Review
Tumor cell- and microenvironment-specific roles of cysteine cathepsins in mouse models of human cancers

https://doi.org/10.1016/j.bbapap.2020.140423Get rights and content

Highlights

  • Papain-like cysteine peptidases have distinct roles in driving cancer progression in various mouse models.

  • A tumor-promoting role of cathepsins in the tumor microenvironment has been established.

  • Cathepsin L exhibited a tumor suppressor-like function in some tumor types.

Abstract

The human genome encodes for 11 papain-like endolysosomal cysteine peptidases, collectively known as the cysteine cathepsins. Based on their biochemical properties and with the help of experiments in cell culture, the cysteine cathepsins have acquired a reputation as promotors of progression and metastasis of various cancer entities. However, tumors are known to be complex tissues in which non-cancerous cells are also critical for tumorigenesis. Here we discuss the results of the intense investigation of cathepsins in mouse models of human cancers. We focus on models in immunocompetent mice, because only such models allow for analysis of cathepsins in a fully functional tumor microenvironment. An important outcome of those studies was the identification of cancer-promoting cathepsins in tumor-associated macrophages. Another interesting outcome of these animal studies was the identification of a homeostatic tumor-suppressive role for cathepsin L in skin and intestinal cancers. Taken together, these in vivo findings provide a basis for the use of cysteine cathepsins as therapeutic targets, prodrug activators, or as proteases for imaging tumors.

Introduction

The lysosome hosts a large variety of more than 60 degradative enzymes [1]. Among the lysosomal proteases the Clan CA family C1 peptidases, the so called cysteine cathepsins, comprise the largest group with 11 members [2]. These are cathepsin B (CtsB), cathepsin C (CtsC), cathepsin F (CtsF), cathepsin H (CtsH), cathepsin K (CtsK), cathepsin L (CtsL), cathepsin O (CtsO), cathepsin S (CtsS), cathepsin V (CtsV, also cathepsin L2), cathepsin W (CtsW), and cathepsin Z (CtsZ, also cathepsin X). Mice lack CtsV but express eight cathepsin L-like cathepsins specifically in the placenta [3,4]. The cysteine cathepsins have been investigated extensively for their diverse roles in cancer [5,6]. Cell-based assays revealed their involvement in regulation of proliferation, death, and motility of cancer cells. Proteomic analyses have started to identify the relevant substrates of the cathepsin proteases. Furthermore, clinical as well as histopathological studies have evaluated the role of cathepsins as markers for prognosis and therapy response. The important link between mechanistic in vitro experiments and clinical relevance has been provided by animal studies, mostly employing laboratory mice as model system. Systematic preclinical cancer treatment trials with cathepsin inhibitors have been rarely performed because of limited selectivity and poor pharmacokinetics of the existing small molecules for cathepsin inhibition [[7], [8], [9], [10]]. Hence, genetic approaches have been widely used to study the in vivo roles of cathepsin proteases in cancer cells and in cells of the tumor microenvironment. Here we will review the evidence obtained for the differential roles of cysteine cathepsins in murine cancer models during the past 15 years.

There are two principal approaches to study cancer in mice [11,12]. In the first approach – so called de novo carcinogenesis models – normal cells of living mice are gradually transformed to invasive and eventually metastatic cancer cells upon the action of mutagenic chemicals or by genetic activation or inactivation of oncogenes or tumor suppressors, respectively (Table 1). Both genetic and chemical induction of tumors allow to study the classical stepwise carcinogenesis process including the gradual adaptation of the tumor microenvironment concerning matrix remodeling, angiogenesis, and tumor-associated inflammation. These models, however, are usually laborious and often require extensive time periods of one year or even longer to achieve end-stage tumors.

The second approach to study cancer in mice is the transfer of cancer cells into the animals (Fig. 1). Those models have the advantage of a well-defined start of the experiment, highly reproducible growth rates and usually well manageable tumor growth periods of a few weeks. Their disadvantage is that the injected cancer cells have already undergone the full malignant transformation process. Therefore, the co-evolution of cancer cells and their environment cannot be readily studied in those models. The transfer of human cancer cells or tumor specimen into mice is generally complicated by an unwanted clearance of the foreign cells by the mouse immune system. To allow for tumor growth, human material is usually injected into immunocompromised mice lacking at least T- and B-cells. Naturally, these so called xenograft tumor models do not mimic the immunological situation in patients with an intact immune system. The more widespread use of immunologically humanized mice is likely to overcome these limitations [13]. The injection of mouse cancer cells into recipient mice of the same genetic background does not face the challenge of immunological graft rejection. Therefore, these syngeneic allograft models have been popular for the study of host responses to a cancer (Fig. 1). The matter is further complicated by the fact that the cancer cells are often injected subcutaneously to the flanks of mice, an anatomical site noticeably different to the tissue from which the initial tumor originated. Those heterotopic models are clearly inferior to orthotopic models, in which the cells are injected into the tissue of origin thereby mimicking the site-specific conditions far more accurately [14].

Many published reports used genetic tools to downregulate or overexpress cathepsins in cancer cells and subjected them to xenograft in vivo studies. Those studies provided valuable insights into cancer cell autonomous functions of the cathepsins and some explored even combinatory effects of CtsB and matrix metalloprotease 9 (MMP9) or CtsB and urokinase-type plasminogen activator receptor (uPAR) downregulations in orthotopic models [15,16]. However, due to the limitations of the xenograft approach we will focus this review on the models of de novo carcinogenesis and to syngeneic allograft approaches that allowed to address the role of cathepsins in non-malignant tumor-associated cells.

Section snippets

Cysteine cathepsins in invasive growth and angiogenesis of pancreatic cancers

The Rip1-Tag2 mouse model is a seminal transgenic model to study oncogene-induced de novo carcinogenesis [17]. The rat insulin promoter (Rip1) directs expression of the Simian virus 40 large T-antigen (Tag2) specifically to the endocrine β-cells of the pancreas. This causes induction of pancreatic neuroendocrine tumors that develop progressively towards invasive pancreatic β-cell carcinomas [18]. In the first study on cathepsins in this model, collective cysteine cathepsin inhibition by the

Exploring the role of stroma-derived cathepsins in the tumor microenvironment using tumor cell transplantation models

It is commonly accepted that epithelial cells bearing oncogenic mutations are not solely responsible for carcinoma progression [[66], [67], [68], [69], [70]]. Rather, tumors develop in a complex relationship of cancer cells with their surrounding stroma. The stroma consists of endothelial and other vasculature cells, resident cells like fibroblasts, and tumor-associated immune cells including leukocytes and macrophages [68,69] as well as ECM components [71]. This tumor microenvironment actively

Concluding remarks

The efforts to elucidate specific in vivo roles of cysteine cathepsins in complex immunocompetent models revealed in many instances their tumor-promoting role in cancer cells, tumor-associated immune cells, or both. However, in some models the genetic manipulation of cathepsins had no influence on cancer growth or metastasis (Table 1). Even worse, knock-out of CtsL had a detrimental effect in some cancer models (see Section 2.4; Table 1). Hence, for upholding cathepsin inhibition as therapeutic

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

The Deutsche Forschungsgemeinschaft (DFG) SFB 850 subproject B7, DFG-grant RE1584/6-2 and the German Cancer Consortium (DKTK) program Oncogenic Pathways project L627 support the work in the Reinheckel laboratory.

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