Recent discoveries enabled us to divide medulloblastoma into molecular sub-groups and uncover novel mutations in these tumors. However, except for superior survival of the WNT sub-group, the prognostic and therapeutic implications of these observations remain unclear. TP53 mutations which confer radioresistance revealed conflicting clinical relevance in different studies. We hypothesized that the effect of TP53 mutations on survival is modulated through molecular sub-grouping. This is especially important since therapeutic targeting of WNT can be achieved with administration of lithium. Here we first confirmed that TP53 mutant tumors confers unfavorable outcome only in SHH subgroup, but not in WNT. We demonstrated that while TP53 mutations cause radioresistance, activation of WNT/β-catenin signaling radiosensitizes medulloblastoma cells. We demonstrated that lithium activates the WNT pathway and effectively sensitize medulloblastoma cells to radiation. Furthermore, lithium did not sensitize normal neural stem cells to radiation, suggesting its potential as an effective radiosensitizer for medulloblastoma.

Recent discoveries enabled us to divide medulloblastoma into molecular sub-groups and uncover novel mutations in these tumors. However, except for superior survival of the WNT sub-group, the prognostic and therapeutic implications of these observations remain unclear. TP53 mutations which confer radioresistance revealed conflicting clinical relevance in different studies. We hypothesized that the effect of TP53 mutations on survival is modulated through molecular sub-grouping. This is especially important since therapeutic targeting of WNT can be achieved with administration of lithium. Here we first confirmed that TP53 mutant tumors confers unfavorable outcome only in SHH subgroup, but not in WNT. We demonstrated that while TP53 mutations cause radioresistance, activation of WNT/β-catenin signaling radiosensitizes medulloblastoma cells. We demonstrated that lithium activates the WNT pathway and effectively sensitize medulloblastoma cells to radiation. Furthermore, lithium did not sensitize normal neural stem cells to radiation, suggesting its potential as an effective radiosensitizer for medulloblastoma.

Glioblastomas are among the most devastating of human cancers with a median survival of only 9-12 months. This type of brain tumor is incurable, largely due its remarkable proliferative capacity and resistance to current treatments. High levels of the Wilms' Tumor 1 (WTI) gene have been identified in glioblastomas, suggesting an oncogenic function. Moreover, known WT1 target genes have been implicated in resistance to radiation. To determine the role of WT1 in radiation resistance, two glioblastoma cell lines expressing WT1 were treated with siRNAs to silence this gene. Confirmation of WT1 knockdown was achieved through real-time PCR and Western blot. After treatment with siRNA, cells were irradiated, and cell survival was assessed using a luminescent ATP assay and clonogenic survival assay. We demonstrate that treatment with WT1 siRNA increased the radiation sensitivity in both cell lines. These findings suggest that WT1 functions to protect glioblastoma cells from radiation-induced death.

A vibration-based sound power (VBSP) measurement method is appealing because of its potential versatility in application compared to pressure- and intensity-based methods. The VBSP method is based on the well-known elementary radiators approach and is reliant on the acoustic radiation resistance matrix. Previous research has developed and validated the VBSP method for flat plates and cylinders. This thesis details work on extending the VBSP method to arbitrarily-curved structures. The approach of computing surface normal velocities from 3D velocity data measured by a scanning laser Doppler vibrometer (SLDV) is presented. This approach is validated with experimental sound power results of a cylindrical shell using the VBSP method with 3D velocity and geometry data. The sound power results are shown to have good agreement with ISO 3741 results. Experimental sound power results from three simple-curved plates using the VBSP and ISO 3741 methods are shown to have good agreement. These experimental results indicate that the VBSP method is less sensitive to background noise than the ISO 3741 method. An overview of exploring inherent symmetry in the radiation resistance matrix is presented for the purpose of increasing efficiency in applying the VBSP method. Sound power sensitivity to the formulation of the radiation resistance matrix is explored as another relevant option for increasing the efficiency of the VBSP method for many cases and for extending the method to more complex structures. The results of the radiation resistance matrix exploration enable the VBSP method to apply to arbitrarily-curved structures. Experimental sound power results using the VBSP method with the simple-curved plate radiation resistance matrix and the ISO 3741 method are compared for two arbitrarily-curved panels and are shown to have good agreement. The VBSP method based on the simple-curved plate form of the radiation resistance matrix is shown to have excellent agreement with numerical results from boundary element models, which inherently use the appropriate form of the radiation resistance matrix.

<p><strong><em>Background</em></strong>: Chemotherapy and radiation resistance are major causes of failure in cancer treatment. The response to treatment in cancer cells depends on several mechanisms and pathways such as Janus kinases-signal transducers and activators of transcription JAK/STAT pathway. STAT1 was the first described transcription factor in the STAT family. STAT1 is activated by stimulation of signaling proteins such as type II interferon (IFN- γ) and the activated STAT1 translocates from cytoplasm to nucleus. The translocation of STAT1 would result in transcription and changes in the cell activity in terms of apoptosis, proliferation and angiogenesis. Overexpression of STAT1 is suggested to be involved in the development of resistance to chemotherapy and radiation. In this study, we were interested in finding an inhibitor of the STAT1 translocation. <strong><em>Material and methods</em></strong>: The cervix carcinoma cell line, HeLa, was exposed to test compounds for 2h and were then stimulated with IFN-γ to induce the translocation of STAT1. To detect STAT1-protein and the nucleus, the cells were stained with fluorescent antibodies and Hoescht 33324, respectively, using a STAT1 activator assay. The difference in fluorescence intensity between cytoplasm and nucleus was measured using a high-content microscope, ArrayScan^®. <strong><em>Results</em></strong>: β-lapachone and CRA-1 were found to be inhibitors of STAT1 translocation.</p>

Background: Chemotherapy and radiation resistance are major causes of failure in cancer treatment. The response to treatment in cancer cells depends on several mechanisms and pathways such as Janus kinases-signal transducers and activators of transcription JAK/STAT pathway. STAT1 was the first described transcription factor in the STAT family. STAT1 is activated by stimulation of signaling proteins such as type II interferon (IFN- γ) and the activated STAT1 translocates from cytoplasm to nucleus. The translocation of STAT1 would result in transcription and changes in the cell activity in terms of apoptosis, proliferation and angiogenesis. Overexpression of STAT1 is suggested to be involved in the development of resistance to chemotherapy and radiation. In this study, we were interested in finding an inhibitor of the STAT1 translocation. Material and methods: The cervix carcinoma cell line, HeLa, was exposed to test compounds for 2h and were then stimulated with IFN-γ to induce the translocation of STAT1. To detect STAT1-protein and the nucleus, the cells were stained with fluorescent antibodies and Hoescht 33324, respectively, using a STAT1 activator assay. The difference in fluorescence intensity between cytoplasm and nucleus was measured using a high-content microscope, ArrayScan®. Results: β-lapachone and CRA-1 were found to be inhibitors of STAT1 translocation.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18869号 / 医博第3980号 / 新制||医||1008(附属図書館) / 31820 / 京都大学大学院医学研究科医学専攻 / (主査)教授 武田 俊一, 教授 小西 郁生, 教授 小松 賢志 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Fatty acid synthase (FASN), the sole cytosolic mammalian enzyme for de novo lipid synthesis, is crucial for cancer cell survival and associates with poor prognosis. FASN overexpression has been found to cause resistance to genotoxic insults. Here we tested the hypothesis that FASN regulates DNA repair to facilitate survival against genotoxic insults and found that FASN suppresses NF-κB but increases specificity protein 1 (SP1) expression. NF-κB and SP1 bind to a composite element in the poly(ADP-ribose) polymerase 1 (PARP-1) promoter in a mutually exclusive manner and regulate PARP-1 expression. Up-regulation of PARP-1 by FASN in turn increases Ku protein recruitment and DNA repair. Furthermore, lipid deprivation suppresses SP1 expression, which is able to be rescued by palmitate supplementation. However, lipid deprivation or palmitate supplementation has no effect on NF-κB expression. Thus, FASN may regulate NF-κB and SP1 expression using different mechanisms. Altogether, we conclude that FASN regulates cellular response against genotoxic insults by up-regulating PARP-1 and DNA repair via NF-κB and SP1.

<p>Renal cell carcinoma (RCC) is the most frequently lethal of urological cancers. It arises in the lining of the proximal convoluted tubule of the kidney and is most common in men ages 50–70. Often, partial or radical nephrectomy is needed to effectively treat the disease, leaving patients with reduced kidney function. RCC frequently displays significant radiation resistance, limiting the usefulness of traditional radiation therapy which might spare patients’ normal tissue. The enzyme carbonic anhydrase 9 (CA9), a product of the hypoxia pathway, is found upregulated in the majority of RCC, particularly the clear cell type. It catalyses the dissolution of carbon dioxide into water as bicarbonate and has been linked to increased invasion and migration in RCC tumour cells. The radiation resistance of two RCC cell lines 786-O (human CCRCC) and RAG (murine renal adenocarcinoma) was investigated by the clonogenic assay in the presence of a CA9 inhibitor or silencing RNA. The interference with CA9 by either of these methods significantly sensitizes 786-O cells to the effects of ionizing radiation <em>in vitro</em>. Moreover, fractionation of the dose delivered can increase this sensitization effect. It is hoped that current targeting of CA9 can make radiation therapy a more feasible option in the treatment of RCC.</p> / Master of Science (MSc)

<p>In this thesis, a model of tissue oxygenation is presented, that takes into account the heterogeneous nature of tumor vasculature. Even though the model is rather simple, the resulting oxygen distributions agree very well with clinically observed oxygen distributions for most tumors and healthy normal tissues. The model shows that the vascular density may not describe the oxygenation of a tissue sufficiently well, unless the heterogeneity of the vascular system is taken into account. Based on the oxygen distributions from the tissue model, the associated radiation response at low and high doses can be determined. </p><p>The radiation response of heterogeneous tumors should preferably be described by two clonogen compartments, one resistant and one sensitive, dominating the response at high and low radiation doses, respectively. Furthermore, each compartment should be characterized by the effective radiation resistance and the effective clonogen number. The resistant-sensitive model of radiation response has been analyzed in great detail. It accurately describes the response of severely heterogeneous tumors, both at low and high doses and LET values. The effective response parameters are given as integrals, averaged over the whole spectrum of radiation resistance. The parameters can also be determined from clinically established dose-response relations. </p><p>The main properties of the dose-response relation for a generally heterogeneous tumor is described in some detail. The normalized dose-response gradient has been generalized to take heterogeneities in both dose delivery and radiation response into account. This quantity is important for accurate treatment plan optimization using intensity modulated radiation therapy for individual patients. </p>