<p>Background: Stereotactic body radiation therapy (SBRT) is a relatively new method which has been employed e.g. in the treatment of liver tumours. Little dosimetric data has been published for SBRT in the liver. The aim of this retrospective study was to quantify the dosimetric parameters that influence the toxicity of the healthy liver, and the effect on the tumour, for SBRT to liver tumours in patients treated at Karolinska University Hospital. A comparison was made to relating published studies.</p><p>Patients and Methods: The patient group to be studied were treated at Karolinska University Hospital for liver metastases with SBRT between July 1993 and October 2004. There were 64 patients treated with 71 treatment plans for 81 tumours. Differential dose volume histograms were collected for the clinical target volume (CTV), the planning target volume (PTV) and the liver excluding the CTV, from all dose plans. Since different fractionation schedules were used, the doses were normalised using the linear quadratic model, to be comparable. The doses to the uninvolved liver were evaluated with the mean liver dose, the Lyman-Kutcher-Burman (LKB) effective volume normal tissue complication probability (NTCP) model as well as the critical volume NTCP-model. A comparison was made to the studies of Dawson et al (2002) and Schefter et al (2005). The doses to the CTV were evaluated using the equivalent uniform dose tumour control probability (TCP) model, and related to target size and date of treatment.</p><p>Results: When the mean doses to the uninvolved liver (the liver volume without tumour tissue) were compared to Dawson and Ten Haken’s results (2005), 20 treatments out of 71 were predicted to give a risk of radiation induced liver disease (RILD) higher than 50%. The effective volume calculations predicted that 18 treatments gave a risk of RILD higher than 50%, when compared to the results of Dawson et al (2002). According to the critical volume model and the parameter values of Schefter et al (2005), our data predict that 10 of the treatments gave a risk of liver function failure, to an unspecified risk level. Treatments of large tumours resulted in higher doses to the liver. The doses to the CTV showed that the maximum prescribed dose decreased with increasing CTV.</p><p>Discussion and Conclusions: An evaluation of clinical data is necessary to make a full analysis of the treatments of this study. Such an analysis is planned for the future.</p>

<p>Positron Emission Tomography (PET) has gained an important roll in clinical for diagnosis, staging and prognosis of a range of cancer types. Utilization of PET for monitoring and evaluation of cancer treatment is an attractive but almost new concept. The proper choice of PET-tracer as a biomarker for treatment follow-up is crucial. The important characteristic for a suitable tracer is its ability to reflect the response to a treatment at an early stage, before any morphologically changes occurs. It would be an advantage to screen a battery of PET tracers in a preclinical model and introduce a few potential tracers in clinical trial. </p><p>The most conventional pre-clinical approach in PET-oncology utilizes xenografts in mice or rats and requires a large number of subjects. It would be a great advantage to introduce a less demanding but still reliable preclinical method for a more efficient planning of studies in animal model and then in human trials. </p><p>The Multicellular Tumour Spheroid (MTS) system represents an intermediary level between cells growing as monolayer and solid tumours in experimental animals or patients. It mimics the growth of naturally occurring human tumours before neovascularization and appears to be more informative than monolayer and more economical and more ethical than animal models.</p><p>The aim of this work was to establish, refine and evaluate the application of MTS model as a preclinical approach in PET oncology. The vision was to introduce a preclinical method to probe and select PET tracer for treatment monitoring of anticancer drugs, which can hopefully be applied for optimization in breast cancer treatment.</p><p>In this thesis, a number of basic experiments were performed to explore the character of 2-[fluorine-18]-fluoro-2-deoxy-d-glucose (FDG) uptake in MTS. FDG as the most established PET tracer was an obvious initial option for the evaluation of the model. For further assess-ment, we studied effects on FDG uptake in MTS treated with five routinely used chemother-apy agents. For association of PET tracer uptake to size change of MTS, we developed a reliable and user-friendly method for size determination of MTS. The next step was to apply the MTS model to screen PET tracers for analysis of early response of chemotherapy in breast cancer. Finally the method was utilized for translational imaging exemplified with a new chemotherapy agent.</p><p>The results were encouraging and the MTS model was introduced and evaluated as a preclini-cal tool in PET oncology. The method was implicated to in vitro quickly assess a therapy profile of existing and newly developed anticancer drugs in order to investigate the effects of candidate drugs on tumour-growth, selection of appropriate PET tracer for treatment monitor-ing and finally understanding relation between growth inhibition and biomarkers as part of translational imaging activities.</p>

Background: This is a retrospective study of patients treated with stereotactic body radiation therapy (SBRT) with the stereotactic body frame for centrally located lung tumours. The purpose was to investigate the doses to the different structures of the tracheobronchial tree and to relate these doses to the incidence of atelectasis. The goal was to estimate a tolerance dose for the bronchi. Materials: The patient material consisted of 71 patient treated at the Karolinska University Hospital for a total of 102 tumours between November 1993 and March 2004. The patient group consisted of 36 men and 35 women with a mean age at the treatment of 67 years (range 34-87). The group was a mixture of patients with primary lung cancer and pulmonary metastases. Methods: After rereading and reactivating the dose plans for the patients in the treatment planning system (TPS) the different tracheobronchial structures (trachea, right mainstem bronchus, right superior bronchus, right intermedius bronchus, right medius bronchus, right inferior bronchus, left mainstem bronchus, left superior bronchus, left intermedius bronchus, left inferior bronchus) were outlined. The dose distribution in each structure was calculated and a dose-volume histogram (DVH) was created. Patients were allocated to four groups, i.e. patients with right sided tumours (22), left sided tumours (14), mediastinal tumours (23) and bilateral tumours (10). After that the maximum and mean doses to all structures were analysed. An oncologist reviewed the medical records for the patients and especially looked for atelectasis. The doses were related to the incidence of atelectasis. Results and Conclusions: For the patient group with right sided tumours it seems like the maximum doses to the bronchi are higher for the patients with atelectasis in comparison with patients without atelectasis. A better correlation between atelectasis and maximum doses rather than mean doses was observed for these patients. At this moment the results are too preliminary, so it is not possible to suggest a tolerance dose for the bronchi. What can be said is that the maximum doses to the bronchi for patients with right sided tumours without atelectasis are below 250 Gy3 expressed in biologically equivalent dose (BED) with α/β=3Gy, while at least one bronchi structure in the atelectasis patients received a maximum dose above 250 Gy3.

Background: Stereotactic body radiation therapy (SBRT) is a relatively new method which has been employed e.g. in the treatment of liver tumours. Little dosimetric data has been published for SBRT in the liver. The aim of this retrospective study was to quantify the dosimetric parameters that influence the toxicity of the healthy liver, and the effect on the tumour, for SBRT to liver tumours in patients treated at Karolinska University Hospital. A comparison was made to relating published studies. Patients and Methods: The patient group to be studied were treated at Karolinska University Hospital for liver metastases with SBRT between July 1993 and October 2004. There were 64 patients treated with 71 treatment plans for 81 tumours. Differential dose volume histograms were collected for the clinical target volume (CTV), the planning target volume (PTV) and the liver excluding the CTV, from all dose plans. Since different fractionation schedules were used, the doses were normalised using the linear quadratic model, to be comparable. The doses to the uninvolved liver were evaluated with the mean liver dose, the Lyman-Kutcher-Burman (LKB) effective volume normal tissue complication probability (NTCP) model as well as the critical volume NTCP-model. A comparison was made to the studies of Dawson et al (2002) and Schefter et al (2005). The doses to the CTV were evaluated using the equivalent uniform dose tumour control probability (TCP) model, and related to target size and date of treatment. Results: When the mean doses to the uninvolved liver (the liver volume without tumour tissue) were compared to Dawson and Ten Haken’s results (2005), 20 treatments out of 71 were predicted to give a risk of radiation induced liver disease (RILD) higher than 50%. The effective volume calculations predicted that 18 treatments gave a risk of RILD higher than 50%, when compared to the results of Dawson et al (2002). According to the critical volume model and the parameter values of Schefter et al (2005), our data predict that 10 of the treatments gave a risk of liver function failure, to an unspecified risk level. Treatments of large tumours resulted in higher doses to the liver. The doses to the CTV showed that the maximum prescribed dose decreased with increasing CTV. Discussion and Conclusions: An evaluation of clinical data is necessary to make a full analysis of the treatments of this study. Such an analysis is planned for the future.

Positron Emission Tomography (PET) has gained an important roll in clinical for diagnosis, staging and prognosis of a range of cancer types. Utilization of PET for monitoring and evaluation of cancer treatment is an attractive but almost new concept. The proper choice of PET-tracer as a biomarker for treatment follow-up is crucial. The important characteristic for a suitable tracer is its ability to reflect the response to a treatment at an early stage, before any morphologically changes occurs. It would be an advantage to screen a battery of PET tracers in a preclinical model and introduce a few potential tracers in clinical trial. The most conventional pre-clinical approach in PET-oncology utilizes xenografts in mice or rats and requires a large number of subjects. It would be a great advantage to introduce a less demanding but still reliable preclinical method for a more efficient planning of studies in animal model and then in human trials. The Multicellular Tumour Spheroid (MTS) system represents an intermediary level between cells growing as monolayer and solid tumours in experimental animals or patients. It mimics the growth of naturally occurring human tumours before neovascularization and appears to be more informative than monolayer and more economical and more ethical than animal models. The aim of this work was to establish, refine and evaluate the application of MTS model as a preclinical approach in PET oncology. The vision was to introduce a preclinical method to probe and select PET tracer for treatment monitoring of anticancer drugs, which can hopefully be applied for optimization in breast cancer treatment. In this thesis, a number of basic experiments were performed to explore the character of 2-[fluorine-18]-fluoro-2-deoxy-d-glucose (FDG) uptake in MTS. FDG as the most established PET tracer was an obvious initial option for the evaluation of the model. For further assess-ment, we studied effects on FDG uptake in MTS treated with five routinely used chemother-apy agents. For association of PET tracer uptake to size change of MTS, we developed a reliable and user-friendly method for size determination of MTS. The next step was to apply the MTS model to screen PET tracers for analysis of early response of chemotherapy in breast cancer. Finally the method was utilized for translational imaging exemplified with a new chemotherapy agent. The results were encouraging and the MTS model was introduced and evaluated as a preclini-cal tool in PET oncology. The method was implicated to in vitro quickly assess a therapy profile of existing and newly developed anticancer drugs in order to investigate the effects of candidate drugs on tumour-growth, selection of appropriate PET tracer for treatment monitor-ing and finally understanding relation between growth inhibition and biomarkers as part of translational imaging activities.

We recently discovered a high-level amplicon spanning the chr19q13.41 microRNA cluster in CNS Primitive Neuroectodermal Tumour, which results in striking upregulation of miR-520g. Constitutive over-expression of miR-520g in untransformed human neural stem cells enhanced cell growth, restricted differentiation down the neuronal lineage, and promoted expression of neural stem/progenitor cell markers. We thus hypothesize that ectopic miR-520g expression promotes tumourigenesis in part by inhibiting cellular differentiation. Consistent with this proposition, miR-520g is silenced upon embryonic stem cell differentiation and its expression is absent from most adult tissues. Moreover, expression analysis of miR-520g overexpressing cells revealed significant dysregulation of developmental signalling pathways. Further efforts focused on elucidating mechanisms of miR-520g function led to the identification of a cell cycle inhibitor, p21, as an important candidate target. These findings collectively suggest that miR-520g may modulate differentiation by regulating developmental signalling pathways and cell cycle exit of neural stem/progenitor cells.

Medulloblastoma is the most common malignant paediatric brain tumour. Similar to other tumours, medulloblastoma pathogenesis involves abnormal regulation of several developmental growth pathways. As my thesis project, I studied the effects of aberrant HGF/MET signalling on medulloblastoma formation in two ways. In my first objective, I investigated the role that mutations play in activated HGF/MET signalling in medulloblastoma by searching for mutations in HGF/MET pathway genes, SPINT1, SPINT2, and MET, within primary medulloblastoma specimens. This screen identified several single nucleotide polymorphisms (SNPs) and two novel variations, one in each SPINT1 and SPINT2 genes. In my second objective, I generated a transgenic mouse model with cerebellar-specific aberrant MET signalling. These mice developed extensive cerebellar abnormalities but formed no tumours. These results indicate that mutations in the HGF/MET pathway components alone are not sufficient to initiate medulloblastoma formation and must coincide with additional genetic insults to promote tumour formation, maintenance, and progression.

We recently discovered a high-level amplicon spanning the chr19q13.41 microRNA cluster in CNS Primitive Neuroectodermal Tumour, which results in striking upregulation of miR-520g. Constitutive over-expression of miR-520g in untransformed human neural stem cells enhanced cell growth, restricted differentiation down the neuronal lineage, and promoted expression of neural stem/progenitor cell markers. We thus hypothesize that ectopic miR-520g expression promotes tumourigenesis in part by inhibiting cellular differentiation. Consistent with this proposition, miR-520g is silenced upon embryonic stem cell differentiation and its expression is absent from most adult tissues. Moreover, expression analysis of miR-520g overexpressing cells revealed significant dysregulation of developmental signalling pathways. Further efforts focused on elucidating mechanisms of miR-520g function led to the identification of a cell cycle inhibitor, p21, as an important candidate target. These findings collectively suggest that miR-520g may modulate differentiation by regulating developmental signalling pathways and cell cycle exit of neural stem/progenitor cells.

Medulloblastoma is the most common malignant paediatric brain tumour. Similar to other tumours, medulloblastoma pathogenesis involves abnormal regulation of several developmental growth pathways. As my thesis project, I studied the effects of aberrant HGF/MET signalling on medulloblastoma formation in two ways. In my first objective, I investigated the role that mutations play in activated HGF/MET signalling in medulloblastoma by searching for mutations in HGF/MET pathway genes, SPINT1, SPINT2, and MET, within primary medulloblastoma specimens. This screen identified several single nucleotide polymorphisms (SNPs) and two novel variations, one in each SPINT1 and SPINT2 genes. In my second objective, I generated a transgenic mouse model with cerebellar-specific aberrant MET signalling. These mice developed extensive cerebellar abnormalities but formed no tumours. These results indicate that mutations in the HGF/MET pathway components alone are not sufficient to initiate medulloblastoma formation and must coincide with additional genetic insults to promote tumour formation, maintenance, and progression.

The type II tumour suppressor ING3 has been shown to modulate transcription, cell cycle control, and apoptosis. To investigate the putative role of ING3 in melanoma development, we examined the expression of ING3 in 58 dysplastic nevi, 114 primary melanomas, and 50 metastatic melanomas with tissue microarray and immunohistochemistry. Overall ING3 was reduced in metastatic melanomas compared with dyslastic nevi and primary melanomas. Reduced nuclear ING3 staining also correlated with melanoma progression, increased cytoplasmic ING3 level, tumour location at sun-exposed sites, and a poorer disease-specific 5-year survival of patients with primary melanoma. Multivariate analysis revealed that nuclear ING3 staining can independently predict patient outcome in primary melanomas. In melanoma cells, ING3 expression was rapidly induced by UV irradiation. Using stable clones of melanoma cells overexpressing ING3, we showed that ING3 significantly promoted UV-induced apoptosis. Unlike its homologues ING1b and ING2, ING3-enhanced apoptosis upon UV irradiation was independent of functional p53. Furthermore, ING3 did not affect the expression of mitochondrial proteins but increased the cleavage of Bid and caspases. Moreover, ING3 upregulated Fas expression and ING3-mediated apoptosis was blocked by inhibiting caspase-8 or Fas activation. Knockdown of ING3 expression decreased UV-induced apoptosis remarkably, suggesting that ING3 plays a crucial role in cellular response to UV radiation. To explore how ING3 is deregulated in advanced melanomas, we examined ING3 expression in metastatic melanoma cells and found that ING3 was downregulated due to a rapid protein turnover in these cells. Further studies demonstrated that ING3 undergoes degradation via the ubiquitin-proteasome pathway. We also demonstrate that ING3 interacts with the SCF (Skp1/Cul1/Roc1/Skp2) E3 ligase complex. Knockdown of Cul1 or Skp2 significantly stabilized ING3 in melanoma cells. In addition, lysine residue 96 is essential for ING3 ubiquitination as its mutation to arginine completely abrogated ING3 turnover and enhanced ING3-stimulatd apoptosis upon UV irradiation. Taken together, ING3 is deregulated in melanomas as a result of both nucleus-to-cytoplasm shift and rapid degradation. The level of ING3 in the nucleus may be an important marker for human melanoma progression and prognosis. Restoration of ING3 expression significantly sensitizes melanoma cells to UV radiation through the activation of Fas/caspase-8 pathway.