GPCR이 포함된 나노디스크를 통한 약물항체 스크리닝

학위논문(석사) -- 서울대학교대학원 : 공과대학 협동과정 바이오엔지니어링전공, 2022.2. 박태현.G 단백질 연결 수용체 (G protein-coupled receptors, GPCR)를 타겟으로 하는 약물의 개발은 제약 산업에서 중요한 주제이며, 현재까지 대부분의 경우 소분자 합성 의약품들이 개발되어 사용되어지고 있다. 그러나 바이오의약품, 특히 항체의 개발은 큰 진전이 없이 2021년 기준 단 2종만이 미국 FDA에 승인받은 상태이다. 이는 항체 약물개발에 항원인 GPCR을 자연적인 형태로 만들어 개발에 사용하기 어려움이 있고 설사 어렵게 만들었다 하여도 그 방식들이 대량생산을 하는데 어려움을 가졌기 때문이다. 본 연구는, E.coli-발현 시스템을 사용하여 나노디스크 플랫폼으로 재조합한 GPCR을 기반으로 한 키트를 제작하고 항체를 스크리닝 하였다. 이 키트는 간접적 효소면역측정법 (Indirect ELISA) 방식을 기반으로 설계되어, 항원인 GPCR이 포함된 나노디스크가 표면에 붙어있는 상태에서 확인하고자 하는 항체들을 분별한 뒤 특정 파장을 유도하는 표시 항체를 후처리 하는 방식으로 진행하였다. 제작된 키트는 미국 FDA에서 승인받은 항체인 Mogamulizumab (MOG)과 6 종류의 대-케모카인 수용체 항체들, 그리고 타겟이 되는 GPCR인 cc-chemokine receptor type 4를 항원으로 사용하여 그 기능을 확인하였다. 대량생산이 가능한 나노디스크 방식의 재조합 GPCR은 약물개발 방식에 새로운 패러다임을 여는데 기여할 것이다.G protein-coupled receptor (GPCR) targeting drug development has been a major topic in the pharmaceutical industry over decades, while mainly chemical molecules are currently employed in the clinical field due to complexity in development schemes for contemporary biopharmaceutics. In developing a therapeutic antibody against GPCR, antigen preparation and its mass production are the main bottlenecks of the scheme. Here, we design E.coli-expressed reconstituted GPCR using a nanodisc (ND) platform, assemble a kit to screen antibodies and derivatives, and perform its capability to . For antibody screening, indirect ELISA is the methodological basis in which antibody to identify serves as primary antibody, and an enzyme-conjugated secondary antibody serves as an indicator. We validated this kit by identifying a well-known therapeutic antibody, Mogamulizumab (MOG), to the cc-chemokine receptor 4 (CCR4)-embedded ND from a pool of antibodies against the CCR family. This new approach suggests that any type of fully reconstituted GPCR can be mass-produced and used for screening purposes, shifting to the new paradigm in antibody drug discovery.Abstract Table of Contents List of Figures and Table 1. Introduction 1 2. Materials and Methods 4 2.1 Expression and purification of CCR4 4 2.2 Expression and purification of MSP1E3D1 7 2.3 Assembly of CCR4-embedded nanodisc (CCR4-ND) 10 2.4 Antibody screening using CCR4-ND 12 3. Result and Discussion 13 3.1 Production of CCR4 and His-tag truncated MSP1E3D1 13 3.2 Reconstitution of CCR4-ND 18 3.3 Kit validation 21 3.3.1 Selection of target antibody 21 3.3.2 Antibody-to-absorbance relationship 25 3.3.3 Reliability test 27 4. Conclusions 30 5. Reference 31 요약 (국문 초록) 37석

Knowledge and Anxiety Related to Hereditary Ovarian Cancer in Serous Ovarian Cancer Patients

Purpose: The awareness of hereditary breast and ovarian cancer (HBOC) and BRCA testing is in creasing in Korea. Compared to the sizable research on HBOC knowledge among breast cancer women, studies in the ovarian cancer population are limited. This paper aimed to investigate the level of knowledge of hereditary ovarian cancer and anxiety in women diagnosed with serous ovarian cancer in Korea and determine differences in the knowledge and anxiety according to whether genetic testing was undertaken and whether BRCA1 or BRCA2 mutations were present. Methods: Using a descriptive research design, a cross-sectional survey was conducted on 100 women diagnosed with serous ovarian cancer at N hospital in Gyeonggi-do, Korea, from July to November 2018. The collected data were analyzed by descriptive statistics, independent t-tests, one-way analysis of variance, and Pearson's correlation coefficient using the SPSS 21.0 program. Results: The hereditary ovarian cancer-related knowledge score was mid-level (mean score 8.90±3.29 out of a total of 17), as was the state anxiety level was mid-level (mean score 47.96±3.26 out of possible score range of 20–80). Genetic knowledge of hereditary ovarian cancer was asso ciated with age, education, occupation, genetic counseling, and BRCA mutations. There were no statistically significant factors related to anxiety and there were no statistically significant correla tions between knowledge level and anxiety. Conclusion: More comprehensive education on gene-related cancer is needed for ovarian cancer patients, especially for items with low knowledge scores. A genetic counseling protocol should be developed to allow more patients to alleviate their anxiety through genetic counseling.ope

Revisiting The Meaning of A Good Nurse

Background: The health care paradigm is being challenged to put greater emphasis on promoting a healthy lifestyle and provide patient-customized care in a rapidly changing Korean society. Within such changes, it is worthwhile to identify how the meaning of a good nurse may be changing. Objective: This study explores the meaning of a good nurse from the perspective of nurses themselves. Methods: In-depth interviews were conducted with ten nurses who worked at senior general hospitals or governmental agencies using purposeful sampling. Individual interviews were done until data saturation was reached. The key question was “What does a good nurse mean to you?” After the interview, content analysis was done. Results: Six main themes and 16 subthemes were identified. The main themes showed that a good nurse was someone who ‘leans in toward the patient’, ‘provides professional care’, ‘keeps the cornerstone of one’s mind with a positive attitude’, ‘maintains a good relationship with colleagues’, ‘matures with her/his nursing philosophy and conviction’, ‘contributes to society’. Conclusion: This study provides basic data for exploring nurses’ self-image as a professional. Specifically, social role and macro level responsibilities were identified with nurses’ internal aspects such as a positive attitude, and nursing philosophy. The necessity of socio-environmental support for nurses was also emphasized. In addition, the results of this study can be used at the policy level to provide a better balance for development of good nursing.restrictio

Effects of a Coaching-based Childbirth Program on Anxiety and Childbirth Self-efficacy among Primigravida Women

PURPOSE: Childbirth self-efficacy plays an important role in women's ability to cope with labor and delivery. Coaching has been gaining popularity as a way to promote cognitive, emotional and behavioral change. This study aimed to test the effects of a Coaching-based childbirth program on anxiety and childbirth self-efficacy among primigravida women. METHODS: The study design was a quasi-experimental pre-post design. A coaching-based childbirth program was developed synthesizing concepts and techniques from the literature on coaching and was verified by an expert panel. It consisted of four weekly 2-hour small group sessions. Pregnant women were recruited from H hospital in Seoul. Childbirth self-efficacy and state anxiety were measured before and after the program. RESULTS: Although there was a no significant reduction in anxiety, there were statistically significant increase for childbirth self-efficacy in the experimental group when compared to the control group. CONCLUSION: The program appears to increase childbirth self-efficacy for pregnant women. Future studies may benefit from using mixed coaching modalities and consider measuring health behaviors and obstetric outcomes to gain insights on its long-term impact.ope

Effects of a Medication Management Program for Cancer Patients Receiving Oral Chemotherapy

Purpose: The purpose of this study was to assess the effects of a medication management program on oral chemotherapy patients. Methods: A nonequivalent control group ppretest-posttest design was used. Participants were 60 cancer patients (intervention group: 30, control group: 30). The medication management program was provided for 6 weeks. Collected data were analyzed using the SPSS/WIN 21.0 program. Results: Although there were no statistically significant differences, scores of self-efficacy (t=-0.12, p=.902), knowledge (t=0.62, p=.537), medication adherence (t=0.51, p=.610), and staff satisfaction (t=1.44, p=.156) were higher in the experimental group than in the control group. Conclusion: There was no significant difference in self-efficacy, knowledge, symptom experience, medication adherence and staff satisfaction. This can be explained by both groups having already received initial instruction concerning basic care when they started to receive chemotherapy. Considering the positive outcome of the medication management program, a specialist nursing effort is needed to improve symptoms and medication adherence. Furthermore, a medication counseling hotline is needed to support the medical staff.ope

Predicting Openness of Communication in Families With Hereditary Breast and Ovarian Cancer Syndrome: Natural Language Processing Analysis

Background: In health care research, patient-reported opinions are a critical element of personalized medicine and contribute to optimal health care delivery. The importance of integrating natural language processing (NLP) methods to extract patient-reported opinions has been gradually acknowledged over the past years. One form of NLP is sentiment analysis, which extracts and analyses information by detecting feelings (thoughts, emotions, attitudes, etc) behind words. Sentiment analysis has become particularly popular following the rise of digital interactions. However, NLP and sentiment analysis in the context of intrafamilial communication for genetic cancer risk is still unexplored. Due to privacy laws, intrafamilial communication is the main avenue to inform at-risk relatives about the pathogenic variant and the possibility of increased cancer risk. Objective: The study examined the role of sentiment in predicting openness of intrafamilial communication about genetic cancer risk associated with hereditary breast and ovarian cancer (HBOC) syndrome. Methods: We used narratives derived from 53 in-depth interviews with individuals from families that harbor pathogenic variants associated with HBOC: first, to quantify openness of communication about cancer risk, and second, to examine the role of sentiment in predicting openness of communication. The interviews were conducted between 2019 and 2021 in Switzerland and South Korea using the same interview guide. We used NLP to extract and quantify textual features to construct a handcrafted lexicon about interpersonal communication of genetic testing results and cancer risk associated with HBOC. Moreover, we examined the role of sentiment in predicting openness of communication using a stepwise linear regression model. To test model accuracy, we used a split-validation set. We measured the performance of the training and testing model using area under the curve, sensitivity, specificity, and root mean square error. Results: Higher openness of communicationscores were associated with higher overall net sentiment score of the narrative, higher fear, being single, having nonacademic education, and higher informational support within the family. Our results demonstrate that NLP was highly effective in analyzing unstructured texts from individuals of different cultural and linguistic backgrounds and could also reliably predict a measure of openness of communication(area under the curve=0.72) in the context of genetic cancer risk associated with HBOC. Conclusions: Our study showed that NLP can facilitate assessment of openness of communication in individuals carrying a pathogenic variant associated with HBOC. Findings provided promising evidence that various features from narratives such as sentiment and fear are important predictors of interpersonal communication and self-disclosure in this context. Our approach is promising and can be expanded in the field of personalized medicine and technology-mediated communication. ©Vasiliki Baroutsou, Rodrigo Cerqueira Gonzalez Pena, Reka Schweighoffer, Maria Caiata-Zufferey, Sue Kim, Sharlene Hesse-Biber, Florina M Ciorba, Gerhard Lauer, Maria Katapodi, CASCADE Consortium.ope

Effects of an Integrated Self-Management Program on Self-Management, Glycemic Control, and Maternal Identity in Women with Gestational Diabetes Mellitus

Purpose: The purpose of the study was to investigate the effects of an integrated self-management program on self-management, glycemic control, and maternal identity in women with gestational diabetes mellitus (GDM). Methods: A non-equivalent control group non-synchronized quasi-experimental design was used. A total of 55 women with GDM were recruited from Cheil General Hospital, Seoul, Korea and were assigned to an experimental (n=28) or control group (n=27). The participants were 24-30 weeks pregnant women who had been diagnosed with GDM as of July 30, 2010. The program was conducted as a 1 hour small group meeting 3 out of 5 times and by telephone-counseling 2 out of 5 times. The integrated self-management program was verified by an expert panel. Results: Although there was no significant reduction in HbA1c (U= -1.17, p=.238), there were statistically significant increases in self-management (U= -3.80, p<.001) and maternal identity (U= -4.48, p<.001), and decreased 2-h postprandial glucose levels (U= -2.43, p<.015) in the experimental group compared to the control group. Conclusion: These findings suggest that an integrated self-management program for women with GDM improves self-management, maternal identity, and glycemic control. Further studies are needed to identify the effects of an integrated self-management program on pregnancy and neonatal outcomes.ope

Psychometric properties of the Korean version of the Pittsburgh Fatigability Scale in breast cancer survivors

Background: Fatigability has recently emerged in oncology as a concept that anchors patients' perceptions of fatigue to defined activities of specified duration and intensity. This study aimed to examine the psychometric properties of the Korean version of the Pittsburgh Fatigability Scale (K-PFS) for women with breast cancer. Methods: This cross-sectional study involved 196 women with breast cancer recruited from a tertiary hospital in Seoul, Korea. Reliability was evaluated using Cronbach's alpha, and confirmatory factor analysis was conducted to examine the factor structure of the K-PFS. Four goodness-of-fit values were evaluated: (1) the comparative fit index (CFI), (2) the Tucker-Lewis index (TLI), (3) the root mean square error of approximation (RMSEA), and (4) the standardized root mean square residual (SRMR). Results: Of the 196 survivors, 71.1% had greater physical fatigability (K-PFS Physical score ≥ 15) and 52.6% had greater mental fatigability (K-PFS Mental score ≥ 13). The Cronbach's alpha coefficient for the total K-PFS scale was 0.926, and the coefficients for the physical and mental fatigability domains were 0.870 and 0.864, respectively. In the confirmatory factor analysis for physical fatigability, the SRMR value (0.076) supported goodness of fit, but other model fit statistics did not (CFI = 0.888, TLI = 0.826, and RMSEA = 0.224). For mental fatigability, although three goodness-of-fit values were acceptable (CFI = 0.948, TLI = 0.919, and SRMR = 0.057), the RMSEA value (0.149) did not indicate good model fit. However, each item coefficient was statistically significant (> 0.5), and the K-PFS was therefore found to be valid from a theoretical perspective. Conclusion: This study provides meaningful information on the reliability and validity of the K-PFS instrument, which was developed to meet an important need in the context of breast cancer survivors. Additional research should examine its test-retest reliability and construct validity with performance measures.ope

Pro-inflammatory Cytokine Levels and Cancer-related Fatigue in Breast Cancer Survivors: Effects of an Exercise Adherence Program

Purpose: This study aimed to determine the effect of an exercise intervention on subjective cancer-related fatigue (CRF) and pro-inflammatory cytokine levels in breast cancer survivors (BCS). Methods: BCS with greater than moderate CRF (≥ 4) were recruited and randomly assigned to experimental or control groups. The experimental group participated in a 12-week exercise adherence program (Better Life after Cancer - Energy, Strength, and Support; BLESS). Interleukin (IL)-6 and tumor necrosis factor-α (TNF-α) levels were determined at 3 time points (M1: baseline, M2: post-intervention, and M4: 6 months after intervention). Subjective fatigue was measured using the Korean version of the revised Piper Fatigue Scale. Results: In this analysis of participants with physiological fatigue measures available (19 experimental, 21 control), there were no statistically significant differences in IL-6 (F = 1.157, p = 0.341), TNF-α levels (F = 0.878, p = 0.436), and level of fatigue (F = 2.067, p = 0.118) between the 2 groups at baseline. Fatigue in the experimental group showed statistically significant improvement compared to the control only at M2 (p = 0.022). There was no significant relationship between subjective and physiological fatigue at the 3 measurement points. Conclusion: The BLESS intervention improved CRF in BCS immediately at post-intervention, and this study presents clinical feasibility for the management of CRF in BCS in the early survivorship phase who are already experiencing fatigue.ope

Privacy and utility of genetic testing in families with hereditary cancer syndromes living in three countries: the international cascade genetic screening experience

Background: Hereditary breast and ovarian cancer and Lynch syndrome are associated with increased lifetime risk for common cancers. Offering cascade genetic testing to cancer-free relatives of individuals with HBOC or LS is a public health intervention for cancer prevention. Yet, little is known about the utility and value of information gained from cascade testing. This paper discusses ELSI encountered during the implementation of cascade testing in three countries with national healthcare systems: Switzerland, Korea, and Israel. Methods: A workshop presented at the 5th International ELSI Congress discussed implementation of cascade testing in the three countries based on exchange of data and experiences from the international CASCADE cohort. Results: Analyses focused on models of accessing genetic services (clinic-based versus population-based screening), and models of initiating cascade testing (patient-mediated dissemination versus provider-mediated dissemination of testing results to relatives). The legal framework of each country, organization of the healthcare system, and socio-cultural norms determined the utility and value of genetic information gained from cascade testing. Conclusion: The juxtaposition of individual versus public health interests generates significant ELSI controversies associated with cascade testing, which compromise access to genetic services and the utility and value of genetic information, despite national healthcare/universal coverage. Copyright © 2023 Barnoy, Dagan, Kim, Caiata-Zufferey, Katapodi, the CASCADE and the K-CASCADE Consortia.ope