Low Tissue Inhibitor Of Metalloproteinases-1 Research Articles

MP03-02 ASSESSMENT OF MATRIX METALLOPROTEINASE-9 AND TISSUE INHIBITOR OF METALLOPROTEINASE-1 VIA M2 MACROPHAGE-SEEDED COLLAGEN MESH FOR TREATMENT OF STRESS URINARY INCONTENCE

You have accessJournal of UrologyCME1 Apr 2023MP03-02 ASSESSMENT OF MATRIX METALLOPROTEINASE-9 AND TISSUE INHIBITOR OF METALLOPROTEINASE-1 VIA M2 MACROPHAGE-SEEDED COLLAGEN MESH FOR TREATMENT OF STRESS URINARY INCONTENCE Ilaha Isali, Phillip McClellan, Thomas R. Wong, James M. Anderson, Ozan Akkus, and Adonis Hijaz Ilaha IsaliIlaha Isali More articles by this author , Phillip McClellanPhillip McClellan More articles by this author , Thomas R. WongThomas R. Wong More articles by this author , James M. AndersonJames M. Anderson More articles by this author , Ozan AkkusOzan Akkus More articles by this author , and Adonis HijazAdonis Hijaz More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003214.02AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Over the past decade, there have been several innovations with respect to macrophage-based therapies and biomaterials. Patients suffering from stress urinary incontinence (SUI) exhibit a significant increase in matrix metalloproteinase-9 (MMP-9) and a significant decrease in tissue inhibitor of metalloproteinase-1 (TIMP-1) expression. These findings are consistent with elevated rates of collagen breakdown and may play a critical role in the onset and development of SUI. Our recent study demonstrated improved biomechanical and histological outcomes following incorporation of M2 macrophages into a collagen mesh developed for tissue repair and offers future strategies focused on connective tissue regeneration. This study aims to assess the regulation of MMP-9 and TIMP-1 by M2 macrophages in terms of regenerating connective tissue in a rat model. METHODS: Macrophages were harvested from rat bone marrow, and macrophage subtypes were identified by flow cytometry. Electrochemically aligned collagen threads were filament wound to produce small collagen meshes and seeded with macrophages and cultured for up to 3 days. Macrophage subtype-seeded meshes were implanted subcutaneously in rats and harvested at 3 weeks and 3 months, respectively. Explanted meshes were analyzed using histology and immunohistochemistry. Specimens stained separately for MMP-9 and TIMP-1, and quantified to determine overall presence of MMP-9 and TIMP-1 in the harvested tissues. RESULTS: Immunohistochemical staining demonstrated a reduction of MMP-9 with addition of M2 macrophages, whereas the M1-seeded group continued to exhibit elevated MMP-9 after 3 months. TIMP-1 presence was elevated with the addition of M2 macrophages, whereas the M1-seeded group showed low TIMP-1 after 3 months (Figure 1). CONCLUSIONS: Delivery of M2 macrophages via collagen mesh can reduce MMP-9 while increasing TIMP-1, which may limit collagen breakdown associated with the development of SUI. This study shows the potential for improved restoration of extracellular matrix following the incorporation of M2 macrophages into a genipin-crosslinked collagen mesh for tissue repair and remodeling. Source of Funding: NIH grant: R21HD095439 © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e21 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ilaha Isali More articles by this author Phillip McClellan More articles by this author Thomas R. Wong More articles by this author James M. Anderson More articles by this author Ozan Akkus More articles by this author Adonis Hijaz More articles by this author Expand All Advertisement PDF downloadLoading ...

TIMP-1-Mediated Chemoresistance via Induction of IL-6 in NSCLC.

Elevated tissue inhibitor of metalloproteinase-1 (TIMP-1) is a negative prognosticator in non-small cell lung carcinoma NSCLC patients. This study sought to identify mechanisms whereby TIMP-1 impacts anticancer therapy. Using NSCLC cells and their TIMP-1 knockdown clones, we examined the chemoresistance against two chemotherapeutic agents, Gemcitabine and Cisplatin, as identified by increased apoptosis in the knockdown clones. A bead-based cytokine screening assay identified interleukin-6 (IL-6) as a key factor in chemoresistance. Exogenous human recombinant rhTIMP-1 or rhIL-6 resulted in reduced apoptosis. IL-6 expression was closely correlated with TIMP-1 kinetics and was upregulated by the addition of exogenous TIMP-1 while TIMP-1 neutralizing antibodies delayed IL-6 elevation. IL-6 production was regulated by TIMP-1, exerting its effect via activation of downstream signal transducer and activator of transcription 3 (STAT3) signaling. Both molecules and their documented transcription factors were upregulated and activated in chemoresistant NSCLC cells, confirming the roles of TIMP-1 and IL-6 in chemoresistance. To examine the role of these genes in patients, survival data from lung adenocarcinoma (LUAD) patients was curated from the cancer genome atlas (TCGA) database. Kaplan-Meier analysis found that individuals expressing low TIMP-1 and IL-6 have a higher survival rate and that the two-gene signature was more significant than the single-gene status. We define for the first time, a regulatory relationship between TIMP-1 and IL-6 in NSCLCs, suggesting that the TIMP-1/IL6 axis may be a valuable prognostic biomarker. Therapeutic interventions directed at this dual target may improve overall prognosis while negatively affecting the development of chemoresistance in NSCLC.

Expressions of Matrix Metalloproteinases-9 and Tissue Inhibitor of Metalloproteinase-1 in Pituitary Adenomas and Their Relationships with Prognosis.

To investigate the expression levels of matrix metalloproteinases-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in pituitary adenomas (PAs), and to analyze the relationship of the expressions of the two with the prognosis of patients. A total of 108 patients with PAs diagnosed in our hospital from May 2010 to May 2012 were selected and divided into the invasive PA (IPA) group (n = 58) and the non-IPA group (n = 50) according to the invasiveness of PAs. Hematoxylin and eosin (H&E) staining was used to observe the pathological state of patients. The expression levels of MMP-9 and TIMP-1 were measured by immunohistochemistry and western blotting at protein level and reverse transcription-polymerase chain reaction at gene level, respectively. The expression levels of MMP-9 and TIMP-1 in serum of patients before operation were tested using enzyme-linked immunosorbent assay, and patients with PAs after operation were followed up. The positive expression rate of MMP-9 in IPAs was significantly higher than that in non-IPAs, whereas that of TIMP-1 was relatively high in non-IPAs, and the differences were statistically significant (p < 0.05). At both protein and gene levels, MMP-9 was highly expressed in IPAs, whereas TIMP-1 was highly expressed in non-IPAs, and the differences were statistically significant (p < 0.05 in all comparisons). Before operation, the expression level of MMP-9 in serum of patients with IPAs was relatively high, whereas that of TIMP-1 in serum of patients with non-IPAs was relatively high, and the differences were statistically significant (p < 0.05 in all comparisons). The postoperative survival rate of patients with highly expressed MMP-9 was relatively low, whereas that of patients with highly expressed TIMP-1 was relatively high. The abnormal expressions of MMP-9 and TIMP-1 play important roles in the invasion process of PAs. The prognoses of patients with low expression MMP-9 and high expression TIMP-1 are more positive.

Survival Outcomes According to TIMP1 and EGFR Expression in Heavily Treated Patients with Advanced Non-small Cell Lung Cancer who Received Biweekly Irinotecan Plus Bevacizumab.

Heavily treated patients with non-small cell lung cancer (NSCLC) have few treatment options, while irinotecan and bevacizumab have proven synergistic action in preclinical studies. A total of 49 patients with heavily treated NSCLC were enrolled from 2011-2014 and treated with irinotecan and bevacizumab. Treatment response along with mutational status of epidermal growth factor receptor (EGFR), and tissue inhibitor of metalloproteinases-1 (TIMP1) and EGFR expression were evaluated. Progression-free (PFS) and overall (OS) survival were monitored. Median follow-up was 13.2 months. Twenty-three patients had received three or more prior therapy lines. Overall response rate was 32% [95% confidence interval (CI)=22%-39%] and 26% of patients achieved stable disease. Median PFS was 4.4 (95% CI=2.8-8.3) months and median OS 18.0 (95% CI=16.2-30.7) months. Nine patients harboring EGFR mutations had a long-lasting partial response. A shorter OS was found in patients with a higher TIMP1 expression (p=0.006). Irinotecan combined with bevacizumab had favorable antitumor activity in heavily pretreated patients with NSCLC. These results suggest this is a reasonable strategy, particularly for patients with low TIMP1 expression.

Overexpression of TIMP-1 and Sensitivity to Topoisomerase Inhibitors in Glioblastoma Cell Lines.

The multifunctional protein - tissue inhibitor of metalloproteinases-1 (TIMP-1) - has been associated with a poor prognosis in several types of cancers including glioblastomas. In addition, TIMP-1 has been associated with decreased response to chemotherapy, and especially the efficacy of the family of topoisomerase (TOP) inhibitors has been related to TIMP-1. As a second line treatment of glioblastomas, the vascular endothelial growth factor (VEGF) antibody bevacizumab is administered in combination with the TOP1 inhibitor irinotecan and glioblastoma cell levels of TIMP-1 could therefore potentially influence the efficacy of such treatment. In the present study, we aimed to investigate whether a high TIMP-1 expression in glioblastoma cell lines would affect the sensitivity to TOP inhibitors, and whether TIMP-1 overexpressing cells would have alterered growth and invasion. We established TIMP-1 overexpressing subclones from two human glioblastoma cell lines. TIMP-1 overexpressing U87MG cells were significantly more resistant than low TIMP-1 expressing clones and parental cells when exposed to SN-38 (TOP1 inhibitor) or epirubicin (TOP2 inhibitor). No significant differences were observed for the TIMP-1 transfected A172 cells. Implantation of both U87MG and A172 spheroids into organotypic brain slice cultures revealed a reduced growth of TIMP-1 overexpressing U87MG spheroids, however, no significant differences in invasion were observed. The present study suggests that TIMP-1 overexpression reduces the effect of TOP inhibitors in glioblastoma. TIMP-1 also appeared to reduce spheroid growth, but did not influence invasion. Whether TIMP-1 plays a role in irinotecan resistance and has a predictive potential in glioblastoma patients remains to be elucidated.

Phosphorylation of Rab37 by protein kinase C alpha inhibits the exocytosis function and metastasis suppression activity of Rab37.

We previously identified a novel Rab small GTPase protein, Rab37, which plays a critical role in regulating exocytosis of secreted glycoproteins, tissue inhibitor of metalloproteinases 1 (TIMP1) to suppress lung cancer metastasis. Patients with preserved Rab37 protein expression were associated with better prognosis. However, a significant number of the patients with preserved Rab37 expression showed poor survival. In addition, the molecular mechanism for the regulation of Rab37-mediated exocytosis remained to be further identified. Therefore, we investigated the molecular mechanism underlying the dysregulation of Rab37-mediated exocytosis and metastasis suppression. Here, we report a novel mechanism for Rab37 inactivation by phosphorylation. Lung cancer patients with preserved Rab37, low TIMP1, and high PKCα expression profile correlate with worse progression-free survival examined by Kaplan-Meier survival, suggesting that PKCα overexpression leads to dysfunction of Rab37. This PKCα-Rab37-TIMP1 expression profile predicts the poor outcome by multivariate Cox regression analysis. We also show that Rab37 is phosphorylated by protein kinase Cα (PKCα) at threonine 172 (T172), leading to attenuation of its GTP-bound state, and impairment of the Rab37-mediated exocytosis of TIMP1, and thus reduces its suppression activity on lung cancer cell motility. We further demonstrate that PKCα reduces vesicle colocalization of Rab37 and TIMP1, and therefore inhibits Rab37-mediated TIMP1 trafficking. Moreover, Phospho-mimetic aspartate substitution mutant T172D of Rab37 significantly promotes tumor metastasis in vivo. Our findings reveal a novel regulation of Rab37 activity by PKCα-mediated phosphorylation which inhibits exocytic transport of TIMP1 and thereby enhances lung tumor metastasis.

Abstract 5906: TIMP-1 expression is inversely correlated with miRNA125a-5p and let-7e in non-small-cell lung carcinoma

Abstract Epigenetic alterations are emerging as significant elements in our understanding the biology of lung cancer which remains the leading cause of cancer deaths worldwide. In earlier studies we have shown that tissue inhibitor of metalloproteinase 1 (TIMP-1) overexpression in NSCLC cells results in aggressive tumors in mice. We have also demonstrated TIMP-1’s anti-apoptotic activity in these cells. It is well documented that miRNAs are involved in many physiological and neoplastic processes, including apoptosis. We have previously shown that knocking down TIMP-1 in A549 NSCLC cells alters the miRNA profile in these cells with upregulation of miR-125a-5p. Subsequently, we have identified that miR let-7e is also upregulated in TIMP-1 KD clones. The present study is focused on confirming the involvement of miR-125a-5p with TIMP-1 by using mimics and antagomirs of 125a-5p. Furthermore, we have sought to validate this relationship in vivo by examining these patterns in clinical samples. We added mimics of miR-125a-5p into the parental A549 cells, which resulted in down regulation of TIMP-1 levels. We were then able to rescue TIMP-1 downregulation by adding antagomirs of miR-125a-5p to the same cells. MiR-125a-5p binding sites were identified in TIMP-1 3’ UTR and using luciferase assay, we have confirmed that TIMP-1 is indeed a bona fide target of miR-125a-5p. Translational studies were carried out using archived NSCLC adenocarcinoma tissues from the Surgical Pathology archives at AU Medical Center. We first measured expression level of TIMP-1 by RT-PCR in 20 tumors and paired normal adjacent tissues and divided the samples into 2 groups, those with low and high TIMP-1 expression levels. Tissues were then interrogated for TIMP-1 and miR-125a-5p and let- 7e miRNA expression using chromogenic in situ hybridization. We identified negative correlation pattern between tumor and paired normal adjacent tissue samples. Generally, TIMP-1 was highly expressed in the tumor region, while miRNA expression was relatively decreased when compared with adjacent normal tissues. miRNA 125a-5p and let- 7e are known tumor suppressors in NSCLC. Analyzing secondary data (adenocarcinoma) by KM plotter we found that high TIMP1 correlated with lower patient survival with a hazard ratio of 3.17 (2.3-4.37) and a log rank p value of 8.1e-14. These studies provide additional clarity and further extend our understanding of the relationship between these miRNA families and TIMP-1 expression. Citation Format: Ammar Kutiyanawalla, Sampa Ghoshal-Gupta, Byung R. Lee, Ashis Mondal, Ravindra Kolhe, Amyn M. Rojiani, Mumtaz V. Rojiani. TIMP-1 expression is inversely correlated with miRNA125a-5p and let-7e in non-small-cell lung carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5906. doi:10.1158/1538-7445.AM2017-5906

Small GTPase Rab37 targets tissue inhibitor of metalloproteinase 1 for exocytosis and thus suppresses tumour metastasis.

Rab small GTPases are master regulators of membrane trafficking and guide vesicle targeting. Recent publications show that Rab-controlled trafficking pathways are altered during tumorigenesis. However, whether any of the Rabs plays a metastasis suppressor role is least explored. Here we address the metastasis suppressive function of human Rab37 (hRAB37) using secretomics, cell, animal and clinical analyses. We show that tissue inhibitor of metalloproteinase 1 (TIMP1), a secreted glycoprotein that inhibits extracellular matrix turnover, is a novel cargo of hRAB37. hRAB37 regulates the exocytosis of TIMP1 in a nucleotide-dependent manner to inactivate matrix metalloproteinase 9 (MMP9) migration axis in vitro and in vivo. Dysfunction of hRAB37 or TIMP1 abrogates metastasis suppression. Lung cancer patients with metastasis and poor survival show low hRAB37 protein expression coinciding with low TIMP1 in tumours. Our findings identify hRAB37 as a novel metastasis suppressor Rab that functions through the TIMP1-MMP9 pathway and has significant prognostic power.

TIMP1 overexpression mediates resistance of MCF-7 human breast cancer cells to fulvestrant and down-regulates progesterone receptor expression

High levels of Tissue Inhibitor of Metalloproteinases-1 (TIMP1) are associated with poor prognosis, reduced response to chemotherapy, and, potentially, also poor response to endocrine therapy in breast cancer patients. Our objective was to further investigate the hypothesis that TIMP1 is associated with endocrine sensitivity. We established a panel of 11 MCF-7 subclones with a wide range of TIMP1 mRNA and protein expression levels. Cells with high expression of TIMP1 versus low TIMP1 displayed significantly reduced sensitivity to the antiestrogen fulvestrant (ICI 182,780, Faslodex®), while TIMP1 levels did not influence the sensitivity to 4-hydroxytamoxifen. An inverse correlation between expression of the progesterone receptor and TIMP1 was found, but TIMP1 levels did not correlate with estrogen receptor levels or growth-promoting effects of estrogen (estradiol, E2). Additionally, the effects of fulvestrant, 4-hydroxytamoxifen, or estrogen on estrogen receptor expression were not associated with TIMP1 levels. Gene expression analyses revealed associations between expression of TIMP1 and genes involved in metabolic pathways, epidermal growth factor receptor 1/cancer signaling pathways, and cell cycle. Gene and protein expression analyses showed no general defects in estrogen receptor signaling except from lack of progesterone receptor expression and estrogen inducibility in clones with high TIMP1. The present study suggests a relation between high expression level of TIMP1 and loss of progesterone receptor expression combined with fulvestrant resistance. Our findings in vitro may have clinical implications as the data suggest that high tumor levels of TIMP1 may be a predictive biomarker for reduced response to fulvestrant.

P4-01-17: TIMP-1 Over-Expression Confers Resistance of MCF-7 Breast Cancer Cells to Fulvestrant.

Abstract Background Endocrine resistance represents a major challenge in the management of estrogen receptor (ER) positive breast cancer. Currently no predictive biomarkers for endocrine resistance in ERpositive breast cancer patients are in clinical use. In a clinical study, patients with metastatic breast cancer and high levels of serum Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) had less benefit from endocrine therapy than patients with a lower level of serum TIMP-1 [1]. Therefore, we evaluated the association between TIMP-1 and response to endocrine therapy using an in vitro approach. We have previously presented initial results on TIMP-1 and response to endocrine therapy [2]. Materials and Methods: MCF-7 cells were stably transfected with pcDNA3.1(Hyg)-TIMP-1 plasmid, and a panel of 11 subclones with different expression levels of TIMP-1 was generated. TIMP-1 expression levels were confirmed using enzyme-linked immunosorbent assay (ELISA). Four subclones with high or low TIMP-1 expression were analyzed for the growth response to estrogen, 4-hydroxytamoxifen and fulvestrant. These four subclones were analyzed for protein expression by western blotting. All subclones were analyzed by whole human genome oligo microarrays 4×44K for determination of gene expression levels. Data were analyzed using the limma R/Bioconductor package. Paired-end Solexa sequencing was applied to selected subclones with high and low TIMP-1 levels to identify transcriptomic changes. Results: High expression of TIMP-1 was associated with resistance to fulvestrant, whereas growth response to either estrogen or 4-hydroxytamoxifen was independent of TIMP-1 expression levels. High expression of TIMP-1 protein and mRNA was associated with undetectable levels of progesterone receptor (PgR) protein and mRNA whereas ER protein and mRNA levels were unaffected by TIMP-1. To characterize the potential role of TIMP-1 in estrogen signaling we analyzed the expression of reported estrogen-responsive genes and no general change was observed. We identified genes that correlated positively or negatively to TIMP-1 expression. Among the identified genes was PgR, which is a direct target for ER. Conclusion: Our data suggest that a high expression of TIMP-1 in vitro is associated with resistance to fulvestrant but not to 4-hydroxytamoxifen. Estrogen-regulated genes are not generally affected by changes in TIMP-1 expression levels and therefore TIMP-1 appears to affect endocrine resistance through other mechanisms than globally regulating ER signaling. However, high expression of TIMP-1 is associated with loss of PgR and this may be related to the resistance towards fulvestrant.

Serum TIMP-1 Predicts Survival Outcomes of Invasive Breast Carcinoma Patients: A Meta-analysis

Tissue inhibitor of metalloproteinase-1 (TIMP-1) is a small secretory glycoprotein with multifunctional activity including anti-apoptosis and the inhibition of matrix metalloproteinase in invasive breast carcinomas. There have been contradictory results as to whether TIMP-1 is a poor or good prognostic factor in breast cancer patients. To address this controversy, we conducted a meta-analysis for the relationship between TIMP-1 levels and prognostic parameters in the breast cancer. The relevant published studies were pooled according to the defined selection criteria. The effect sizes of overall survival and prognostic parameters were calculated by a hazard ratio (HR) or an odds ratio (OR). HRs or ORs were combined using a random-effects model. Survival outcomes between high or elevated and low or normal serum TIMP-1 levels were compared by uni- and multivariate analyses involving 886 and 844 breast cancer patients, respectively. Patients with high or elevated serum TIMP-1 levels had unfavorable survival outcomes compared to patients with low or normal serum TIMP-1 levels in the uni- and multivariate analyses (HR, 1.7 and 2.4; p <0.001 and p= 0.033, respectively). However, no survival difference was evident in the data from tissue TIMP-1 levels by enzyme-linked immunosorbent assay and the expression of tissue TIMP-1 mRNA. The high or positive immunohistochemical expression of tissue TIMP-1 protein was not related to adjusted and unadjusted HRs, lymph node metastasis, and clinical stages. This meta-analysis indicates that serum TIMP-1 levels may be useful for predicting survival outcomes of invasive breast cancer patients.

TIMP-1 Status Measured by Immunohistochemistry in 268 ER Positive Postmenopausal High-Risk Breast Cancer Patients Treated with Adjuvant Tamoxifen: Association to Recurrence Free Survival.

Abstract Background: A proportion of ER positive breast cancer patients do not benefit from adjuvant tamoxifen treatment. Tamoxifen has been shown to induce apoptosis in human breast cancer cells in vitro and Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) protects cancer cells against apoptosis. We therefore raised the hypothesis that ER positive breast cancer cells with high TIMP-1 immunoreactivity would show reduced benefit to tamoxifen treatment.Material and methods: Tumor tissue from 268 high-risk postmenopausal ER positive breast cancer patients was used for this study. All patients underwent surgery for primary invasive breast cancer. The majority of the patients were lymph node positive: 92% (246/268). Patients were allocated to 1, 2 or 5 years of adjuvant tamoxifen as mono-therapy. Recurrent disease was diagnosed in 33% (89/268) of the patients. Median time to recurrence was 3.1 years and the median time of follow-up was 12.4 years. Archival formalin-fixed and paraffin-embedded primary tumor tissue was used to generate tissue microarrays comprising two, 2 mm cores from each tumor. TIMP-1 immunoreactivity was evaluated using the mouse monoclonal antibody (clone VT7) raised against recombinant human TIMP-1. This antibody exclusively reacts with TIMP-1. A central assessment of TIMP-1 status was performed. Tumors were analysed by bright field microscopy and scored semi-quantitatively as 0 - 3 with regard to the number of stained tumour cells and intensity of the staining. An index combining the number of positive tumor cells and staining intensity was created: low (0-2), median (3-4) and high expression (5-6).Results: IHC was successfully performed in 266 cases. TIMP-1 IHC negative cases were observed in 39% (105/266) of the tumors and 61% (161/266) were TIMP-1 IHC positive. The index combining the number of positive tumor cells and staining intensity showed low expression in 59% (157/266), median expression in 24% (65/266) and high expression in 17% (44/266) of the tumors. TIMP-1 negativity and low TIMP-1 index showed no significant association with tumor grade, tumor size or lymph node status. When performing Kaplan-Meier plots, TIMP-1 expression was not significantly associated with recurrence free survival in univariate analyses (p = 0.1).Conclusion: This study did not confirm our hypothesis that TIMP-1 immunoreactivity in ER positive breast cancer cells are associated with reduced benefit from adjuvant tamoxifen treatment. This is partly in contrast with prior studies on endocrine therapy of metastatic breast cancer (Lipton et al 2008) where high serum TIMP-1 was associated with reduced response to endocrine therapy. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 2025.

Low expression of tissue inhibitor of metalloproteinases-1 (TIMP-1) in glioblastoma predicts longer patient survival

In colorectal cancer and breast cancer a high TIMP-1 level has been shown to correlate with a shorter overall patient survival and it has been suggested that TIMP-1 is involved in tumour invasion, proliferation and apoptosis in different types of cancers. TIMP-1 is known to be expressed in gliomas but whether TIMP-1 is a prognostic marker in gliomas has not previously been investigated. In the present study, the TIMP-1 expression was investigated immunohistochemically in 112 formalin-fixed paraffin embedded astrocytomas and related to tumour grade and overall patient survival by scoring the TIMP-1 immunoreactivity of both tumour cells and blood vessels. Moreover, TIMP-1 in situ hybridisation was performed on ten of the glioblastomas. In the vast majority of the tumours TIMP-1 protein was expressed in both tumour cells and blood vessels. In situ hybridisation for TIMP-1 mRNA on glioblastomas confirmed the immunohistochemical expression of TIMP-1. The percentage of TIMP-1 positive tumour cells and blood vessels as well as the staining intensity varied between tumours of the same grade, but the total staining score increased with tumour grade. The multivariate Cox regression test showed that glioblastoma patients with the lowest TIMP-1 expression had a significantly longer overall survival (HR (95% CI) = 3.2 (1.5-6.7), P = 0.004) when compared to the patients with higher TIMP-1 protein expression. In conclusion, this study showed that low TIMP-1 immunohistochemical expression predicts longer overall survival in glioblastoma patients, suggesting a role for TIMP-1 as a biomarker in glioblastoma.

Tumor tissue levels of Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) and outcome following adjuvant chemotherapy in premenopausal lymph node-positive breast cancer patients: A retrospective study

BackgroundWe have previously demonstrated that high tumor tissue levels of TIMP-1 are associated with no or limited clinical benefit from chemotherapy with CMF and anthracyclines in metastatic breast cancer patients. Here, we extend our investigations to the adjuvant setting studying outcome after adjuvant chemotherapy in premenopausal lymph node-positive patients. We hypothesize that TIMP-1 high tumors are less sensitive to chemotherapy and accordingly that high tumor tissue levels are associated with shorter survival.MethodsFrom our original retrospectively collected tumor samples we selected a group of 525 pre-menopausal lymph node-positive patients (adjuvant treatment: CMF, 324 patients; anthracycline-based, 99 patients; no adjuvant chemotherapy, 102 patients). TIMP-1 levels were measured using ELISA in cytosolic extracts of frozen primary tumors. TIMP-1 was analyzed as a continuous variable and as a dichotomized one using the median TIMP-1 concentration as a cut point between high and low TIMP-1 groups. We analyzed the benefit of adjuvant CMF and anthracyclines in univariate and multivariable survival models; endpoints were disease-free (DFS) and overall survival (OS).ResultsIn this selected cohort of high-risk patients, and in the subgroup of patients receiving no adjuvant therapy, TIMP-1 was not associated with prognosis. In the subgroup of patients treated with anthracyclines, when analyzed as a continuous variable we observed a tendency for increasing TIMP-1 levels to be associated with shorter DFS (multivariable analysis, HR 1.75, 95% CI 1.00-3.07, P = 0.05) and a significant association between increasing TIMP-1 and shorter OS in both univariate (HR 3.52, 95% CI 1.54-8.06, P = 0.003) and multivariable analyses (HR 4.19, 95% CI 1.67-10.51, P = 0.002). No statistically significant association between TIMP-1 and DFS was observed in the CMF-treated patients although high TIMP-1 was associated with shorter OS when analyzed as a dichotomized variable (HR 1.64, 95% CI 1.02-2.65, P = 0.04).ConclusionIn the subgroup of patients receiving adjuvant chemotherapy we found an association between shorter survival after treatment in TIMP-1 high patients compared with TIMP-1 low patients, especially in patients receiving anthracycline-based therapy. This suggests that high tumor tissue levels of TIMP-1 might be associated with reduced benefit from classical adjuvant chemotherapy. Our findings should be validated in larger prospective studies.

Correlation of plasma TIMP-1 levels and disease-free survival in primary colorectal cancer independent of serum CEA levels

e22040 Background: Introduction of independent prognostic markers may play a significant role in future treatment of early stage colorectal cancer (CRC). CEA is still the only recommended (ASCO and EGTM) serological marker in CRC. However, Tissue Inhibitor of Metallo Proteinases-1 (TIMP-1), which is a glycoprotein that acts as an inhibitor of most of the active matrix metalloproteinases, has previously been shown to carry independent prognostic information in patients with primary CRC. The purpose of the present study was to assess the combination of preoperative serum CEA and plasma TIMP-1as prognostic markers in patients undergoing resection for primary CRC. Methods: In the present prospective study serum and plasma samples were collected before surgery from 422 patients with primary CRC stage I-III. CEA was determined in serum by a commercial assay and TIMP-1 was determined in plasma using a thoroughly validated, in-house ELISA. Time to recurrence or death of CRC was registered and the association to serum CEA and plasma TIMP-1 levels were studied in a Cox multivariate model including age, gender, disease stage and tumor location. Hazard ratios and 95% confidence intervals (HR (95%CI)) for disease free survival (DFS) were calculated. Results: An event was recorded in 186 patients, 75 had local recurrence, 103 had a distant metastases (28 of these patients had both local recurrence and distant metastases) and 36 died from their cancer without a registered recurrence. Scoring CEA and TIMP-1 as continuous variables on a logarithmic scale (base 2), both serum CEA and plasma TIMP-1 were statistically significant in a multivariable analysis with HR=1.12 (1.03–1.21) and HR=1.51 (1.12–2.04), respectively. Additional calculations of low CEA plus low TIMP-1, high CEA plus low TIMP-1, low CEA plus high TIMP-1 and high CEA plus high TIMP-1 showed that high plasma TIMP-1 levels carried prominent information of poor prognosis independent of CEA. Conclusions: Preoperative serum CEA and plasma TIMP-1 levels were independent predictors of disease free survival for patients with primary CRC. Combination of the two proteins showed that TIMP-1 was a prominent predictor of poor DFS independent of CEA. [Table: see text]

Tumor tissue levels of tissue inhibitor of metalloproteinases-1 (TIMP-1) and survival following adjuvant chemotherapy in pre-menopausal lymph node-positive breast cancer patients (N=525).

Abstract Abstract #6054 Predictive markers are needed to guide planning of adjuvant therapy for breast cancer patients. We have recently shown that high tumor tissue levels of TIMP-1 are associated with decreased response to chemotherapy in metastatic breast cancer (Schrohl et al, Clin Cancer Res, 2006) suggesting that TIMP-1 may be a predictive marker in breast cancer patients.&amp;#x2028; Purpose: This study investigates the association of tumor tissue TIMP-1 levels with response to adjuvant chemotherapy with CMF (cyclophosphamide/methotrexate/5-fluorouracil) or an anthracycline-based regimen.&amp;#x2028; Patients and Methods: 525 pre-menopausal lymph node-positive patients were included; 324 patients received adjuvant CMF, 99 received an adjuvant anthracycline-containing regimen and 102 had no adjuvant chemotherapy. Total TIMP-1 levels were measured using ELISA in cytosolic extracts of frozen primary tumors. Using the untreated patient group as a reference group we analyzed the benefit of adjuvant CMF and anthracyclines in TIMP-1 high and low patients, respectively. The median TIMP-1 concentration was used to dichotomize patients into high and low TIMP-1 groups. End points were disease-free and overall survival (DFS, OS).&amp;#x2028; Results: The median TIMP-1 level in the total patient group was 12,54 ng/mg of total protein (range, 0 – 112,9 ng/mg). TIMP-1 levels in subgroups according to adjuvant therapy were not significantly different (P=0,20). In a multivariate model including basic clinico-pathological parameters, TIMP-1 low and high patients benefited differentially from adjuvant CMF and anthracyclines when compared to untreated patients. In particular, patients with high tumor levels of TIMP-1 had little benefit from adjuvant anthracyclines. Hazard ratios (HR) and 95% confidence intervals (CI) are given in the table for the analysis of DFS. A similar pattern was seen in the analyses of OS. In the CMF-treated group, both TIMP-1 low and high patients had significantly better survival than untreated patients (P&amp;lt;0,01). Among anthracycline-treated patients those with TIMP-1 low tumors appeared to benefit more from the adjuvant therapy than TIMP-1 high patients although these results did not reach significance in the present analyses.&amp;#x2028; &amp;#x2028; Conclusion: This study suggests that high tumor tissue TIMP-1 levels are associated with decreased benefit from adjuvant chemotherapy. Especially in the group treated with anthracycline-based therapy there is a strong tendency for TIMP-1 high tumors to be less sensitive to the treatment. This group, however, is small and should be enlarged to confirm our results. In the group treated with CMF all patients benefit from the therapy with TIMP-1 high patients having slightly less benefit than TIMP-1 low patients. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 6054.

High serum TIMP-1 correlates with poor prognosis in breast carcinoma – A validation study

A number of studies have demonstrated that high tumor tissue levels of tissue inhibitor of metalloproteinases-1 (TIMP-1) are associated with a poor prognosis in breast cancer, suggesting that TIMP-1 could be a valid prognostic marker in this disease. Recently, our laboratories have presented results showing that TIMP-1 also carries prognostic information when measured in serum. This is an important finding, since serum is a much more preferable material compared with tumor tissue extracts. The aim of the present study was to validate the previous results concerning the prognostic value of TIMP-1 in serum obtained preoperatively from 68 patients with primary breast cancer. This was done by measuring the same serum samples as in the previous study but in a different laboratory using a different ELISA assay. We confirmed that patients with the highest serum levels of TIMP-1 (> 197.7 ng/ml) had significantly shorter disease-specific survival compared with patients with low serum TIMP-1 levels. In the group of node-negative patients, 53% of the patients with high levels of TIMP-1 survived after 10 years of follow-up compared to 92% of the patients with low levels. This study thus confirms the reproducibility across laboratories of the results concerning the prognostic value of TIMP-1 in serum. We also investigated whether measurements of the specific fraction of uncomplexed TIMP-1 improved the prognostic value of TIMP-1 in serum, as has been shown to be the case for tumor tissue extracts. However, including information of the level of uncomplexed TIMP-1 did not seem to provide additional prognostic information to that already provided by total TIMP-1.

TIMP-1 Is Significantly Associated with Objective Response and Survival in Metastatic Colorectal Cancer Patients Receiving Combination of Irinotecan, 5-Fluorouracil, and Folinic Acid

Tissue inhibitor of metalloproteinase-1 (TIMP-1) is known to protect cells against apoptosis. We raised the hypothesis that elevated tumor tissue levels and thereby plasma levels of TIMP-1 would predict resistance to apoptosis-inducing chemotherapy. Ninety patients with metastatic colorectal cancer were included in the study. Plasma TIMP-1 and serum carcinoembryonic antigen (CEA) were measured in samples obtained before the first cycle of chemotherapy. Analysis of best objective response (complete or partial response versus stable or progressive disease) showed that patients with low plasma TIMP-1 had higher probability of obtaining an objective response [odds ratio (OR), 3.5; 95% confidence interval (95% CI), 1.4-8.5, P=0.007]. CEA treated as a continuous variable was also a statistically significant predictor of no response (OR, 1.3; 95% CI, 1.0-1.7, P=0.02, area under the curve 0.66) but much less so. Plasma TIMP-1 was the only significant covariate in a multivariable analysis of best objective response (OR, 3.6; 95% CI, 1.4-9.5; P=0.001). Plasma TIMP-1 scored as a continuous variable on the log scale (log(e)) was significantly associated with overall survival [OS; hazard ratio (HR), 3.8; 95% CI, 2.4-5.9; P<0.0001] and with time to progression (TTP; HR, 1.5; 95% CI, 1.0-2.3; P=0.048). Multivariable analysis showed that plasma TIMP-1 was significant for OS when including routine clinical baseline covariates (HR, 3.5; 95% CI, 2.1-5.8; P<0.0001). A multivariable analysis including TTP instead of OS showed that only plasma TIMP-1 was retained in the model (HR, 1.5). CEA was not significantly associated with TTP or OS when TIMP-1 was included in the model. This study shows that plasma TIMP-1 levels are significantly and independently associated with objective response, TTP, and OS in patients with metastatic colorectal cancer receiving combination chemotherapy.

Serum levels of tissue inhibitor of metalloproteinases-1 (TIMP-1) in colorectal cancer patients.

Expression of tissue inhibitor of metalloproteinases (TIMP)-1 in colorectal cancer tissue is known to be related to disease progression; however, the clinical significance of measuring the blood level of TIMP-1, which we evaluate herein, has not yet been clarified. The serum level of TIMP-1 was measured by a one-step enzyme immunoassay in 123 patients who underwent resection of primary colorectal cancer. An elevated level of serum TIMP-1 was associated with advanced Dukes' stage ( P = 0.03), greater diameter of the primary tumor ( P = 0.03), more lymph node metastasis ( P = 0.04), and liver metastasis ( P < 0.001). There was a weakly positive correlation between the serum carcinoembryonic antigen (CEA) level and the serum TIMP-1 level. In patients who underwent potentially curative resection, the disease-free survival was not different between those with a high TIMP-1 level (>=203.5 ng/ml, n = 32) and those with a low TIMP-1 level (<203.5 ng/ml, n = 66, P = 0.62). In patients with Dukes' stage D cancer who underwent noncurative resection, the survival times were not different between those with a high TIMP-1 level ( n = 13) and those with a low TIMP-1 level ( n = 10, P = 0.20). Elevated levels of serum TIMP-1 reflect the extent of colorectal cancer, without a close correlation with the serum CEA level. These findings suggest that measuring the serum TIMP-1 level would not help to predict the prognosis of patients with colorectal cancer.

Evaluation of matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 levels in gingival fibroblasts of cyclosporin A-treated patients.

Cyclosporin A (CsA) is a potent immunosuppressant used to prevent organ transplant rejection and to treat various autoimmune diseases. CsA-induced gingival overgrowth (CsA GO) is the most widely seen side effect of this drug; its pathogenesis is not completely understood. The aim of this study was to identify and compare matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) levels in gingival fibroblast cultures of tissues derived from renal transplant patients receiving CsA and exhibiting gingival overgrowth and from periodontally healthy control subjects. Gingival overgrowth samples were obtained from patients undergoing therapy with CsA, and control tissues were obtained from systemically healthy donors. Gingival fibroblasts were grown using explant cultures. Three different study groups were identified: 1) CsA GO fibroblast culture; 2) CsA-treated healthy gingival fibroblast culture (H+CsA); and 3) healthy gingival fibroblast culture (H). The levels of MMP-1 and TIMP-1 in these groups of gingival fibroblasts were analyzed by enzyme-linked immunoabsorbent assay (ELISA). The levels of TIMP-1 were significantly lower in CsA GO than H (P < 0.05). There was no statistically significant difference in the levels of MMP-1 between H and CsA GO (P = 0.505). The ratio of MMP-1 to TIMP-1 was significantly higher in CsA GO than H (P < 0.05). The results of this study indicate that CsA therapy does not have a significant effect on MMP-1 levels. However, low TIMP-1 levels can be an important factor in the pathogenesis of CsA GO, since the balance between MMP-1 and TIMP-1 levels was changed by CsA.