新規TGF-β標的因子TUFT1のmTORシグナルにおける機能解析

学位の種別: 課程博士審査委員会委員 : （主査）東京大学教授 饗場 篤, 東京大学教授 武川 睦寛, 東京大学教授 吉川 雅英, 東京大学教授 水島 昇, 東京大学教授 柴田 龍弘University of Tokyo(東京大学

Early Intensive Nutrition Intervention with Dietary Counseling and Oral Nutrition Supplement Prevents Weight Loss in Patients with Advanced Lung Cancer Receiving Chemotherapy: A Clinical Prospective Study

【Background】 Weight loss in patients with cancer is caused by cancer cachexia and chemotherapy-induced nausea and vomiting (CINV). Recent developments in antiemetic drugs have substantially improved CINV, but nutritional intervention did not improve body weight. This study aimed to investigate the effects of nutrition intervention with appropriate antiemetic treatment in patients with non-small-cell lung cancer during chemotherapy. 【Methods】 Patients received individualized nutrition counseling by a registered dietitian and were provided with oral supplements for 90 days. Body weight and other parameters were measured at baseline and after 90-day intervention. To evaluate this nutrition intervention, patients were also retrospectively set as control, and then body weight change was compared with inverse probability of treatment weights (IPTW) analysis. 【Results】 Ten patients received individualized nutrition counseling and were provided with oral supplements for 90 days. Of them, 7 patients consumed nutritional supplements, and the mean intake was 130 kcal/day. After 90-day intervention, the patients did not show significant weight and BMI loss during the course of cytotoxic chemotherapy. A total of 38 patients were retrospectively enrolled as controls. The number of the patients who gain the body weight after 90 days in the study cohort was significantly larger than that in the retrospective controls with the IPTW analysis (Odds Ratio (OR) = 8.4; 95% Confidence Interval (CI): 1.6-42; P = 0.01). 【Conclusion】 Early intensive nutrition intervention with appropriate antiemetic treatment prevents weight loss. Nutrition interventions might be also beneficial for quality of life, treatment response and survival

TUFT1 interacts with RABGAP1 and regulates mTORC1 signaling

The mammalian target of rapamycin (mTOR) pathway is commonly activated in human cancers. The activity of mTOR complex 1 (mTORC1) signaling is supported by the intracellular positioning of cellular compartments and vesicle trafficking, regulated by Rab GTPases. Here we showed that tuftelin 1 (TUFT1) was involved in the activation of mTORC1 through modulating the Rab GTPase-regulated process. TUFT1 promoted tumor growth and metastasis. Consistently, the expression of TUFT1 correlated with poor prognosis in lung, breast and gastric cancers. Mechanistically, TUFT1 physically interacted with RABGAP1, thereby modulating intracellular lysosomal positioning and vesicular trafficking, and promoted mTORC1 signaling. In addition, expression of TUFT1 predicted sensitivity to perifosine, an alkylphospholipid that alters the composition of lipid rafts. Perifosine treatment altered the positioning and trafficking of cellular compartments to inhibit mTORC1. Our observations indicate that TUFT1 is a key regulator of the mTORC1 pathway and suggest that it is a promising therapeutic target or a biomarker for tumor progression.UTokyo FOCUS Articles掲載「がんの増殖・転移を促進する新規因子の同定 小胞輸送を標的とする新しいがん治療戦略への可能性」 https://www.u-tokyo.ac.jp/focus/ja/articles/z0508_00119.htmlUTokyo FOCUS Articles "Possible target for future cancer treatment : Deregulation of system to move molecules in the cell may promote tumor growth, metastasis" https://www.u-tokyo.ac.jp/focus/en/articles/z0508_00120.htm

Comprehensive Biostatistical Analysis of CpG Island Methylator Phenotype in Colorectal Cancer Using a Large Population-Based Sample

The CpG island methylator phenotype (CIMP) is a distinct phenotype associated with microsatellite instability (MSI) and BRAF mutation in colon cancer. Recent investigations have selected 5 promoters (CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1) as surrogate markers for CIMP-high. However, no study has comprehensively evaluated an expanded set of methylation markers (including these 5 markers) using a large number of tumors, or deciphered the complex clinical and molecular associations with CIMP-high determined by the validated marker panel. METHOLODOLOGY/PRINCIPAL FINDINGS: DNA methylation at 16 CpG islands [the above 5 plus CDKN2A (p16), CHFR, CRABP1, HIC1, IGFBP3, MGMT, MINT1, MINT31, MLH1, p14 (CDKN2A/ARF) and WRN] was quantified in 904 colorectal cancers by real-time PCR (MethyLight). In unsupervised hierarchical clustering analysis, the 5 markers (CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1), CDKN2A, CRABP1, MINT31, MLH1, p14 and WRN were generally clustered with each other and with MSI and BRAF mutation. KRAS mutation was not clustered with any methylation marker, suggesting its association with a random methylation pattern in CIMP-low tumors. Utilizing the validated CIMP marker panel (including the 5 markers), multivariate logistic regression demonstrated that CIMP-high was independently associated with older age, proximal location, poor differentiation, MSI-high, BRAF mutation, and inversely with LINE-1 hypomethylation and beta-catenin (CTNNB1) activation. Mucinous feature, signet ring cells, and p53-negativity were associated with CIMP-high in only univariate analysis. In stratified analyses, the relations of CIMP-high with poor differentiation, KRAS mutation and LINE-1 hypomethylation significantly differed according to MSI status.Our study provides valuable data for standardization of the use of CIMP-high-specific methylation markers. CIMP-high is independently associated with clinical and key molecular features in colorectal cancer. Our data also suggest that KRAS mutation is related with a random CpG island methylation pattern which may lead to CIMP-low tumors

Photodynamic inactivation of oral bacteria with silver nanoclusters/rose bengal nanocomposite

Antimicrobial photodynamic therapy (a-PDT) is a promising anti-infective technique for generation of singlet oxygen (1O2) to target dental disease. However, conventional organic photosensitizers have problems for clinical use in terms of cytotoxicity, quenching of a-PDT activity by self-dimerization, and the lack of long-term antibacterial effect. We herein propose silver nanoclusters/rose bengal nanocomposite (AgNCs/RB) as a novel photosensitizer with two primary antibacterial effects: (1) 1O2 generation by irradiated RB and (2) Ag+ ion release from AgNCs. AgNCs/RB irradiated with white light-emitting diode (LED) for a short irradiation time of 1 min significantly decreased the bacterial turbidity of Streptococcus mutans, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans (P < 0.05). In SEM, TEM and LIVE/DEAD staining images, photoexcited AgNCs/RB reduced S. mutans colonization, destroyed the cell membrane, and increased the number of dead cells. The antibacterial efficiency of photoexcited AgNCs/RB was greater than that of AgNCs or RB alone (P < 0.05), suggesting a synergistic effect of 1O2 and Ag+ ions from photoexcited AgNCs/RB. By contrast, photoexcited AgNCs/RB did not affect WST-8 and LDH activities and morphology of NIH3T3 mammalian cells, indicating low cytotoxicity. Interestingly, the antibacterial activity of AgNCs/RB on S. mutans was maintained even after the cessation of LED irradiation, indicating a long-term antibacterial effect due to released Ag+ ions. The present AgNCs/RB photosensitizers provide effective synergistic antibacterial effects for dental a-PDT via 1O2 and Ag+ ions coupled with low cytotoxicity

Antibacterial and Antibiofilm Photodynamic Activities of Lysozyme-Au Nanoclusters/Rose Bengal Conjugates

Antibacterial photodynamic therapy (aPDT) utilizes reactive oxygen species such as singlet oxygen (1O2) and free radicals via photosensitizers, which are light and light-sensitive agents, to reduce bacterial infections. It has been utilized as a treatment for dental diseases in place of antibiotic therapies. However, aPDT does not always cause the desired therapeutic effect due to the instability of organic photosensitizers and the formation of bacterial biofilms. To promote the antibacterial and antibiofilm effects of aPDT, we have proposed a lysozyme (Lys)-gold nanoclusters (Au NCs)/rose bengal (Lys-Au NCs/RB) conjugate as a novel photosensitizer. This conjugate was found to effectively impede the growth of both gram-positive and gram-negative bacteria when exposed to white light-emitting diode (LED) irradiation. The photoexcited Lys-Au NCs/RB showed significantly higher antibacterial activity than photoexcited Lys-Au NCs or RB alone. The synergistic effect is a result of the combination of Lys (an antibacterial protein) and enhanced 1O2 generation related to resonance energy transfer (RET) in the Au NCs/RB conjugate. Photoexcited Lys-Au NCs/RB increased the effects of aPDT in a dose- and time-dependent manner. Furthermore, the photoexcited Lys-Au NCs/RB successfully decreased Streptococcus mutans biofilm formation. However, in contrast, it did not have a negative effect on the proliferation, adhesion, or spread of mammalian cells, indicating low cytotoxicity. Lys-Au NCs/RB is a novel photosensitizer with low cytotoxicity that is capable of bacterial inactivation and the suppression of biofilm formation, and could help to improve dental treatments in the future

Antimicrobial photodynamic activity and cytocompatibility of Au25(Capt)18 clusters photoexcited by blue LED light irradiation

Antimicrobial photodynamic therapy (aPDT) has beneficial effects in dental treatment. We applied captopril-protected gold (Au25(Capt)18) clusters as a novel photosensitizer for aPDT. Photoexcited Au clusters under light irradiation generated singlet oxygen (1O2). Accordingly, the antimicrobial and cytotoxic effects of Au25(Capt)18 clusters under dental blue light-emitting diode (LED) irradiation were evaluated. 1O2 generation of Au25(Capt)18 clusters under blue LED irradiation (420–460 nm) was detected by a methotrexate (MTX) probe. The antimicrobial effects of photoexcited Au clusters (0, 5, 50, and 500 μg/mL) on oral bacterial cells, such as Streptococcus mutans, Aggregatibacter actinomycetemcomitans, and Porphyromonas gingivalis, were assessed by morphological observations and bacterial growth experiments. Cytotoxicity testing of Au clusters and blue LED irradiation was then performed against NIH3T3 and MC3T3-E1 cells. In addition, the biological performance of Au clusters (500 μg/mL) was compared to an organic dye photosensitizer, methylene blue (MB; 10 and 100 μg/mL). We confirmed the 1O2 generation ability of Au25(Capt)18 clusters through the fluorescence spectra of oxidized MTX. Successful application of photoexcited Au clusters to aPDT was demonstrated by dose-dependent decreases in the turbidity of oral bacterial cells. Morphological observation revealed that application of Au clusters stimulated destruction of bacterial cell walls and inhibited biofilm formation. Aggregation of Au clusters around bacterial cells was fluorescently observed. However, photoexcited Au clusters did not negatively affect the adhesion, spreading, and proliferation of mammalian cells, particularly at lower doses. In addition, application of Au clusters demonstrated significantly better cytocompatibility compared to MB. We found that a combination of Au25(Capt)18 clusters and blue LED irradiation exhibited good antimicrobial effects through 1O2 generation and biosafe characteristics, which is desirable for aPDT in dentistry