A meta-analysis of previous falls and subsequent fracture risk in cohort studies.

The relationship between self-reported falls and fracture risk was estimated in an international meta-analysis of individual-level data from 46 prospective cohorts. Previous falls were associated with an increased fracture risk in women and men and should be considered as an additional risk factor in the FRAX® algorithm. INTRODUCTION: Previous falls are a well-documented risk factor for subsequent fracture but have not yet been incorporated into the FRAX algorithm. The aim of this study was to evaluate, in an international meta-analysis, the association between previous falls and subsequent fracture risk and its relation to sex, age, duration of follow-up, and bone mineral density (BMD). METHODS: The resource comprised 906,359 women and men (66.9% female) from 46 prospective cohorts. Previous falls were uniformly defined as any fall occurring during the previous year in 43 cohorts; the remaining three cohorts had a different question construct. The association between previous falls and fracture risk (any clinical fracture, osteoporotic fracture, major osteoporotic fracture, and hip fracture) was examined using an extension of the Poisson regression model in each cohort and each sex, followed by random-effects meta-analyses of the weighted beta coefficients. RESULTS: Falls in the past year were reported in 21.4% of individuals. During a follow-up of 9,102,207 person-years, 87,352 fractures occurred of which 19,509 were hip fractures. A previous fall was associated with a significantly increased risk of any clinical fracture both in women (hazard ratio (HR) 1.42, 95% confidence interval (CI) 1.33-1.51) and men (HR 1.53, 95% CI 1.41-1.67). The HRs were of similar magnitude for osteoporotic, major osteoporotic fracture, and hip fracture. Sex significantly modified the association between previous fall and fracture risk, with predictive values being higher in men than in women (e.g., for major osteoporotic fracture, HR 1.53 (95% CI 1.27-1.84) in men vs. HR 1.32 (95% CI 1.20-1.45) in women, P for interaction = 0.013). The HRs associated with previous falls decreased with age in women and with duration of follow-up in men and women for most fracture outcomes. There was no evidence of an interaction between falls and BMD for fracture risk. Subsequent risk for a major osteoporotic fracture increased with each additional previous fall in women and men. CONCLUSIONS: A previous self-reported fall confers an increased risk of fracture that is largely independent of BMD. Previous falls should be considered as an additional risk factor in future iterations of FRAX to improve fracture risk prediction.

Evaluation of collagen turnover biomarkers as an objective measure for efficacy of treatment with rurioctocog alfa pegol in patients with hemophilia A: a secondary analysis of a randomized controlled trial.

BACKGROUND: Patients with hemophilia who have recurrent hemarthroses develop hemophilic arthropathy (HA). Regular prophylaxis with factor (F) VIII (FVIII) can reduce HA, but there is a need for objective outcome measures to evaluate treatment efficacy. OBJECTIVES: Evaluate and assess collagen turnover biomarkers in patients with hemophilia A to determine the efficacy of rurioctocog alfa pegol treatment and understand their potential as tools for guiding treatment decisions and monitoring outcomes. METHODS: Joint remodeling was assessed by analyzing serum levels of collagen remodeling products at baseline and months 3, 6, 9, and 12 in a 98 patient subset receiving pharmacokinetics-guided prophylaxis with rurioctocog alfa pegol, targeting FVIII trough levels of 1 to 3 International Units (IU)/dL or 8 to 12 IU/dL (PROPEL study, NCT0285960). RESULTS: Basement membrane metabolism-related type 4 collagen remodeling products (C4M and PRO-C4) decreased after 3 months at all time points by up to 25% at 1 to 3 IU/dL (P = .049, P < .0001) and 8 to 12 IU/dL FVIII trough levels (P = .0002, P < .0001). Interstitial tissue metabolism-related type 3 (C3M) and 5 (PRO-C5) collagen remodeling products decreased after 3 months, by up to 19% at 1 to 3 IU/dL FVIII trough level (P = .0001, P = .009) and 23% at 8 to 12 IU/dL FVIII trough level (P = .0002, P = .001). An increase of up to 12% was seen for cartilage metabolism-related type 2 collagen product (PRO-C2, not C2M) after 6 months at both trough levels (P = .01, P = .005). When stratified by prior treatment, changes in C3M (P = .03) and C4M (P = .02) levels were observed between trough levels for prior on-demand treatment but not for prophylaxis prior to study entry. CONCLUSION: Joint improvement measured by collagen remodeling biomarkers specific to the basement membrane, interstitial matrix, and cartilage was seen with pharmacokinetics-guided prophylaxis. These collagen remodeling biomarkers warrant further exploration as biomarkers to guide treatment toward improvement in HA.

Citrullinated and MMP-degraded vimentin is associated with chronic pulmonary diseases and genetic variants in PADI3/PADI4 and CFH in postmenopausal women.

Citrullinated vimentin has been linked to several chronic and autoimmune diseases, but how citrullinated vimentin is associated with disease prevalence and genetic variants in a clinical setting remains unknown. The aim of this study was to obtain a better understanding of the genetic variants and pathologies associated with citrullinated and MMP-degraded vimentin. Patient Registry data, serum samples and genotypes were collected for a total of 4369 Danish post-menopausal women enrolled in the Prospective Epidemiologic and Risk Factor study (PERF). Circulating citrullinated and MMP-degraded vimentin (VICM) was measured. Genome-wide association studies (GWAS) and phenome wide association studies (PheWAS) with levels of VICM were performed. High levels of VICM were significantly associated with the prevalence of chronic pulmonary diseases and death from respiratory and cardiovascular diseases (CVD). GWAS identified 33 single nucleotide polymorphisms (SNPs) with a significant association with VICM. These variants were in the peptidylarginine deiminase 3/4 (PADI3/PADI4) and Complement Factor H (CFH)/KCNT2 gene loci on chromosome 1. Serum levels of VICM, a marker of citrullinated and MMP-degraded vimentin, were associated with chronic pulmonary diseases and genetic variance in PADI3/PADI4 and CFH/ KCNT2. This points to the potential for VICM to be used as an activity marker of both citrullination and inflammation, identifying responders to targeted treatment and patients likely to experience disease progression.

Tetrathiomolybdate (TM)-associated copper depletion influences collagen remodeling and immune response in the pre-metastatic niche of breast cancer.

Tetrathiomolybdate (TM) is a novel, copper-depleting compound associated with promising survival in a phase II study of patients with high-risk and triple-negative breast cancer. We sought to elucidate the mechanism of TM by exploring its effects on collagen processing and immune function in the tumor microenvironment (TME). Using an exploratory cohort, we identified markers of collagen processing (LOXL2, PRO-C3, C6M, and C1M) that differed between those with breast cancer versus controls. We measured these collagen biomarkers in TM-treated patients on the phase II study and detected evidence of decreased collagen cross-linking and increased degradation over formation in those without disease compared to those who experienced disease progression. Preclinical studies revealed decreased collagen deposition, lower levels of myeloid-derived suppressor cells, and higher CD4+ T-cell infiltration in TM-treated mice compared with controls. This study reveals novel mechanisms of TM targeting the TME and immune response with potential applications across cancer types.

Endotrophin is associated with chronic multimorbidity and all-cause mortality in a cohort of elderly women.

BACKGROUND: The signalling peptide endotrophin is derived through proteolytic cleavage of the carboxyl-terminal during formation of type VI collagen. It is expressed by most descendants of the mesenchymal stem cells lineage, including adipocytes and fibroblasts, and have been proposed to be a central extracellular matrix hormone associated with several age-related diseases. We aimed to assess the association of endotrophin with chronic disease incidence and death in older women. METHODS: 5,602 elderly Danish women from the observational, prospective cohort: The Prospective Epidemiological Risk Factor (PERF) study were included in the analysis which covered baseline (BL) and follow-up (FU) 14 years later. An elastic net was used to investigate the relative importance of 58 variables to serum endotrophin-levels. 20 chronic diseases were defined on the basis of clinical variables available along with diagnoses extracted from both the National Patient Register, the National Diabetes Register and the Danish Cancer Registry. The cross-sectional associations between endotrophin-levels and these 17 chronic age-related diseases were investigated using logistic regression and a set-analysis explored disease-combinations within multimorbidity. The association of endotrophin with mortality was assessed by Cox proportional hazard models. FINDINGS: Formation of type III collagen (PRO-C3), age and creatine-levels were the most influential variables of endotrophin-levels. Several chronic diseases were significantly associated with endotrophin-levels independent of age and BMI including chronic kidney disease (BL OR=3.7, p < 0.001; FU OR = 7.9 p < 0.001), diabetes (BL OR = 1.5, p = 0.0015, FU OR=1.6, p = 0.004) and peripheral arterial disease (BL OR = 1.3, p = 0.029; FU OR=2.4, p < 0.001). Lastly, endotrophin-levels were significantly rising with number of morbidities (p < 0.001) and a predictor of death after adjusting for age and BMI (BL HR=1.95; FU HR = 2.00). INTERPRETATION: Endotrophin was associated with death and increased with number of morbidities. Endotrophin may be a central hormone of fibroblast that warrant investigation and possible targeted intervention in several chronic diseases. FUNDING: The funder of the PERF study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.

Blood-based extracellular matrix biomarkers as predictors of survival in patients with metastatic pancreatic ductal adenocarcinoma receiving pegvorhyaluronidase alfa.

BACKGROUND: Extensive extracellular matrix (ECM) remodeling is a hallmark of metastatic pancreatic ductal adenocarcinoma (mPDA). We investigated fragments of collagen types III (C3M, PRO-C3), VI (PRO-C6), and VIII (C8-C), and versican (VCANM) in plasma as biomarkers for predicting progression-free survival (PFS) and overall survival (OS) in patients with mPDA treated with pegvorhyaluronidase alfa, a biologic that degrades the ECM component hyaluronan (HA), in a randomized phase 2 study (HALO109-202). METHODS: HALO109-202 comprised a discovery cohort (Stage 1, n = 94) and a validation cohort (Stage 2, n = 95). Plasma ECM biomarkers were analyzed by ELISAs. Univariate Cox regression analysis and Kaplan-Meier plots evaluated predictive associations between biomarkers, PFS and OS in patients treated with pegvorhyaluronidase alfa plus nab-paclitaxel/gemcitabine (PAG) versus nab-paclitaxel/gemcitabine (AG) alone. RESULTS: PFS was improved with PAG vs. AG in Stage 1 patients with high C3M/PRO-C3 ratio (median cut-off): median PFS (mPFS) 8.0 vs. 5.3 months, P = 0.031; HR = 0.40; 95% CI 0.17-0.92). High C3M/PRO-C3 ratio was validated in Stage 2 patients by predicting a PFS benefit of PAG vs. AG (mPFS: 8.8 vs. 3.4 months, P = 0.046; HR = 0.46; 95% CI 0.21-0.98). OS was also improved in patients with high C3M/PRO-C3 ratio treated with PAG vs. AG (mOS 13.8 vs 8.5 months, P = 0.009; HR = 0.35; 95% CI 0.16-0.77). Interestingly, high C3M/PRO-C3 ratio predicted for a PFS benefit to PAG vs. AG both in patients with HA-low tumors (HR = 0.36; 95% CI 0.17-0.79) and HA-high tumors (HR = 0.20; 95% CI 0.06-0.69). CONCLUSIONS: The C3M/PRO-C3 ratio measuring type III collagen turnover in plasma has potential as a blood-based predictive biomarker in patients with mPDA and provides additional value to a HA biopsy when applied for patient selection. TRIAL REGISTRATION: NCT01839487. Registered 25 April 2016.

Matrix metalloproteinase-degraded type I collagen is associated with APOE/TOMM40 variants and preclinical dementia.

OBJECTIVE: Dysregulation of type I collagen metabolism has a great impact on human health. We have previously seen that matrix metalloproteinase-degraded type I collagen (C1M) is associated with early death and age-related pathologies. To dissect the biological impact of type I collagen dysregulation, we have performed a genome-wide screening of the genetic factors related to type I collagen turnover. METHODS: Patient registry data and genotypes have been collected for a total of 4,981 Danish postmenopausal women. Genome-wide association with serum levels of C1M was assessed and phenotype-genotype association analysis performed. RESULTS: Twenty-two genome-wide significant variants associated with C1M were identified in the APOE-C1/TOMM40 gene cluster. The APOE-C1/TOMM40 gene cluster is associated with hyperlipidemia and cognitive disorders, and we further found that C1M levels correlated with tau degradation markers and were decreased in women with preclinical cognitive impairment. CONCLUSIONS: Our study provides elements for better understanding the role of the collagen metabolism in the onset of cognitive impairment.

Collagen biology and non-invasive biomarkers of liver fibrosis.

There is an unmet need for high-quality liquid biomarkers that can safely and reproducibly predict the stage of fibrosis and the outcomes of chronic liver disease (CLD). The requirement for such markers has intensified because of the high global prevalence of diseases such as non-alcoholic fatty liver disease (NAFLD). In particular, there is a need for diagnostic and prognostic tools, as well as predictive biomarkers that reflect the efficacy of interventions, as described by the BEST criteria (Biomarkers, EndpointS, and other Tools Resource). This review covers the various liver collagens, their functional role in tissue homeostasis and delineates the common nomenclature for biomarkers based on BEST criteria. It addresses the common confounders affecting serological biomarkers, and describes defined collagen epitope biomarkers that originate from the dynamic processes of extracellular matrix (ECM) remodelling during liver injury.

Atrial fibrillation is associated with lean body mass in postmenopausal women.

This study investigated the association between body composition and risk of atrial fibrillation (AF) in postmenopausal women. In a retrospective analysis we assessed data from 5704 postmenopausal women (age 70.7 ± 6.5 yrs.) who in 1999-2001 participated in The Prospective Epidemiological Risk Factor study with body composition assessed by dual-energy X-ray absorptiometry. Outcomes were obtained from Danish Health Registries and body composition association to risk of AF was evaluated by univariable and multivariable Cox Hazard regression. 850 women developed AF after baseline. High lean body mass was associated with increased risk of AF in multivariable analyses, adjusting for body mass index (BMI), height or weight (adjusted for: BMI, hazard ratio (HR) 1.49, 95% Confidence Interval (1.22-1.80); height, HR 1.27 (1.03-1.56); weight, 1.33 (1.06-1.65)). Height and weight were associated with increased risk of AF in multivariable analyses adjusting for body composition measures. When adjusting for total lean mass, only height remained statistically significant (HR 1.34 (1.09-1.64)). In a cohort of elderly Caucasian women, high lean body mass, height and weight were associated with increased risk of AF and the variables remained significant after adjusting for age and other known risk factors of AF.

Low bone turnover levels predict increased risk of cancer.

BACKGROUND: Several epidemiological studies have shown an association between bone mineral density (BMD) and risk of breast cancer in postmenopausal women, but it remains unknown whether bone turnover is associated with increased risk of cancer. The aim of this study was to investigate if markers of bone formation and resorption are associated with increased risk of cancer. MATERIAL AND METHODS: The study population included 5855 postmenopausal Danish women enrolled in the Prospective Epidemiologic Risk Factor (PERF) study. Cancer diagnosis was obtained from the Danish Cancer Registry. Baseline spine, femur, and whole-body BMD were evaluated by DXA-scanners. Baseline bone turnover (CTX-1 and osteocalcin) were measured in serum. Multivariate Cox analysis was performed with 3, 6 and 12 years of follow-up. All continuous variables were transformed into z-score for the cox analyses. RESULTS: 252 developed cancer after 3 years, 462 developed cancer after 6 years, and 881 developed cancer with 12 years of follow-up. CTX-1, osteocalcin and spine BMD were all predictors of cancer at all time points (3 years of follow-up: Spine BMD, HR = 1.20, p = 0.003; CTX-1, HR = 0.82, p = 0.005; osteocalcin, HR = 0.75, p < 0.001). After adjusting for cancer risk factors and other bone measures CTX-1 and osteocalcin remained independent predictors of cancer (3 years of follow-up: CTX-1, HR = 0.82, p = 0.02; osteocalcin, HR = 0.75, p = 0.002). CONCLUSIONS: We found that levels of the bone turnover markers CTX-1 and osteocalcin were inversely associated with risk of cancer independent of BMD and other known cancer risk factors in postmenopausal women.

Incidence of total hip and total knee replacements from the prospective epidemiologic risk factor study: considerations for event driven clinical trial design.

BACKGROUND: Osteoarthritis (OA) leads to joint failure and total joint replacement (TJR, either hip (H) or knee (K)). Worsening of pain and joint space narrowing are believed to be surrogates for joint failure; however, we hypothesize that TJR, as a reflection of joint failure, can be used as an endpoint in event-driven clinical trials within a reasonable duration. We explored the incidence of TJR in the Prospective Epidemiologic Risk Factor (PERF I) study. METHODS: A total of 5855 Danish postmenopausal women aged 49-88 enrolled in the PERF I study during 1999-2001 (baseline). Three-, six- and twelve-year follow-up data from the Danish National Patient Registry was collected, including occurrence of TJR and OA diagnosis. At baseline the women were asked whether they had OA. RESULTS: The women with a TJR diagnosis before or after baseline were on average 1 year older (p < 0.001) and heavier (p < 0.001), compared to women with no TJR. The 3-, 6- and 12-year cumulative incidences were 1.1, 2.4 and 6.0% for TKR, and 2.1, 4.4 and 9.3% for THR. For those with an OA diagnosis at baseline the respective incidences were 2.7, 5.6 and 11.7% and 3.9, 7.2 and 13.6% CONCLUSIONS: Within 3, 6 or 12 years TJR incidences were double for women with an OA diagnosis compared to the all-comer population. TJRs are frequent amongst elderly women with OA and it is, therefore, feasible to conduct event-driven clinical trials where TJR is the endpoint demonstrating clinical benefit of a novel disease-modifying OA drug (DMOAD).

Cancer risk in relation to body fat distribution, evaluated by DXA-scans, in postmenopausal women - the Prospective Epidemiological Risk Factor (PERF) study.

Studies with direct measures of body fat distribution are required to explore the association between central and general obesity to cancer risk in postmenopausal women. This study investigates the association between central obesity and general obesity to overall/site-specific cancer risk in postmenopausal women. The analysis included 4,679 Danish postmenopausal women. Body fat distribution was evaluated by whole-body dual-energy X-ray absorptiometry scanners. Cancer diagnoses were extracted from the Danish Cancer Registry and multivariable Cox regression models explored the association between cancer risk and central obesity after adjusting for BMI. Our results showed that high central obese women had a 50% increased risk of overall cancer relative to low central obese women (Q1vs.Q4: [HR:1.50, CI:1.20-1.88]). For site-specific cancers, central obesity was significantly associated with Respiratory (Q1vs.Q4: [HR:2.01, CI:1.17-3.47]), Gastrointestinal (Q1vs.Q4: [HR:1.55, CI:0.99-2.41]) and Female genital organs (Q1vs.Q4: [HR:1.95, CI:1.00-3.78]) cancer diagnoses. Sub-analyses stratified by smoking-habits found a significant association between central obesity and a cancer diagnosis for current (Q1vs.Q4: [HR:1.93, CI:1.25-2.99]) and former smokers (Q1vs.Q4: [HR:1.90, CI:1.23-2.94]). These analyses suggest that central obesity is associated with some cancers in postmenopausal women independent of BMI.

Matrix Metalloprotease Generated Fragments of Type VI Collagen Have Serum Biomarker Potential in Cancer - A Proof of Concept Study.

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.

Bone and Soft Tissue Turnover in Relation to All-cause Mortality in Postmenopausal Women.

BACKGROUND: The relationship between levels of extracellular matrix (ECM) turnover and mortality is currently unknown. The study aimed to determine if levels of ECM turnover are predictors of all-cause mortality in a large cohort of postmenopausal women. METHODS: 5,855 postmenopausal Danish women enrolled in the Prospective Epidemiologic Risk Factor (PERF) study. Baseline demographics and serum were collected at registration. Dates of death were obtained from the Danish Death Registry. ECM turnover was evaluated by serological biomarkers measuring bone (telopeptide of type I collagen [CTX-1] and osteocalcin) and soft tissue (formation of type VI collagen [PRO-C6], MMP-degraded type IV collagen [C4M], formation of type III collagen [PRO-C3], and MMP-degraded type I collagen [C1M]) turnover. Multivariate Cox analyses were performed with 3, 5, and 15 years of follow-up. RESULTS: The association of bone turnover (CTX-1 and osteocalcin) with all-cause mortality was U-shaped for all time periods. After adjustment for possible confounders, the lowest quintile of bone formation and degradation remained significant for all time periods. We observed J-shaped association between all-cause mortality and PRO-C6, C4M, and PRO-C3, and there was a linear association between C1M and all-cause mortality. After adjustment for possible confounders, the highest quintile of the soft tissue turnover biomarkers (PRO-C6, C4M, PRO-C3, and C1M) remained significantly associated with all-cause mortality for all time periods. CONCLUSION: Both low and high levels of tissue turnover were associated with increased risk of all-cause mortality in postmenopausal women. Overall, these results highlight the importance of bone and soft tissue homeostasis.

Matrix Metalloproteinase Mediated Type I Collagen Degradation is an Independent Predictor of Increased Risk of Acute Myocardial Infarction in Postmenopausal Women.

Acute myocardial infarction (AMI) is often underdiagnosed in women. It is therefore of interest to identify biomarkers that indicate increased risk of AMI and thereby help clinicians to have additional focus on the difficult AMI diagnosis. Type I Collagen, a component of the cardiac extracellular matrix, is cleaved by matrix metalloproteinases (MMPs) generating the neo-epitope C1M. We investigated the association between serum-C1M and AMI and evaluated whether C1M is a prognostic marker for outcome following AMI. This study is based on The Prospective Epidemiological Risk Factor (PERF) Study including postmenopausal women. 316 out of 5,450 women developed AMI within the follow-up period (14 years, median). A multivariate Cox analysis assessed association between serum-C1M and AMI, and re-infaction or death subsequent to AMI. The risk of AMI increased by 18% (p = 0.03) when serum-C1M was doubled and women in the highest quartile had a 33% increased risk compared to those in the low quartiles (p = 0.025). Serum-C1M was, however not related to reinfarction or death subsequent to AMI. In this study C1M was be an independent risk factor for AMI. Measuring MMP degraded type I collagen could be useful for prediction of increased risk of AMI if replicated in other cohorts.

Quantification of altered tissue turnover in a liquid biopsy: a proposed precision medicine tool to assess chronic inflammation and desmoplasia associated with a pro-cancerous niche and response to immuno-therapeutic anti-tumor modalities.

Immuno-therapy has begun to revolutionize cancer treatment. However, despite the significant progress achieved in regard to the duration of clinical benefits, a substantial number of patients do not respond to these therapies. To improve the outcome of patients receiving immuno-therapy, there is a need for novel biomarkers that can predict and monitor treatment. Tumor microenvironment alterations, more specifically the state of chronic inflammation and desmoplasia (tumor fibrosis), are important factors to consider in this context. Here, we discuss the potential for quantification of altered tissue turnover in a liquid biopsy as a proposed precision medicine tool to assess chronic inflammation and desmoplasia in the immuno-oncology (IO) setting. We highlight the need for novel non-invasive biomarkers in IO and the importance of addressing tumor microenvironment alterations. We focus on desmoplasia and extracellular matrix (ECM) remodeling, and how the composition of the ECM defines T-cell permissiveness in the tumor microenvironment and opens up the possibility for associated liquid biopsy biomarkers. Moreover, we address the importance of the assessment of chronic inflammation, primarily macrophage activity, in a liquid biopsy.

Excessive matrix metalloprotease-mediated degradation of interstitial tissue (type I collagen) independently predicts short-term survival in an observational study of postmenopausal women diagnosed with cancer.

Extensive tissue remodeling mediated by matrix metalloproteases (MMPs) is an important part of cancer. The aim of this study was to investigate whether serum biomarkers reflecting MMP-mediated degradation of type I collagen (C1M), type IV collagen (C4M) and citrullinated vimentin (VICM) were predictive of cancer-specific mortality. Between 1999 and 2001, 5855 Danish postmenopausal women participated in The Prospective Epidemiologic Risk Factor (PERF I) study. Demographics and serum samples were collected at enrolment. Cancer diagnosis, and cause and time of death were obtained from Danish registries. C1M, C4M and VICM were measured by ELISA. Hazard ratios (HR) and Kaplan-Meier curves were applied to assess mortality at 3 and 12 years of follow-up for women diagnosed with cancer within 3 years from blood sampling. Within 3 years from blood sampling, 250 women had been diagnosed with cancer. C1M and VICM were associated with survival over time at 3 years of follow-up. Only C1M was predictive of mortality at 3 years follow-up: the adjusted HR was 2.65 [95% CI: 1.08-6.51]. In conclusion, C1M and VICM are associated with survival in postmenopausal women with cancer, and C1M is an independent risk factor for cancer-specific mortality. Thus, quantification of tissue remodeling is important in cancer.

Unique insight into microenvironmental changes in colorectal cancer: Ex vivo assessment of matrix metalloprotease-mediated molecular changes in human colorectal tumor tissue and corresponding non-neoplastic adjacent tissue.

Matrix metalloprotease (MMP)-mediated tissue remodeling is one of the malignant changes driving colorectal cancer. Measurement of altered MMP expression/activity is not sufficient to fully understand the effect of MMP-mediated tissue remodeling. Biomarkers are required that specifically reflect the dynamic processes of the MMP-mediated degradation of signature proteins from colorectal tissue. The aim of the present study was to profile and characterize the release of MMP-degraded type III collagen (C3M) and citrullinated and MMP-degraded vimentin (VICM) from tumor tissue and corresponding non-neoplastic adjacent tissue (NAT) in a human colorectal cancer ex vivo model. Colorectal tumor tissue and NAT biopsies from tissue removed during resection of colorectal cancer patients (n=13) were cut into pieces of 2 mm2 and cultured for 24 h in growth medium. C3M and VICM were evaluated by ELISA. As part of the characterization, C3M was determined subsequent to the tumor tissue being cleaved with recombinant MMP-2/-9 and trypsin. C3M was generated by MMP-2/-9, but not by trypsin. In addition, the level of C3M was significantly elevated in the conditioned medium from tumor tissues (3.7 ng/ml) compared with that observed in the conditioned medium from the NATs (2.2 ng/ml) and in the growth medium alone (1.9 ng/ml). The level of VICM was significantly elevated in the tumor tissues (0.51 ng/ml) and NATs (0.52 ng/ml) compared with that in the growth medium alone (0.03 ng/ml). No differences were detected between the tumor tissues and NATs. No correlation was observed between biomarker levels from the tumor tissue and corresponding NAT, and the biomarker levels did not correlate with tumor stage. In conclusion, the present study provided support of the concept that C3M and VICM are applicable as tools to investigate dynamic tissue changes of colorectal tumor tissue and corresponding NAT. By the assessment of these specific MMP-mediated molecular changes, the present study provides novel and relevant insight into the dynamic changes of colorectal tumor tissue and corresponding NAT.

Nidogen-1 Degraded by Cathepsin S can be Quantified in Serum and is Associated with Non-Small Cell Lung Cancer.

Loss of basement membrane (BM) integrity is typically associated with cancer. Nidogen-1 is an essential component of the BM. Nidogen-1 is a substrate for cathepsin-S (CatS) which is released into the tumor microenvironment. Measuring nidogen-1 degraded by CatS may therefore have biomarker potential in cancer. The aim of this study was to investigate if CatS-degraded nidogen-1 was detectable in serum and a possible biomarker for cancer, a pathology associated with disruption of the BM. A competitive enzyme-linked immunosorbent assay (NIC) was developed with a monoclonal mouse antibody specific for a CatS cleavage site on human nidogen-1. Dilution and spiking recovery, inter- and intravariation, as well as accuracy were evaluated. Serum levels were evaluated in patients with breast cancer, small cell lung cancer (SCLC), and non-SCLC (NSCLC) and in healthy controls. The results indicated that the NIC assay was specific for nidogen-1 cleaved by CatS. Inter- and intraassay variations were 9% and 14%, respectively. NIC was elevated in NSCLC as compared to healthy controls (P<.001), breast cancer (P<.01), and SCLC (P<.5). The diagnostic power (area under the receiver operating characteristics) of NIC for NSCLC as compared to all other samples combined was 0.83 (95% confidence interval: 0.71-0.95), P<.0001. In conclusion, nidogen-1 degraded by CatS can be quantified in serum by the NIC assay. The current data strongly suggest that cathepsin-S degradation of nidogen-1 is strongly associated with NSCLC, which needs validation in larger clinical cohorts.

Remodeling of the Tumor Microenvironment Predicts Increased Risk of Cancer in Postmenopausal Women: The Prospective Epidemiologic Risk Factor (PERF I) Study.

BACKGROUND: An altered tumor microenvironment is one of the earliest signs of cancer and an important driver of the disease. We have seen previously that biomarkers reflecting tumor microenvironment modifications, such as matrix metalloproteinase (MMP)-degraded type 1 collagen (C1M), MMP-degraded type IV collagen (C4M), and citrullinated and MMP-degraded vimentin (VICM), were higher in the serum of cancer patients than in healthy controls. However, it is not known if these biomarkers could predict an increased risk of cancer. The aim of this study was to investigate whether C1M, C4M, and VICM were elevated prior to diagnosis of solid cancers in a large prospective study. METHODS: Between 1999 and 2001, 5,855 postmenopausal Danish women ages 48 to 89 years enrolled in the Prospective Epidemiologic Risk Factor study. Baseline demographics and serum were collected at the time of registration. Follow up cancer diagnoses were obtained from the Danish Cancer Registry in 2014. Serum C1M, C4M, and VICM levels were measured by competitive ELISAs. RESULTS: A total of 881 women were diagnosed with solid cancers after baseline. C1M, C4M, and VICM levels were significantly elevated in women diagnosed less than 1 year after baseline. C1M and VICM, but not C4M, were independent predictors of increased risk of cancer. CONCLUSION: C1M, C4M, and VICM are elevated prior to cancer diagnosis. C1M and VICM are both independent predictors of increased cancer risk. IMPACT: C1M and VICM are predictors for increased risk of cancer. Cancer Epidemiol Biomarkers Prev; 25(9); 1348-55. ©2016 AACR.