Evaluation of novel CEX resin for continous processing of MAb purification

We have evaluated novel cation exchange resin and membrane which is potentially suited for continuous process in biologics manufacturing. Typically cation exchange chromatography (CEX) is used in a bind/elute (B/E) mode for the MAb process since MAb act as cation at the buffer condition used for CEX due to the pI from neutral to weak basic. Generally, purpose of CEX chromatography is to remove aggregates and other impurities like HCP and DNA. Especially, aggregate removal is of interest to the industries. A continuous process in MAb downstream process can solve several bottlenecks in typical batch process. However its capability cannot be fully utilized if some step of chromatography is used in a B/E mode since it will take a longer process time due to the posing of product stream for binding and washing and it will take a more cost due to the requirement of large amount of resins as we use at the batch process. In such a case, chromatography in flowthrough mode has a potential to overcome those issues for both cost and time and enables us to develop more streamlined continuous process. We will present the result of our study to evaluate novel CEX resin developed by Merck Millipore by comparing with the existing process of Mab A and we obtained a conclusion that this resin is fitted to the continuous processing very much. The novel resin showed a better impurity clearance than our existing process. For example, over 65% removal of aggregate by the novel resin was obtained in contrast to no removal by the existing process. A 20-fold better clearance for DNA was confirmed for the novel resin than the existing process. This indicates an additional polishing step can be omitted and this new chromatography can be a strong option if we need to reduce such impurities further. Also the resin cost is expected to be reduced down to 1/10 in maximum since those impurity clearance results were obtained about 10-times larger load than the typical B/E mode operation of CEX. Considering those aspects, we conclude that this resin showed a better fit for a continuous process. We will also discuss an expected effect of the novel CEX resin on cost and process time savings by a continuous proces

Modeling SARS-CoV-2 infection and its individual differences with ACE2-expressing human iPS cells

ACE発現ヒトiPS細胞を用いたSARS-CoV-2感染の個人差再現と原因究明. 京都大学プレスリリース. 2021-04-19.Stem cells show gender differences in COVID-19 risk. 京都大学プレスリリース. 2021-04-19.Genetic differences are a primary reason for differences in the susceptibility and severity of COVID-19. As induced pluripotent stem (iPS) cells maintain the genetic information of the donor, they can be used to model individual differences in SARS-CoV-2 infection in vitro. We found that human iPS cells expressing the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) (ACE2-iPS cells) can be infected w SARS-CoV-2. In infected ACE2-iPS cells, the expression of SARS-CoV-2 nucleocapsid protein, budding of viral particles, and production of progeny virus, double membrane spherules, and double-membrane vesicles were confirmed. We performed SARS-CoV-2 infection experiments on ACE2-iPS/ embryonic stem (ES) cells from eight individuals. Male iPS/ES cells were more capable of producing the virus compared with female iPS/ES cells. These findings suggest that ACE2-iPS cells can not only reproduce individual differences in SARS-CoV-2 infection in vitro but also are a useful resource to clarify the causes of individual differences in COVID-19 due to genetic differences

Activities of the EMEA project during 1999-2005

金沢大学大学院自然科学研究科The EMEA Project, Kanazawa University2005 International Symposium on Environmental Mornitoring in East Asia -Remote Sensing and Forests-,Hosted The EMEA Project, Kanazawa University 21st=Century COE Program -Environmental Monitoring and Predicition of Long- and Short- Term Dynamics of Pan-Japan Sea Area- ,予稿集, EMEA 2005 in Kanazawa, 国際学術研究公開シンポジウム『東アジアの環境モニタリング』－リモートセンシングと森林－,年月日：200511月28日～29日, 場所：KKRホテル金沢, 金沢大学自然科学研究科, 主催：金沢大学EMEAプロジェクト, 共催：金沢大学21世紀COEプログラム「環日本海域の環境変動と長期・短期変動予測

6.EMEA International Symposium in Kanazawa, Japan

The EMEA Project, Kanazawa UniversityProject Number 14404021, Peport of Research Project ; Grant-in-Aid for Scientific Research(B)(2), from April 2002 to March 2006, Edited by Muramoto,Ken-ichiroKamata, NaotoKawanishi, TakuyaKubo, MamoruLiu, JiyuanLee, Kyu-Sung , 人工衛星データ活用のための東アジアの植生調査、課題番号14404021, 平成14年度～平成17年度科学研究費補助金, 基盤研究（B)(2)研究成果報告書, 研究代表者：村本, 健一郎, 金沢大学自然科学研究科教

Characterization of the quinol-dependent nitric oxide reductase from the pathogen Neisseria meningitidis, an electrogenic enzyme

Abstract Bacterial nitric oxide reductases (NORs) catalyse the reduction of NO to N2O and H2O. NORs are found either in denitrification chains, or in pathogens where their primary role is detoxification of NO produced by the immune defense of the host. Although NORs belong to the heme-copper oxidase superfamily, comprising proton-pumping O2-reducing enzymes, the best studied NORs, cNORs (cytochrome c-dependent), are non-electrogenic. Here, we focus on another type of NOR, qNOR (quinol-dependent). Recombinant qNOR from Neisseria meningitidis, a human pathogen, purified from Escherichia coli, showed high catalytic activity and spectroscopic properties largely similar to cNORs. However, in contrast to cNOR, liposome-reconstituted qNOR showed respiratory control ratios above two, indicating that NO reduction by qNOR was electrogenic. Further, we determined a 4.5 Å crystal structure of the N. meningitidis qNOR, allowing exploration of a potential proton transfer pathway from the cytoplasm by mutagenesis. Most mutations had little effect on the activity, however the E-498 variants were largely inactive, while the corresponding substitution in cNOR was previously shown not to induce significant effects. We thus suggest that, contrary to cNOR, the N. meningitidis qNOR uses cytoplasmic protons for NO reduction. Our results allow possible routes for protons to be discussed

Vegatation Measured at Three Different Scales at Kanazawa University, Kakuma Campus

Proceedings of EMEA 2000 in Kanazaw

A Proteomic Approach for the Diagnosis of ‘Oketsu’ (blood stasis), a Pathophysiologic Concept of Japanese Traditional (Kampo) Medicine

‘Oketsu’ is a pathophysiologic concept in Japanese traditional (Kampo) medicine, primarily denoting blood stasis/stagnant syndrome. Here we have explored plasma protein biomarkers and/or diagnostic algorithms for ‘Oketsu’. Sixteen rheumatoid arthritis (RA) patients were treated with keishibukuryogan (KBG), a representative Kampo medicine for improving ‘Oketsu’. Plasma samples were diagnosed as either having an ‘Oketsu’ (n = 19) or ‘non-Oketsu’ (n = 29) state according to Terasawa's ‘Oketsu’ scoring system. Protein profiles were obtained by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) and hierarchical clustering and decision tree analyses were performed. KBG treatment for 4 or 12 weeks decreased the ‘Oketsu’ scores significantly. SELDI protein profiles gave 266 protein peaks, whose expression was significantly different between the ‘Oketsu’ and ‘non-Oketsu’ states. Hierarchical clustering gave three major clusters (I, II, III). The majority (68.4%) of ‘Oketsu’ samples were clustered into one cluster as the principal component of cluster I. The remaining ‘Oketsu’ profiles constituted a minor component of cluster II and were all derived from patients cured of the ‘Oketsu’ state at 12 weeks. Construction of the decision tree addressed the possibility of developing a diagnostic algorithm for ‘Oketsu’. A reduction in measurement/pre-processing conditions (from 55 to 16) gave a similar outcome in the clustering and decision tree analyses. The present study suggests that the pathophysiologic concept of Kampo medicine ‘Oketsu’ has a physical basis in terms of the profile of blood proteins. It may be possible to establish a set of objective criteria for diagnosing ‘Oketsu’ using a combination of proteomic and bioinformatics-based classification methods

Activities of the EMEA project during 1999-2001

Proceedings of EMEA 2001 in Kanazawa, 25-27 September 2001 in Kanazawa (Japan

Field Survey Trip of EMEA Project in China: From Russian Border to North Korean Border

Proceedings of EMEA 2001 in Kanazawa, 25-27 September 2001 in Kanazawa (Japan