Secretion Of Pregnancy-associated Plasma protein-A Research Articles

Persistent DNA Damage and Senescence in the Placenta Impacts Developmental Outcomes of Embryos.

Cohesin is an evolutionarily conserved chromosome-associated protein complex essential for chromosome segregation, gene expression, and repair of DNA damage. Mutations that affect this complex cause the human developmental disorder Cornelia de Lange syndrome (CdLS), thought to arise from defective embryonic transcription. We establish a significant role for placental defects in the development of CdLS mouse embryos (Nipbl and Hdac8). Placenta is a naturally senescent tissue; we demonstrate that persistent DNA damage potentiates senescence and activates cytokine signaling. Mutant embryo developmental outcomes are significantly improved in the context of a wild-type placenta or by genetically restricting cytokine signaling. Our study highlights that cohesin is required for maintaining ploidy and the repair of spontaneous DNA damage in placental cells, suggesting that genotoxic stress and ensuing placental senescence and cytokine production could represent a broad theme in embryo health and viability.

Characterization of mouse pericardial fat: regulation by PAPP-A

Although implicated in cardiovascular disease, little is known about the fat surrounding the heart. In humans, epicardial fat is the visceral fat depot of the heart, which directly contacts the myocardium. This strategically placed fat depot is thought to produce bioactive molecules that could affect cardiac function. A major limitation in understanding the biology of epicardial fat is its restricted access in humans and its seeming absence in commonly-used experimental animal models. Although laboratory mice do not have epicardial fat per se, they do have a fat depot around the heart. In this study, we found that mouse pericardial fat has the molecular signature, small adipocyte size, and resistance to differentiation consistent with visceral fat. In addition, we show that mouse pericardial fat is regulated by pregnancy-associated plasma protein-A (PAPP-A), a key modulator of local insulin-like growth factor bioavailability. PAPP-A is highly expressed in mouse pericardial fat at levels equivalent to those in mesenteric visceral fat and 10-fold higher than in subcutaneous inguinal fat (P = .0003). Cultured pre-adipocytes isolated from pericardial fat show 2-fold increased PAPP-A secretion compared to pre-adipocytes isolated from inguinal fat. Furthermore, PAPP-A knock-out mice fed a high fat diet for 20 weeks have significantly reduced pericardial fat (by 60%; P < .0001) compared to wild-type littermates. There was no significant difference in inguinal fat between wild-type and PAPP-A knock-out mice. These data characterize a new mouse model of visceral-like pericardial fat and lay a foundation for understanding its role in human heart disease.

Abstract 1597: Inhibition of the novel therapeutic target pregnancy associated plasma protein A (PAPP-A) in Ewing sarcoma enhances efficacy of IGF1R targeting in vivo

Abstract Background: Despite intensive treatment regimens, Ewing sarcoma (EWS) patients with metastatic or relapsed disease still face a 5-year overall survival of less than 20%, indicating a clear need for novel targeted therapies. Utilizing ribosomal depleted RNA sequencing of 122 EWS samples (49 cell lines and 73 tumor samples) and 96 normal samples of variable tissues, we previously reported PAPP-A as one of the top 5 genes overexpressed in EWS (&amp;gt; than 4-fold) compared to normal tissue. Notably, PAPP-A showed substantial expression in tumors (median log2FPKM = 3.933, n=122) and minimal expression in normal tissue (median log2FPKM = -1.564, n=96). Pregnancy Associated Plasma Protein A (PAPP-A) is a secreted zinc metalloproteinase, that anchors to cell surface of heparan sulfate proteoglycans and enhances local IGF signaling. Through the release of IGFs from IGFBP-4, IGFBP-2 and IGFBP-5, free, bioactive IGF is being increased in close proximity to its receptors. In pregnancy PAPP-A is highly expressed by placental trophoblasts and represents a key regulator of fetal growth. The impact of IGF signaling in EWS is illustrated by the modest activity of IGF-1R inhibition as a single agent treatment in clinical trials, an approach that is limited by the rapid development of resistance. Methods: In an attempt to investigate the role of PAPP-A in Ewing sarcoma, we used CRISPR/Cas9 technology to target the PAPP-A locus in EWS cell lines and generated PAPP-A knockout clones (EW8, TC32) for further investigation in vitro and in vivo. Additionally, we tested inhibition of PAPP-A proteolytic function in a xenograft NSG mouse model, as a single agent treatment as well as in conjunction with IGF1R inhibition. Results: Knockout of PAPP-A in EWS cell lines utilizing CRISPR/Cas9 technology completely abrogated PAPP-A secretion and metalloproteinase activity in single cell clones, resulting in significantly increased complexed IGFBP-4 and diminished bioactive IGF-1, together with decreased cell growth in vitro. This phenotype could be rescued by recombinant, soluble PAPP-A. In vivo experiments showed that treatment with a monoclonal PAPP-A neutralizing antibody that inhibits the IGFBP-4 proteolytic activity of PAPP-A delays tumor growth of xenografted EWS tumor cells (EW8) in NSG mice. Furthermore, in conjunction with IGF1R inhibition (mAB h7C10), PAPP-A neutralization significantly decreased tumor growth and resulted in prolonged survival of mice (p=0.021).Conclusions: PAPP-A is a novel biologically relevant and highly tumor specific cell surface target in Ewing sarcoma. Our data suggest that neutralization of PAPP-A is a new therapeutic option that merits further evaluation for the treatment of Ewing's Sarcoma. Citation Format: Sabine Heitzeneder, John F. Shern, Lee J. Helman, Javed Khan, Crystal L. Mackall. Inhibition of the novel therapeutic target pregnancy associated plasma protein A (PAPP-A) in Ewing sarcoma enhances efficacy of IGF1R targeting in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1597. doi:10.1158/1538-7445.AM2017-1597

Abstract 571: Pregnancy associated plasma protein A (PAPP-A) is a potential novel therapeutic target in Ewing sarcoma

Abstract Background: Ewing sarcoma (EWS) is the second most common bone malignancy in the pediatric population. Despite intensive treatment regimens, 5-year overall survival for patients with metastatic or relapsed disease is less than 20%, indicating a clear need for novel targeted therapies. The importance of IGF signaling in EWS is illustrated by activity of antagonistic targeting of IGF-1R in clinical trials, although this approach is plagued by rapid development of resistance. Pregnancy Associated Plasma Protein A (PAPP-A) anchors to cell surface proteoglycans and mediates metalloproteinase activity, that cleaves IGFs from IGFBP-4, IGFBP-2 and IGFBP-5, thus increasing local bioavailability of bioactive, free IGF. PAPP-A is expressed at high levels by placental trophoblasts, where it induces high levels of IGFs, which are required for fetal growth. Methods: In an attempt to identify novel targetable cell surface antigens in EWS, we performed ribosomal depleted RNA sequencing in 122 EWS samples (49 cell lines and 73 tumor samples) and 96 normal samples of variable tissues. Results: PAPP-A was identified as one of the top 5 membrane associated proteins overexpressed in EWS (&amp;gt; than 4-fold) compared to normal tissue (p = 3.89E-30). PAPP-A showed substantial expression in tumors (median log2FPKM = 3.933, n = 122) and minimal expression in normal tissue (median log2FPKM = -1.564, n = 96). We confirmed cell surface expression of PAPP-A by flow cytometry on 10 EWS cell lines tested, and identified PAPP-A in the supernatant of 12 EWS cell lines, measured by ELISA. Targeting the PAPP-A locus in the EWS cell line EW8 utilizing CRISPR/Cas9 completely abrogated PAPP-A secretion in single cell clones and significantly diminished bioactive IGF-1 levels in EWS cultures, while increasing IGF-1 bound to IGFBP-4, results which confirm absent PAPP-A metalloproteinase activity. Gene knockout of PAPP-A in EW8 resulted in decreased cell growth in vitro. Conclusions: We have identified PAPP-A as a novel potential cell surface target in Ewing sarcoma, characterized by high level tumor expression, limited normal tissue expression and biological significance in this tumor. Ongoing work is focused on studying growth of PAPP-A KO cell lines, and assessing whether PAPP-A neutralizing antibodies may mediate antitumor effects in EWS. Citation Format: Sabine Heitzeneder, John F. Shern, Javed Khan, Crystal L. Mackall. Pregnancy associated plasma protein A (PAPP-A) is a potential novel therapeutic target in Ewing sarcoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 571.

Pattern of secretion of pregnancy-associated plasma protein-A (PAPP-A) during pregnancies complicated by fetal aneuploidy, in vivo and in vitro.

BackgroundPregnancy-associated placental protein-A (PAPP-A) is a metalloprotease which circulates as an hetero-tetramer in maternal blood. Its maternal serum concentration in fetal trisomy 21 is decreased during the first trimester, so that PAPP-A is a useful screening biomarker. However, the regulation of PAPP-A placental secretion is unclear. We therefore investigated the secretion of PAPP-A in pregnancies complicated by fetal aneuploidies, both in vivo and in vitro.MethodsMaternal serum collected between 10 WG and 33 WG during 7014 normal pregnancies and 96 pregnancies complicated by fetal trisomy 21, 18, and 13 were assayed for PAPP-A using the Immulite 2000xpi system®. The pregnancies were monitored using ultrasound scanning, fetal karyotyping and placental analysis. Villous cytotrophoblasts were isolated from normal and trisomic placenta and cultured to investigate PAPP-A secretion in vitro (n = 6).ResultsAn increased nuchal translucency during the first trimester is a common feature of many chromosomal defect but each aneuploidy has its own syndromic pattern of abnormalities detectable at the prenatal ultrasound scanning and confirmed at the fetal examination thereafter. PAPP-A levels rise throughout normal pregnancy whereas in trisomy 21, PAPP-A levels were significantly decreased, but only during the first trimester. PAPP-A levels were decreased in trisomy 13 and sharply in trisomy 18, whatever the gestational age. In vitro, PAPP-A secretion was decreased in aneuploidy, and associated with decreased hCG secretion in Trisomy 21 and 18. These biochemical profiles did not appear to be linked to any specific histological lesions affecting the placenta.ConclusionsThese profiles may reflect different quantitative and qualitative placental dysfunctions in the context of these aneuploidies.

Protein Secretion Is Required for Pregnancy-Associated Plasma Protein-A to Promote Lung Cancer Growth In Vivo

Pregnancy-associated plasma protein-A (PAPPA) has been reported to regulate the activity of insulin-like growth factor (IGF) signal pathway through proteolytic degradation of IGF binding proteins (IGFBPs) thereby increasing the local concentration of free IGFs available to receptors. In this study we found that PAPPA is secreted from two out of seven lung cancer cell lines examined. None of immortalized normal bronchial epithelial cells (HBE) tested secrets PAPPA. There is no correlation between expression level and secretion of PAPPA in these cells. A cell line over-expressing PAPPA accompanied with secretion shows no notable changes in proliferation under cell culture conditions in vitro, but displays significantly augmentation of tumor growth in vivo in a xenograft model. In contrast, a cell line over-expressing PAPPA without secretion exhibits reduction of tumor growth both in vitro and in vivo. Down-regulation of PAPPA expression and secretion by RNAi knockdown decreases tumor growth after implanted in vivo. The tumor promoting activity of PAPPA appears to be mediated mainly through augmentation of the IGF signaling pathway as indicated by notable increases in downstream Akt kinase phosphorylation in tumor samples. Our results indicate that PAPPA secretion may play an important role in lung cancer growth and progression.

Maternal Serum ADAM12 Levels Correlate with PAPP-A Levels during the First Trimester

Background: We wished to investigate the correlation between a new Down screening marker ADAM12 (a disintegrin and metalloproteinase-12) and pregnancy-associated plasma protein-A (PAPP-A) and free β-human chorionic gonadotropin (fβ-hCG) during the first trimester of pregnancy. Methods: ADAM12, PAPP-A and fβ-hCG were measured in 225 serum samples of randomly chosen pregnancies with completely normal outcome. The samples were taken between pregnancy weeks 9+0 and 12+6. Results: The ADAM12 levels tended to increase with advanced gestational age and the highest levels were detected at pregnancy week 12. The ADAM12 levels correlated with PAPP-A levels. After weight correction and logarithmic transformation the multiples of median (MoM) of ADAM12 still correlated with the MoMs of PAPP-A and also with the MoMs of fβ-hCG. Smokers had lower ADAM12 levels than non-smokers. Conclusion: The secretion of ADAM12 seems to resemble the secretion of PAPP-A in the end of the first trimester. Accordingly ADAM12 appears not to be a separate marker independent of PAPP-A. It remains to be assessed whether adding ADAM12 in Down screening risk calculation will reduce the false positive rate during the first trimester of pregnancy.

Expression of Pregnancy-associated Plasma Protein-A (PAPP-A) During Human Villous Trophoblast Differentiation In Vitro

Pregnancy-associated placental protein-A (PAPP-A), first isolated from maternal serum, has been identified as a metalloprotease cleaving insulin-like growth factor binding protein-4 (IGFBP-4). The source of PAPP-A during pregnancy is unclear. We therefore investigated PAPP-A expression during in vitro human villous cytotrophoblast cell (CT) differentiation into syncytiotrophoblast (ST). CT were isolated from normal first trimester, second trimester and term placentae (n=10) and cultured to form ST. PAPP-A mRNA was quantified by real-time PCR, and PAPP-A protein expression was studied by immunocytochemistry and TRACE technology with specific monoclonal antibodies. PAPP-A mRNA expression in total placental extracts increased during the course of pregnancy. PAPP-A protein was detected in the cytoplasm of both CT and ST. ST formation in vitro was associated with a 19-fold increase in PAPP-A mRNA expression and an 8-fold increase in PAPP-A secretion into the culture medium. No significant difference in PAPP-A production was observed between cultured cells isolated from early and term placentae.In conclusion, PAPP-A production in vitro, is associated to the differentiation of villous cytotrophoblast cells into syncytiotrophoblast, independantly of the age of gestation.

Secretion of pregnancy-associated plasma protein-A by cultured human placental villous trophoblasts

Secretion of pregnancy-associated plasma protein-A by cultured human placental villous trophoblasts

Immunohistochemical localization of pregnancy-associated plasma protein-A in the male genital tract

Pregnancy-associated plasma protein-A (PAPP-A) has been found in seminal fluid at concentrations 10-20 times higher than in plasma. This observation prompted us to undertake a morphological study of the male genital tract with the aid of the immunoperoxidase technique. Immunoreactive cells were found among the Leydig cell population, as well as among the epithelium of the rete testis, the head of the epididymis and the seminal vesicles. The relatively small number of weakly staining epithelial cells leaves open the question of whether PAPP-A is excreted by these cells or whether an active transfer mechanism - possibly in the seminal vesicles - is at work, the Leydig cells increasing the plasma concentration locally, by secretion of PAPP-A into the circulation.

Pregnancy-associated plasma protein-A (PAPP-A): concentration in uterine fluid and immunohistochemical localization in the endometrium.

Pregnancy-associated plasma protein-A (PAPP-A) was localized immunohistochemically in the endometrium and measured in uterine fluid of non-pregnant women. The variations of PAPP-A concentrations in uterine fluid during the menstrual cycle paralleled those found in the endometrium. In patients receiving hormone therapy there was a significant correlation between the uterine fluid PAPP-A concentration and the progestogen to oestrogen potency ratio of the hormonal treatment. The presence of PAPP-A in the uterine fluid cannot simply be explained by blood contamination or cell damage. These results are interpreted as indirect evidence for an exocrine as well as an endocrine secretion of PAPP-A by the endometrium which might be influenced by hormones.

Pregnancy-associated plasma protein-A (PAPP-A) and hCG in early pregnancy.

Pregnancy-associated plasma protein-A (PAPP-A) is a recently described glycoprotein of unknown biological function. The development of a radioimmunoassay enabled us to measure plasma levels of PAPP-A and human chorionic gonadotrophin (hCG) in 12 non-pregnant volunteers and in 159 women in early pregnancy attending the outpatient clinic for legal abortion. PAPP-A but not hCG was measurable in all non-pregnant women. In pregnant patients (with 36 to 86 days of amenorrhea) hCG reached a peak value (163.1-197.6 ng/ml) between the 9th and the 13th week whereas PAPP-A steadily increased throughout this period of pregnancy. Between the 6th and the 13th week after the last menstrual period, levels of PAPP-A increased proportionally more than hCG. This work provides the first evidence of a PAPP-A production in non-pregnant subjects and the very early marked increase of PAPP-A secretion during pregnancy.