Recombinant cathepsin S propeptide attenuates cell invasion by inhibition of cathepsin L-like proteases in tumor microenvironment

Human cathepsin L along with cathepsin S, K, and V are collectively known as cathepsin L-like proteases due to their high homology. The overexpression and aberrant activity of each of these proteases has been implicated in tumorigenesis. These proteases contain propeptide domains that can potently inhibit both their cognate protease and other proteases within the cathepsin L-like subfamily. In this investigation, we have produced the cathepsin S propeptide recombinantly and have shown that it is a potent inhibitor of the peptidolytic, elastinolytic, and gelatinolytic activities of the cathepsin L-like proteases. In addition, we show that this peptide is capable of significantly attenuating tumor cell invasion in a panel of human cancer cell lines. Furthermore, fusion of an IgG Fc-domain to the COOH terminus of the propeptide resulted in a chimeric protein with significantly enhanced ability to block tumor cell invasion. This Fc fusion protein exhibited enhanced stability in cell-based assays in comparison with the unmodified propeptide species. This approach for the combined inhibition of the cathepsin L-like proteases may prove useful for the further study in cancer and other conditions where their aberrant activity has been implicated. Furthermore, this strategy for simultaneous inhibition of multiple cysteine cathepsins may represent the basis for novel therapeutics to attenuate tumorigenesis.

Evaluation of the antiangiogenic potential of AQ4N.

Purpose: A number of cytotoxic chemotherapy agents tested at low concentrations show antiangiogenic properties with limited cytotoxicity, e.g., cyclophosphamide, tirapazamine, and mitoxantrone. AQ4N is a bioreductive alkylaminoanthraquinone that is cytotoxic when reduced to AQ4; hence, it can be used to target hypoxic tumor cells. AQ4N is structurally similar to mitoxantrone and was evaluated for antiangiogenic properties without the need for bioreduction.

Experimental Design:The effect of AQ4N and fumagillin on human microvascular endothelial cells (HMEC-1) was measured using a variety ofin vitro assays, i.e., 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide, wound scrape, tubule formation, rat aortic ring, and invasion assays. Low-dose AQ4N (20 mg/kg) was also given in vivo to mice bearing a tumor in a dorsal skin flap.

Results:AQ4N (10-11to10-5mol/L) hadno effect on HMEC-1viability. AQ4N (10-9to10-5mol/L) caused a sigmoidal dose-dependent inhibition of endothelial cell migration in the wound scrape model. Fumagillin showed a similar response over a lower dose range (10-13 to 10-9 mol/L); however, the maximal inhibition was less (25% versus 43% for AQ4N). AQ4N inhibited HMEC-1 cell contacts on Matrigel (10-8 to 10-5 mol/L), HMEC-1 cell invasion, and sprouting in rat aorta explants. Immunofluorescence staining with tubulin, vimentim, dynein, and phalloidin revealed that AQ4N caused disruption to the cell cytoskeleton. When AQ4N (20 mg/kg) was given in vivo for 5 days, microvessels disappeared in LNCaP tumors grown in a dorsal skin flap.

Conclusions:This combination of assays has shown that AQ4N possesses antiangiogenic effects in normoxic conditions, which could potentially contribute to antitumor activity

Hypoxia selects for androgen independent LNCaP cells with a more malignant geno- and phenotype.

Hypoxia confers resistance to common cancer therapies, however, it has also has been shown to result in genetic alterations which may allow a survival advantage and increase the tumorigenic properties of cancer cells. Additionally, it may exert a selection pressure, allowing expansion of tumor cells with a more aggressive phenotype. To further assess the role of hypoxia in malignant progression in prostate cancer we exposed human androgen dependent prostate cancer cells (LNCaP) to cycles of chronic hypoxia and isolated a subline, LNCaP-H1. This article describes the partial characterization of this cell line. The LNCaP-H1 subline showed altered growth characteristics and exhibited androgen independent growth both in vitro and in vivo. Furthermore, these cells were resistant to mitochondrial-mediated apoptosis, probably since the endogenous levels of Bax was lower and Bcl-2 higher than in the parental LNCaP cells. Microarray analysis revealed that a complex array of pathways had differential gene expression between the 2 cell lines, with LNCaP-H1 cells exhibiting a genetic profile which suggests that they may be more likely metastasize to distant organs, especially bone. This was supported by an in vitro invasion assay, and an in vivo metastasis study. This study shows that hypoxia can select for androgen independent prostate cancer cells which have a survival advantage and are more likely to invade and metastasize.

Survival patterns among lymphoma patients with a family history of lymphoma

Purpose: Genetic factors are important in the etiology and pathogenesis of chronic lymphocytic leukemia (CLL), Hodgkin's lymphoma (HL), and non-Hodgkin's lymphoma (NHL). Only a few small studies have assessed clinical characteristics and prognosis for familial patients, with inconsistent findings. Methods: Using population-based registries from Sweden and Denmark, 7,749 patients with CLL, 7,476 patients with HL, and 25,801 patients with NHL with linkable first-degree relatives were identified. Kaplan-Meier curves were constructed to compare survival in patients with lymphoma with and without a family history of lymphoma. The risk of dying was assessed using adjusted Cox proportional hazard models. Results: We found 85 patients with CLL (1.10%), 95 patients with HL (1.28%), and 206 patients with NHL (0.80%) with a family history of any lymphoma. Five-year mortality was similar for patients with CLL (hazard ratio [HR], 1.28; 95% CI, 0.95 to 1.72), HL (HR, 0.78; 95% CI, 0.49 to 1.25), and NHL (HR, 0.91; 95% CI, 0.74 to 1.12) versus without a family history of any lymphoma. Mortality was also similar for patients with versus without a family history of the same lymphoma. T-cell/anaplastic lymphoma patients with a family history of NHL had poorer outcome 5-years after diagnosis (HR, 5.38; 95% CI, 1.65 to 17.52). Results were similar for 10 years of follow-up. Conclusion: With the exception of T-cell/anaplastic lymphoma, survival patterns for patients with CLL, HL, and NHL with a family history of lymphoma were similar to those for sporadic patients, suggesting that most familial lymphomas do not have an altered clinical course. Our findings provide no evidence to modify therapeutic strategies for patients with CLL, HL, or NHL based solely on family history. © 2008 by American Society of Clinical Oncology.


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Dietary glycaemic index, glycaemic load and endometrial and ovarian cancer risk: a systematic review and meta-analysis.

Long-term consumption of a high glycaemic index (GI) or glycaemic load (GL) diet may lead to chronic hyperinsulinaemia, which is a potential risk factor for cancer. To date, many studies have examined the association between GI, GL and cancer risk, although results have been inconsistent, therefore our objective was to conduct a systematic review of the literature. Medline and Embase were systematically searched using terms for GI, GL and cancer to identify studies published before December 2007. Random effects meta-analyses were performed for endometrial cancer, combining maximally adjusted results that compared risk for those in the highest versus the lowest category of intake. Separate analysis examined risk by body mass index categories. Five studies examining GI and/or GL intake and endometrial cancer risk were identified. Pooled effect estimates for endometrial cancer showed an increased risk for high GL consumers (RR 1.20; 95% CI: 1.06-1.37), further elevated in obese women (RR 1.54; 95% CI: 1.18-2.03). No significant associations were observed for GI. Only two studies examined ovarian cancer and therefore no meta-analysis was performed, but results indicate positive associations for GL also. A high GL, but not a high GI, diet is positively associated with the risk of endometrial cancer, particularly among obese women. © 2008 Cancer Research UK All rights reserved.