Modulation of the granzyme B inhibitor proteinase inhibitor 9 (PI-9) by activation of lymphocytes and monocytes <i>in vitro</i> and by Epstein–Barr virus and bacterial infection

Classen, CF; Bird, Phillip I.; Debatin, Klaus‐Michael

doi:10.1111/j.1365-2249.2006.03006.x

Cited by 25 publications

(29 citation statements)

References 26 publications

(54 reference statements)

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“…Besides protecting the CTLs, PI-9 may also defend bystander cells or antigenpresenting cells likely to be exposed to GrB during an immune response. The expression of PI-9 in B cells (3), monocytes (23), mast cells (24), endothelial and mesothelial cells (25), smooth muscle cells (26) and dendritic cells (21,27) is consistent with such a role. Cells at immune-privileged sites including the eye lens capsula, testis, ovaries, placenta, embryonic stem cells also upregulate the PI-9 expression (20,25,28).…”

Section: Introductionsupporting

confidence: 57%

Expression of proteinase inhibitor-9/serpinB9 in non-small cell lung carcinoma cells and tissues

Krepela¹

2009

Int J Oncol

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Abstract. Human proteinase inhibitor-9 (PI-9)/serpinB9 is an intracellular ovalbumin-family serpin with nucleocytoplasmic distribution which is expressed in certain normal cell types and cancer cells of different origin. Due to binding and inactivating of granzyme B (GrB), PI-9 can protect the cells from GrB-mediated apoptosis. High levels of PI-9 expression in certain cancer cells may contribute to their resistance against the immune mediated killing. So far, it is not known whether non-small cell lung cardinomas (NSCLCs) express PI-9 mRNA and a functional PI-9 protein. Herein we report for the first time that NSCLC cells express both PI-9 mRNA and protein and that there is a subset of NSCLC cells with upregulated PI-9 mRNA and protein expression. Futhermore, our work revealed that the PI-9 protein expressed in NSCLC cells can inhibit the active GrB. Finally, analysis of PI-9 mRNA expression in NSCLC tumours from surgically treated patients showed that the expression of this transcript is upregulated in the less-differentiated lung adenocarcinomas. We suggest, that the upregulated expression of PI-9 in NSCLC cells may serve to protect them from apoptosis induced by GrB.

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Section: Introductionsupporting

confidence: 57%

Expression of proteinase inhibitor-9/serpinB9 in non-small cell lung carcinoma cells and tissues

Krepela¹

2009

Int J Oncol

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“…1 AML cell line HL60 was exposed to each of the granzyme B constructs and we found that both the wildtype and mutant versions were cytotoxic with similar IC 50 values. Annexin V staining and caspase 3/7 assays confirmed the induction of apoptosis.…”

Section: Discussionmentioning

confidence: 99%

Targeted ex vivo reduction of CD64‐positive monocytes in chronic myelomonocytic leukemia and acute myelomonocytic leukemia using human granzyme B‐based cytolytic fusion proteins

Schiffer

Rosinke

Jost

et al. 2014

Intl Journal of Cancer

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CMML (chronic myelomonocytic leukemia) belongs to the group of myeloid neoplasms known as myelodysplastic and myeloproliferative diseases. In some patients with a history of CMML, the disease transforms to acute myelomonocytic leukemia (AMML). There are no specific treatment options for patients suffering from CMML except for supportive care and DNA methyltransferase inhibitors in patients with advanced disease. New treatment strategies are urgently required, so we have investigated the use of immunotherapeutic directed cytolytic fusion proteins (CFPs), which are chimeric proteins comprising a selective domain and a toxic component (preferably of human origin to avoid immunogenicity). The human serine protease granzyme B is a prominent candidate for tumor immunotherapy because it is expressed in cytotoxic T lymphocytes and natural killer cells. Here, we report the use of CD64 as a novel target for specific CMML and AMML therapy, and correlate CD64 expression with typical surface markers representing these diseases. We demonstrate that CD64-specific human CFPs kill CMML and AMML cells ex vivo, and that the mutant granzyme B protein R201K is more cytotoxic than the wild-type enzyme in the presence of the granzyme B inhibitor PI9. Besides, the human CFP based on the granzyme B mutant was also able to kill AMML or CMML probes resistant to Pseudomonas exotoxin A.The clonal hematopoietic stem cell disorder CMML (chronic myelomonocytic leukemia), is a highly aggressive and resistant form of leukemia. It is characterized by persistent monocytosis in the peripheral blood, at least one dysplasia component in the bone marrow, and <20% promonocytes and blasts in peripheral blood and bone marrow. 1 CMML can be classified further according to blast numbers, that is, CMML1 (<5% blasts in peripheral blood and <10% blasts in bone marrow) and CMML2 (<20% blasts in peripheral blood and 10-19% blasts in bone marrow).2,3 The disease occurs mainly in elderly people with a median survival of 15-20 months and leukemic transformation rates of 15-30%, both factors depending on the disease subtype. 4 Other diagnostic criteria include immunophenotyping based on the overlapping antigen expression patterns in CMML and acute myelomonocytic leukemia (AMML). The expression of CD56 combined with reductions in the levels of myeloid markers such as CD15 and CD13 is a unique signature of CMML monocytes.5 Furthermore, CD14 is expressed at higher levels on bone marrow monocytes in CMML compared with reactive monocytosis and normal marrow samples. 5 High levels of CD33 have also been reported. 6 Clonal cytogenetic abnormalities are found in 20-40% of CMML patients, but these do not appear to be specific. [7][8][9] The close relationship between CMML and other myelodysplastic (MDS)/MPNs makes correct diagnosis and treatment a significant challenge. Novel molecular markers described more recently include specific alleles of TET2 and CBL, but these

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“…This ratio may be controlled or influenced by a number of immunomodulatory cytokines or infections that alter SERPINB9 expression. 21,[55][56][57] Selectivity of the SERPINB9-gzmB Interaction Granzyme B seems to be the sole protease target of SERPINB9. In a prime example of the importance of P1 residue in determining the specificity for protease inhibition, the SERPINB9 P1 residue is a glutamate, rather than the aspartate preferred by gzmB.…”

Section: Serpinb9 and Granzyme Bmentioning

confidence: 99%

Control of granzymes by serpins

2009

Self Cite

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Although proteolysis mediated by granzymes has an important role in the immune response to infection or tumours, unrestrained granzyme activity may damage normal cells. In this review, we discuss the role of serpins within the immune system, as specific regulators of granzymes. The well-characterised human granzyme B-SERPINB9 interaction highlights the cytoprotective function that serpins have in safeguarding lymphocytes from granzymes that may leak from granules. We also discuss some of the pitfalls inherent in using rodent models of granzyme-serpin interactions and the ways in which our understanding of serpins can help resolve some of the current, contentious issues in granzyme biology. Cell Death and Differentiation (2010) 17, 586-595; doi:10.1038/cdd.2009; published online 6 November 2009As described elsewhere in this series, granzymes are key mediators of cytotoxic lymphocyte (CL) function, exhibiting both intracellular and extracellular functions. Thus, it is imperative that their activities be tightly controlled. Too little activity reduces the effectiveness of the immune system, resulting in infection and cancer. Conversely, overactivity can lead to disease caused by tissue destruction and cellular apoptosis. This review focusses on the manner in which granzyme activity is controlled at the posttranslational level by members of the serpin superfamily.The serpin superfamily is an ever-expanding group of structurally related proteins, currently comprising approximately 1000 members across all kingdoms of life. 1,2 Serpins function as intracellular or extracellular regulators of a wide range of physiological processes such as complement activation, blood coagulation and apoptosis. Within the vertebrate immune system, they control proteases of the classical and innate complement systems, and are also potent inhibitors of many leukocyte granule proteases. Serpin StructureStructures of almost 100 serpins from viruses to humans have been determined and all conform to the same basic architecture first described in 1984. The fold comprises nine a-helices, three b-sheets and a variable reactive centre loop (RCL), which is the primary site of interaction with target proteases (Figure 1a). The first structure determined was human SERPINA1 cleaved within the RCL. 3 Surprisingly, it was found that the residues around the cleavage point were separated by almost 70 Å , with the N-terminal portion of the RCL inserted into b-sheet A to form a fully antiparallel sixstranded sheet. It has subsequently been shown that, in the native state, the RCL is solvent exposed, and that its insertion into b-sheet A is a consequence of the inhibitory mechanism. The RCL is flanked by two highly conserved motifs (the proximal and distal hinges) that permit its insertion into b-sheet A. Insertion is also facilitated by the breach and shutter regions that assist the opening of b-sheet A and by the movement of helix F. 4 The RCL is a critical determinant of serpin inhibitory specificity, acting as a pseudosubstrate and cleavage site for the...

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Modulation of the granzyme B inhibitor proteinase inhibitor 9 (PI-9) by activation of lymphocytes and monocytes in vitro and by Epstein–Barr virus and bacterial infection

Cited by 25 publications

References 26 publications

Expression of proteinase inhibitor-9/serpinB9 in non-small cell lung carcinoma cells and tissues

Expression of proteinase inhibitor-9/serpinB9 in non-small cell lung carcinoma cells and tissues

Targeted ex vivo reduction of CD64‐positive monocytes in chronic myelomonocytic leukemia and acute myelomonocytic leukemia using human granzyme B‐based cytolytic fusion proteins

Control of granzymes by serpins

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