Summary In order to evaluate the role of cysteine peptidase cathepsin H (Cath H) in human lung cancer its protein levels were determined in 148 pairs of lung tumour tissue and adjacent non-tumourous lung parenchyma using the enzyme-linked immunosorbent assay technique. Additionally, Cath H levels were determined in sera of 171 patients with malignant tumours, 34 patients with benign lung diseases and 47 healthy controls. The median level of Cath H in tumour tissue was 0.64 times that in the corresponding lung parenchyma. Relating tumour levels with histological type we found higher Cath H levels in small-cell and adenocarcinomas and lower levels in squamous cell carcinoma, large-cell carcinoma and secondary tumours. A significant difference in Cath H level between lung tumour tissue and non-tumourous lung parenchyma was associated with the group of cigarette smokers (156 vs 263 ng mg -1 protein, P < 0.001). For this group of patients Cath H tumour levels correlated with the survival rate, while for the entire patient population this was not the case. Smokers with high tumour levels of Cath H experienced poor survival. Cath H was significantly higher in sera of patients with malignant and benign lung diseases than in control sera (P < 0.001). The increase was significant for all histological types, being the highest in small-cell and squamous cell carcinomas. Our study reveals that in lung tumours there is different behaviour of Cath H compared with other cysteine peptidases, e.g. cathepsin B and cathepsin L. Variations between tissue and serum levels of Cath H indicate either reduced expression or enhanced secretion of this enzyme in lung tumours.

Antigen-presenting cells (APC) play a pivotal role in the initiation of the T cell-mediated and antigen-specific immune response. The suggested role of endogenous inhibitor cystatin C (CyC) is to modulate cysteine proteases (cathepsins) present in human APC. To test this hypothesis, dendritic cells (DC) were generated in vitro from isolated monocytes, and changes in content, localization, and secretion of CyC and cathepsins S, L, and H (CatS, -L, and -H, repsectively) were followed in response to interleukin-4, enabling monocyte differentiation, and to tumor necrosis factor alpha (TNF-alpha), enabling DC maturation. A large increase in intracellular CyC accompanied the differentiation of monocytes to immature DC, also shown by strong immunolabeling of Golgi in immature DC. On DC maturation, intracellular CyC levels decreased, and CyC was mostly absent from the Golgi. On prolonged incubation of mature DC with TNF-alpha, CyC was found located in the proximity of the plasma membrane, indicating that the transport of CyC from Golgi was not blocked as the result of the arrested exocytosis in mature DC. The secretion of CyC ceased, consistent with the peak of the surface expression of phenotypic markers (CD40, CD54, CD80, CD83, CD86, and major histocompatibility complex class II), characteristic for the mature DC stage, whereas the secretion of cathepsins did not correlate with the maturation stage. The difference in localization of CyC and of CatS, -L, and -H in immature and mature DC shows that the regulatory potential of CyC toward CatS, -L, and -H inside DC is limited. However, these interactions may occur extracellularly in lymph, as suggested by the large excess of CyC over secreted CatS, -L, and -H, and they may facilitate DC migration to lymph nodes.

Stefin B (cystatin B) is an inhibitor of lysosomal cysteine cathepsins and does not inhibit cathepsin D, E (aspartic) or cathepsin G (serine) proteinases. In this study, we have investigated apoptosis triggered by camptothecin, staurosporin (STS), and anti-CD95 monoclonal antibody in the thymocytes from the stefin B-deficient mice and wild-type mice. We have observed increased sensibility to STS-induced apoptosis in the thymocytes of stefin B-deficient mice. Pretreatment of cells with pan-caspase inhibitor z-Val-Ala-Asp(OMe)-fluoromethylketone completely inhibited phosphatidylserine externalization and caspase activation, while treatment with inhibitor of calpains- and papain-like cathepsins (2S,3S)-trans-epoxysuccinyl-leucylamido-3-methyl-butane ethyl ester did not prevent caspase activation nor phosphatidylserine exposure. We conclude that sensitization to apoptosis induced by STS in thymocytes of stefin B-deficient and wild-type mice is not dependent on cathepsin inhibition by stefin B.

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SUMMARYActivation of the CD4 + T-cell mediated immune response relies on the proteolytic capacity of enzymes involved in modulating major histocompatibility complex (MHC) II-associated antigen presentation in antigen-presenting cells (APC). The MHC II-associated chaperone molecule p41 isoform of invariant chain (inhibitory p41 Ii) has been suggested to regulate stability and activity of cathepsin L in these APC. In the present study the human lymph node distribution of noninhibitory p31 Ii and inhibitory p41 Ii have been compared by differential labelling, using two specific monoclonal antibodies. The distribution of p41 Ii, but not p31 Ii, matched the distribution of cathepsins L and H in subcapsular and cortical sinuses and germinal centres. Co-localization of p41 Ii with cathepsin H was confirmed in strongly CD68 + sinus-lining macrophages, acting as APC. Furthermore, p41 Ii was determined together with cathepsins L and H in tingible body macrophages, highly phagocytic, but not antigen-presenting cells inside germinal centres. With respect to the physiological function that these two populations of macrophages have in human lymph nodes, our results support a regulatory function of p41 Ii towards cathepsins L and H in human macrophages, associated with the processes of phagocytosis rather than antigen presentation.

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Cathepsin H is a lysosomal cysteine protease that may participate in tumor progression. In order to evaluate its potential as a prognostic marker, its protein levels were measured by ELISA in preoperative sera from 324 patients with colorectal cancer. The level of cathepsin H was significantly increased in patient sera, the median level was 8.4 ng/mL versus 2.1 ng/mL in 90 healthy blood donors (p < 0.0001). A weak association of cathepsin H levels was found with patient age (p = 0.02) but not with Dukes' stage, sex, or the level of carcinoembryonic antigen (CEA). In survival analysis a significant difference was found between the group of patients with low cathepsin H (first tertile) who had a poor prognosis and the remaining patients (p = 0.03). The risk of patients was further stratified when cathepsin H levels were combined with CEA. Patients with high CEA and low cathepsin H had the highest risk of death with a hazard ratio of 2.72 (95% CI 1.73-4.28), p < 0.0001. Our results show that the prognostic information of cathepsin H differs from that of the related cathepsins B and L and suggest different roles during the progression of malignant disease.

Cathepsin H is a lysosomal cysteine protease that may participate in tumor progression. In order to evaluate its potential as a prognostic marker, its protein levels were measured by ELISA in preoperative sera from 324 patients with colorectal cancer. The level of cathepsin H was significantly increased in patient sera, the median level was 8.4 ng/mL versus 2.1 ng/mL in 90 healthy blood donors (p < 0.0001). A weak association of cathepsin H levels was found with patient age (p = 0.02) but not with Dukes' stage, sex, or the level of carcinoembryonic antigen (CEA). In survival analysis a significant difference was found between the group of patients with low cathepsin H (first tertile) who had a poor prognosis and the remaining patients (p = 0.03). The risk of patients was further stratified when cathepsin H levels were combined with CEA. Patients with high CEA and low cathepsin H had the highest risk of death with a hazard ratio of 2.72 (95% CI 1.73-4.28), p < 0.0001. Our results show that the prognostic information of cathepsin H differs from that of the related cathepsins B and L and suggest different roles during the progression of malignant disease.