| 1. |
- Holmgren, Klas, 1989-
(författare)
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Permanent stoma after anterior resection for rectal cancer : prevalence and mechanisms
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- While sphincter-saving surgery constitutes standard treatment for rectal cancer, anterior resection still harbours a significant risk of a permanent stoma in the long run. Although anastomotic leakage plays a major role in this surgical dilemma, the exact mechanisms are not known, while surveys indicate a stoma-free outcome is essential for a majority of patients. To address this issue, the overall aim of the present thesis was to investigate the permanent stoma prevalence in patients undergoing anterior resection for rectal cancer in Sweden, and to identify plausible mechanisms that impede prospects of a stoma-free outcome.In a population-based cohort, chart review of patients who had anterior resection for rectal cancer in the Northern healthcare region in Sweden between 2007 and 2013 showed that 75 out of 316 (24%) patients ended up with a permanent stoma. Of 274 patients (87%) primarily defunctioned with a stoma, 229 underwent stoma closure, 21 (9%) of whom suffered major complications that required return to theatre or worse. A permanent stoma was shown to be more common among patients with anastomotic leakage and an advanced tumour stage.A registry-based method to estimate nationwide stoma outcome after anterior resection for rectal cancer was developed, using data from the Swedish Colorectal Cancer Registry and the National Patient Registry. With a chart-reviewed cohort as reference, stoma outcome was assessed with a positive predictive value of 85.1%, and a negative predictive value of 100.0%. In patients operated in Sweden between 2007 and 2013, the registry-based method determined that 942 out of 4768 (19.8%) had a permanent stoma, while stoma rates varied substantially between different healthcare regions.In a 1:1 matched case-control study of 82 patients who had curative resection for non-disseminated colorectal cancer, a subgroup analysis of 34 patients with rectal cancer displayed biomarker aberrations in serum measured preoperatively in those with anastomotic leakage. Compared to complication-free controls, 15 proteins related to inflammation were elevated, of which two (C-X-C motif chemokine 6, and C-C motif chemokine 11) remained significant after adjustment for multiple testing.Based on a cohort of 4529 patients who had anterior resection, tumour height served as a proxy to determine the extent of mesorectal excision, while long-term stoma outcome was classified using a previously validated registry-based method. Defunctioning stomas significantly decreased chances of a stoma-free outcome, especially in patients undergoing partial mesorectal excision; for these patients, faecal diversion was also least beneficial in terms of reducing anastomotic leakage.In conclusion, every fifth patient undergoing anterior resection for rectal cancer in Sweden eventually ends up with a permanent stoma. Although construction of a defunctioning stoma decreases the risk of symptomatic anastomotic leakage, subsequent takedown surgery carries a substantial risk of major complications, while chances of a long-term stoma-free outcome become significantly reduced. To facilitate selective use of faecal diversion, novel markers to identify high-risk anastomoses prior to surgery have been identified, but require validation in larger prospective settings. Anterior resection without a defunctioning stoma should be considered in appropriately informed patients for whom a stoma-free outcome is of importance. In particular, this holds true for patients eligible for partial mesorectal excision, where anastomotic dehiscence is less frequent and the advantageous effects of a defunctioning stoma are limited.
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| 2. |
- Bayadsi, Haytham, 1987-
(författare)
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Tumour stromal and demographical factors affecting the metastatic aggressiveness of small differentiated papillary thyroid cancers in Sweden
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Background: The incidence of papillary thyroid cancer (PTC) has been increasing over the recent decades, especially that of small papillary thyroid cancers (sPTCs) (≤ 20mm in size). sPTCs are generally classified as low risk cancers with a very favourable diagnosis, yet some of these cancers still cause locoregional and distant metastasis, recurrence and even death.Aims: To investigate the role of tumour stromal, environmental and demographical factors affecting the metastatic aggressiveness of sPTCs in Sweden.Material & Methods: Selected tumour stromal proteins (Types I (Col1) and IV (Col4) collagens, alpha smooth muscle actin (a-SMA) and matrix metallopeptidase 9 (MMP-9)) were analysed for their role in metastatic disease (Paper I). Demographic and clinicopathological differences regarding recurrence between metastasized vs. non-metastasized sPTCs in Sweden were studied in 2 registry-based retrospective observational cohort studies (Papers II & III). The geographic distribution of patients with sPTC in Sweden was pinpointed and layered with maps of gamma radiation deposits of radionuclides Caesium-137 (Cs-137), Thorium-232 (Th-232), Uranium-238 (U-238) and Potassium-40 (K-40) using different spatial analysis methods (Paper IV).Results: Col1 and Col4 were significantly more expressed in the non-metastatic tumours compared with metastatic ones. Patients with N1b disease were younger, had a smaller tumour size and higher recurrence rates compared to patients with N0 and N1a disease. The mean number of metastatic LNs at initial surgery was higher in the N1b group than the N1a group and correlated with more recurrent disease. The prevalence of metastatic sPTC was associated with significantly higher levels of gamma radiation from Th-232, U-238 and K-40.Conclusions: The higher expression of Col1 and Col4 in the non-metastasized tumours indicates a potential protective role in tumour progression. LN stage N1b at diagnosis, and having five or more metastatic nodes, are strong risk factors for cancer recurrence and decreased disease-free survival in sPTC. Environmental factors such as gamma radiation are believed to play a major role in the pathogenesis of the PTC. These findings altogether underscore the importance of LN evaluation, tumour biological as well as environmental factors in sPTC patients, suggesting that the management of these patients should be based on an individual risk stratification instead of a “one size fits all” approach.
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| 3. |
- Jansson, Malin, 1978-
(författare)
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The role of stroma-derived substances in breast cancer progression and their function as tumour markers
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Background: In 2020, more than 2,260,000 women were diagnosed with breast cancer. Most patients are cured with surgery and adjuvant treatment, but despite that, approximately 700,000 women die of the disease every year. The historical focus on breast cancer progression has been on the malignant epithelial cell. However, cancer cells do not grow in isolation. In recent years, the importance of the tumour microenvironment in cancer progression has been highlighted. Perlecan and type IV collagen are basement membrane (BM) proteins in the normal mammary gland, and type I collagen is the main fibrillar collagen in the interstitial extracellular matrix (ECM). In cancer development, perlecan and type IV collagen have multifunctional roles and when degraded from the BM, bioactive substances and other fragments are released in the circulation. Significant ECM changes also occur that lead to an accumulation of fibrillar collagens. Given their abundance in the ECM; perlecan, type IV and type I collagen are of interest for breast cancer progression and may be of importance as new biomarkers to monitor disease, predict patient outcome and the treatment effect.Aim: In this thesis, the protein and mRNA expression of perlecan, type IV and I collagen in breast cancer tissue is studied. The aim is to characterize the expression pattern of these proteins in breast cancer tissue and to see whether there is a correlation to known prognostic biomarkers and to the patient prognosis. Moreover, to evaluate circulating perlecan and type IV collagen as diagnostic and prognostic biomarkers in breast cancer patients.Methods: In this thesis project, eight different patient cohorts were used. In freshly frozen normal breast and breast cancer tissue, perlecan protein expression was visualized using immunofluorescence. Type IV and I collagen protein expression were studied with immunohistochemistry in formalin-fixed, paraffin embedded primary breast cancer tissue, and type IV collagen in metastatic breast cancer tissue. For gene expression analysis, mRNA and clinicopathological data were extracted from the Cancer Genome Atlas and cBioportal database. Circulating plasma levels of perlecan were analysed in breast cancer patients and controls, circulating levels of CA15-3 and type IV collagen in patients with primary and metastatic breast cancer as well as controls. Perlecan and type IV collagen were measured with ELISA assays, and CA15-3 were using an electrochemiluminescence immunoassay.Results: In breast cancer tissue, perlecan and type IV collagen protein expression in the epithelial BM was fragmented or completely lost, and perlecan and type IV collagen was expressed to varying extent in the tumour stroma. The mRNA analysis confirmed that type IV collagen mRNA was expressed in primary breast cancer tissue and highly expressed in metastatic tissue. Type I collagen was mostly highly expressed in the tumour stroma. Low type I collagen protein and mRNA expression correlated with biomarkers for aggressive breast cancer, but no effect on survival could be seen. Among patients receiving chemotherapy, low stromal type I collagen protein expression was associated with better survival compared to high expression, even after adjusting for other relevant factors. There was no correlation of perlecan or type IV collagen protein expression to clinically used prognostic biomarkers, but an oestrogen receptor dependent correlation between mRNA expression of perlecan and several matrix-degrading enzymes were found. Survival analysis showed that high stromal type IV collagen protein and mRNA expression in the primary tumour was significantly associated with a poorer survival, and high protein expression with a risk of developing distant metastasis. Metastatic breast cancer patients had higher levels of circulating type IV collagen compared to healthy controls and patients with primary breast cancer. High circulating type IV collagen levels correlated with poorer survival in metastatic breast cancer patients, and was superior to CA15-3 at detecting metastatic breast cancer.Conclusions: The protein expression pattern of perlecan, type IV collagen and type I collagen become abnormal during breast cancer development. Stromal type IV collagen protein and mRNA in the primary tumour correlates to poorer prognosis, most likely due to a higher risk of developing metastatic disease. Circulating type IV collagen can function as a biomarker for detecting metastatic disease in patients with primary breast cancer and is prognostic in patients with metastatic breast cancer. Low stromal type I collagen is a marker for an aggressive breast cancer disease and can predict chemotherapy response.
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| 4. |
- Nyström, Hanna, 1980-
(författare)
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Stromal collagens in colorectal cancer and in colorectal liver metastases : tumour biological implications and a source for novel tumour markers
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Background: Colorectal cancer (CRC) remains one of the leading causes of cancer-related mortality. About 50 % of patients with CRC will develop subsequent liver metastases (CLM). The survival for untreated CLM is only a few months and liver resection provides the only chance for a lasting cure. It is therefore essential to detect CLM early, enabling successful surgical resection and achieving a long-term cure. There are no optimal tumour markers for CRC or CLM. The best marker available is Carcinoembryonic Antigen (CEA), a marker found elevated in about 50-60% of patients with CLM, but also in many other conditions. The main focus of cancer research has been on the malignant cancer cell. However, a tumour consists of more than cancer cells. A major part of all solid tumours is made up by the stroma. The tumour stroma is defined as the non-malignant cells of a tumour such as fibroblasts, the cells of the vascular and immune systems as well as the extracellular matrix (ECM). The basement membrane (BM) is a specialized form of the ECM in which type IV collagen is the major protein component. All epithelial cells need a contact to the BM and the definition of an invasive cancer is the degradation of the BM and the spread of cancer cells beyond this structure. Different metastatic growth patterns of CLM have previously been described, namely the desmoplastic, pushing and replacement type of CLM. These differ in their stromal reaction in the border, which separates the tumour from the normal liver. In this thesis the tumour stroma of CRC and CLM is studied with a special emphasis on stromal collagens. The aim is to investigate whether stromal collagens/ circulating type IV collagen can be used as tumour markers for CRC and CLM, and to compare this to the conventional marker CEA. The circulating type IV collagen level is also measured in liver metastases from other primary tumours than CRC. Furthermore, the differences between the stroma of a primary CRC that metastasizes to the liver when compared to a CRC that never spreads are analysed. Additionally, the metastatic growth pattern of CLM is studied in relation to the primary tumour, stromal components and survival. We also sought out to find whether CRC cell lines possess the trait to produce ECM proteins endogenously, and in response to a normal liver stroma in a novel organotypic model for CLM.Methods: Expression patterns of type I, III and IV collagen were studied by immunofluorescence (IF), chemical staining and immunohistochemistry (IHC) in normal colorectal tissue, normal liver, CRC, CLM, benign liver lesions and in liver metastases of other origin than CRC. Circulating plasma levels of type IV collagen were analysed in healthy controls, patients with CRC (T stage I-III) and in patients with CLM. Samples were analysed at the time of diagnosis, during and after oncological and surgical treatment and at the time of relapsing or progressive disease. Additionally, circulating levels were analysed in patients with benign liver lesions and in liver metastases of other origin than CRC. The metastatic growth pattern of CLM was classified according to earlier descriptions. CRC cell lines were studied regarding their production of type IV collagen. The growth, invasiveness and stromal production in CRC cell lines were also investigated in a new organotypic model for CLM using human liver specimens.Results: Circulating type IV collagen levels are increased in patients with CLM and other epithelial-derived liver metastases, and is found normal in patients with primary CRC (stage I-III), with liver metastases from tumours of non-epithelial origin, benign liver lesions and in healthy controls. The type IV collagen levels in patients with CLM reflect the tumour burden in the liver, decreases in response to therapy and is found increased in progressive or relapsing disease. The combination of circulating type IV collagen and CEA increased the sensitivity and specificity for detecting CLM. Livermetastatic CRC displayed an increased stromal production when compared to non-metastatic CRC, with an increased type IV collagen expression in the direct vicinity of the CRC cells. The earlier described growth patterns of CLM were verified, with the pushing type of CLM associated with a short survival and poor outcome. Furthermore, CRC cell lines possess the trait of endogenously producing type IV collagen. The novel organotypic liver model revealed that CRC cell lines grown in the context of normal liver stroma, devoid of other cells, does not elicit a desmoplastic reaction.Conclusion: Circulating type IV collagen is a promising tumour marker for CLM, where the levels reflect the hepatic tumour burden and can detect disease relapse after liver surgery. The combination of the tumour markers CEA and type IV collagen is superior to CEA alone. The stromal composition of primary CRC predicts the risk of subsequent CLM and the metastatic growth pattern of CLM is related to survival.
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| 5. |
- Bendrik, Christina, 1964-
(författare)
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Angiogenesis regulation in hormone dependent breast- and ovarian cancer
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Angiogenesis is a key event in tumor progression and a rate-limiting step in the establishment of a clinical cancer disease. The net balance of pro- and anti-angiogenesis mediators in the tissue dictates the angiogenic phenotype of a tumor. Matrix metalloproteinases (MMPs) are major regulators of extracellular matrix turnover and have for long been associated with pro-tumorigenic activities due to their tissue degradation capacities. However, broad-spectra MMP inhibitors as anti-tumor therapy in clinical trials have failed, and it has now become evident that several MMPs may induce biological activities beneficial to the host, such as suppressed angiogenesis. In this thesis the protective role of specific MMPs in breast and ovarian tumor tissues was further demonstrated.The process of angiogenesis is essential for physiological functions in the female reproductive tract, where sex steroids regulate new blood vessel formation and regression in each cycle. Despite progress made during the past years, our knowledge in sex steroid regulation of angiogenesis in hormone-dependent tumor tissues remains limited. Tamoxifen is a cornerstone in the treatment of estrogen receptor (ER)-positive breast cancer. The therapeutic value of tamoxifen in the treatment of ER-positive ovarian cancer is to date less investigated. The results presented in this thesis suggest that tamoxifen may induce anti-tumorigenic responses in ER-positive ovarian cancer by means of both anti-proliferative and anti-angiogenic mechanisms. In experimental models of human ovarian cancer in vitro and in vivo, tamoxifen treatment increased extracellular levels of MMP-9 and enhanced generation of the angiogenesis inhibitor endostatin which resulted in significantly decreased angiogenesis and tumor growth. Low levels of MMP-9 and endostatin in ascites collected from ovarian cancer patients suggest a possibility to therapeutically enhance MMP-9 by administration of tamoxifen, and thereby counteract angiogenesis in ovarian tumors by increased generation of anti-angiogenesis fragments, such as endostatin.The significance of enhanced MMP activities in tumor tissues was further investigated by experimental models of intratumoral MMP gene transfer to human breast tumor xenografts, which were assessed by using microdialysis. Treatment of tumors with MMP-9 or MMP-3 resulted in dose-dependent inhibition of tumor growth. Low dose of either MMP induced tumor stasis whereas a higher dose induced significant tumor regression. MMP-9 and tamoxifen exerted synergistic therapeutic effects on breast tumor angiogenesis and growth whereas gene transfer of the MMP-inhibitor TIMP-1 counteracted the beneficial effects induced by tamoxifen.Further on, we confirm the pro-angiogenic potential of estradiol by demonstrating a significant correlation between local levels of estradiol and the pro-angiogenic cytokine IL-8 in normal human breast tissues and in ER/PgR-positive breast cancers of women. Estradiol-induced IL-8 secretion was additionally confirmed in normal human whole breast biopsies in culture and in experimental human breast cancer in vitro and in vivo.In conclusion, the results of this thesis may hopefully increase the overall understanding of several mechanisms involved in angiogenesis regulation and may additionally be useful in the development of novel approaches for targeted therapy in the treatment of hormone-sensitive breast- and ovarian cancer.
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| 6. |
- Borgmästars, Emmy, 1990-
(författare)
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In search of early biomarkers in pancreatic ductal adenocarcinoma using multi-omics and bioinformatics
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Background: Pancreatic ductal adenocarcinoma (PDAC) is a very aggressive malignancy with a 5-year survival of 10 %. Surgery is the only curative treatment. Unfortunately, few patients are eligible for surgery due to late detection. Thus, we need ways to detect the disease at an earlier stage and for that good screening biomarkers could be used. Previous studies have analyzed circulating analytes in prospective studies to identify early PDAC signals. One such class is microRNAs (miRNAs). MicroRNAs are non-coding RNAs of around 22 nucleotides that act as post- transcriptional regulators by interaction with messenger RNAs (mRNAs). The function of a miRNA can be elucidated by target prediction, to identify its potential targets, followed by enrichment analysis of the predicted targets. Challenges with this approach includes a lot of false positives being generated and that miRNAs can perform their role in a tissue- or disease-specific manner. Other classes of analytes that have previously been studied in prospective PDAC cohorts are metabolites and proteins. Aims: This thesis has three aims. First, to build a miRNA functional analysis pipeline with correlation support between miRNA and its predicted target genes. Second, to identify potential circulating biomarkers for early detection of PDAC using multi-omics. Third, to identify potential prognostic metabolites in a prospective PDAC cohort.Methods: We used publicly available data from the cancer genome atlas-pancreatic adenocarcinoma (TCGA-PAAD) and pre-diagnostic plasma samples from the Northern Sweden Health and Disease Study. We built a pipeline in R including miRNA, mRNA, and protein expression data from TCGA-PAAD for in silico miRNA functional analysis. Pre- diagnostic plasma samples from future PDAC patients as well as matched healthy controls were analyzed using multi- omics. Tissue polypeptide specific antigen (TPS) was analyzed by enzyme linked immunosorbent assay in 267 future PDAC samples and 320 healthy controls. Metabolomics and clinical biomarkers (carbohydrate antigen (CA) 19-9, carcinoembryonic antigen (CEA), and CA 15-3) were profiled in 100 future PDAC samples and 100 healthy controls using liquid chromatography-mass spectrometry (MS), gas chromatography-MS, and multi-plex technology. Of these, a subset of 39 future PDAC patients and 39 healthy controls were profiled for 2083 microRNAs using targeted sequencing and 644 proteins using proximity extension assays. Circulating levels of multi-omics analytes were analyzed using conditional or unconditional logistic regression. Least absolute shrinkage and selection operator (LASSO) in combination with 500 bootstrap iterations identified the most informative variables. The prognostic value of metabolites was assessed using cox regression. Multi-omics factor analysis (MOFA) and data integration analysis for biomarker discovery using latent components (DIABLO) were used for multi-omics integration analyses.Results: An automated pipeline was built consisting of 1) miRNA target prediction, 2) correlation analyses between miRNA and its targets on mRNA and protein expression levels, and 3) functional enrichment of correlated targets to identify enriched Kyoto encyclopedia of genes and genomes (KEGG) pathways and gene ontology (GO) terms for a specific miRNA. The pipeline was run for all microRNAs (~700) detected in the TCGA-PAAD cohort. These results can be downloaded from a shiny app (https://emmbor.shinyapps.io/mirfa/). TPS was not altered in pre-diagnostic PDAC patients up to 24 years prior to diagnosis, but increased at diagnosis (OR = 1.03, 95 % CI: 1.01-1.05). Internal area under curves of 0.74, 0.80, and 0.88 were achieved for five metabolites, two proteins, and two miRNAs that were selected by LASSO and bootstrap iterations, in combination with CA 19-9. Neither MOFA nor DIABLO separated well between future PDAC cases and healthy controls. Conclusions: Our bioinformatics pipeline for in silico functional analysis of microRNAs successfully identifies enriched KEGG pathways and GO terms for miRNA isoforms. The investigated plasma samples are heterogeneous, but among the analyzed variables, we identified five metabolites, two proteins, and two microRNAs with highest potential for early PDAC detection. CA 19-9 levels increased closer to diagnosis. We identified five fatty acids that could be studied in a diagnostic PDAC cohort as prognostic biomarkers.
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| 7. |
- Grahn, Oskar, 1985-
(författare)
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Modulating the inflammatory response after colorectal cancer surgery : friend or foe?
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Colorectal cancer was the second most deadly and third most common cancer globally in 2020. In Sweden, more than 5,000 new colonic cancer cases and more than 2,000 rectalcancer cases were reported in 2021, making colorectal cancer the third most common in Sweden (excluding skin malignancies).Anastomotic leakage after colorectal cancer surgery is a feared complication that confers substantial morbidity, including a higher risk of permanent stoma and cardiovascular morbidity, but can also impart an increased risk of recurrence and mortality; the reason why leakage might cause this is not established. Perioperative inflammation including upregulation of cyclooxygenase-enzymes, which is further increased by anastomotic leakage, can possibly modulate both anastomotic healing as well as impact minimal residual disease. Non-steroidal anti-inflammatory drugs (NSAIDs) act by inhibiting COX-enzymes and can be part of a postoperative multimodal analgesia protocol. However, their postoperative use has been debated, with fears of NSAIDs possibly increasing anastomotic leakage rates.Study I was a retrospective cohort study on 1,341 patients who had undergone anterior resection for rectal cancer. Exposure was at least two days with NSAIDs during the first postoperative week, and the primary outcome was recurrence-free survival. A Cox regression model could not demonstrate a significant association with a hazard ratio (HR) of 1.02 (95% confidence interval (CI): 0.79–1.33) and neither did a propensity score-matched analysis. An instrumental variable analysis displayed a tentative improvement in recurrence-free survival in the NSAID-exposed (HR 0.61; 95% CI 0.38–0.99), but the core assumptions to perform such an analysis were not fully satisfied.Study II was a protocol-based retrospective cohort study with a total of 6,945 patients resected for colorectal cancer with a primary anastomosis formed. NSAID-exposure was determined by each individual hospital’s postoperative analgesia protocol, while patient data and outcomes were retrieved from the Swedish colorectal cancer registry. Some 3,996 (58%) patients were treated at a hospital with NSAIDs included in their postoperative analgesia protocol. No significant association with recurrence-free survival was seen (HR 0.97, 95% CI0.87–1.09). However, a reduction in cancer recurrence was demonstrated (HR 0.83, 95% CI0.72‒0.95), with an increased risk reduction for locoregional (HR 0.68, 95% CI 0.48–0.97) in comparison to distant recurrence (HR 0.85, 95% CI 0.74–0.98). Anastomotic leakage was less frequent as well, mainly because of a reduction in the group with colorectal or ileorectal anastomoses (HR 0.47, 95% CI 0.33–0.68).In Study III, the aim was to explore proteomic and biological pathway alterations in patients with peritoneal infection. This was a 1:1 matched cohort study on patients resected for colorectal cancer with a primary anastomosis formed, including 32 cases who suffered a postoperative peritoneal infection matched with 32 controls with a complication-free postoperative stay. Serum samples were retrieved from their first postoperative visit and at one year postoperatively. Out of a total of 270 proteins tested, 77 were differentially expressed at the first postoperative visit at a median sampling time of 41 days after surgery. Many of the top hub proteins are known actors in colorectal cancer progression, including survival and invasiveness, potentially enhancing minimal residual disease. Over-represented pathways were related to cardiomyopathy, cell-adhesion, extracellular matrix, phosphatidylinositol-3-kinase/Akt (PI3K-Akt) and transforming growth factor beta (TGF-Beta) signalling.In Study IV, the aim was to evaluate the frequency of a known polymorphism of the COX-2 gene promotor -765G>C in a Swedish cohort of colorectal cancer patients, and whether a previously reported association between this gene variant with an increase in anastomotic leakage could be reproduced. This was a 1:1 matched case-control study on a total of 94 patients who were resected for colorectal cancer with a subsequent primary anastomosis, with cases suffering a peritoneal infection. Preoperative blood and serum samples were genotyped and analysed using pre-defined protein panels. Of the 94 patients in total, one in each group were homozygous for the minor allele C/C, and ten cases and 14 controls were heterozygous with G/C and the rest were homozygous for the major allele. Thus, there were fewer individuals with the minor allele in the case group, with a non-significant odds ratio of 0.71(p=0.413), ultimately not replicating the finding of the previous study. The protein quantitative trait loci analysis rendered no associations of interest.In conclusion, no statistically significant effects on recurrence-free survival from postoperative NSAIDs in patients resected for colorectal cancer could be demonstrated in study I, whereas significant associations between NSAID use and reduction in frequency of anastomotic leaks as well as cancer recurrence could be shown in study II. In study III, numerous proteins were differentially expressed in patients suffering a postoperative peritoneal infection, even after more than a month’s duration, potentially stimulating minimal residual disease. The over-representation analysis found pathways related to cardiomyopathy, which could help explain the increase in cardiovascular morbidity in patients suffering anastomotic leakage. Study IV could not reproduce the potentially marked increase in anastomotic leak frequency in carriers of the COX-2 gene promotor -765G>C polymorphism in a Swedish sample. Whether to include NSAIDs or not in postoperative multimodal analgesia is a question still not answered, and it may depend on the genotype, the patient’s preoperative inflammatory state, tumour location, the specific NSAID used, and whether a leak has already occurred. NSAIDs might have effects on both morbidity including cardiovascular and anastomotic leakage as well as minimal residual disease including recurrence and mortality. This thesis suggests potential protective effects regarding both anastomotic leakage as well as cancer recurrence, but it seems to depend on at least some of the aforementioned factors. The proteomic landscape regarding postoperative peritoneal infection has been investigated, and where it has also been demonstrated that the duration of said alterations can be greater than was earlier suspected. Finally, even if a replication attempt was not successful considering the relation between a COX-2 gene promotor polymorphism and anastomotic leakage, it could be worthwhile to attempt further replication studies.
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| 8. |
- Holsti, Mari, 1963-
(författare)
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Vascular remodelling and circulating basement membrane fragments in abdominal aortic aneurysm
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- An abdominal aortic aneurysm (AAA) is a degenerative disease, characterized by advanced inflammation and extracellular matrix (ECM) remodelling. Enhanced protease activity mediated by cytokines results in the degradation of ECM proteins, leading to the generation of different bioactive fragments. Some of these generated fragments are released from the vascular basement membrane (VBM), a highly specialized ECM. VBM provides mechanical and structural stability and regulates many important cellular functions of the vascular system. Type IV and XVIII collagens are two structural proteins in VBM, with crucial roles in maintaining of the VBM integrity and vascular architecture. Circulating levels of type IV and XVIII collagen fragments are found physiologically, but have also been associated with many diseases. Remodelling of VBM and expression of its components has not been as well studied in AAA as that of the interstitial ECM.Here we investigate these VBM collagens, their expression and possible association with aortic diameter and expansion rate in individuals with an AAA in comparison with different control groups. Further we study whether there is a link between the circulating VBM collagen fragments and several inflammatory markers, all highly involved in AAA pathogenesis. Lastly, we study the impact of surgical intervention on plasma levels of VBM collagens in patients treated by either open surgical repair (OSR) or endovascular aortic aneurysm repair (EVAR).Methods: Circulating levels of type IV and XVIII collagen fragments were analysed in individuals with an AAA and compared with healthy controls and patients with peripheral artery disease (paper I). A possible association between VBM collagen fragments and the aortic diameter and expansion was studied in a large population-based cohort of 615 men stratified into three aortic diameter groups based on initial maximum aortic diameter (paper II). Furthermore, 159 individuals were followed up over time with repeated measurements of aortic diameter and blood samples. The follow up cohort were divided into two subgroups based on expansion rate of AAA. Moreover, the location of VBM collagens in tissue from aortic wall in individuals with an AAA was characterized and the expression pattern was compared with normal aorta (paper II). In paper III, the association between the plasma levels of VBM collagens and inflammatory markers; IL-1 (IL-1α and IL-1β), IL-6, IL-8, TNF-α INF-γ and hs-CRP were studied in same cohort as paper II. Finally, the effect of surgical intervention on circulating levels of VBM collagen fragments was investigated in AAA patients who had undegone either OSR or EVAR by comparison of plasma levels before and after AAA repair.Ultrasound technique was used for measurements of aortic diameter (paper I, II, III and IV). Analysis of circulating VBM collagens and inflammatory markers were performed by ELISA-assay (Paper I, II, III and IV) and Multiplex-assays, respectively (paper III). Aortic wall tissues were analysed by haematoxylin and eosin (H&E) and immunofluorescence staining (Paper II).Results: There were significantly increased plasma levels of VBM collagen fragments in individuals with an AAA, compared with healthy controls and individulas with a peripheral artery disease (PAD), (Paper I). The levels of type IV collagen in AAA patients did not differ from the group with PAD, and there were no significant differences between the control groups regarding plasma levels of both VBM collagen fragments (Paper I). The increased levels of VBM collagen fragments were significantly associated with aortic diameter with highest levels in the group with an AAA (Paper II). Altered expression of the VBM collagens and fragmentation of elastic fibres were observed in tissue from AAA patients (Paper II). A significant association between the levels of pro-inflammatory cytokines IL-6 and IL-8, and VBM collagens was found. Additionally, there were a significant association between the plasma levels of IL-8, TNF-α and hs-CRP and an AAA (Paper III). Aneurysms with faster expansion rate had significantly higher levels of IL-6, IL-1β, and type XVIII/endostatin collagen. Additionally, IL-6, type XVIII/endostatin collagen and baseline-aortic diameter were significantly associated with expansion rate (Paper III). AAA repair was associated with changes in plasma levels of VBM collagens (Paper IV).Conclusion: Circulating levels of VBM collagens were increased in patients with an AAA, and significantly associated with aortic diameter and expansion rate. The expression of VBM collagens was altered in AAA tissue compared with normal aorta. In addition, plasma levels of several inflammatory markers were associated as with VBM collagens, aortic diameter and expansion rate. The levels of both VBM collagens were altered at short and long time after AAA repair.
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| 9. |
- Lindgren, Moa, 1990-
(författare)
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Type IV collagen in breast and colorectal cancer : a potential biomarker of metastatic disease
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Background: Metastatic colorectal cancer (mCRC) and metastatic breast cancer (mBC) are two leading causes of cancer-related mortality worldwide. Early detection of metastatic disease is critical, and sensitive, easily accessed and cost-effective biomarkers that can diagnose mBC and mCRC at an early stage would have high clinical value. The best circulating markers: CEA and CA 15-3 are suboptimal, but a combination with other proteins can improve their potential to detect metastatic disease. One potential source of new biomarkers is the tumor stroma, including the extracellular matrix (ECM), vasculature, and stromal cells like immune cells and fibroblasts. Both the tumor cell and stromal compartment are vital for cancer progression. A combination of biomarkers from both compartments could likely best reflect the heterogeneous nature of the metastatic disease. Stromal type IV collagen (COL IV) is the main constituent of the basement membrane of healthy tissues. COL IV is upregulated with some cancers, including colorectal liver metastases (CLM), and is considered a potential biomarker for CLM. The origin of elevated levels of COL IV in CLM is not known but may result from both increased ECM production and ECM degradation associated with cancer-related tumor stroma remodeling. Aims: In this thesis, COL IV and its potential to be used as a biomarker for mCRC and mBC is studied, with a specific emphasis on liver metastases. The aims are to compare levels of circulating COL IV (cCOL IV) in mCRC and mBC patients with controls and evaluate its diagnostic and prognostic value; to evaluate the combination of cCOL IV with other proteins; to determine the cellular origin of COL IV in CLM and study COL IV expression in cell lines; to study the expression of COL IV degrading proteases in CLM and to evaluate tissue expression of COL IV in bone and liver metastases from BC patients. Methods: Plasma levels of cCOL IV, CA 15-3, CEA, and other cancer-related proteins were analyzed with ELISA, ECLIA and Multiplex assay in mCRC and mBC patients, healthy controls and patients with primary CRC or BC as controls. The cellular origin of COL IV expression in CLM was examined with in situ hybridization, and the expression of COL IV in mBC tissue and COL IV degrading proteases (MMP -2, -7, -9 and -13) in CLMs were studied with immunohistochemistry. COL IV expression in CRC and fibroblast cell lines was analyzed with immunofluorescence.Results: cCOL IV is elevated in mBC patients and correlates with poor survival. The combination of cCOL IV with CA 15-3 and cCOL IV alone are superior to CA 15-3 at detecting mBC. COL IV is highly expressed in the tissue of liver- and bone BC metastases. Circulating COL IV, CEA, OPN, CYFRA 21-1, IL-8, HGF, and MIF are elevated, and TRAIL is lower in mCRC patients compared with controls. COL IV, CEA, OPN, CYFRA 21-1, and IL-8 were higher, and TRAIL was lower in mCRC patients with liver metastases compared to patients with extrahepatic disease. Circulating CEA, OPN, and HGF are very good, and cCOL IV is acceptable at distinguishing mCRC patients from patients with primary CRC. The combination of OPN + CEA is superior to CEA alone at detecting mCRC. High HGF and cCOL IV (one cohort) in mCRC patients correlate to poor prognosis. cCOL IV is elevated in CLM patients compared to healthy controls and is very good at discriminating between healthy controls and CLM patients. COL IV is expressed in CLM by cancer-associated fibroblasts, and COL IV degrading proteases are expressed primarily by stromal cells in CLM. COL IV is expressed by fibroblasts, not tumor cells, in vitro. Conclusion: cCOL IV is a promising tumor marker of metastatic BC and CRC and circulating HGF and OPN are potential biomarkers of mCRC. Our results show that the metastatic site can impact the circulating levels of numerous cancer-related proteins, which aligns with our hypothesis that combining biomarkers instead of using one might be best for detecting metastatic cancer through blood analysis. COL IV is expressed by stromal cells, not tumor cells, in CLM tissue and in vitro.
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- Wadsten, Charlotta, 1964-
(författare)
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DCIS of the breast : aspects on treatment and prognosis
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Breast cancer is the most common cancer form and a leading cause of death in women worldwide. Ductal breast carcinoma in situ (DCIS) is characterized by a proliferation of malignant cells confined within the mammary ducts and is a potential precursor of invasive breast cancer. The risk estimations of a DCIS to develop into invasive cancer over a 10 year period range from 30-50%. In the past 25 years, concomitant with the implementation of screening mammography, the incidence of DCIS has increased dramatically and presently almost 1 000 women are diagnosed with DCIS each year in Sweden. The increased incidence poses concerns of overtreatment and current research aim at identifying clinical or pathological markers that can reliably distinguish hazardous from harmless DCIS. The overall aim of this thesis was to explore the prognostic significance of clinical and tumourbiological characteristics of DCIS and to assess the benefits and harms of adjuvant treatment.In a population-based cohort of 2 952 women with primary DCIS, we analysed trends in incidence, treatment and outcome over a 20-year period (paper I). Information was obtained from the regional breast cancer register in Uppsala-Örebro healthcare region between 1992 and 2012. A validation of 300 randomly selected women revealed high overall completeness and reliability of most key variables, whereas follow-up data were of moderate quality with only 65% of the recurrences reported to the register.The major finding of the study was a trend towards more intensified treatment over time. The frequency of mastectomy increased from 23.0% to 39.0% and the proportion of patients receiving adjuvant radiotherapy after breast-conserving surgery increased from 30.1% to 67.6%. This did not, however, translate into any noteable improvements in outcome. Relative survival was >97% after 10 years with no significant variation over time. In conclusion, these results may reflect adequate treatment selection, but may also indicate a significant overtreatment.In paper II and III, a nested case-control study was conducted from a cohort of 6 964 women with primary DCIS to identify clinical characteristics in DCIS associated with subsequent breast cancer death. Ninety-six women who later died from breast cancer were compared to 318 controls selected by incidence density sampling. Information was obtained from medial records and histopathology reports.Tumour size over 25 mm or multifocal DCIS (OR 2⋅55; 95%CI 1⋅53 to 4⋅25), a positive or uncertain margin status (OR 3⋅91; 95%CI 1⋅59 to 9⋅61) and detection outside the screening programme (OR 2⋅12; 95%CI 1⋅16 to 3⋅86) increased the risk of death from breast cancer. In the multivariable analysis, tumour size (OR 1⋅95; 95%CI 1⋅06 to 3⋅67) and margin status (OR 2⋅69; 95%CI 1⋅15 to 7⋅11) remained significant. More extensive treatment was not associated with lower risk, which may be due to confounding by indication, or indicate that some DCIS have an inherent potential for metastatic spread. In paper III, to further explore the association of tumour biology and risk of breast cancer death, archival tumour blocks were collected. Freshly cut hematoxylin and eosin (H&E) stained sections of the primary DCIS were histopathologically evaluated for nuclear grade, presence of comedonecrosis and lymphocytic infiltration (LI). Tissue microarrays were constructed for immunohistochemical analysis (IHC) of oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and Ki67. Using the results of the IHC analyses, tumours were classified into surrogate molecular subtypes.Presence of intense periductal LI was associated with an increased risk of subsequent breast cancer death (OR 2.25; 95%CI 1.02 to 4.99). None of the other biomarkers were individually related to breast cancer death, nor were there any statistically significant differences in risk between the molecular subtypes. In multivariable analysis, stepwise adjusting for age, tumour size and treatment, PR negativity in combination with LI; PR negativity, LI and presence of comedonecrosis and the combination of PR negativity, LI, comedonecrosis and HER2 positivity were all independently associated with increased risk of breast cancer death. The significance of features in the peritumoral stroma need further investigation and may have implications for targeted treatments.In paper IV, we studied the risk of ischemic heart disease (IHD) after treatment for DCIS. Postoperative radiotherapy (RT) in DCIS reduces recurrence rates by half but confers no benefits in terms of survival. It is thus of major importance to consider long-term adverse effects. Left-sided breast irradiation may involve exposure of the heart to ionising radiation with an associated risk of subsequent cardiovascular disease. The cumulative incidence of IHD was analysed in a population-based cohort of 6270 women with DCIS compared 31 257 women without a history of breast cancer. Of the women with DCIS, 38.9% had received adjuvant RT.After a median follow-up of 8 years, there was no increased risk of IHD for women with DCIS versus the comparison cohort. The risk was lower for women with DCIS allocated to RT compared to non-irradiated women and to the comparison cohort, probably due to patient selection. Comparison of RT by laterality did not show any over-risk for irradiation of the left breast. These results are reassuring, but longer follow-up may be warranted considering the continuously increasing use of RT in DCIS management.
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