<p>Infrainguinal bypass grafting is an established method in the treatment of patients with femoropopliteal and crural occlusive disease leading to critical lower limb ischaemia (CLI). However, complications related to surgical procedure are not negligible and percutaneous transluminal angioplasty (PTA) has emerged as an alternative. The present thesis covers some aspects of infrainguinal PTA in patients with chronic severe lower limb ischaemia.</p><p>The records of 217 patients undergoing 272 PTA procedures at various infrainguinal arterial segments were analysed. The indication for intervention was subcritical ischaemia in 76 limbs and critical ischaemia in 177 limbs. The role of duplex ultrasound examination in the selection of patients for PTA was retrospectively evaluated following a prospective validation of the method against angiography.</p><p>A technically successful PTA was achieved in 89%. The overall 30-day mortality was 2.7%. No patient underwent amputation directly related to failed PTA. The primary success rates at 12 and 60 months following femoropopliteal PTA were 40% and 27% compared, to 51% and 36% in limbs undergoing crural artery PTA. Primary success rate in limbs with SFA occlusion longer than 5 cm was only 12% after 5 years, compared to 32% if the occlusion was equal or less than 5 cm in length (p<0.01). In patients undergoing distal PTA through patent infrainguinal grafts, the primary and primary assisted patency rates at 3 years were 32% and 53%, respectively. The sensitivity of duplex scanning in the selection of lesions for PTA was less satisfactory in the popliteal and crural arteries compared to the superficial femoral arteries.</p><p>In conclusion, the results of infrainguinal PTA performed for treatment of subcritical or CLI seemed to be inferior to the results of surgical interventions reported in the literature. However, due to the fact that the PTA procedure does not preclude the performance of bypass grafting, it might be an alternative to surgical intervention in limbs with stenotic or short occlusive lesions.</p>

<p>Weight loss and other nutritional problems are common in cancer patients. The problems are of importance for response to treatment and survival and the well-being of the patients.</p><p>Nutritional support can be carried out in different ways. The efforts considered in this thesis are; assessment of nutritional status to find the patients who are at risk to become or already are malnourished, assessment of dietary intake, dietary advice, information and support to the families, information and education to the caregivers, and supplementation with drugs that possibly could influence the weight development. The Swedish version of the Patient Generated Subjective Global assessment of nutritional status, PG-SGA, is useful in assessment of nutritional status in cancer patients. Dietary advice and support to patients and their families combined with information and education to the staff, at the hospital and in the home care, turned out to have a positive influence at the weight development and other parameters related to nutrition. The effects were seen in consecutive patients with small cell lung cancer in comparison with a historical control group, and in patients in a randomised trial. Fish oil and melatonin could stabilise weight development in patients with advanced gastrointestinal cancer, but had no marked influence on factors reflecting cachexia. Problems with nutrition in cancer patients are possible to recognise and various interventions may be beneficial.</p>

<p>Radioactive substances are used <i>in vivo</i> to localize and characterize malignant tumours, generally by scintigraphic methods. In this context positron emission tomography (PET) in combination with radiolabelled monoclonal antibodies (mAbs) may provide a sensitive and specific method for detection of cancer. Individual dose calculations, based on such PET measurements, may be carried out to predict the possible use of mAbs labelled with therapeutic nuclides. The positron emitter ⁷⁶Br, with a half-life of 16 h, is a well-suited candidate for radiolabelling and PET imaging. One drawback of radiobromine is that bromide, the ultimate catabolite after degradation of brominated mAb, is only tardily excreted from the body and is evenly distributed throughout the extracellular space, thereby increasing the background radioactivity. The aim of this work was to produce ⁷⁶Br-mAb preparations with high accumulation and retention in tumour tissue together with a quick clearance of ⁷⁶Br-labelled catabolites. Furthermore, the possibility to use brominated or iodinated mAbs in combination with PET to predict ²¹¹At-mAb dosimetry was evaluated.</p><p>Monoclonal Abs directed against colorectal cancer were labelled with ⁷⁶Br using the direct Chloramine-T-method or indirectly by labelling the precursor molecule N-succinimidyl para-(tri-methylstannyl) benzoate with ⁷⁶Br, which was subsequently conjugated to the mAbs. Monoclonal Ab A33 labelled with ⁷⁶Br using the two labelling protocols was characterized in vitro and in vivo in a rat tumour xenograft model. The mAb A33 was also labelled with ¹²⁵I for comparison. In addition, mAb A33 was labelled with ²¹¹At, ¹²⁵I and ⁷⁶Br using the indirect labelling protocol and the mAb pharmacokinetics was studied in normal rats in order to estimate if data from brominated or iodinated mAb could be used for dosimetry of ²¹¹At in healthy organs and tissue.</p><p>In conclusion, both direct and indirect labelling resulted in high yields and mAbs with preserved immunoreactivity. <i>In vivo</i> characterization of ⁷⁶Br-brominated mAb A33 showed that the indirect labelling method makes ⁷⁶Br-brominated mAb A33 a promising candidate for tumour imaging with PET due to the faster excretion of radiolabelled catabolites compared with direct bromination. Finally, mAb A33 labelled with ⁷⁶Br and ^124/125I can be used to predict the ²¹¹At dose of astatinated mAb A33 in most organs given that a correction factor is applied for organs with varying uptake.</p>

<p>The relationship between Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) is not entirely elucidated and a clonal relation may be present more often than previously believed. Mechanisms of tumour progression and resistance to therapy are poorly understood.</p><p>Between 1974 and 1994 all individuals in Sweden with both HL and NHL were identified. Thirty-two cases were studied using clinical, histopathological and immunohistochemical methods. The second lymphoma often appeared in an aggressive clinical form and a significant correlation between the expression of p53 and LMP-1 in the first and second lymphoma was demonstrated.</p><p>The treatment outcome for 307 patients with advanced stages of HL, in an unselected population was in accordance with the treatment results of large centres world-wide. Some patients were successfully selected for a shorter chemotherapy-regimen without inferior treatment results.</p><p>In 124 patients with relapse, the survival of those primarily treated with radiotherapy according to the National guidelines was in accordance with the survival of patients of initially advanced stages. A worse outcome was found for those who received both chemotherapy and radiotherapy initially, probably because of a higher frequency of bulky disease in this group. </p><p>Immunohistochemical analysis of the tumour suppressor protein p53 and retinoblastoma protein (Rb) of paired samples at diagnosis and at relapse in 81 patients did not reveal any specific staining pattern affecting survival.</p><p>A novel B-cell line (U-2932) was established from a patient with a diffuse large B-cell lymphoma previously treated for advanced stage and subsequent relapses of HL. An identical rearranged IgH gene was demonstrated in tumour cells from the patient and in U-2932. A p53 point mutation was detected and over-expression of the p53 protein was found. A complex karyotype with high-level amplifications of the chromosomal regions 18q21 and 3q27, i.e. the loci for <i>bcl-2</i> and <i>bcl-6</i> were demonstrated. </p>

<p>Members of the transforming growth factor-β (TGF-β) superfamily of growth and differentiation factors regulate a vast array of biological functions in the adult, and are of great importance in governing cell fate determination and patterning in the developing embryo. The TGF-β signal is propagated intracellularly by Smad proteins resulting in transcriptional responses. Smad6 and Smad7 are inhibitory Smads known to downregulate the TGF-β signal and thereby possibly modulating the biological response. This thesis describes a functional analysis of the inhibitory Smad7 from an <i>in vitro </i>and <i>in vivo </i>perspective<i>.</i></p><p>The prostate gland is dependent on androgens for its growth and differentiation. Androgen withdrawal can cause regression and apoptosis in normal and malignant prostate. Previous studies suggest a role for TGF-β in the apoptotic mechanism. We investigated the expression levels of Smad proteins in the rat ventral prostate as well as in an androgen sensitive prostate tumor model (Dunning R3327 PAP) by immunohistochemistry. We observed an increased immunoreactivity for Smad3, Smad4 and phosphorylated Smad2 in the rat ventral prostate epithelial cells after castration, as well as in the prostate tumor cells. Expression of inhibitory Smad6 and Smad7 were also increased in both normal and malignant prostate in response to castration. </p><p>Several studies have shown that Smad7 is upregulated in response to TGF-β stimuli, suggesting a role in a negative feedback loop attenuating the TGF-β response. We investigated the molecular mechanism behind that response by studying the transcriptional regulation of the Smad7 gene. We identified a palindromic Smad binding element (SBE) in the promoter. Point mutations introduced into the SBE abolished transcriptional activation via TGF-β. We also observed that mutating or deleting binding motifs for Sp1 and AP-1, led to an attenuation of the TGF-β mediated transcriptional induction as well as the basal promoter activity.</p><p>Gene ablation of Smad proteins has revealed specific physiological and developmental roles. We analysed mice targeted on the Smad7 locus. The mice appeared viable and fertile with a slight reduction in litter size, suggesting a perinatal loss. Biochemical analysis of mouse embryonic fibroblasts (MEFs) showed no major difference between wild type and mutant MEFs. </p>

<p>The proline-rich tyrosine kinase (Pyk2) together with focal adhesion kinase (FAK) define a family of non-receptor protein tyrosine kinases that are regulated by diverse stimuli. Activation of Pyk2 has been implicated in multiple signaling events, including modulation of ion channels, activation of MAP kinase cascades and apoptotic cell death. This thesis investigates the role of Pyk2 in the regulation of mitogenic signals and cell cytoskeleton.</p><p>We identified a hematopoietic isoform of Pyk2 (designated Pyk2-H)that is generated by alternative RNA splicing and is mainly expressed in thymocytes, B cells and natural killer cells. In addition, we demonstrated that engagement of antigen receptors in lymphocytes leads to rapid tyrosine phosphorylation of Pyk2-H suggesting a potential role in host immune responses. These findings were corroborated by defects in B cell-mediated immune responses of Pyk2-/- mice. </p><p>Several reports have previously indicated that Pyk2 acts as an upstream regulator of ERK and JNK MAP kinase cascades in response to numerous extracellular signals. Which MAP kinase pathway is activated by Pyk2 depends on arrays of effector proteins associated with Pyk2. We proposed a model where the formation of Pyk2-Src complexes results in phosphorylation of Shc, p130Cas and Pyk2. This creates binding sites for the SH2 domains of adaptor proteins Grb2 and Crk, which in turn recruit exchange factors for Ras and Rho GTPases that specifically activate ERK or JNK.</p><p>Integration of signaling pathways initiated by receptor tyrosine kinases and integrins is essential for growth factor-mediated biological responses. We described neuronal cellular models where activation of both growth factor receptors and integrins is required for neurite outgrowth. In these cells, Pyk2 and FAK associate with integrin-linked complexes containing EGF receptors via their C- and N-terminal domains. Inhibition of Pyk2/FAK functions was sufficient to block neurite outgrowth and effectors of the C-terminal domain of Pyk2/FAK, including paxillin, were shown to regulate neurite outgrowth independently of ERK/MAP kinase in these cells. We thus proposed that Pyk2 and FAK play important roles in signal integration proximal to the integrin-growth factor receptor complexes.</p>

<p>The role of platelet-derived growth factor (PDGF) in malignancies involves both autocrine and paracrine stimulation of cells within the tumor. The interstitial fluid pressure (IFP) is one of the forces that govern the transvascular flow of fluids. In both experimental and clinical cancers, the IFP is elevated and is thought to act as a barrier for delivery of drugs. Increasing evidence points to PDGF as a positive regulator of the interstitial fluid pressure in loose connective tissue. In this thesis, the effect of PDGF receptor inhibition on the tumor IFP, transvascular transport and efficacy of anti-cancer drugs is investigated.</p><p>All studies were performed using tumor models that display extensive tumor stroma and PDGF receptor expression restricted to stroma cells. Blocking of PDGF receptor signaling significantly reduced the tumor IFP in various tumor models. In parallel, pre-treatment with PDGF antagonists increased the tumor content of cytotoxic agents without affecting the uptake in other organs. Moreover, combination treatment with PDGF receptor inhibitors and chemotherapeutic agents dramatically enhanced the anti-tumor effects of the cytotoxic drugs, whereas treatment with only PDGF receptor inhibitors did not affect the growth of the tumors. Beneficial effects on the tumor reponse to radioimmunotherapy were also produced after concomitant administration of PDGF antagonists. Importantly, anti-angiogenic effects, changes in cell composition and increased tumor cell sensitivity to cytotoxic agents were ruled out as the cause for the synergistic effects. </p><p>Studies with different temporal scheduling of PDGF receptor inhibitors demonstrated a perfect correlation between a reduced IFP, an increased transvascular transport and an enhanced therapeutic effect of cytotoxic drugs, strongly suggesting that the phenomena are causally linked.</p><p>The studies presented herein illustrate for the first time the potential of cells in the stroma compartment as a target for efforts to treat cancer. In conclusion, a novel, possibly general, strategy to enhance the effects of conventional anti-cancer drugs has been identified.</p>

<p>In this thesis, two different vaccines were evaluated as adjuvant therapy for patients with colorectal cancer. The ability of the two candidate vaccines to generate antigen-specific cellular and humoral responses, respectively, was studied. The effectiveness of granulocyte colony stimulating factor (GM-CSF) as a cytokine adjuvant to augment the immune response was also examined.</p><p>The first vaccination strategy involved immunization with the recombinant tumor-associated protein, carcinoembryonic antigen (CEA). Recombinant CEA was administered at 4 different dose levels 7 times during one year. Peripheral blood samples were regularly analyzed during 36 months. This vaccination regimen induced a strong immunoglobulin 1 (IgG1) and IgG4 response, a moderate IgG2 response and a weak IgG3 response against CEA. GM-CSF markedly augmented the effect on IgG1 and IgG4 as well as the T cell response. In contrast, dose of rCEA had no or modest effect on induced immune responses. The response gradually increased during the 12 months immunization period. Responses of all three IgG subclasses and of T cells were protracted up to 36 months. The anti-CEA IgG titers related significantly to survival. Functional HLA-DR epitopes of CEA could be defined. These major histocompatibility class II epitopes may serve as putative components of a peptide-based vaccination strategy. </p><p>The other vaccine strategy consisted of the tumor-associated antigen epithelial cell adhesion molecule (Ep-Cam) expressed as a transgene in a viral vector, ALVAC. Patients were immunized subcutaneously/intradermally 3 times over 6 weeks and monitored for immune responses for 46 weeks. No anti-Ep-Cam specific humoral response was induced, but Ep-Cam specific type 1 T cells (interpheron-gamma production) were induced, mainly in the GM-CSF group. The cytotoxic cellular response appeared late, or a few months after the last immunization.</p><p>Both vaccines were well tolerated. Since GM-CSF was an important component for both regimens, immungenicity of this cytokine was assessed. Multiple immunizations with low dose GM-CSF were associated with a low incidence of GM-CSF antibodies that did not neutralize the biological effect of GM-CSF. </p><p>In conclusion, both vaccines are promising candidate vaccines. GM-CSF is necessary to induce a strong humoral and cellular immune response. Large clinical trials are urgently warranted to evaluate the clinical efficacy.</p>

<p>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.</p><p>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. </p><p>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.</p><p>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.</p><p>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. </p><p>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. </p><p> 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. </p>

<p>Interaction between mercury and selenium has previously been observed both in man and in animals. The aim of this work was to study expressions of interaction between mercury and selenium in human K-562 cells. Inorganic and organic forms of mercury and selenium were used and cells were either pre-treated with selenium or simultaneously exposed to selenium and mercury. Concentrations of selenium and mercury chosen were indicated by a study of growth inhibition in the individual compounds: a low concentration of selenium and selenomethionine induced slight cell growth inhibition, while a high concentration resulted in a notable growth inhibition. Two mercury concentrations were chosen: one with minimal toxicity and another with high cell toxicity. In addition, uptake and retention patterns of selenomethionine and selenite differed in both selenocompounds.</p><p>All simultaneous treatments with 3.5 μM methylmercury produced a reduction in cellular mercury with increased selenium concentration. This was particularly obvious in selenite treatments. Growth curves from the simultaneous 3.5 μM methylmercury and selenite treatments indicated protection with increased selenite concentrations. In both exposure protocols, the 5 μM methylmercury treatments were toxic to the cells. </p><p>In both study protocols, cells exposed to selenite and mercuric chloride manifested increased cellular mercury uptake with increased selenium concentration. In all selenite and 35 μM mercuric chloride treatments, no inhibition of growth was observed, while the 50 μM mercuric chloride treatments were toxic to the cells. Selenite-dependent protection was achieved in both exposure protocols when considering the cellular uptake of mercury. With few exceptions, selenomethionine produced similar effects as selenite on mercuric chloride uptake and growth inhibition.</p>