The extracellular matrix (ECM) plays an important role in cancer progression. It can be divided into the basement membrane (BM) that supports epithelial/endothelial cell behavior and the interstitial ...matrix (IM) that supports the underlying stromal compartment. The major components of the ECM are the collagens. While breaching of the BM and turnover of e.g. type IV collagen, is a well described part of tumorigenesis, less is known regarding the impact on tumorigenesis from the collagens residing in the stroma. Here we give an introduction and overview to the link between tumorigenesis and stromal collagens, with focus on the fibrillar collagens type I, II, III, V, XI, XXIV and XXVII as well as type VI collagen. Moreover, we discuss the impact of the cells responsible for this altered stromal collagen remodeling, the cancer associated fibroblasts (CAFs), and how these cells are key players in orchestrating the tumor microenvironment composition and tissue microarchitecture, hence also driving tumorigenesis and affecting response to treatment. Lastly, we discuss how specific collagen-derived biomarkers reflecting the turnover of stromal collagens and CAF activity may be used as tools to non-invasively interrogate stromal reactivity in the tumor microenvironment and predict response to treatment.
The extracellular matrix (ECM) plays a vital role in maintaining normal tissue function. Collagens are major components of the ECM and there is a tight equilibrium between degradation and formation ...of these proteins ensuring tissue health and homeostasis. As a consequence of tissue turnover, small collagen fragments are released into the circulation, which act as important biomarkers in the study of certain tissue-related remodeling factors in health and disease. The aim of this study was to establish an age-related collagen turnover profile of the main collagens of the interstitial matrix (type I and III collagen) and basement membrane (type IV collagen) in healthy men and women. By using well-characterized competitive ELISA-assays, we assessed specific fragments of degraded (C1M, C3M, C4M) and formed (PINP, Pro-C3, P4NP7S) type I, III and IV collagen in serum from 617 healthy men and women ranging in ages from 22 to 86. Subjects were divided into 5-year age groups according to their sex and age. Groups were compared using Kruskal-Wallis adjusted for Dunn's multiple comparisons test and Mann-Whitney t-test. Age-specific changes in collagen turnover was most profound for type I collagen. PINP levels decreased in men with advancing age, whereas in women, the level decreased in early adulthood followed by an increase around the age of menopause (age 40-60). Sex-specific changes in type I, III and IV collagen turnover was present at the age around menopause (age 40-60) with women having an increased turnover. In summary, collagen turnover is affected by age and sex with the interstitial matrix and the basement membrane being differently regulated. The observed changes needs to be accounted for when measuring ECM related biomarkers in clinical studies.
BACKGROUND: Type VI collagen (COL6) is associated with several pro-tumorigenic events. COL6 is primarily composed of three alpha-chains (a1-a3) forming a specialized microfibrillar network to support ...tissue architecture. COL6 homeostasis is lost in the tumor due to increased COL6 synthesis by activated fibroblast and altered proteolytic degradation by matrix metalloproteases (MMPs). Consequently, pathology-specific COL6 fragments are released to the circulation. This study evaluates four COL6 fragments measured in serum as potential biomarkers for cancer. METHODS: C6Ma1 (MMP-generated neo-epitope on the a1 chain), C6Ma3 (MMP-generated neo-epitope on the a3 chain), PRO-C6 (C-terminal of the a3 chain) and IC-6 (internal epitope on the a1 chain) were measured by ELISA in serum from patients with various stage 1–4 cancer indications (n = 4–11 per indication, total n = 65) and healthy controls (n = 13). RESULTS: C6Ma1 and C6Ma3 were significantly elevated in most cancer types compared to controls; PRO-C6 and IC6 were not. No significant differences were seen according to age, gender and TNM stage. Comparing cancer patients to controls, the AUROC was 0.90 (P < .0001), 0.87 (P < .0001), 0.59 (P = .311) and 0.53 (P = .747) for C6Ma1, C6Ma3, PRO-C6 and IC-6, respectively. Only C6M and C6Ma3 correlated significantly (Spearman, r = 0.74, P < .0001). CONCLUSIONS: MMP-generated COL6 fragments (C6Ma1, C6Ma3) were elevated in serum from cancer patients compared to controls and had promising diagnostic accuracy. This supports that MMP-mediated COL6 remodeling is important in tumorigenesis and indicate cancer biomarker potential of quantifying COL-6 fragments in serum. Future studies should determine biological and clinical applicability of the COL-6 serum biomarkers in relation to cancer.
It has been known for decades that the tumor extracellular matrix (ECM) is dysfunctional leading to loss of tissue architecture and promotion of tumor growth. The altered ECM and tumor fibrogenesis ...leads to tissue stiffness that act as a physical barrier to immune cell infiltration into the tumor microenvironment (TME). It is becoming increasingly clear that the ECM plays important roles in tumor immune responses. A growing body of data now indicates that ECM components also play a more active role in immune regulation when dysregulated ECM components act as ligands to interact with receptors on immune cells to inhibit immune cell subpopulations in the TME. In addition, immunotherapies such as checkpoint inhibitors that are approved to treat cancer are often hindered by ECM changes. In this review we highlight the ways by which ECM alterations affect and regulate immunity in cancer. More specifically, how collagens and major ECM components, suppress immunity in the complex TME. Finally, we will review how our increased understanding of immune and immunotherapy regulation by the ECM is leading towards novel disruptive strategies to overcome immune suppression.
Despite the overall clinical success of immune checkpoint inhibitors (ICIs) for treating patients with solid tumors, a large number of patients do not benefit from this approach. Consequently, there ...is a need for predictive biomarkers. The most prevalent biomarkers such as PD-L1 expression and tumor mutational burden (TMB) do not reliably predict response to ICIs across different solid tumor types suggesting that a broader view of regulating factors in the tumor microenvironment is needed. Emerging evidence indicates that one central common denominator of resistance to ICIs may be fibrotic activity characterized by extracellular matrix (ECM) and collagen production by cancer-associated fibroblasts (CAFs). A fibroblast-and collagen-rich stroma attenuates immunotherapy response by contributing to inhibition and exclusion of T cells. Here we review opportunities and limitations in the utilization of the most prevalent biomarkers for ICIs and elaborate on the unique opportunities with biomarkers originating from the activated fibroblasts producing an impermeable ECM. We propose that ECM and collagen biomarkers measured non-invasively may be a novel and practical approach to optimize treatment strategies and improve patient selection for ICI therapy.
Autoantibodies have the potential as cancer biomarkers as they may associate with the outcome and immune-related adverse events (irAEs) following immunotherapy. Cancer and other fibroinflammatory ...diseases, such as rheumatoid arthritis (RA), are associated with excessive collagen turnover leading to collagen triple helix unfolding and denaturation with exposure of immunodominant epitopes. In this study, we aimed to investigate the role of autoreactivity against denatured collagen in cancer. A technically robust assay to quantify autoantibodies against denatured type III collagen products (anti-dCol3) was developed and then measured in pretreatment serum from 223 cancer patients and 33 age-matched controls. Moreover, the association between anti-dCol3 levels and type III collagen degradation (C3M) and formation (PRO-C3) was investigated. Anti-dCol3 levels were significantly lower in patients with bladder (
= 0.0007), breast (
= 0.0002), colorectal (
< 0.0001), head and neck (
= 0.0005), kidney (
= 0.005), liver (
= 0.030), lung (
= 0.0004), melanoma (
< 0.0001), ovarian (
< 0.0001), pancreatic (
< 0.0001), prostate (
< 0.0001), and stomach cancers (
< 0.0001) compared to controls. High anti-dCol3 levels were associated with type III collagen degradation (C3M,
= 0.0002) but not type III collagen formation (PRO-C3,
= 0.26). Cancer patients with different solid tumor types have downregulated levels of circulating autoantibodies against denatured type III collagen compared to controls, suggesting that autoreactivity against unhealthy type III collagen may be important for tumor control and eradication. This autoimmunity biomarker may have the potential for studying the close relationship between autoimmunity and cancer.
BackgroundInterleukin-8 (IL-8) is a cytokine which promote tumor progression, epithelial-to-mesenchymal transition and suppress immune activation. Therefore, blockage of IL-8 has been proposed as an ...immunotherapy target. However, immunotherapies, of any kind, might be insufficient if the stromal fibrotic barrier surrounding the tumors is not bypassed, as immune cells have been shown to be inactivated in the tumor fibrotic space. Many cytokines have shown to module the extracellular matrix (ECM). Thus, in this study we investigated if IL-8 have ECM-modulatory properties, and could potentially act as a combinational immune-stroma therapy target.MethodsHumane pancreatic fibroblasts were cultured in Ficoll-media. Fibroblasts were treated w/wo TGF-β (1ng/mL) or IL-8 (1 and 10 ng/mL). The fibrotic activity of the fibroblasts was investigated by measuring the formation of type III collagen in the supernatant (PRO-C3) at days 3, 6, 9 and 12. Cell-viability was evaluated by Alamar-blue.ResultsThe formation of type III collagen (PRO-C3) was significantly increased when fibroblasts were treated with TGF-β at days 6, 9 and 12 compared to no treatment. At day 9 and 12 PRO-C3 was also significantly increased in supernatant from fibroblasts treated with IL-8 compared to no treatment.ConclusionsIn this study we show that IL-8 stimulate type III collagen production in fibroblasts similar to the pro-fibrotic cytokine TGF-β. Therefore, inhibiting IL-8 could potentially be two-sided and act as both an immune-modulatory and an anti-tumor fibrotic drug.
Collagens are a primary component of the extracellular matrix and are functional ligands for the inhibitory immune receptor leukocyte-associated immunoglobulin-like receptor (LAIR)-1. LAIR-2 is a ...secreted protein that can act as a decoy receptor by binding collagen with higher affinity than LAIR-1. We propose that collagens promote immune evasion by interacting with LAIR-1 expressed on immune cells, and that LAIR-2 releases LAIR-1-mediated immune suppression. Analysis of public human datasets shows that collagens, LAIR-1 and LAIR-2 have unique and overlapping associations with survival in certain tumors. We designed a dimeric LAIR-2 with a functional IgG1 Fc tail, NC410, and showed that NC410 increases human T cell expansion and effector function in vivo in a mouse xenogeneic-graft versus-host disease model. In humanized mouse tumor models, NC410 reduces tumor growth that is dependent on T cells. Immunohistochemical analysis of human tumors shows that NC410 binds to collagen-rich areas where LAIR-1
+
immune cells are localized. Our findings show that NC410 might be a novel strategy for cancer immunotherapy for immune-excluded tumors.
Excessive extracellular matrix (ECM) remodeling and a reactive stroma can affect T-cell infiltration and T-cell activity in the tumor and hereby influence response to immune checkpoint inhibitors ...(ICI). In the pursuit of finding biomarkers that predict treatment response, we evaluated the association between serum biomarkers of collagen and vimentin turnover and outcomes in metastatic melanoma patients treated with the anti-CTLA-4 antibody ipilimumab (IPI).
Type III collagen formation (PRO-C3), MMP-degraded type I, type III and type IV collagens (C1M, C3M and C4M), and citrullinated and MMP-degraded vimentin (VICM) were measured with ELISAs in serum from metastatic melanoma patients before (n = 66) and 3 weeks after (n = 52) initiation of IPI treatment. Biomarker levels were associated with Disease Control Rate (DCR) and survival outcomes.
We found that baseline levels of PRO-C3 (p = 0.011), C1M (p = 0.003), C3M (p = 0.013) and C4M (p = 0.027) were significantly elevated in patients with progressive disease (PD). Univariate Cox regression analysis identified high PRO-C3 (p = 0.021) and C4M (p = 0.008) as predictors of poor overall survival (OS) and the biomarkers remained significant when evaluated with other covariates (PRO-C3 (p = 0.049) and C4M (p = 0.046)). Multivariate analysis identified VICM as a predictor of longer OS (p = 0.026). Similarly, a high C3M/PRO-C3 ratio predicted for increased OS (p = 0.034). Only C3M (p = 0.003) and VICM (p < 0.0001) increased 3 weeks after treatment.
ECM and tissue remodeling quantified in pre-treatment serum were associated with response and survival outcomes in metastatic melanoma patients treated with IPI. This highlights the importance of addressing the ECM and stromal component non-invasively in future ICI studies.
Protein citrullination and degradation by matrix metalloproteinases (MMP) plays a central role in the pathology of rheumatoid arthritis (RA). Autoantibodies are known to target citrullinated ...vimentin. The aim of this study was to investigate the relationship between the blood levels of MMP-degraded and citrullinated vimentin (VICM), as compared with the levels of MMP-degraded and non-citrullinated vimentin (VIM), and the standard anti-CCP biomarker in RA patients undergoing treatment. Thus, VIM, VICM and anti-CCP were quantified by ELISA in serum samples from baseline and week 8 of patients (n = 257) with RA, treated with either tocilizumab (8 mg/kg), methotrexate (7.5−15 mg/kg) or a placebo and compared with a reference cohort (n = 64). The three biomarkers were elevated in RA serum compared with the reference cohort: medians were 1.7 vs. 0.8 ng/mL (p < 0.05) for VIM; 7.5 vs. 0.7 ng/mL (p < 0.0001) for VICM; 57 vs. 4 RU/mL (p < 0.001) for anti-CCP. VICM was decreased in response to tocilizumab (2.9-fold, p < 0.0001) and to methotrexate (1.5-fold, p < 0.05) compared with the placebo, while anti-CCP was not. Serum VIM was also modulated by both drugs, although to a lesser degree. A high baseline level of VICM was predictive of a low disease activity response at week 8. In conclusion, VICM can differentiate between RA and healthy donors in a similar manner to anti-CCP; furthermore, VICM is also a pharmacodynamic marker.
PDF
