| 1. |
- Andersson, Bengt-Åke
(författare)
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Circulating Biomarkers in Patients with Head and Neck Cancer and the Influence of Cigarette Smoking
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Head and neck cancer (HNC) is a collective name for heterogeneous tumors located in the head and neck regions for which smoking, alcohol and human papillomavirus (HPV) are documented risk factors. The survival of HNC patients has only improved marginally during the last decade. The most important prognostic factors are tumor size, local spread and distant metastases, tumor node metastasis (TNM) staging. Prognostic biomarkers are needed as a complement to TNM staging.The aim for this thesis was to investigate rapid and low cost blood based biomarkers which could indicate the risk of HNC, recurrence of the disease or the survival of HNC patients. Furthermore, the aim was to examine how cigarette smoking influences the levels of biomarkers.In paper I, a possible role of plasma cytokines or proteins associated with immune response or inflammation, as biomarkers for the survival of HNC patients was investigated. Higher levels of C-reactive protein (CRP) and tumor necrosis factor alpha (TNF-α) were detected in plasma of the patients compared with the levels in the controls. The elevated levels of these two biomarkers detected in patients were associated with decreased survival.In paper II, the influence of 45 single nucleotide polymorphisms (SNPs) located in 41 genes associated with cell cycle progression, cell death, DNA repair or immune response on cancer risk, tumor recurrence and survival in HNC patients were investigated. SNPs in immune response genes were associated with risk for HNC, an elevated risk for recurrence and a decreased survival in HNC patients.In paper III, the influence of cigarette smoking on levels of inflammatory cells, proteins or cytokines/chemokines, microRNAs (miRNAs) and SNPs was analysed in healthy smokers and non-smokers. Higher levels of total white blood cells (WBCs), neutrophils, monocytes, lymphocytes, neutrophil to lymphocyte ratio (NLR), CRP, monocyte chemoattractant protein- 1 (MCP-1) and interferon gamma (IFN-γ) were detected in smokers compared to non-smokers and indicate an inflammatory response. Also, a lower level of oncomiRNA miR-21was detected in smokers. This alteration, in combination with the elevated levels of IFN-γ in smokers could be a protective response to cigarette smoke. The higher levels of IFN-γ in smokers compared to non-smokers were however only detected in individuals with SNP rs2069705 genotype AG/GG. This indicates a genetic association of the levels of IFN-γ.In paper IV, the separate effects of cigarette smoking and HNC on inflammatory or immune biomarkers and the impact of high risk human papillomavirus, age and gender were investigated. Comparisons of circulating levels of WBCs and its subpopulations, plasma proteins or cytokines/chemokines between smoking and non-smoking patients, smoking and non-smoking controls and between the patient and control groups were analysed. Smoking had highest impact on elevated levels of WBCs, IFN-γ and MCP-1, and HNC had highest impact on elevated levels of neutrophils, monocytes, NLR, CRP, macrophage inflammatory protein 1 beta and TNF-α.In conclusion, host immune response associated parameters could be suitable as biomarkers for the risk of HNC, risk of recurrence or in predicting survival of HNC patients. This thesis show that HNC are associated with systemic inflammatory response and upregulated CRP and TNF-α is related to shorter survival in HNC patients. Additionally, SNPs in immune response genes such as rs1800629 in the TNF-α gene indicates a risk for HNC or an elevated risk for recurrence and a decreased survival in HNC patients. These rapid and low cost blood based biomarkers could be used in combination or as a supplement to established biomarkers in the clinic for a more personalized treatment modality.
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| 2. |
- Ansell, Anna, 1983-
(författare)
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Identification of Tumor Cell- and Stroma Derived Biomarkers of Treatment Response in Head and Neck Cancer
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Head and neck squamous cell carcinoma (HNSCC) poses a major health problem in the world with approximately 600 000 new cases yearly. Treatment resistance is a major problem within this patient group and despite advances in treatment strategies the overall survival rate has unfortunately not increased.One of the major components of the tumor microenvironment is the cancer associated fibroblasts (CAFs) which can modulate the treatment sensitivity, tumor growth, and the invasive potential of tumor cells.The aim of this thesis was to identify predictive markers for treatment response in HNSCC and to study the crosstalk between tumor cells and CAFs that may underlie treatment resistance.In paper I, we identified gene expression differences between one cisplatin sensitive cell line and two cisplatin resistant cell lines, by microarray analysis, and found that a high expression of matrix metalloproteinase (MMP) -7 was associated with resistance to cisplatin. In paper II, the epidermal growth factor (EGF) receptor ligands EGF, amphiregulin, and epiregulin were evaluated regarding their potential use as predictive biomarkers for cetuximab treatment response in tongue cancer cell lines and it was shown that EGF may serve as a marker for poor cetuximab response. In paper III and IV, we investigated the influence of CAFs on the proliferation, migration, gene expression, and cetuximab response of tumor cells. It was found that CAFs induced resistance to cetuximab in a MMP-dependent manner. In addition, a microarray analysis, comparing tumor cells co-cultured with CAFs and tumor cells cultured alone, revealed that CAFs induced multiple gene expression changes in tumor cells some of which are related to epithelial to mesenchymal transition. Some of these changes were found to be dependent on cell-cell contact.Taken together, we here suggest MMP-7 and EGF to be predictive markers of cisplatin and cetuximab response, respectively. We also show that CAFs protect HNSCC cells from cetuximab treatment; however, the factor responsible for the protective effect is yet to be discovered.
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| 3. |
- 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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| 4. |
- Bendrik, Christina, et al.
(författare)
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MMP-3 and MMP-9 Gene Transfer Decrease Growth and Angiogenesis in Breast Cancer Xenografts In Vivo
- 2009
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Matrix metalloproteinases (MMPs) are largely implicated in tumor behaviour due to their extracellular matrix (ECM) remodelling capacities. Although MMP activity generally is discussed in terms of facilitating tumor invasion, MMP inhibition in clinical trials has failed. Increasing amounts of data show that MMPs may inhibit tumor progression by generating anti-angiogenic factors such as endostatin from the tumoral stroma. We have previously shown that intratumoral gene transfer of MMP-9 induced tumor regression and reduced angiogenesis of breast cancer in vivo. Whether MMP activities induce tumor progression or regression may depend on type of MMP and the expression level in the tumor tissue. In this study we treated established breast cancers in nude mice with adenovirus vectors carrying the human genes of MMP-3 or MMP-9 in low or high dose. Microdialysis was used to sample endostatin in situ and tumor growth was monitored for 35 days. Tumors in mice treated with low-dose of either MMP-3 or MMP-9 vectors exhibited tumor stasis throughout the experiment whereas high-dose gene transfer of either MMP-3 or MMP-9 induced significant tumor regression compared to controls treated with empty vectors. The extracellular in vivo levels of endostatin were increased in tumors that received either high or low MMP-3 or MMP-9 gene transfer and these tumors exhibited decreased microvessel area compared to controls. Our results propose that increased expression of MMP-3 and MMP-9 have therapeutic effects of established breast cancer in a dose dependent manner where a slight increase of MMP expression results in tumor stasis and a high expression of either MMP-3 or MMP-9 by gene transfer results in a potent tumor regression.
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| 5. |
- Dabrosin, Charlotta
(författare)
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Effects of sex steroids on normal human breast : studies in vivo using microdialysis and in vitro in cell culture
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Prolonged exposure to sex steroids may constitute a risk factor for the development of breast cancer. The biological mechanisms involved in breast carcinogenesis are not well understood.Basic knowledge of sex steroid effects on the normal human breast is still limited, one reason being the lack of an available in vivo technique for investigations of breast tissue metabolism.In this study, the microdialysis technique was developed and evaluated as a method for measurements of tissue-specific concentrations of amino acids, lactate, pyruvate and glutathione in normal human breast tissue during the menstrual cycle. The technique was successfully applied to breast tissue and it was observed that the concentrations of several amino acids as well as glutathione changed during the menstrual cycle. Oxidative damage to cells is one of the mechanisms which may be involved in the development of breast cancer. Normal aerobic metabolism generates potentially dangerous oxidants which are controlled by a variety of antioxidant systems. The exact regulatory mechanisms of these systems are not yet fully understood. We studied the effects of estradiol and progesterone on antioxidative activity in normal human breast tissue, in vivo with the microdialysis technique, and in vitro using normal human breast epithelial cells in culture. The in vivo levels of the antioxidant glutathione were measured early and late in the menstrual cycle in breast tissue and subcutaneous fat. The glutathione levels were higher late in the menstrual cycle in both tissues, when the serum levels of estradiol and progesterone were high. In vitro, breast epithelium exposed to estradiol and progesterone exhibited decreased activity of the antioxidative enzymes catalase and glutathione reductase, whereas the activity of glutathione peroxidase tended to increase compared with cells grown in medium without added sex hormones. The vulnerability to oxidative stress, induced by hydrogen peroxide, increased in cells grown with estradiol and progesterone present in the media. α-Tocopherol, and α-tocopherol in combination with ascorbic acid, but not ascorbic acid alone, protected from cell death induced by hydrogen peroxide. This effect was not dependent on estradiol and progesterone exposure.In conclusion, the data suggest an effect of estradiol and progesterone on antioxidative activity in normal human breast tissue both in vivo and in vitro.Microdialysis will be a useful tool in future research of these and other aspects concerning human breast tissue.
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| 6. |
- Garvin, Stina, 1977-
(författare)
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Effects of sex steroids and tamoxifen on VEGF in the breast
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Sex steroid exposure constitutes a risk factor for breast cancer, but little is known about the effects of sex steroids on factors mediating angiogenesis, the development of new blood vessels, in normal and malignant breast tissue. In this thesis we have investigated the effects of estradiol, progesterone, and the nonsteroidal anti-estrogen tamoxifen on vascular endothelial growth factor (VEGF) and its receptors (VEGFR-1 and VEGFR-2) in normal human breast tissue, endothelial cells, and breast cancer. We have applied the technique of microdialysis to provide in situ sampling of estradiol and VEGF in tumors and normal breast tissue of breast cancer patients in vivo. Furthermore, we present a novel method of culturing normal human breast tissue ex vivo.Our results suggest a pro-angiogenic effect of estradiol and an anti-angiogenic effect of tamoxifen in the breast. Estradiol increased extracellular levels of VEGF in normal human breast tissue and breast cancer cells in vitro. In addition, estradiol decreased sVEGFR-1 in breast cancer cells and indirectly increased VEGFR-2 in endothelial cells. Compared to estradiol treatment alone, estradiol + tamoxifen increased sVEGFR-1 and decreased VEGF in breast cancer cells in vitro. Furthermore, estradiol + tamoxifen decreased tumor VEGF levels and tumor vasculature in human breast cancer xenografts in vivo. In breast cancer patients, a significant correlation was found between in vivo levels of estradiol and VEGF sampled by microdialysis in normal human breast tissue, suggesting that estradiol may be a potent regulator of VEGF in the breast in vivo. Tumor levels of VEGF were significantly higher than in normal breast tissue in vivo, supporting the role of VEGF in tumor angiogenesis. For studies of normal human breast, whole breast tissue may be cultured in vitro for up to one week with preserved morphology. Using this method, estradiol, and not progesterone, appears to be the main sex steroid regulator of extracellular VEGF in normal breast tissue. In conclusion, the data suggest that sex steroids and tamoxifen exert pro- and anti-angiogenic effects in normal breast tissue and breast cancer.
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| 7. |
- Lindahl, Gabriel, 1971-
(författare)
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The effects of flaxseed and tamoxifen on the inflammatory microenvironment in normal breast tissue and in breast cancer
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Breast cancer is the most common cancer among women worldwide today. Nearly 9000 women are diagnosed with breast cancer in Sweden yearly and despite advantages in diagnostics and treatments approximately 1400 women still die from their disease every year. Breast cancer has a diverse etiology and hormonal factors and life-style factors contribute to an increased breast cancer risk. High mammographic density is also considered a risk factor but the underlying mechanisms are not fully understood. Inflammation is associated with poor survival in several malignancies and is considered a hallmark of cancer. There is evidence indicating that increased inflammation is associated with dense breast tissue and may contribute to an increased risk of breast cancer in these patients.There is an urgent need to find risk reduction strategies in breast cancer prevention. Several studies have shown that antiestrogens significantly reduce breast cancer incidence in women with high risk of developing breast cancer and can be used for chemoprevention. These drugs may have potentially severe side effects and other strategies are needed. Dietary interventions may influence breast cancer risk without any major side effects. Studies indicate that dietary phytoestrogens may reduce breast cancer risk. The most common phytoestrogens in Western populations are lignans, mainly found in flaxseed, but results from several studies with lignans for breast cancer prevention have been inconsistent.In this thesis we investigated the effects of tamoxifen and flaxseed on inflammatory mediators in normal breast tissue and in breast cancer. We used the microdialysis technique to sample proteins from the extracellular space in vivo. This technique gives us the opportunity to study proteins in their bioactive compartment in situ and to study changes in protein levels at different time points without affecting the tissue of interest. We also used experimental models and cell cultures to study tumor growth of human breast cancer xenografts, cancer cell proliferation and angiogenesis.In paper I, we investigated whether tamoxifen, flaxseed, enterolactone or genestein reduced growth of human breast cancer xenografts and their association with pro-inflammatory cytokine interleukin 1β (IL-1β) and its antagonist interleukin 1 receptor antagonist (IL-1Ra). In paper II, we investigated whether tamoxifen and flaxseed exerted similar effects on inflammatory mediators in normal breast tissue in vivo. In paper III, we investigated whether osteopontin (OPN), a pro-inflammatory cytokine, was associated with dense breast tissue and breast cancer and if tamoxifen and flaxseed could alter OPN levels in normal breast tissue in vivo. We also investigated the correlation between OPN and inflammatory mediators in normal breast tissue and in breast cancer in vivo. In conclusion, we showed that tamoxifen and flaxseed affected breast cancer growth in an experimental model and may exert an anti-inflammatory effect in breast cancer and normal breast tissue by increasing the IL-1Ra/IL-1β ratio in vivo. We showed that dense breast tissue and breast cancer were associated with increased levels of OPN. Circulating estrogen did not correlate to OPN and tamoxifen and flaxseed did not affect OPN levels suggesting an estrogen independent regulation of OPN in vivo. These finding contributes to our understanding of how tamoxifen and flaxseed affects inflammation and the role of inflammation in the pathogenesis of breast cancer.
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| 8. |
- Morad, Vivian, et al.
(författare)
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Correlation between vascular endothelial growth factor and leptin in normal human breast tissue in vivo
- 2015
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Introduction: Events in the microenvironment are important for carcinogenesis of the breast. Adipocytes, which produce adipokines with paracrine effects, are the most abundant cell type in breast tissue. Exposure to sex steroids affects the risk of breast cancer. It has previously been shown that estrogen regulates the extracellular levels of leptin, adiponectin, IL-1β, and VEGF in normal human breast tissue in vivo.Objective: We aimed to determine if there were any relationships between leptin, adiponectin, IL-1β, and/or VEGF in normal human breast tissue in vivo and to elucidate the role of adipocytes in the regulation of these factors.Design and methods: Microdialysis was used to sample proteins of normal human breast tissue and abdominal subcutaneous (s.c.) fat in situ in pre-and postmenopausal women. An in vitro co-culture model of breast cancer cells and primary mature human adipocytes was used.Results: In vivo, in normal breast tissue, significant positive correlations between VEGF and leptin, and VEGF and leptin/adiponectin ratio were detected. No correlations were found in s.c. abdominal fat tissue. Co-culture of adipocytes and breast cancer cells per se increased the secretion of VEGF and leptin and enhanced the effects of estradiol compared to culture of either cell type alone. In vitro, inhibition of VEGF diminished the release of leptin while inhibition of leptin had no influence on VEGF secretion. In breast tissue, significant correlations between IL-1β and leptin and VEGF were revealed.Conclusions: Our results suggest that VEGF regulates leptin in normal human breast tissue. Moreover, physical contact between adipocytes and breast cancer cells, induces phenotypic changes and enhances the effects of estradiol. These mechanisms may be involved in breast cancer progression.
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| 9. |
- Morad, Vivian, 1982-
(författare)
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Hormonal regulation of immune modulators in human breast tissue
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Breast cancer is the most common form of cancer and the second leading cause of malignancy-associated death in women worldwide. Estrogens are the main sex hormones in women. They are essential for the development and function of normal breast mammary glands; however, prolonged exposure to estrogens increases the risk of breast cancer development and progression. Approximately two-thirds of all breast cancer patients are positive for estrogen receptor (ER), but only 50% of those cases can benefit from antiestrogen therapy.In this thesis we investigated the effects of estrogen, diet modification, and anti-estrogen drugs on several immune modulators in normal human breast tissue. We used the microdialysis technique to sample the immune modulators in situ in normal human breast tissue, in malignant breast tissue, and in tumor tissue from both the immune competent mice with murine breast cancer and immune deficient mice bearing human breast tumors. Furthermore, we also used ex vivo culture of normal breast tissue and in vitro cell culture of breast cancer cell lines. A combined cell culture (co-culture) of breast cancer cell lines, together with the primary mature adipocytes, was also used in this thesis.In Paper I and Paper II, our results suggested that estrogen exerted both proinflammatory and pro-tumorigenic effects in normal human breast tissue. Estradiol increased extracellular interleukin-1β (IL-1β) and leptin levels and decreased IL-1Ra and adiponectin levels in normal human breast tissue. In contrast, tamoxifen decreased IL-1β and leptin levels and increased IL-1Ra and adiponectin levels, shifting the environment towards an antiinflammatory and antitumorigenic state. Diet modification with flaxseed for 30 days also increased IL-1Ra levels, creating an anti-inflammatory environment in normal breast tissue. In the breast cancer tissue, we found that extracellular IL-1β levels and leptin levels were significantly higher, whereas adiponectin levels were significantly lower, compared with normal adjacent breast tissue, which suggested a more proinflammatory state.In the third paper, our in vivo investigation of normal breast tissue revealed significant correlations between vascular endothelial growth factor (VEGF) and leptin, IL-1β and leptin, and between VEGF and IL-1β. No correlations were found in the abdominal subcutaneous (s.c.) fat tissue. Our in vitro inhibition experiments suggested that VEGF was a potent regulator of leptin, but that leptin was not a potent regulator of VEGF. Co-culture per se altered the release of VEGF and leptin and enhanced the effects of estradiol, compared with monocultures of the included cell types.In conclusion, the results presented in this thesis will increase the overall understanding of the role of estrogens in breast cancer, which may be useful in future treatment studies.
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| 10. |
- Nilsson, Ulrika W., 1977-
(författare)
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Effects of sex steroids and tamoxifen on matrix metalloproteinase activity and generation of endostatin in the breast
- 2007
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Sex steroids are inevitable in women. However, long-term exposure to sex steroids increases the risk of breast cancer. A complete understanding of sex steroid control of the breast and how it relates to breast cancer risk is still lacking. Angiogenesis and proteolytic enzyme activity are crucial for the process by which tumors evolve into a vascularized, invasive phenotype. Matrix metalloproteinases are potent matrixdegrading enzymes that affect several steps in tumor progression including angiogenesis. In the female reproductive organs, sex steroids regulate angiogenesis and MMP activity, yet little is known how sex steroids affect these crucial events in normal and malignant breast tissue.This thesis elucidates a link between sex steroids, MMP activity, and angiogenesis. It is shown that estradiol down-regulates while tamoxifen up-regulates the protein expression and activity of MMP-2 and MMP-9 in human breast cancer cells in vitro and in human breast cancer xenografts in vivo. The results further suggest that a biological consequence of this regulation may be modulation of tumor angiogenesis. The net effect of adding tamoxifen to estradiol treatment was an increase in extracellular levels of the endogenous angiogenesis inhibitor endostatin and decreased levels of the tumor promoter TGF-β1 compared to estradiol treatment only. This was accompanied by reduced vasculature and decreased tumor growth. Similarly, a regulatory effect of estradiol and tamoxifen on endostatin generation was observed in normal human breast tissue by whole-tissue culture and microdialysis in human breast tissue in situ.In conclusion, the results presented in this thesis suggest previously unknown mechanisms of action of estradiol and tamoxifen in breast cancer and in normal human breast tissue, and novel means by which estradiol may tip the scale to favor angiogenesis. This knowledge may be important for the understanding of sex steroid dependent breast carcinogenesis and in the future development of tissue-specific preventive as well as therapeutic strategies against breast cancer.
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