One-carbon metabolism, a network of enzymatic reactions involving the transfer of methyl groups, depends on B-vitamins as cofactors, folate as a methyl group carrier, and amino acids, betaine, and choline as methyl group donors. One-carbon metabolism influences many processes in cancer initiation and development such as DNA synthesis, genome stability, and histone and epigenetic methylation. To study markers of one-carbon metabolism and inflammation in relation to colorectal cancer (CRC) risk, we used prediagnostic plasma samples from over 600 case participants and 1200 matched control participants in the population-based Northern Sweden Health and Disease Study cohort. This thesis studies CRC risk with respect to the following metabolites measured in pre-diagnostic plasma samples: 1) folate, vitamin B12, and homocysteine; 2) components of one-carbon metabolism (choline, betaine, dimethylglycine, sarcosine, and methionine); and 3) three markers of different aspects of vitamin B6 status. In addition, this thesis examines three homocysteine ratios as determinants of total B-vitamin status and their relation to CRC risk. In two previous studies, we observed an association between low plasma concentrations of folate and a lower CRC risk, but we found no significant association between plasma concentrations of homocysteine and vitamin B12 with CRC risk. We have replicated these results in a study with a larger sample size and found that low folate can inhibit the growth of established pre-cancerous lesions. Using the full study cohort of over 1800 participants, we found inverse associations between plasma concentrations of the methionine cycle metabolites betaine and methionine and CRC risk. This risk was especially low for participants with the combination of low folate and high methionine versus the combination of low folate and low methionine. Well-functioning methionine cycle lowers risk, while impaired DNA synthesis partly explains the previous results for folate. We used the full study cohort to study associations between CRC risk and the most common marker of vitamin B6 status, pyridoxal' 5-phosphate (PLP), and two metabolite ratios, PAr (4-pyridoxic acid/(PLP + pyridoxal)) estimating vitamin B6 related inflammatory processes and the functional vitamin B6 marker 3-hydroxykynurenine to xanthurenic acid (HK:XA). Increased vitamin B6-related inflammation and vitamin B6 deficiency increase CRC risk. Inflammation was not observed to initiate tumorigenesis. Total B-vitamin status can be estimated by three different recently introduced homocysteine ratios. We used the full study cohort to relate the ratios as determinants of the total B-vitamin score in case and control participants and estimated the CRC risk for each marker. Sufficient B-vitamin status as estimated with homocysteine ratios was associated with a lower CRC risk. These studies provide a deeper biochemical knowledge of the complexities inherent in the relationship between one-carbon metabolism and colorectal tumorigenesis.

Glioblastoma is the most common primary brain tumor in humans and has a poor prognosis. Current therapies are not curative. Oncolytic viruses (OVs) are being investigated as tools to induce immunogenic cell death (ICD), cell death capable of activating the immune system. Semliki Forest Virus (SFV) strain 4 is an OV being investigated to treat glioblastoma. Previous studies in our lab have shown that SFV4 can induce ICD in human osteosarcoma (HOS) cells and ongoing in vivo studies show that SFV4 infected GL261 cell vaccination providesprotective immunity in mouse models. This study aimed to characterize the ICD induced by SFV4in glioblastoma cell lines, namely GL261, SB28 and CT2A, and to explain some of our in vivoobservations, namely why vaccination with SFV infected GL261 provides protective immunity but vaccination with infected CT2A and SB28 does not. Our in vitro studies found that GL261 is resistant to SFV4 while SB28 and CT2A are susceptible. We show that the virus can replicate in all three cell lines as seen by the presence of dsRNA, but that viral translation is delayed or inhibited in GL261 cells as not all cells positive for dsRNA were positive for SFV4 protein. Additionally, the type I interferon (IFN) pathway, responsible for antiviral defense, was highly upregulated in CT2A and SB28 but not as much in GL261 after infectionas seen by surveying IFIT1, IFITM3 and IFN-beta genes. Interferon stimulated genes (ISG) like CXCL10, a chemoattractant, was highly upregulated in GL261 after infection and might account for the protective immunity seen in vivo after vaccination. PDL1, an interferon stimulated gene responsible for self-tolerance, was highly upregulated in CT2A after infection. The IFN-beta ELISA revealed that both infected and uninfected GL261 cells produce IFN-beta suggesting a constitutively active pathway. Our DC phagocytosis assay showed that SFV4 infection of CT2A and SB28 cells induced a significant increase in DC phagocytosis and SFV4 infection of all three cell lines significantly increased DC maturation. We conclude that SFV4 infection of GL261 cells may induce ICD in vivo through a persistent viral infection and increased expression of CXCL10.

In recent years, digitalization and computer-based technologies have greatly revolutionized the field of bioinformatics. Advance research and development of computer-based programs have enhanced various DNA sequencing technologies. This advancement has significantly broadened our understanding of genomic evolution and has widely contributed to the application of clinical genomics. Cancer has been one of the major causes of death across the world. Cancer is mainly caused due to the damage or changes in DNA that affect the function of genes which contain a set of instructions that control various functions of cells. This damage in genes that maintain DNA repair mechanism may lead towards genome instability allowing rapid growth of cancer.   Microsatellite instability (MSI) is one such condition characterized due to genomic alteration leading towards the failure of DNA repair mechanism in cancerous cells. MSI is found in various types of cancer but is most often found in colorectal cancer, gastric cancer, and endometrial cancer. Hence, detection of this MSI can greatly contribute towards cancer therapies and enable to plan for the best treatment. This study mainly focuses on evaluating the performance of MSI calling algorithms using targeted sequencing methods.   The literature provides a detailed outline of various topics related to MSI detection. Moreover, different computational methods like MSIsensor, MSIsensor-ct, MSIsensor-pro, MSings, MiMSI, and MSIsensor2 were used in this study for the detection of MSI in selected samples are thoroughly discussed in the methodology section. Finally, the findings of this study conclude that the MSI calling algorithms mentioned above provide accurate detection of MSI in the chosen samples. Also, these algorithms enable us to determine the MSI status of the chosen samples more precisely

Bakgrund: Ordet cancer kan vara ett väldigt laddat ord för många människor, cancer hos barn är ännu mer laddat eftersom det gäller just ett barn. Då ett barn får diagnosen cancer är det inte bara barnet som drabbas, utan även familjen och stora delar av omgivningen såsom skola och kompisar. Idag finns det goda chanser att barn överlever sin cancersjukdom och det finns välgjorda riktlinjer för hur vården ska bedrivas under aktiv behandling och hur uppföljningen av barnet ska ske efter avslutad behandling. Men finns det någon plan för att uppföljning av föräldrarna och hur är deras mående tas omhand efter en påfrestande tid som denna? Syfte: Syftet var att undersöka föräldrars erfarenheter av livet efter att barnet har avslutat en lyckad cancerbehandling. Metod: En litteraturstudie genomfördes och resultatet baseras på åtta vetenskapliga kvalitativa och kvantitativa artiklar. Resultat: Fyra övergripande kategorier identifierades: Erfarenhet av känslomässiga reaktioner; Erfarenhet av påverkan på livet; Betydelsen av stöd och relationer; Att återgå till det normala. Slutsats: Resultat i studien visade att många föräldrar upplever ständig närvaro av oro, ångest eller depression även efter barnets avslutade behandling. De föräldrar som uppfattar sina barn som mer sårbara och bräckliga blir mer påverkade av den negativa stress som sjukdomen och dess behandling har utgjort. Vidare gick att utläsa att den ständiga jakten efter eventuella symtom blir ett stresspåslag och utgör en påminnelse i det vardagliga livet. Allt detta bidrar till okontrollerbara tankar om återfall eller död vilket leder till ett hinder för familjen att bearbeta det de varit med om och gå vidare. En förlorad nära kontakt med vårdpersonal efter avslutad behandling får vissa föräldrar att känna ångest.

Introduction: Rituximab is a monoclonal antibody used to treat hematological malignancies. The antibody depletes CD20+ B cells via cytotoxic immune mechanisms, such as complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC), which is mainly induced by natural killer (NK) cells. Rituximab is mostly well-tolerated but has been reported to induce the release of large amounts of cytokines in blood, thus causing systemic inflammatory response. Aim: To study rituximab-induced B cell depletion and cytokine release in blood from healthy volunteers and how this was affected by Fc modified versions of the antibody. Methods and materials: Fresh blood from healthy donors (n=3) was incubated with rituximab and Fc modified versions that influence the antibody’s target functions, namely ADCC and CDC, for 4 hours in a blood loop system. Results were measured using multicolor flow cytometry, except for cytokine release in plasma which was measured by enzyme-linked immunosorbent assay (ELISA). Results: Of all treatments, rituximab wild type (WT) showed superior B cell depletion than Fc mutant rituximab. The C1q knock-out variant (rituximab-P331S) and the variant with improved affinity to Fc receptor CD16 (rituximab-GASDALIE) did not differ in depletion. A cytokine release was not detected with the treatments, however, a cytokine stimulation in NK cells was observed. Rituximab-GASDALIE had the most prominent cytokine stimulation and CD107a (marker of NK cell functional activity) expression on NK cells. Rituximab-WT and rituximab-P331S had a minor and similar cytokine stimulation and CD107a expression between each other. Rituximab-IgG2 had minimal B cell depletion, CD107a expression and cytokine stimulation. Conclusions: Rituximab depleted B cells without inducing measurable cytokine release for healthy individuals. Among the treatments, Fc mutant rituximab seem to induce less B cell depletion. Moreover, rituximab-GASDALIE appear to elicit an enhanced NK cell activation. Further studies should include more donors as supplement and the results should be interpreted as complementary data to future data analyzed by performing the loop experiment using blood from patients.

Cancer is a global health issue and is estimated to be the second leading cause of death worldwide. Within cancer treatment, it has become attractive to introduce precision medicine to harness the body’s own immune system to fight cancer. Personalised peptide cancer vaccine can activate the immune system and elicit an immune response by using the patient’s own blood and tumour cells.   The aim of this study was to gather information to better understand the production of individual-specific peptides and the challenges when producing these peptides with focus from a GMP perspective. The methodology for conducting this study and retrieve information for the result was gathered by three different data bases; EMA, European Commission and FDA, and by qualitative semi-structured interviews.   The findings show that these personalised peptides should be manufactured by using solid-phase peptide synthesis, cleavage from the resin, HPLC purification, and final salt exchange. The traditional GMP requirements must be used as a basis to build a pragmatic program. In terms of delivery time, the peptides can be delivered within a few weeks. The main challenges will be to manufacture successful peptides since the peptide sequences vary from patient to patient and due to that fact have different production efficacy, deliver within the set timeframe, and have flexibility in the manufacture process.   Multiple guidelines need to be followed in order to set up a process that consistently delivers the intended product. When producing oncology drugs on-demand it is essential to keep the timeline since the patients are severely ill. However, further studies are needed to determine how the manufacture of personalised peptides could be performed to harmonise with regulatory guidelines.

BAKGRUND: I Sverige är cancer en av de vanligaste sjukdomarna. Bröstcancer är den vanligaste cancersjukdomen hos kvinnor. Bröstcancer kan sprida sig i kroppen och bilda en sekundär cancer som kallas metastas. Transformerande tillväxtfaktor-β-receptorer (TGF-β-receptorer) har en viktig roll i tumörutveckling. SYFTE: Syftet med denna studie är att studera rollen av TGF-β-receptorer i cancer, särskild bröstcancer med fokus på metastaserna. METOD: Den är en litteraturstudie, som är baserad på sex vetenskapliga artiklar. Dessa artiklar hämtas från databas PubMed genom användning av olika specifika sökord. RESULTAT: Resultat visar att TGF-β-receptorer har två motsatta rollar i cancerutveckling. Första fungerar TGF-β-receptorer som en tumörundertryckare i början av cancerutveckling. Däremot bidrar TGF-β-receptorer till utveckling av maligna celler lite senare under cancerutveckling. Det gör det genom att förbättra metastatiska potential samt undertrycka antitumörimmunitet. Metastaser bildas via Epithelial to mesenchymal transition med hjälp av vissa andra faktorer såsom cancerassocierade fibroblaster. Epithelial to mesenchymal transition och andel av cancerassocierade fibroblaster kan stimuleras av TGF-β-receptorer, som kan i sin tur leda till ökning av metastaseringen. Ett exempel på metastas är skelettcancer. DISKUSSION: Transformerande tillväxtfaktor-β receptorer, som kan påverka cancer, betraktas som ett bra fynd i läkemedelsutveckling mot cancer. TGF-β-receptorer har en viktig roll i tumörutveckling, särskilt sekundär cancer. Sekundär cancer/metastas är farligare än primär cancer, därför är det bättre att försöka förstå hela mekanismen bakom TGF-β-receptorer på ett tydligt sätt, för att kunna behandla patienter i god tid. SLUTSATS: Transformerande tillväxtfaktor- β -receptorer är en av de viktigaste parametrarna för att få den optimala effekten av medicineringen mot cancer.

Background: Neuroblastoma (NB) is a pediatric tumor in infants and young children. The survival rate is only around 50 percent for high-risk NB despite advanced and intense multi-modal therapy. Current research aims to find new effective treatment additional to modern therapy to improve prognosis of high-risk NB in children. As such, SU11248 may be a valuable approach for improving treatment and survival as growth factors have crucial roles in tumor growth, angiogenesis, and metastasis. Aim: The aim of this investigation was to examine tissues from SU11248 treated and nontreated tumor-bearing animals on the abundance of tumor-associated macrophages (TAMs) in metastases found on vital organs. Our hypothesis is that if SU11248 could cause “deactivation” or “silencing” of the stroma of metastases particularly by acting on stromal immune cells such as TAMs. Methods: Paraffin-embedded metastases developed in an orthotopic xenograft model in beige SCID mice were stained with a monoclonal rat anti-mouse antibody as a marker of TAMs. Morphological analysis of tissue slides, and macrophage quantification was performed using a microscope. Statistical analysis was achieved using an unpaired two tailed t-test.  Results: Macrophages were stained nicely, but the number of macrophages in the metastases were not statistically different between the vehicle treated controls and SU11248 treated metastases. Conclusion: In patients with high-risk NB, SU11248 may be a useful therapeutic supplement. We believe that further research into mechanisms that target critical factors for angiogenesis and metastasis in NB, such as TAMs, is an important step toward improving patient outcomes in high-risk NB.

Background: As a nurse, when caring for children with cancer, you need to offer the best care for the child and in the meantime focus on family centered care in order to meet the thoughts and wishes the family carries. Caring for children with cancer might contain many obstacles and difficulties. Communication, education and to understand one's role as a nurse is important when caring for these children. Aim: To identify nursing experience when caring for children with cancer. Method: A literature overview has been done and ten studies were gathered through the databases PubMed and Cinahl. The analysis of these studies created four categories with associated subcategories. Results: It came through those nurses faced difficulties in pediatric care in different situations. These situations created categories which were: Ethical issues, Education, Cooperation with the family and Nurses emotions and support. Conclusion: Factors found in this study were nurses' experience of lacking the necessary preparation, the need of emotional support and the importance of working close to the families. It is found that nurses felt unprepared for their work and dealing with tough situations. The relationship between the nurse and the patient's family is important to provide high quality care. Nurses expressed the need for support from colleagues and family members in order to grow as a nurse and as a person.

Rhabdomyosarcoma (RMS) is a type of soft tissue sarcoma that mainly occurs in children. RMS can be divided into two subtypes embryonal (ERMS) and alveolar (ARMS). The ARMS subtype can be especially aggressive when a balanced chromosomal translocation is present. This translocation results in the expression of a PAX3/7-FOXO1 fusion protein, an oncogenic transcription factor. PAX3-FOXO1 positive RMS has an especially bad prognosis and survival rate. In the cell painting assay relevant organelles are stained and morphological features are extracted on a cellular level. Based on these features, morphological profiles of each cell type and a similarity score can be calculated. The morphological profiles of ERMS cell lines RD and RD18 as well as the ARMS cell lines RH30 and CW9019 were obtained. RD and RD18 are most similar to each other followed by RH30. CW9019 has a very different profile from the other cell lines. Since ERMS is associated with a better survival rate a drug that reprograms the phenotype from ARMS to a more ERMS-like phenotype might sensitize the cells to the standard treatment of RMS. ARMS patients might then benefit from a combination therapy of such a drug and the standard treatment. To find such a drug a drug screen was conducted. Drugs were selected for the screen that either target fusion-protein stability or overexpressed targets of the fusion protein. Phenotypic reference compounds were included to get a first idea of the mechanisms and involved organelles of the screened drugs. In total, 30 compounds and 9 phenotypic reference compounds were screened for changing morphological profiles of RMS cell lines with the cell painting assay. 15 compounds were identified that change the phenotype of the ARMS cell line RH30. In addition, 7 of these compounds shift the phenotype of RH30 towards an ERMS-like phenotype. If that morphological resemblance of RH30 cells to ERMS cells translates into a change of a more ERMS-like behavior and sensitizes the cells for the standard treatment of RMS remains to be investigated. The reprogramming hits showed high similarity with phenotypic reference compounds that increase nucleus size which might suggest changed behavior of the fusion protein. Especially, Bosutinib, Midostaurin and Alisertib are promising new compounds for ARMS treatment. They shifted the ARMS phenotype towards an ERMS-like phenotype in the drug screen. This shift is likely a result of the interaction with PLK1 or Aurora-kinase A that are shown to have an influence on fusion-protein stability.