Atlas of Genetics and Cytogenetics

in Oncology and Haematology OPEN ACCESS JOURNAL AT INIST-CNRS

Gene Section Review

SERPINB5 ( peptidase inhibitor, clade B (), member 5) Jim Heighway, Shirley Smith, Naomi Bowers, Daniel Betticher Target Identification Group, Roy Castle International Centre for Lung Cancer Research, University of Liverpool, 200 London Rd, Liverpool L3 9TA, UK (JH, SS, NB); Institute of Medical Oncology, University of Bern, 3010 Bern, Switzerland (DCB)

Published in Atlas Database: June 2004 Online updated version: http://AtlasGeneticsOncology.org/Genes/SerpinB5ID42267.html DOI: 10.4267/2042/38104 This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 2.0 France Licence. © 2004 Atlas of Genetics and Cytogenetics in Oncology and Haematology

normal cells that express the . The analysis of the Identity SERPINB5 promoter in a range of primary cell cultures Other names: ; Protease Inhibitor 5, PI5 provided the first formal demonstration that cytosine HGNC (Hugo): SERPINB5 methylation is important in the regulation of normal cell-type specific gene expression. Location: 18q21.33 Pseudogene Local order: BCL2; FVT1; VPS4B; SERPINB5; SERPINB12; SERPINB13. No known pseudogene. Note: SERPINB5 is the most centromeric member of a cluster of 10 SERPIN which encode products with homology to known serine protease inhibitors. Description DNA/RNA SERPINB5 encodes a 375 amino acid 42kDa protein, maspin, which shows to of Description the ovalbumin-type subfamily. In common with other family members, the protein is predicted to fold in such The SERPINB5 gene comprises seven exons and six a way as to expose a short reactive site loop (amino introns which commonly encode a 2.6kb mRNA. Two acids 331-345) on the surface. However, it is not alternatively processed splice variants have been currently clear whether maspin actually acts as a true reported but their significance is unknown. The ATG serine protease inhibitor and there are some suggestions start is located in exon 2 with the stop codon in exon 7. that the RSL may represent primarily a serine protease Transcription substrate. Maspin is therefore generally classified as a Transcriptional control is complex. SERPINB5 may be non-inhibitory serpin. A number of isoforms of maspin induced by p53. In addition to a p53 consensus binding have been detected in primary tissue samples and site, the promoter also contains functional Ets and AP-1 cultures. These may reflect processed or modified sites. In oestrogen receptor-positive breast cancer cells, forms of the protein or else may represent degradation expression may also be induced either directly or products. indirectly by the oestrogen antagonist, Tamoxifen. The Expression cell-type specific expression of SERPINB5 is heavily Maspin is expressed strongly in specific populations of influenced by the level of cytosine methylation in a cells within a range of normal tissues: for example, in region of sequence immediately upstream of exon 1. In the myoepithelial cells of the breast or the basal normal cells that do not express SERPINB5, this region epithelial cells of the bronchial airways. Other cell is: heavily methylated, associated with hypoacetylated types within such tissues may generally show a histones and the local chromatin structure is complete lack of expression. Particular cell-type inaccessible. The converse appears to be true for

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specific expression levels are likely to be controlled in thyroid. Furthermore, in LA7 cells, a well characterised the normal situation by strikingly different levels of rat adenocarcinoma in vitro model of mammary gland cytosine promoter methylation. Many studies have differentiation, maspin was shown to negatively reported differences in the expression and methylation regulate dome formation, possibly through the status of SERPINB5 between tumours and matched perturbation of cell adhesion. This observation suggests normal tissue. However, given the complex pattern of that at least in some situations, maspin expression can cell-type specific expression/regulation of this gene and block differentiation processes. our general lack of knowledge of the identity and Homology expression characteristics of the tumour progenitor cells, the pathological significance of these data are Maspin is an ovalbumin (ov-) serpin, located in one of currently difficult to interpret. the two ov-serpin clusters (18q21.33, 6p25) in the . Phylogenetic analysis has suggested Localisation that maspin is most closely related to the members of Sub-cellular localisation is variable depending on cell the 6q group and SERPINB8 on 18q. type. Consistent with predicted function, maspin may be found in certain cell types in cytoplasmic and peri- Mutations cellular locations. However, more recent studies have shown that maspin may be localised predominantly to Somatic the nucleus of certain normal or malignant cells. The No somatic coding sequence mutations have been role of the protein in the nucleus is not yet defined. described for SERPINB5. Minimally, three validated Function non-synonymous and one synonymous cSNP are located within the gene at amino acids: 176 (C/T, The exact cellular role of maspin is not currently clear. Phe/Ser), 187 (C/G, Leu/Val), 298 (C/T, Ser/Ser) and Whilst the protein can specifically influence certain 319 (A/G, Ile/Val). aspects of cell behaviour, often tumour and normal The gene is not thought to be a frequent tumour tissue derived expression data are at least superficially amplification or translocation target. contradictory. This likely reflects our incomplete understanding of the function of the protein in different Epigenetics situations: that is, in the context of different cell types, However, there are number of reports highlighting differentiation states and gene expression backgrounds. differences in promoter methylation status in primary SERPINB5 was originally described as a breast tumour human tumours compared to matched normal tissues. suppressor, a gene which was active in normal breast Whilst these studies are difficult to interpret given that epithelial cells and which was down-regulated cytosine methylation is used appropriately to control progressively towards malignancy with increasing the cell-type expression of SERPINB5 in normal tissue degrees of tissue disorder being associated with less (and therefore probably in the tumour cell progenitor), frequent instances of expression. Consistent with such a the striking observation of allele-specific differences in tumour suppressor function, work in vitro and in vivo promoter methylation in pre-neoplastic gastric lesions suggested that maspin suppressed angiogenesis, suggests that pathological epigenetic alteration may be reduced tumour invasiveness, growth, and meta-stasis a significant factor in deregulating (or failing to and sensitised cells to apoptosis. It was suggested that appropriately inactivate) the gene in certain cancers. maspin exerted these effects, at least in part, through modulation of plasminogen activation. In a breast Implicated in cancer cell line, the maspin RSL was deemed to be critical for the inhibition of tumour cell invasion and Oncogenesis the promotion of cell adhesion to extracellular matrix Maspin was classified as a breast cancer suppressor on molecules. the basis of functional and observational However, recent studies have subsequently painted a (immunohistochemical-IHC) studies. However, as more complex and perhaps contradictory picture. more model systems, tumour types and corresponding Maspin appears to have a critical role in early normal tissues were analysed, the situation became embryonic development. Homozygous loss of more complex. Several microarray analyses of different expression in mice knockouts is lethal at the peri- human cancers indicated that maspin was frequently implantation stage. The absence of the protein (-/-) strongly expressed in tumour over normal tissue. disrupts the formation of the endodermal cell layer Furthermore, strong and frequent expression was seen whilst maspin heterozygote knockout (+/-) endodermal in some instances in pre-neoplastic lesions. Clearly, cells grow more slowly than wild-type (+/+) cells. This some tumours in a range of tissues express maspin and is particularly interesting in the context of high-level some do not, the subcellular localisation of the protein maspin expression in tumours arising from organs of is variable, as is the methylation status of the promoter. endodermal origin, such as the GI tract, lungs and Expression may correlate to some degree with the

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clinical behaviour or characteristics of particular not exclude the possibility that maspin is associated tumours but in many cases the data from different with some aspect of the multipotent stem cell studies of the same diseases and tissues appear to be phenotype nor that it might be involved in the actual contradictory. One point that must be borne in mind is differentiation process of particular cell types. that a gene can be associated with good prognostic Lung cancer factors and improved survival and yet it can still be driving the malignant phenotype in the tumours in Note which it is expressed. In summary, many questions Lung cancers generally arise from a component of the concerning the patholo-gical role of maspin in human bronchial epithelium (BE). This pseudostratified lining cancer remain unanswered. of the airways comprises multipotent basal cells and specialised differentiated apical cells. The basal cells Breast cancer represent a reserve component of the epithelium which Note can differentiate into each of the mature cell types. The terminal duct-lobular system of the breast Alveolar, stromal and normal differentiated epithelial comprises two specialised epithelial cell types, inner cells are all typically negative for maspin expression. luminal secretory cells and outer contractile However, strong nuclear staining is seen in all airway myoepithelial cells. It is thought that most breast basal cells (Figure: A, B). Maspin appears to be tumours arise from cell types located within this strongly expressed, generally in the cytoplasm and structure. SERPINB5 is strongly expressed in the outer nucleus, in >95% of squamous cell carcinomas (Figure oestrogen receptor (ER) negative myoepithel-ial cells, C) and 30-50% of adenocarcinomas. The expression of where it is frequently located in both the nucleus and maspin is linked to the degree of promoter methylation the cytoplasm. The luminal cells appear not to express and allele-specific transcript analysis suggests that in maspin. The largest study (1068) of maspin expression approximately 50% of lesions, expression in the tumour in breast tumours scored a nuclear signal in 96% of is predominantly driven from one chromosomal allele. carcinomas and a cytoplasmic signal in 35%. The Maspin is strongly and presumably inappropriately nuclear staining was correlated with ER and expressed in a significant fraction of pre-neoplastic progesterone receptor (PR) positivity whilst the bronchial lesions and a small fraction of histologically cytoplasmic staining was associated with ER and PR non-neoplastic epithelia (Figure D). The observation negativity. It has been suggested that breast cancer that maspin is strongly expressed apparently initiates and is maintained from an aberrant adult stem appropriately in the normal stem cell-like basal cell. Maspin expression levels in tumours may component of the BE and that this expression appears therefore reflect a tendency for the tumour initiating to be down-regulated in the differentiated BE cells stem cell component to differentiate towards cells might suggest that the protein has a role in some aspect which lack or retain maspin expression. These lesions of maintenance of the basal cell (perhaps lung stem may behave differently clinically in a way which is or cell) phenotype and that a failure to inactivate maspin is not directly related to maspin expression. This would appropriately during differentiation may contribute to bronchial neoplasia.

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The images show an immunohistochemical analysis of maspin expression in histologically normal lung (A, B, D) and NSCLC (C) tissues. The tumour cells (C) show intense nuclear and cytoplasmic staining. Panels A and B show typical staining patterns for normal bronchial tissue. Stromal cells are negative, basal epithelial cells in the airway epithelia in the normal sections show strong predominantly nuclear staining whilst apical epithelial cells are negative. A small number of hyperplastic epithelia show strong nuclear/cytoplasmic expression of maspin (D).

Gastric cancer the protein was found to be expressed frequently in high grade prostate intraepithelial neoplasia, an Note observation which would be consistent with a role for IHC reports are to some extent contradictory and this maspin in pre-malignancy of the prostate. may reflect differing aetiologies. However, several studies suggest that maspin is strongly expressed in Melanoma gastric carcinoma over normal mucosa. It has been Note reported that 80% of primary tumours and all gastric Maspin appears to be expressed in a small fraction of normal mucosa (GNM) with intestinal metaplasia (IM) primary melanomas but not normal melanocytes. This show dense and diffuse cytoplasmic immunoreactivity expression is correlated with the methylation status of while in contrast, GNM without IM show only weak or the promoter. no staining. They further demonstrated that the GNM Disease with IM show maspin promoter hypermethylation of one parental allele which would be consistent with Melanoma. expression occurring from a single allele in a clonal Thyroid expansion, either as a result of an aberrant Note demethylation event or else as a consequence of a Maspin expression has been reported in a large fraction failure to methylate appropriately one parental allele at of: papillary thyroid carcinomas, undifferentiated some time during the differentiation process from a carcinomas and poorly differentiated carcinomas but maspin expressing precursor. An alternative only rarely in well differentiated tumours and not at all explanation, that SERPINB5 is developmentally in normal thyroid, follicular adenomas or follicular imprinted in a GM with IM precursor, seems less likely carcinomas. The expression of maspin in thyroid but is nevertheless possible. Both alleles were carcinomas was therefore closely associated with a lack hypermethylated in GNM without IM and both were of differentiation of the tumour cells. generally hypomethylated in tumours. Ovarian cancer Pancreatic cancer Note Note Whilst normal ovarian surface epithelia have low levels Maspin is strongly expressed in most if not all of expression, maspin appears to be strongly expressed pancreatic adenocarcinomas whereas normal pancreatic in a large fraction of primary tumours (37%). Tumours tissue appears to be negative or only weakly positive. with high levels of maspin were more likely to be Prostate cancer invasive and show cytoplasmic staining. Maspin over- Note expression was further associated with higher tumour Whilst maspin expression in prostatic cancer was grade, the presence of ascites and shorter survival. reportedly correlated with a less aggressive However, somewhat paradoxically, introduction of pathological and histological tumour grade, wild-type maspin into two ovarian cancer cell lines reduced their invasiveness.

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Bladder cancer Reis-Filho JS, Milanezi F, Schmitt FC. Maspin is expressed in the nuclei of breast myoepithelial cells. J Pathol. 2002 Note Jun;197(2):272-3; author reply 273-4 Maspin does not appear to be expressed in normal Sood AK, Fletcher MS, Gruman LM, Coffin JE, Jabbari S, transitional cells of the bladder. However, a sizable Khalkhali-Ellis Z, Arbour N, Seftor EA, Hendrix MJ. The fraction of tumours show strong nuclear and paradoxical expression of maspin in ovarian carcinoma. Clin cytoplasmic expression; which is significantly Cancer Res. 2002 Sep;8(9):2924-32 correlated with muscle-invasive over non-invasive Akiyama Y, Maesawa C, Ogasawara S, Terashima M, Masuda bladder cancer. T. Cell-type-specific repression of the maspin gene is disrupted frequently by demethylation at the promoter region in gastric intestinal metaplasia and cancer cells. Am J Pathol. 2003 Breakpoints Nov;163(5):1911-9 Note Mohsin SK, Zhang M, Clark GM, Craig Allred D. Maspin expression in invasive breast cancer: association with other Maspin lies 158 kb telomeric to BCL2, which is prognostic factors. J Pathol. 2003 Apr;199(4):432-5 targeted in the t(14;18) translocation in follicular lymphoma. Ngamkitidechakul C, Warejcka DJ, Burke JM, O'Brien WJ, Twining SS. Sufficiency of the reactive site loop of maspin for induction of cell-matrix adhesion and inhibition of cell invasion. References Conversion of ovalbumin to a maspin-like molecule. J Biol Chem. 2003 Aug 22;278(34):31796-806 Zhang M, Maass N, Magit D, Sager R. Transactivation through Ets and Ap1 transcription sites determines the expression of Smith SL, Watson SG, Ratschiller D, Gugger M, Betticher DC, the tumor-suppressing gene maspin. Cell Growth Differ. 1997 Heighway J. Maspin - the most commonly-expressed gene of Feb;8(2):179-86 the 18q21.3 serpin cluster in lung cancer - is strongly expressed in preneoplastic bronchial lesions. Oncogene. 2003 Scott FL, Eyre HJ, Lioumi M, Ragoussis J, Irving JA, Nov 27;22(54):8677-87 Sutherland GA, Bird PI. Human ovalbumin serpin evolution: phylogenic analysis, gene organization, and identification of Gao F, Shi HY, Daughty C, Cella N, Zhang M. Maspin plays an new PI8-related genes suggest that two interchromosomal and essential role in early embryonic development. Development. several intrachromosomal duplications generated the gene 2004 Apr;131(7):1479-89 clusters at 18q21-q23 and 6p25. Genomics. 1999 Dec Liu Z, Shi HY, Nawaz Z, Zhang M. Tamoxifen induces the 15;62(3):490-9 expression of maspin through estrogen receptor-alpha. Cancer Zou Z, Gao C, Nagaich AK, Connell T, Saito S, Moul JW, Seth Lett. 2004 Jun 8;209(1):55-65 P, Appella E, Srivastava S. p53 regulates the expression of the Ogasawara S, Maesawa C, Yamamoto M, Akiyama Y, Wada tumor suppressor gene maspin. J Biol Chem. 2000 Mar K, Fujisawa K, Higuchi T, Tomisawa Y, Sato N, Endo S, Saito 3;275(9):6051-4 K, Masuda T. Disruption of cell-type-specific methylation at the Maass N, Hojo T, Ueding M, Lüttges J, Klöppel G, Jonat W, Maspin gene promoter is frequently involved in undifferentiated Nagasaki K. Expression of the tumor suppressor gene Maspin thyroid cancers. Oncogene. 2004 Feb 5;23(5):1117-24 in human pancreatic cancers. Clin Cancer Res. 2001 Sugimoto S, Maass N, Takimoto Y, Sato K, Minei S, Zhang M, Apr;7(4):812-7 Hoshikawa Y, Jünemann KP, Jonat W, Nagasaki K. Zucchi I, Bini L, Valaperta R, Ginestra A, Albani D, Susani L, Expression and regulation of tumor suppressor gene maspin in Sanchez JC, Liberatori S, Magi B, Raggiaschi R, Hochstrasser human bladder cancer. Cancer Lett. 2004 Jan 20;203(2):209- DF, Pallini V, Vezzoni P, Dulbecco R. Proteomic dissection of 15 dome formation in a mammary cell line: role of tropomyosin-5b Wada K, Maesawa C, Akasaka T, Masuda T. Aberrant and maspin. Proc Natl Acad Sci U S A. 2001 May expression of the maspin gene associated with epigenetic 8;98(10):5608-13 modification in melanoma cells. J Invest Dermatol. 2004 Futscher BW, Oshiro MM, Wozniak RJ, Holtan N, Hanigan CL, Mar;122(3):805-11 Duan H, Domann FE. Role for DNA methylation in the control Yatabe Y, Mitsudomi T, Takahashi T. Maspin expression in of cell type specific maspin expression. Nat Genet. 2002 normal lung and non-small-cell lung cancers: cellular property- Jun;31(2):175-9 associated expression under the control of promoter DNA Heighway J, Knapp T, Boyce L, Brennand S, Field JK, methylation. Oncogene. 2004 May 20;23(23):4041-9 Betticher DC, Ratschiller D, Gugger M, Donovan M, Lasek A, Rickert P. Expression profiling of primary non-small cell lung This article should be referenced as such: cancer for target identification. Oncogene. 2002 Oct Heighway J, Smith S, Bowers N, Betticher DC. SERPINB5 31;21(50):7749-63 (serpin peptidase inhibitor, clade B (ovalbumin), member 5). Pierson CR, McGowen R, Grignon D, Sakr W, Dey J, Sheng S. Atlas Genet Cytogenet Oncol Haematol. 2004; 8(3):231-235. Maspin is up-regulated in premalignant prostate epithelia. Prostate. 2002 Dec 1;53(4):255-62

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