| 1. |
- Österlund, Emerik, 1995-
(författare)
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Prognostic and Predictive Factors in Metastatic Colorectal Cancer
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The outcome for metastatic colorectal cancer (mCRC) patients has improved substantially in recent decades. This has chiefly been observed in study populations, and predominantly in left-sided primary tumours, which is why we wanted to study if and how survival has improved in the background population. It has also been seen that certain molecular subtypes are more common in population-based materials, and, thus, we studied the prevalence and effects of different molecular alterations.Paper I is a national population-based material of all 19 566 Swedish patients with a diagnosis of mCRC 2007-2016, 55% were male and 70% had synchronous metastases. Median overall survival (OS) for all patients was 14.0 months. An improvement could be seen over time, also in stratified analyses. OS was influenced by presentation of metastases, age, primary tumour location, and sex. All except sex remained statistically significant in a multivariable analysis. Differences of about one month in median OS were seen between healthcare regions, but these diminished over time.Paper II included all 765 patients from the Uppsala Region with a mCRC diagnosis 2010-2020. Right colon primary tumours were seen in 38%, left colon in 27% and rectum in 34%. BRAF-V600E mutations (mt) and deficient mismatch repair (dMMR) had a poor OS and were more common in right colon primary tumours. Primary tumour location did not affect OS in subgroups according to mutations in RAS or BRAF, nor in a multivariable analysis. Molecular alterations seem to be more important than primary tumour location for prognosis.Paper III studied KRAS-G12Cmt in three population-based and one real-world material. KRAS-G12C was seen in 2-4% of all tested and in 4-8% of all KRASmt. No differences in patient characteristics were observed between KRAS-G12C and other KRASmt. No differences in OS were seen between KRAS-G12C and other KRASmt, neither for all patients, nor in different treatment groups.Paper IV studied atypical BRAFmt (aBRAFmt) in two population-based and one real-world cohort. aBRAFmt was seen in 1-4% of the adequately tested patients in the different cohorts. aBRAFmt patients were predominantly male, had dMMR less often, more rectal primary tumours, and less peritoneal metastases compared with BRAF-V600Emt. Serrated adenocarcinomas were seen in about half of the aBRAFmt. OS was significantly better for aBRAFmt than in BRAF-V600Emt, but worse than for RASmt and RAS&BRAFwt patients. Nine aBRAFmt received epidermal growth factor receptor inhibitors without responses.
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| 2. |
- Österlund, Torben, et al.
(författare)
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Domain-structure analysis of recombinant rat hormone-sensitive lipase
- 1996
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Ingår i: Biochemical Journal. - 0264-6021. ; 319:Pt 2, s. 411-420
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Tidskriftsartikel (refereegranskat)abstract
- Hormone-sensitive lipase (HSL) plays a key role in lipid metabolism and overall energy homoeostasis, by controlling the release of fatty acids from stored triglycerides in adipose tissue. Lipases and esterases form a protein superfamily with a common structural fold, called the alpha/beta-hydrolase fold, and a catalytic triad of serine, aspartic or glutamic acid and histidine. Previous alignments between HSL and lipase 2 of Moraxella TA144 have been extended to cover a much larger part of the HSL sequence. From these extended alignments, possible sites for the catalytic triad and alpha/beta-hydrolase fold are suggested. Furthermore, it is proposed that HSL contains a structural domain with catalytic capacity and a regulatory module attached, as well as a structural N-terminal domain unique to this enzyme. In order to test the proposed domain structure, rat HSL was overexpressed and purified to homogeneity using a baculovirus/insect-cell expression system. The purification, resulting in > 99% purity, involved detergent solubilization followed by anion-exchange chromatography and hydrophobic-interaction chromatography. The purified recombinant enzyme was identical to rat adipose-tissue HSL with regard to specific activity, substrate specificity and ability to serve as a substrate for cAMP-dependent protein kinase. The recombinant HSL was subjected to denaturation by guanidine hydrochloride and limited proteolysis. These treatments resulted in more extensive loss of activity against phospholipid-stabilized lipid substrates than against water-soluble substrates, suggesting that the hydrolytic activity can be separated from recognition of lipid substrates. These data support the concept that HSL has at least two major domains.
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| 3. |
- Österlund, Torben
(författare)
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Structure-function relationships of hormone-sensitive lipase
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The primary role of hormone-sensitive lipase (HSL), an 84 kDa enzyme of 768 amino acids (in the rat protein), is to hydrolyse stored triacylglycerols in adipocytes of white adipose tissue. Catecholamines and insulin regulate lipolysis through cellular signalling. A major target of this regulation is HSL, activated through phosphorylation by cAMP-dependent protein kinase (cAMP-PK). Insulin activates a cAMP phosphodiesterase, leading to decreased cAMP levels and deactivation HSL. The structure-function relationships of HSL have been studied using the recombinant rat HSL expressed in COS cells and insect cells. The latter has provided large amounts of HSL protein that was purified to more than 95 % purity. The recombinant protein was shown to be identical to HSL purified from rat adipocytes with respect to different activities, specific activity and phosphorylation/activation by cAMP-PK. The primary structure of HSL from different species was compared to each other and to that of a bacterial lipase. Those parts that aligned to the bacterial lipase were suggested to form the a/b-hydrolase fold core of HSL, a fold common to lipases, esterases and other enzymes. A separate study confirmed this localisation and has provided a three-dimensional model for the catalytic core of HSL. A stretch of ca. 200 amino containing identified phosphorylation sites intervenes this fold and is thought to be a regulatory module. Primary-, secondary- and tertiary structure analysis provided suggestions for the amino acids of the catalytic triad (another common feature of lipases and esterases). All three suggested amino acids were probed by site-directed mutagenesis and found to be essential for activity. Other conserved amino acids, that were chosen as controls, were not important for activity. Thus, the catalytic triad of rat HSL is thought to be Ser-423, Asp-703 and His-733. The sequence analysis also suggested that HSL has at least two structural domains, one having the a/b-hydrolase fold core and regulatory module, and a second N-terminal domain of unknown function. HSL, subjected to denaturation and limited proteolysis, was monitored by measurements of activities, circular dichroism and fluorescence spectroscopy. These studies clearly support the concept that HSL has two structural domains. Cleavage peptides of HSL were identified by mass spectrometry after proteolysis and electrophoretic separation, and confirmed the location of predicted exposed regions, i.e. the regulatory module and a hinge region between the structural domains. Analysis of the hydropathic pattern of HSL, supported these observations, by revealing that hydrophilic regions correlate with the exposed regions. The N-terminal domain is mostly amphipathic whereas the a/b-hydrolase fold core is mostly hydrophobic. Thus, detergent/membrane interaction sites are thought to be located to the N-terminal domain and parts of the hydrolase core. The picture of HSL emerging from these studies is that HSL probably has two structural domains separated by a short hinge region. The N-terminal domain is composed of ca. the 320 first residues and may mediate interactions with the intracellular lipid droplet substrate. The C-terminal domain is composed of ca. the last 430 residues of which half constitutes the a/b-hydrolase fold core and harbours the catalytic triad, whereas the rest is suggested to be a regulatory module containing the phosphorylation sites.
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