in the initiation and development of Ischemic Heart Disease (IHD). The aim of this study was to evaluate the serum levels of several autoantibodies including rheumatoid factor (RF), anti-nuclear antibodies (ANA), anti-small nuclear ribonucleoprotein (anti-Sm), anti-phosphatidylserine (anti-PS) and anti-cardiolipin (anti-CL) antibodies in patients with IHD. A total of 120 patients with IHD with acute myocardial infarction (AMI; n=60) or unstable angina (UA; n=60) and 60 sex- and age-matched healthy subjects were enrolled in this study. Serum samples of participants were tested for the ANA, anti-Sm, anti-PS and anti-CL antibodies by ELISA. Serum level of RF was measured by a turbidometric method. The mean serum levels of RF and anti-PS antibodies in AMI group and UA group were significantly higher than those observed in the control group (p<0.0001). The mean serum levels of RF and anti-PS antibodies in AMI patients were significantly higher than the UA group (p<0.01 and p<0.001, respectively). The mean serum levels of RF in men with AMI or UA diseases were significantly higher as compared to healthy control men (p<0.0001 and p<0.003, respectively). The differences of the serum levels of ANA, anti-Sm and anti-CL antibodies were not significant between AMI, UA and the control groups. There was no difference in the serum levels of RF, ANA, anti-Sm, anti-PS or anti-CL antibodies in patients with traditional risk factors, including hypertension, dyslipidemia, diabetes and smoking, and those without a certain risk factor. Higher serum levels of RF and anti-PS antibody in patients with IHD may be considered as independent risk factors for IHD.

Failure of eculizumab to correct paroxysmal cold hemoglobinuria

Dear Editor, Paroxysmal cold hemoglobinuria (PCH) is a rare cause of intravascular hemolytic anemia in which an antibody against the P antigen develops and hemolysis ensues via a pathway involving thermally dependent complement binding. Previously an association of syphilis due to crossreactivity of antibody, PCH has since predominantly become a disease of childhood following varicella-zoster virus infection. The childhood form is typically acute, selflimiting, and steroid-responsive. Rare cases of PCH in adults have been described in association with chronic lymphocytic leukemia and lymphoma but never previously with myeloma [1]. We describe a case of steroid-refractory PCH in a patient with myeloma that was only partially corrected with eculizumab, a monoclonal antibody against C5 complement. A 51-year-old Nigerian man with Stage III ISS IgA kappa multiple myeloma in very good partial response following treatment with bortezomib, thalidomide, cyclophosphamide, and dexamethasone was admitted to our hospital with severe hemolytic anemia. At admission, lactate dehydrogenase (LDH) was elevated at 543 U/L (100–200 U/L); bilirubin, 164 μmol/L (<20 μmol/L); and hemoglobin was 60 g/L (125–175 g/L). Initial transfusion requirement were 2 to 4 U of red blood cells per day to achieve a target hemoglobin 60–80 g/L. PCH was diagnosed with the Donath–Landsteiner test following positive urinary hemosiderin on day 21 of admission. No complement C3 was detected on his red cells at diagnosis. Blood film was significant for rosette formation of erythrocytes around neutrophils and erythrophagocytosis by neutrophils (Fig. 1). Syphilis serology and cerebrospinal fluid treponemal PCR were negative. Being refractory to oral prednisolone and cyclophosphamide (1 g/m), he received standard induction with eculizumab 600 mg weekly for 4 weeks [2]. Urinary hemosiderin became negative by day 36; however, hemolysis continued albeit at reduced rate, and was finally controlled following a single dose of intravenous cyclophosphamide 4 g/m. The patient became transfusionindependent 31 days post high-dose cyclophosphamide and hemoglobin returned to normal reference range (136 g/L) at 10 weeks, with normalization of LDH at 9 weeks. This is the first reported case of eculizumab use for treatment of PCH. Failure of eculizumab to control hemolysis in this patient suggests that extravascular clearance of red cells in PCH may be induced by earlier components of complement and opsonization with anti-P antibodies. Moreover, the observation of neutrophil erythG. P. Gregory (*) : S. Opat :H. Quach : J. Shortt :H. Tran Department of Clinical Hematology, Monash Medical Centre, Clayton, VIC, Australia e-mail: g.gregory@alfred.org.au

Single anti-P ribosomal antibodies are not associated with lupus nephritis in patients suffering from active systemic lupus erythematosus

Background In clinical practice, it is sometimes difficult to diagnose a relapse in patients suffering from systemic lupus erythematosus (SLE) and lupus nephritis (LN) having potential complications, including renal failure and death. Some immunological markers can help to determine their association with LN and, therefore, diagnose the early onset of complications. Objectives Evaluating the association between systemic and/or kidney activity and anti-P ribosomal and anti-dsDNA antibodies in patients suffering from active SLE. Methods 389 patients were evaluated, 140 of whom were subsequently included in the study. The patients were divided into two groups by means of case–control studies, including Colombian patients having American College of Rheumatology (ACR) classification criteria for SLE (1997). The SLE disease activity index (SLEDAI) was applied and all patients presenting an increase of 5 or more compared to their last evaluation, as well as presenting renal manifestations, were considered to be cases; all patients had an activity score. An ELISA kit and the indirect immunofluorescence method with Crithidia luciliae were used for determining the presence of anti-P ribosomal and anti-dsDNA antibodies, respectively. Results No association was found between anti-P ribosomal antibodies and LN ( p = 0.2971) but anti-P ribosomal antibodies showed association with a > 5 SLEDAI score (OR = 4.87; 1.32–17.98 95% CI; p = 0.008). The coexistence of anti-P ribosomal and anti-dsDNA antibodies was associated with LN (OR = 3.52; 1.07–13.42 95% CI; p = 0.019) and anti-dsDNA was associated with LN ( p = 0.001). Conclusion There was no association between anti-P ribosomal antibodies and LN but anti-P ribosomal antibodies coexisting with anti-dsDNA antibodies was associated with LN, thereby suggesting that the coexistence of two antibodies is nephritogenic to a greater extent. Additional studies are needed to evaluate the coexistence of kidney-specific antibodies in SLE to determine the biological nature of LN.

Use of monoclonal antibodies against Hendra and Nipah viruses in an antigen capture ELISA

BackgroundOutbreaks of Hendra (HeV) and Nipah (NiV) viruses have been reported starting in 1994 and 1998, respectively. Both viruses are capable of causing fatal disease in humans and effecting great economical loss in the livestock industry.ResultsThrough screening of hybridomas derived from mice immunized with γ-irradiated Nipah virus, we identified two secreted antibodies; one reactive with the nucleocapsid (N) protein and the other, the phosphoprotein (P) of henipaviruses. Epitope mapping and protein sequence alignments between NiV and HeV suggest the last 14 amino acids of the carboxyl terminus of the N protein is the target of the anti-N antibody. The anti-P antibody recognizes an epitope in the amino-terminal half of P protein. These monoclonal antibodies were used to develop two antigen capture ELISAs, one for virus detection and the other for differentiation between NiV and HeV. The lower limit of detection of the capture assay with both monoclonal antibodies was 400 pfu. The anti-N antibody was used to successfully detect NiV in a lung tissue suspension from an infected pig.ConclusionThe antigen capture ELISA developed is potentially affordable tool to provide rapid detection and differentiation between the henipaviruses.

Cutaneous vasculitis in systemic lupus erythematosus: association with anti-ribosomal P protein antibody and Raynaud phenomenon

Ninety-one consecutive systemic lupus erythematosus (SLE) patients (American College of Rheumatology criteria) with a history of cutaneous vasculitis were compared to 163 SLE controls without this clinical manifestation from July to December 2007 in order to determine the possible clinical and serological association of this manifestation. Data were obtained in an ongoing electronic database protocol and autoantibodies to anti-double-stranded DNA, anti-Sm, anti-RNP, anti-Ro/SS-A, anti-La/SS-B, and anticardiolipin and ribosomal P protein antibody (anti-P) were detected by standard techniques. Exclusion criteria were the presence of anti-phospholipid syndrome or antibodies, Sjögren syndrome, and a history of thrombosis. The mean age (38.5 ± 11.5 vs. 37.8 ± 11.6years, p = 0.635), disease duration (12.5 ± 7.8 vs. 11.8 ± 7.9years, p = 0.501), and frequency of white race (71.4% vs. 70.5%, p = 0.872) and female sex (96.8% vs. 93.7%, p = 0.272) were comparable in both groups. The vasculitis group had a higher frequency of malar rash (97.9% vs. 87.4%, p = 0.004), photosensitivity (91.4% vs. 81.6%, p = 0.030), and Raynaud phenomenon (RP; 27.7% vs. 7.5%, p < 0.001), whereas all other clinical manifestation including renal and central nervous system involvements were similar to the control group. Laboratorial data revealed that only anti-P (35.1% vs. 12.1%, p < 0.001) was more frequent in patients with vasculitis. In a multivariate logistic regression model, cutaneous vasculitis was associated to the presence of RP (OR = 3.70; 95% confidence interval [CI] = 1.73-8.00) and anti-P (OR = 3.42; 95% CI = 1.76-6.66). In summary, SLE cutaneous vasculitis characterizes a subgroup of patients with more RP and anti-P antibodies but not accompanied by a higher frequency of renal and central nervous system involvements.

Clinical manifestation of systemic lupus erythematosus in patients with antiribosomal P protein antibodies

Antibodies against ribosomal P protein (anti-P) are detected predominantly in patients with systemic lupus erythematosus (SLE). However, the data on their frequency and clinical relevance remain inconclusive. The aim of the study was to assess the frequency as well as clinical and serological relevance of anti-P autoantibodies in Polish patients with SLE and to determine the significance of these antibodies in the diagnosis of SLE. Anti-P antibody levels were measured in the sera of 100 SLE patients using an enzyme-linked immunosorbent assay and Western blotting. All patients underwent a routine clinical and laboratory evaluation. Disease activity was assessed using the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) score. Anti-P antibodies were detected in 14 of 100 patients. When compared to anti-P-negative patients, this group was characterized by earlier onset of SLE, higher disease activity, more frequent occurrence of fever and facial erythema, decreased serum levels of complement, and elevated levels of alanine and aspartate aminotransferases. In 2 cases, anti-P antibodies were the only serological marker of SLE detected in these patients. SLE with the presence of anti-P antibodies is characterized by an early onset and high disease activity.

Male gender results in more severe lupus nephritis

Gender may produce different characteristics in the manifestation of systemic lupus erythematosus (SLE). The present study investigated the influence of gender on clinical, laboratory, autoantibodies and histopathological classes of lupus nephritis (LN). As much as 81 patients diagnosed with SLE (ACR criteria) and active nephritis, who underwent renal biopsy between 1999 and 2004, and who had frozen serum samples and clinical data available from the time of biopsy, were selected for this study. The presence of anti-P and antichromatin antibodies was measured using ELISA, and anti-dsDNA was measured using indirect immunofluorescence. All of the renal biopsies were reviewed in a blinded manner by the same expert renal pathologist. The charts were extensively reviewed for demographic and renal features obtained at the time of the biopsy. Of the 81 patients (13.6%), 11 were male SLE patients. Both male and female lupus patients were of similar age and race, and had similar durations of lupus and renal disease. The female patients had more cutaneous (95.7 vs. 45.5%, P = 0.0001) and haematological (52.9 vs. 18.2%, P = 0.04) involvements than the male SLE patients. In addition, the articular data, central nervous system analyses, serositis findings and SLEDAI scores were similar in both experimental groups. Positivity for anti-dsDNA, anti-ribosomal P and antichromatin did not differ between the two groups, and both groups showed similarly low C3 or C4 serum levels. Our analysis indicated that no histopathological class of LN was predominant in both males and females. Interestingly, the serum creatinine levels were higher in the male SLE patients compared to the female SLE group (3.16 +/- 2.49 vs. 1.99 +/- 1.54 mg/dL, P = 0.03), with an increased frequency of high creatinine (81.8 vs. 47.1%, P = 0.04) as well as renal activity index (7.6 +/- 3.5 vs. 4.8 +/- 3.5, P = 0.02). In addition, whilst the mean levels of proteinuria, cylindruria and serum albumin were markedly altered, they were comparable between both lupus men and women. Moreover, the frequencies of dialysis, renal transplantation and death were similar between the two groups. These data suggest that male patients had a more severe LN compared to women diagnosed with this renal abnormality.

The role of antiphospholipid antibodies toward the protein C/protein S system in venous thromboembolic disease

The association between venous thromboembolism (VTE) and antibodies anti-Protein C (PC)/Protein S (PS) is still uncertain. We performed a case-control study to determine the risk of VTE related to the presence of these auto-antibodies considered independently of the presence of lupus anticoagulant (LAC) or anti-cardiolipin antibodies (ACA). One hundred thirty-five patients with idiopathic VTE and 164 healthy subjects were enrolled. Anti-PC and anti-PS antibodies (both IgG and IgM) were assessed using commercially available ELISA kits. Among cases there was a higher prevalence of elevated anti-PC IgM antibodies than in controls (OR 2.44, 95%CI 1.00-5.94). The presence of anti-PC IgG and anti-PS IgG and IgM antibodies was also higher in cases than in controls, but the difference was not statistically significant. Only five patients had both anti-PC or anti-PS antibodies and LAC or ACA. We performed a stepwise multivariate logistic regression analysis showing that anti-PC IgM>958 percentile was a significant predictor of VTE after adjustment for LAC or ACA (OR 2.52, 95%CI 1.01-6.24)). Larger prospective studies are needed to confirm this finding.

Clinical and diagnostic value of ribosomal P autoantibodies in systemic lupus erythematosus

To analyse prospectively the diagnostic sensitivity and specificity as well as the clinical relevance of ribosomal P (anti-P) autoantibodies in a large cohort of SLE patients. The anti-P autoantibodies were evaluated in the serum of 200 Tunisian SLE patients at disease onset and 130 various control subjects by a sensitive immunodot assay. A complete laboratory evaluation and clinical examination were performed in each SLE patient. During the follow-up, the patients were regularly monitored for clinical parameters. Global SLE activity was measured by the ECLAM. The sensitivity and specificity of anti-P testing for SLE were 23.5 and 98.4%, respectively. The anti-P-positive samples 14/47 (29.8%), 27/47 (57.4%) and 5/47 (10.6%) were negative for anti-dsDNA, anti-Sm or both antibodies, respectively. The anti-P-positive patients showed more active disease activity and a much higher prevalence of arthritis. An association between IgG aCLs and anti-P antibodies was also found. However, anti-P antibodies were not associated with neuropsychiatric manifestations or lupus nephritis. This study does not seem to confirm the described association of anti-P antibodies with neuropsychiatric manifestations of SLE. However, it supports the anti-P antibody association with arthritis and disease activity as well as the presence of aCL. Based on our study and other related studies, we propose that, akin to anti-Sm and anti-dsDNA, anti-P antibodies detected by one agreed method may be considered for inclusion as a criterion for the classification of SLE.

Anti-prothrombin antibodies are associated with thrombosis in children

Introduction This investigation aimed to evaluate thrombotic risk factors in children, with special reference to autoantibodies against prothrombin and protein S. Materials and methods We studied 57 consecutive Swedish children and adolescents referred with a radiologically confirmed acute thrombotic event. Clinical data were collected and a thrombophilia investigation was performed, including analysis of autoantibodies against protein S (anti-PS) and prothrombin (anti-PT). The anti-PS and anti-PT autoantibodies were also investigated in sera from 47 healthy controls. Detection of autoantibodies was performed by quantitative enzyme-linked immunosorbent assays. Results Results for anti-PT antibodies were positive in 21% (12/57) of the patients and 2.1% (1/47) of the controls (OR 12.0, 95% CI 1.7–534; p = 0.005). Seven percent (4/57) of the patients and 2.1% (1/47) of the controls were positive for anti-PS antibodies (OR 3.4, 95% CI 0.3–174; p > 0.30). The FV G1691A mutation was found in 25% (14/57), and 44% (25/57) had 2 or more prothrombotic risk factors. Sixty percent (34/57) of the thrombosis patients were female. Peaks in frequency of thromboembolic events were found in the neonatal and the adolescent periods. Fifty-three percent (30/57) had thrombosis in the lower venous system. Associated clinical conditions occurred in 91% (52/57) : systemic illness in 31% (18/57), infections in 26% (15/57), and oral contraceptive use in 25% (14/57). Four percent (2/57) had no apparent clinical or prothrombotic risk factors. Conclusions This study suggests that anti-PT autoantibodies may be common risk factors for thrombosis in children, and it confirms the multifactorial nature of pediatric thrombosis.

Evaluation of anti-ribosomal P protein immunoassay in Japanese patients with connective tissue diseases: comparison with an indirect immunofluorescence assay

Objectives: To investigate the prevalence of anti-ribosomal P protein (anti-P) antibodies in Japanese patients with connective tissue diseases (CTDs) using enzyme-linked immunosorbent assays (ELISAs) and western blotting (WB) and to evaluate the indirect immunofluorescence (IIF) staining patterns of anti-P-positive sera.Methods: Anti-P antibodies were measured by two different commercially available ELISA kits and WB in 239 outpatients, 99 with systemic sclerosis (SSc), 73 with systemic lupus erythematosus (SLE), 45 with dermatomyositis (DM), and 22 with Sjögren's syndrome (SjS). Sera positive for anti-P antibodies by WB were analysed by IIF.Results: The frequency of positive WB findings in SLE (18/73, 25%) was higher than in other diseases. ELISA kits A and B for anti-P antibodies showed 21% and 43% sensitivity, and 93% and 88% specificity, respectively, for diagnosing SLE, based on the manufacturer's cut-off values. Receiver operating characteristic (ROC) curve analysis, based on positive WB findings, determined a new cut-off threshold but revealed that both ELISA kits still had good diagnostic characteristics. In IIF assays on anti-P antibody positive sera, typical anti-P antibody cytoplasmic staining patterns (n=8) were seen less frequently than other staining patterns (n=17).Conclusions: Routine screening for anti-P antibodies by IIF has low sensitivity. ELISAs using cut-off values established by individual facilities are suitable for detecting anti-P antibodies and provide a tool with good diagnostic characteristics, on a parity with WB.

Antiribosomal-P protein antibodies in a patient with systemic lupus erythematosus and non-Hodgkin’s lymphoma: more than coincidental finding?

Patients with systemic lupus erythematosus (SLE) are at an increased risk of lymphomas, but mechanisms underlying this association are obscure. Recently, it has been shown that antiribosomal-P protein (anti-P) antibodies cross-react with phospholipids and enhance the production of cytokines which may influence lymphomagenesis. We report a 46-year-old woman who suffered high grade diffuse large B-cell non-Hodgkin's lymphoma (DLBCL) 28 months after the diagnosis of SLE. Development of lymphoma was associated with occurrence of serum monoclonal IgM, and pronounced prolongation of phospholipid-dependent clotting tests. Anti-P IgG antibodies were highly positive both on HEp-2 cells and in ELISA test. Anticardiolipin, anti-beta2 glycoprotein I, and antiprothrombin IgM antibodies have also been found in high concentrations. Complete remission of DLBCL and SLE, with normalisation of clotting tests, and disappearance of M component was achieved with administration of rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone. The progression of SLE to DLBCL associated with presence of anti-P antibodies has not been previously reported. This association may not be coincidental, but further investigations are required to confirm this hypothesis.

Thermodynamics and Density of Binding of a Panel of Antibodies to High-Molecular-Weight Capsular Polysaccharides

The interaction between antipolysaccharide (anti-PS) antibodies and their antigens was investigated by the use of isothermal titration calorimetry to determine the thermodynamic binding constant (K), the change in the enthalpy of binding (DeltaH), and the binding density (N) to high-molecular-weight PSs. From these values, the change in the entropy of binding (DeltaS) was calculated. The thermodynamic parameters of binding to high-molecular-weight capsular PSs are reported for two monoclonal antibodies (MAbs) with different specificities for meningococcal serogroup C PS, five MAbs specific for different pneumococcal serotypes, and the Fab fragments of two antipneumococcal MAbs. The K values were in the range of 10(6) to 10(7) M(-1), and these values were 1 to 2 orders of magnitude greater than the previously reported K values derived from antibody-oligosaccharide interactions. The DeltaH associated with binding was favorable for each MAb and Fab fragment. The DeltaS associated with binding was also generally favorable for both the MAbs and the Fab fragments, with the exception of the anti-serotype 14 MAb and its Fab fragment. N provides information regarding how densely MAbs or Fabs can bind along PS chains and, as expressed in terms of monosaccharides, was very similar for the seven MAbs, with an average of 12 monosaccharides per bound MAb. The value of N for each Fab was smaller, with five or seven monosaccharides per bound Fab. These results suggest that steric interactions between antibody molecules are a major influence on the values of N of high-affinity MAbs to capsular PSs.

Progesterone from the Cumulus Cells Is the Sperm Chemoattractant Secreted by the Rabbit Oocyte Cumulus Complex

Sperm chemotaxis in mammals have been identified towards several female sources as follicular fluid (FF), oviduct fluid, and conditioned medium from the cumulus oophorus (CU) and the oocyte (O). Though several substances were confirmed as sperm chemoattractant, Progesterone (P) seems to be the best chemoattractant candidate, because: 1) spermatozoa express a cell surface P receptor, 2) capacitated spermatozoa are chemotactically attracted in vitro by gradients of low quantities of P; 3) the CU cells produce and secrete P after ovulation; 4) a gradient of P may be kept stable along the CU; and 5) the most probable site for sperm chemotaxis in vivo could be near and/or inside the CU. The aim of this study was to verify whether P is the sperm chemoattractant secreted by the rabbit oocyte-cumulus complex (OCC) in the rabbit, as a mammalian animal model. By means of videomicroscopy and computer image analysis we observed that only the CU are a stable source of sperm attractants. The CU produce and secrete P since the hormone was localized inside these cells by immunocytochemistry and in the conditioned medium by enzyme immunoassay. In addition, rabbit spermatozoa express a cell surface P receptor detected by western blot and localized over the acrosomal region by immunocytochemistry. To confirm that P is the sperm chemoattractant secreted by the CU, the sperm chemotactic response towards the OCC conditioned medium was inhibited by three different approaches: P from the OCC conditioned medium was removed with an anti-P antibody, the attractant gradient of the OCC conditioned medium was disrupted by a P counter gradient, and the sperm P receptor was blocked with a specific antibody. We concluded that only the CU but not the oocyte secretes P, and the latter chemoattract spermatozoa by means of a cell surface receptor. Our findings may be of interest in assisted reproduction procedures in humans, animals of economic importance and endangered species.

Humoral autoimmune response to ribosomal P proteins in chronic Chagas heart disease

The C terminal region of a Trypanosoma cruzi ribosomal P protein, encoded by the lambda gt11 JL5 recombinant, defined a major antigenic determinant in chronic Chagas heart disease. Immunopurified anti-JL5 antibodies were tested for anti-human ribosome reactivity by immunoblotting. They recognized the parasite ribosomal P proteins and clearly reacted with the corresponding human P proteins. The peptide R-13, that comprises the 13 C terminal residues of the JL5 recombinant and defines the specificity shared between chronic Chagas heart disease anti-JL5 antibodies and the systemic lupus erythematosus (SLE) anti-P antibodies, was used to study the specificity and the IgG subclass distribution of the anti-R-13 response by ELISA. The R-13 autoepitope is recognized mainly by sera from chagasic patients, but not by sera from malaria patients. Moreover, there was a significant correlation between anti-R-13 antibody levels and anti-T. cruzi antibody titres. The anti-R-13 response was mainly restricted to the IgG1 heavy chain isotype and correlated with the anti-T. cruzi isotype distribution.

A phase Ib safety and pharmacokinetic study of bavituximab plus chemotherapy in patients with refractory advanced solid tumor malignancies

3038 Background: Bavituximab (B) is a first in class anti-phosphatidylserine (PS) monoclonal antibody. This immunotherapeutic agent selectively binds to PS exposed on tumor blood vessels causing an enhanced immune response and shutting down blood flow into the tumor. Pre-clinical data suggest a synergistic action using anti-PS antibodies in combination with chemotherapy. Methods: This was an open-label, non- randomized, multi-center study in pts with refractory advanced solid tumors. B was given weekly for up to 8 weeks, at a dose of 3mg/kg i.v., in combination with docetaxel (D)(75–80mg/m2 every 3 weeks) or gemcitabine (G) (1,000mg/m2 weeks 1–7) or paclitaxel (175mg/m2 every 3 weeks) plus carboplatin (AUC=5) (P+C). Study objectives were to assess safety and tolerability of B and to characterize its PK profile when used in combination with chemotherapeutic agents. Serum levels of B were determined by ELISA. Results: 14 pts were enrolled, mean age was 48.4yr, 78.6% were female. All had a stage IV tumor. 5 pts received B+D, 5 pts received B+G and 4 pts received B+P+C. Pts received a mean of 6 doses of B. AEs were reported by 13 pts. Most common were vomiting (6pts), cough (4 pts), asthenia, pain, headache, neutropenia, prolonged aPTT and pain in extremities, each in 3pts. 3 pts reported SAEs considered to be related to B, 1 pt had non-related SAE. On Days 0 and 21, no significant differences in PK characteristics were seen between the three treatment groups following administration of B. No accumulation of B was seen following multiple dose administration. Using RECIST criteria, week 8 tumor responses were PR (3pts), SD (3pts) and PD (6pts); 2 pts were non-evaluable. Conclusions: The PK of B was not impacted by co-administration of conventional chemotherapy. 50% of all evaluable pts achieved an objective tumor response or stable disease. These findings warrant further efficacy studies of B in pts with refractory solid tumors. Treatment Arm Day 0 (Dose 1) Day 21 (Dose 4) Cmax (μg/mL) Tmax (hr) AUCinf (μ*hr/mL) T1/2 (hr) Cmax (μg/mL) Tmax (hr) AUCinf (μ*hr/mL) T1/2 (hr) B+D 47.5 5 3303 25.5 40.7 2.5 3421 27.9 B+G 56.2 5 4354 26.5 44.6 2.5 4083 28.3 B+P+C 58.5 4.25 3835 25 58.7 1.75 4446 24.4 Values shown are medians Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Expert Testimony Other Remuneration Peregrine Pharmaceuticals, Inc. Peregrine Pharmaceuticals, Inc. Peregrine Pharmaceuticals, Inc.

Autoantibodies and neuropsychiatric events at the time of systemic lupus erythematosus diagnosis: Results from an international inception cohort study

To examine, in an inception cohort of systemic lupus erythematosus (SLE) patients, the association between neuropsychiatric (NP) events and anti-ribosomal P (anti-P), antiphospholipid (lupus anticoagulant [LAC], anticardiolipin), anti-beta2-glycoprotein I, and anti-NR2 glutamate receptor antibodies. NP events were identified using the American College of Rheumatology case definitions and clustered into central/peripheral and diffuse/focal events. Attribution of NP events to SLE was determined using decision rules of differing stringency. Autoantibodies were measured without knowledge of NP events or their attribution. Four hundred twelve patients were studied (87.4% female; mean +/- SD age 34.9 +/- 13.5 years, mean +/- SD disease duration 5.0 +/- 4.2 months). There were 214 NP events in 133 patients (32.3%). The proportion of NP events attributed to SLE varied from 15% to 36%. There was no association between autoantibodies and NP events overall. However, the frequency of anti-P antibodies in patients with central NP events attributed to SLE was 4 of 20 (20%), versus 3 of 107 (2.8%) in patients with other NP events and 24 of 279 (8.6%) in those with no NP events (P = 0.04). Among patients with diffuse NP events, 3 of 11 had anti-P antibodies (27%), compared with 4 of 111 patients with other NP events (3.6%) and 24 of 279 of those with no NP events (8.6%) (P = 0.02). Specific clinical-serologic associations were found between anti-P and psychosis attributed to SLE (P = 0.02) and between LAC and cerebrovascular disease attributed to SLE (P = 0.038). There was no significant association between other autoantibodies and NP events. Clinically distinct NP events attributed to SLE and occurring around the time of diagnosis were found to be associated with anti-P antibodies and LAC. This suggests that there are different autoimmune pathogenetic mechanisms, although low sensitivity limits the clinical application of testing for these antibodies.