1.
Brattström, Daniel, 1966-
(författare)
Angiogenesis Related Markers In Non-Small Cell Lung Cancer
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
This thesis investigated the predictive and the prognostic powers of angiogenesis related markers in both operable and inoperable non-small cell lung cancer (NSCLC) patients.In the first and second study, we investigated the serological fractions of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in 2 cohorts of patients with either operable or inoperable NSCLC. Regarding operable NSCLC, we demonstrated significant correlations between VEGF and tumour volume and overall survival. Regarding bFGF, significant correlations with recurrent disease and survival were demonstrated. VEGF and bFGF correlated to each other and with platelet counts. In multivariate analysis, bFGF proved to be a significantly independent prognostic factor.Regarding inoperable NSCLC, we demonstrated that patients with elevated bFGF levels before any treatment and during chemotherapy had a significantly poorer survival. During chemotherapy, each rise of one unit of bFGF (ng/L) corresponded to a 4 times increased risk of death. Regarding VEGF, elevated levels after radiotherapy corresponded with better survival. All prognostic information demonstrated in this study concerned patients with a, co-sampled, normal platelet count.In the third study, three putative markers, HER-2, EGFR and COX-2, suitable for targeted therapies in resected NSCLC were investigated in a panel of 53 tumours and further investigated for a possible correlation with microvessel density. We demonstrated that HER-2 and COX-2 were mainly expressed in adenocarcinomas, whereas EGFR was only expressed in squamous cell carcinomas. COX-2 showed a trend towards a correlation with microvesssel density. The expression profile, HER-2+/EGFR-, was significantly correlated to poorer survival. In the fourth study, a predictive model for recurrences consisting of p53, CD34 and CD105, and circulating serum fractions of VEGF and bFGF, was investigated. The two endothelial markers correlated with each other. CD105 expression correlated with p53 expression. No other significant correlations between markers could be demonstrated. A significant correlation between p53 overexpression and recurrent disease was demonstrated. The mutational status could not confirm the immunohistochemical correlation between p53 and recurrences. In conclusion, the present thesis demonstrates that the angiogenic factors VEGF and bFGF analysed in sera have both predictive and prognostic information when measured in operable and inoperable NSCLC. Since HER-2 is overexpressed in NSCLC and linked with prognostic information, this marker might be a suitable target for therapy in NSCLC. Furthermore, in patients with operable NSCLC, p53 expression status was linked with recurrent disease and mean MVD.
2.
3.
Fernebro, Eva
(författare)
Rectal Cancer - Tumor Biology and Prognostic Markers
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Colorectal cancer is one of our most common malignancies and the second leading cause of cancer death worldwide. About 1/3 of the tumors are located in the rectum. Treatment advances such as the introduction of the standardized surgical technique total mesorectal excision (TME), pre-operative radiotherapy, and adjuvant chemotherapy have reduced the previously high local recurrence rates and improved survival in rectal cancer patients, but despite these advances about 40% of the patients still die from disseminated disease. Rectal cancer evolves through an accumulation of genetic alterations in the tumor cells, and although multiple such changes have been characterized, the molecular markers have not yet gained widespread clinical use. The prognostic markers currently available fail to identify patients at risk for tumor recurrencies, which implies a need for refined prognostic tools. This thesis focuses on analysis of different molecular events in rectal cancer with correlations to prognosis. Papers I-II evaluate the novel serological markers suPAR and TIMP-1 analyzed in pre-treatment plasma samples from patients with rectal cancer. High suPAR and TIMP-1 values were associated with shorter survival and these markers may thus be of potential prognostic use. In paper III, we evaluated the tissue microarray technique (TMA) for immunohistochemistry (IHC) using the markers p53 and Ki-67. Good quality staining was obtained and TMA could reproduce the results obtained from whole-tissue sections. Paper IV presents IHC tumor profiling using TMA in rectal cancer with correlations to prognosis using the markers Ki-67, p53, Bcl-2, EGFR, ß-catenin, E-cadherin, MLH1, and MSH2. The cell adhesion molecules ß-catenin and E-cadherin significantly correlated with metastatic disease, whereas the other markers did not. In paper V, we studied 30 cancers from patients younger than 50 years at diagnosis with respect to molecular alterations associated with the microsatellite instability (MSI) and the chromosomal instability (CIN) pathways for colorectal cancer development. We found that microsatellite stable (MSS), aneuploid tumors with increased expression of p53 and ß-catenin predominate also among young rectal cancer patients. Only 10% of the tumors displayed MSI, all of which showed loss of expression for MSH2, which suggests presence of a mutation associated with hereditary nonpolyposis colorectal cancer (HNPCC). In summary, this thesis presents confirmatory data on the plasma levels of suPAR and TIMP-1 as presumptive prognostic markers in rectal cancer. In addition, we have demonstrated that the TMA-technique allows high-throughput immunostaining in rectal cancer. A TMA-based molecular profiling demonstrated that aberrant expression patterns frequently occur, and that the expression of the cell-adhesion proteins ß-catenin and E-cadherin correlated with clinical outcome. Finally, the 5% of the rectal cancers that occur in young patients predominantly develop through the CIN pathway, whereas MSI is found in a small, HNPCC-related subset of the tumors. Hence, other mechanisms than defective MMR and HNPCC cause rectal cancer in the majority of young patients.
4.
Karlsson, Richard
(författare)
Mechanisms of survival and maintenance of Hematopoietic Stem Cells and Multipotent Progenitor Cells
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Hematopoietic stem cells and progenitor cells are maintained in the bone marrow microenvironment, where factors including soluble and membrane-bound cytokines influence the processes of self-renewal, proliferation and differentiation. Two nonreduntant cytokines with effects on hematopoiesis, Kit Ligand (KL) and Flt3 Ligand (FL), signal via related tyrosine kinase receptors, c-kit and Flt3. In this thesis, we have studied which signaling pathways that are activated by these cytokines and are important for survival. We have shown that the serine-threonine kinase PKB (also known as Akt) is activated by both cytokines, but the upstream target PI-3 kinase is only necessary for the survival induced by KL and not FL. Also, KL could not prevent apoptosis in cells overexpressing a dominant negative form of PKB. The forkhead transcription factor FoxO3, which is phosphorylated and inactivated by PKB, was further shown to be of central significance for KL-mediated survival. The survival signals mediated by FL was shown to include antiapoptotic Bcl-2 family proteins. When overexpressed in progenitor cell lines, Bcl-2 was more effective in blocking apoptosis than PKB. Although both Bcl-2 and PKB could prevent a decrease in the mitochondrial membrane potential, Bcl-2 was superior with this respect. Some enhancement in survival was also seen when both proteins were overexpressed simultaneously. Finally, we studied the survival of bone marrow-derived multipotent hematopoietic progenitors in the presence of low levels of oxygen, hypoxia. We showed that survival was enhanced by hypoxia and improved the expansion of primitive colony forming cells.
5.
Kinhult, Sara
(författare)
Endothelial and cardiac effects of 5-fluorouracil. An experimental and clinical study.
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
The cytostatic drug 5-fluorouracil (5-FU) has been shown to affect both morphology and function of vascular endothelium. These effects could be part of the pathophysiology for 5-FU induced cardiotoxicity. The present thesis explores the mechanisms of this endothelial toxicity. In an animal model, treatment with thromboprophylactic doses of the low-molecular weight heparin (LMWH) dalteparin could not protect the endothelium from damage caused by 5-FU, although the secondary thrombosis was prevented. A subsequent study with three different LMWH (dalteparin, enoxaparin and tinzaparin) showed a moderate endothelial injury after treatment with LMWH alone for up to 60 days. Probucol, a lipid-lowering drug with strong antioxidant properties, was given as prophylaxis for two weeks before 5-FU treatment. With this drug, the endothelium was protected from the negative effects of 5-FU and had a normal morphology. Patients receiving 5-days infusion of 5-FU, combined with cisplatin, were studied for endothelial and cardiac effects. There was a significant increase in markers for endothelial injury (von Willebrand factor and soluble thrombomodulin) and in malondialdehyde, a marker for increased lipoperoxidation and possibly free radical production. Myocardial and echocardiographic parameters were not changed. Human umbilical vein endothelial cells (HUVEC) were incubated with 5-FU. A dose-dependant increase in secretion of endothelin-1 (a potent vasoconstrictor) was shown. No influence of apoptosis was seen. The results indicate increased oxidative stress caused by 5-FU, followed by endothelial damage and secretion of a vasoactive peptide. This could be part of the pathophysiological mechanisms of 5-FU induced cardiotoxicity.
6.
Lindman, Henrik, 1963-
(författare)
Individually Tailored Toxicity-based Chemotherapy : Studies on Patients with Primary and Metastatic Breast Cancer
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Standard dosing of chemotherapy based on body surface area (BSA) results in large individual differences in toxicity due to a large inter-patient variability in pharmacokinetics (PK) and pharmacodynamics (PD). This results in under-dosing in certain patients with a potentially weaker antitumoral effect.Three clinical studies of individually tailored dosing of chemotherapy, based on haematological toxicity were conducted. In the first study, 26 women with metastatic breast cancer were treated with tailored and dose-escalated 5-fluorouracil, epirubicin and cyclophosphamide, supported by G-CSF (dFEC). In the second study 525 patients with high-risk primary breast cancer were randomised between dFEC and high-dose chemotherapy with autologous bone-marrow transplantation. The feasibility of a FEC regimen with doubled cyclophosphamide dose to mobilise peripheral stem cells was investigated. In the third study, 44 metastatic patients were treated with tailored epirubicin and docetaxel (ET). PK and PD were also investigated in these patients. The potential effects of G-CSF on MRI tumour evaluation were studied in 18 patients with skeletal metastases.Toxicity-based dosing entailed an evenly distributed two- to three-fold range of tolerated doses in all three studies. Efficacy and toxicity were not correlated to tolerated dose-levels. Tailored dFEC resulted in a response rate of 81% and the same regimen resulted in fewer breast cancer relapses compared with standard FEC followed by high-dose therapy. Toxicity was manageable except for an increased rate of secondary leukaemia. The modified FEC could safely mobilise sufficient numbers of stem-cells. Tailored ET resulted in a response rate of 63%. The inter-individual variability in drug clearance was larger than the inter-occasion variability and a semi-physiological model of PK and PD could predict leukocyte nadir and duration. An increased diffuse MR signal in the long TE IR-TSE sequence was observed in normal bone-marrow during G-CSF treatment; this could be mistaken as disseminated metastatic disease and could obscure focal metastases.In conclusion, the concept of individually tailored toxicity-based dosage of chemotherapy was equally feasible in primary and metastatic breast cancer, in two different chemotherapy regimens and in treatment with or without G-CSF support and may provide a pragmatic way of overcoming the shortcomings of standard BSA-based dosing.
7.
Loman, Niklas
(författare)
Clinical Aspects of Hereditary Breast Cancer
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
A positive family history of breast cancer (BC) is one of the strongest predictors of the disease. Two major BC susceptibility genes, BRCA1 and BRCA2 were identified about a decade ago. In this thesis, studies of different biological. clinical and epidemiological aspects of hereditary BC are presented. In the first paper data on the expression of steroid hormone receptors in hereditary BC are presented. We confirm previously published data regarding the paucity of estrogen and progesterone receptors in BRCA1-associated BC, and report new findings in BRCA2-associated tumours and cases with an unidentified hereditary factor. Hormone receptor levels in these two groups do not differ significantly from each other or from an age-matched control population unselected for family history. The second paper is a cohort study of cancer incidence in relatives of BRCA1 and BRCA2 germline mutation carriers. Other cancers than BC and ovarian cancer do not appear to be greatly increased and require specific follow-up in carriers. The third study is a retrospective case-control study on the prognosis and clinical presentation of BRCA2-associated BC. BC in families with an identified BRCA2 mutation seemed to present at a more advanced stage at diagnosis compared with age and year of diagnoses matched controls. This translated into an increased risk to die from BC in this group, but overall survival was not significantly decreased. The fourth study is a population-based study of family history and BRCA1 and BRCA2 mutations in BC cases below the age of 41. A positive family history was very frequent in this group of young BC patients. About one-third of them had a family history including at least one first- or second-degree relative with BC. A positive BRCA1-mutation status was observed in 6.8% of the cases, and BRCA2-mutations were seen in 2.1% of the in women. BRCA1 and BRCA2 mutations were more prevalent in younger women, in women with a positive family history including at least one first-degree relative with breast or ovarian cancer, and in women that had already or did develop a bilateral BC during a median follow-up of 5.5 years. In the fifth study relative and cumulative cancer incidence in the first-degree relatives of the woman of the previous study were calculated and compared with the population. The BC risk was still increased in first-degree relatives of women with early-onset BC (below the age of 41) when BRCA-mutation carriers were excluded, in addition an association with prostate cancer was suggested. The risk of prostate cancer did only appear to be increased in relatives of women with BC below the age of 36. Furthermore, cumulative BC incidences were calculated for women with different hereditary backgrounds.
8.
Melander, Fredrik
(författare)
ß2 integrin-induced signal transduction events in human neutrophils
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Inflammation is a localized protective response in vascularized tissues and can be induced by microbial infection or cell and tissue injury. Polymorphonuclear neutrophils (PMN) are the most common white blood cells and are recognized as major cellular mediators of acute inflammation. PMN are highly mobile cells that possess regulated mechanisms for controlling the expression and activation of adhesion molecules such as selectins and integrins. This is crucial for their tethering and migration into inflammatory sites. Once in the tissue PMN can execute their important bacteriocidal functions that will not only kill microorganisms, but also cause potential tissue damage. This thesis work is focused on the intracellular signalling events that the ß2 integrins induce. In particular we focused on small GTPases of the Rho family, proteins that are key regulators of cytoskeletal rearrangements but also important for the production of bacteriocidal substances. We could show that the small GTPases Rho and Rac display opposing activities in PMN with activated ß2 integrins and we could also identify important signalling proteins responsible for this. Moreover, the ubiquitin ligase c-Cbl was also shown to be phosphorylated and activated in adherent PMNs and is suggested to play a key role in the signalling pathway induced by ß2 integrins. ß2 integrins, or enzymes involved in the signalling cascade this adhesion receptor activate, could represent potential targets to control inflammatory conditions where unwanted and tissue destructive accumulation of leukocytes occurs.
9.
Sand-Dejmek, Janna
(författare)
Role of Wnt-5a in breast cancer
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Breast cancer is one of the most common cancer forms in the industrialized world. Only in Sweden, nearly 6000 women are diagnosed with breast cancer each year and 1/3 of them eventually succumb to the disease. Wnts are a family of genes that have been implicated in many human tumours, so far mainly studied in colorectal cancer where activated Wnt signalling occurs in a vast majority of tumours. Whereas the oncogenic Wnt-1 is the main Wnt protein investigated in breast cancer, very little work has been focused on Wnt-5a, a protein supposedly antagonizing the oncogenic effects of Wnt-1. We found that Wnt-5a protein expression is reduced in many invasive breast carcinomas and that reduced Wnt-5a expression in the primary tumour strongly correlates with an increased risk of metastatic disease and shorter survival. The expression of Wnt-5a protein co-varies with that of Syk, a tyrosine kinase that is lost in up to 30% of invasive breast carcinomas and associated with poor prognosis. Furthermore, patients with tumours expressing both Wnt-5a and Syk had a significantly better prognosis as compared with those displaying loss of either of or both proteins. Despite the co-expression, loss of Wnt-5a and Syk protein appear to be regulated at the translational and transcriptional levels, respectively. In normal breast epithelium, Wnt-5a is important for cell-ECM adhesion and activation of the collagen receptor DDR1. Restitution of Wnt-5a signalling in breast tumour cells confer better adhesion and ability to activate DDR1 as well as a less malignant-looking phenotype. In human breast epithelial cells Wnt-5a activates the canonical b-catenin pathway as well as the non-canonical Ca2+/calmodulin and planar cell polarity (PCP) pathways. The latter involves Src, the RhoGTPase Cdc42 and JNK and was shown to counteract the NFAT activation induced by the Ca2+/calmodulin pathway. The collagen-induced activation of DDR1, which also is Src-dependent, appears to be mediated by the PCP pathway. Wnt-5a signalling is thus involved in the activation of DDR1 as well as in hampering NFAT activity, both of which affect the migratory capacity of tumour cells. Consequently, the increased metastatic potential of breast tumours with low Wnt-5a expression could be due to inactivation of DDR1 or enhanced NFAT activation. The results presented in this thesis thus imply that Wnt-5a contains metastasis-suppressing activity in breast cancer. Depending on the cellular effects through which Wnt-5a mediates its function, reconstitution of Wnt-5a signalling might have future therapeutical implications.
10.
Trollér, Ulrika
(författare)
The role of PKC in neuronal differentiation and in regulation of the actin cytoskeleton during neurite outgrowth
2003
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Protein kinase C (PKC) is a family of serine/threonine kinases subgrouped into classical (α, βI, βII, γ), novel (δ, ε, η, θ) and atypical (ζ, ι/λ) isoforms. PKC can be activated by phorbol esters, and prolonged treatment of neuroblastoma cells has been shown to induce neuronal differentiation with neurite outgrowth and increased expression of neuronal genes. Here we show that a classical PKCβ isoform is important for the phorbol ester-induced gene expression via the MAP kinases Erk 1/2. A previous study has shown that novel PKCε, via its regulatory domain, induces neurites in neuroblastoma cells. In this thesis we show that membrane localization and an actin-binding site in the regulatory domain is important for this effect. Besides neuroblastoma cells, the regulatory domain of PKCε is sufficient to induce cellular processes in both immortalized neural cells and fibroblasts, indicating that it targets commonly used cytoskeletal proteins. Furthermore, the catalytic activity of PKC is not only dispensable, it counteracts the neurite-inducing capacity of the regulatory domain. Overexpression of PKCδ does not induce the formation of long cellular extensions by itself. However, treatment with phorbol ester triggers the formation of processes in PKCδ-overexpressing cells. This provides a model system to study initial cytoskeletal changes, which revealed that stress fiber loss accompanies outgrowth of processes. This indicates that the small GTPase RhoA, which is the main regulator of stress fiber formation, is suppressed. Furthermore, overexpression of a constitutively active RhoA blocked both PKC-mediated stress fiber loss and neurite outgrowth. A possible mechanism for the RhoA suppression is via p190RhoGAP, with which PKCε forms a complex. Cdc42 was further shown to be crucial for the PKC-mediated stress fiber loss, and both Rac1 and Cdc42 are important for PKC-mediated outgrowth of cellular processes.
