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Sequence analysis of SARS-CoV-2 genome reveals features important for vaccine design [article]

Jacob Kames, David Dillon Holcomb, Ofer Kimchi, Michael DiCuccio, Nobuko Hamasaki-Katagiri, Tony Wang, Anton A Komar, Aikaterini Alexaki, Chava Kimchi-Sarfaty
2020 biorxiv/medrxiv  
As the SARS-CoV-2 pandemic is rapidly progressing, the need for the development of an effective vaccine is critical. A promising approach for vaccine development is to generate, through codon pair deoptimization, an attenuated virus. This approach carries the advantage that it only requires limited knowledge specific to the virus in question, other than its genome sequence. Therefore, it is well suited for emerging viruses for which we may not have extensive data. We performed comprehensive in
more » ... ilico analyses of several features of SARS-CoV-2 genomic sequence (e.g., codon usage, codon pair usage, dinucleotide/junction dinucleotide usage, RNA structure around the frameshift region) in comparison with other members of the coronaviridae family of viruses, the overall human genome, and the transcriptome of specific human tissues such as lung, which are primarily targeted by the virus. Our analysis identified the spike (S) and nucleocapsid (N) proteins as promising targets for deoptimization and suggests a roadmap for SARS-CoV-2 vaccine development, which can be generalizable to other viruses.
doi:10.1101/2020.03.30.016832 pmid:32511300 pmcid:PMC7217226 fatcat:7p2bkw3dqzcyll5x3pf22r7vpe

Recent advances in (therapeutic protein) drug development

H.A. Daniel Lagassé, Aikaterini Alexaki, Vijaya L. Simhadri, Nobuko H. Katagiri, Wojciech Jankowski, Zuben E. Sauna, Chava Kimchi-Sarfaty
2017 F1000Research  
doi:10.12688/f1000research.9970.1 pmid:28232867 pmcid:PMC5302153 fatcat:tlv7im4e6fh55nykdczm3zktk4

Pseudovirions as Vehicles for the Delivery of siRNA

Paul E. Lund, Ryan C. Hunt, Michael M. Gottesman, Chava Kimchi-Sarfaty
2009 Pharmaceutical Research  
Over the last two decades, small interfering RNA (siRNA)-mediated gene silencing has quickly become one of the most powerful techniques used to study gene function in vitro and a promising area for new therapeutics. Delivery remains a significant impediment to realizing the therapeutic potential of siRNA, a problem that is also tied to immunogenicity and toxicity. Numerous delivery vehicles have been developed, including some that can be categorized as pseudovirions: these are vectors that are
more » ... irectly derived from viruses but whose viral coding sequences have been eliminated, preventing their classification as viral vectors. Characteristics of the pseudovirions discussed in this review, namely phagemids, HSV amplicons, SV40 in vitro-packaged vectors, influenza virosomes, and HVJ-Envelope vectors, make them attractive for the delivery of siRNA-based therapeutics. Pseudovirions were shown to deliver siRNA effector molecules and bring about RNA interference (RNAi) in various cell types in vitro, and in vivo using immune-deficient and immune-competent mouse models. Levels of silencing were not always determined directly, but the duration of siRNA-induced knockdown lasted at least 3 days. We present examples of the use of pseudovirions for the delivery of synthetic siRNA as well as the delivery and expression of DNA-directed siRNA.
doi:10.1007/s11095-009-0012-2 pmid:19998056 pmcid:PMC2831147 fatcat:dh6vtxmmxrbaxbxrywox6posoa

Sequence analysis of SARS-CoV-2 genome reveals features important for vaccine design

Jacob Kames, David D. Holcomb, Ofer Kimchi, Michael DiCuccio, Nobuko Hamasaki-Katagiri, Tony Wang, Anton A. Komar, Aikaterini Alexaki, Chava Kimchi-Sarfaty
2020 Scientific Reports  
As the SARS-CoV-2 pandemic is rapidly progressing, the need for the development of an effective vaccine is critical. A promising approach for vaccine development is to generate, through codon pair deoptimization, an attenuated virus. This approach carries the advantage that it only requires limited knowledge specific to the virus in question, other than its genome sequence. Therefore, it is well suited for emerging viruses, for which we may not have extensive data. We performed comprehensive in
more » ... silico analyses of several features of SARS-CoV-2 genomic sequence (e.g., codon usage, codon pair usage, dinucleotide/junction dinucleotide usage, RNA structure around the frameshift region) in comparison with other members of the coronaviridae family of viruses, the overall human genome, and the transcriptome of specific human tissues such as lung, which are primarily targeted by the virus. Our analysis identified the spike (S) and nucleocapsid (N) proteins as promising targets for deoptimization and suggests a roadmap for SARS-CoV-2 vaccine development, which can be generalizable to other viruses.
doi:10.1038/s41598-020-72533-2 pmid:32973171 fatcat:uxapxvonyje6hp27umcpr6zfgi

Ethnicity-related polymorphisms and haplotypes in the human ABCB1 gene

Chava Kimchi-Sarfaty, Andrew H Marple, Shiri Shinar, Avraham M Kimchi, David Scavo, M Isabella Roma, In-Wha Kim, Adam Jones, Mili Arora, John Gribar, David Gurwitz, Michael M Gottesman
2007 Pharmacogenomics (London)  
Introduction-The human multi-drug resistance gene (MDR1, ABCB1) codes for P-glycoprotein (P-gp), an important membrane-bound efflux transporter known to confer anti-cancer drug resistance as well as affect the pharmacokinetics of many drugs and xenobiotics. A number of single nucleotide polymorphisms (SNPs) have been identified throughout the ABCB1 gene which may have an effect on P-gp expression levels and function. Haplotype as well as genotype analysis of SNPs is becoming increasingly
more » ... nt in identifying genetic variants underlying susceptibility to human disease. Three SNPs, 1236C>T, 2677G>T, and 3435C>T have been repeatedly shown to predict changes in the function of P-gp. The frequencies with which these polymorphisms exist in a population have also been shown to be ethnically related. Methods-In this study, 95 individuals representative of the entire ethnic make-up of the United States were compared to 101 individuals from an Ashkenazi Jewish population. These individuals were analyzed by genomic sequencing and PCR-RFLP to calculate their genotype frequencies. Results-Twenty-five SNPs were located in the exons of the ABCB1 gene. All of the polymorphisms identified were in parts of the ABCB1 gene product predicted to be intracellular, and 16 appear to be novel as compared to those listed by NCBI. Frequencies of the 1236C>T and 2677G>T/A/C SNPs were similar for the American and Ashkenazi populations (64.2% and 60.4% respectively for 1236C>T -χ 2 is 0.30 p≤1; 55.8% and 64.4% for 2677G>T/A/C χ 2 is 1.49 p≤1), but were different for 3435C>T (24.2% for the American population and 69.3% for the Ashkenazi population χ 2 is 39.927 p<0.001). The 1236T/2677T/3435T haplotype occurred in 23.6% (SE 0.013) of the Ashkenazi population. Conclusion- The SNP at location 3435C>T plays a significant role in the ABCB1 gene. The haplotype and genotype analysis from these data may be used as a basis for studies on the relationship between ABCB1 genotypes and drug efficacy, drug toxicity, disease susceptibility or other phenotypes.
doi:10.2217/14622416.8.1.29 pmid:17187507 pmcid:PMC1876748 fatcat:g4upab5c2vbdhi4xkspr3bkojy

A new and updated resource for codon usage tables

John Athey, Aikaterini Alexaki, Ekaterina Osipova, Alexandre Rostovtsev, Luis V. Santana-Quintero, Upendra Katneni, Vahan Simonyan, Chava Kimchi-Sarfaty
2017 BMC Bioinformatics  
Acknowledgements We thank Nobuko Katagiri and Ryan Hunt from DPPT/OTAT/CBER/FDA, and Paul Kitts and Avi Kimchi from NIH/NLM/NCBI, for fruitful discussions.  ... 
doi:10.1186/s12859-017-1793-7 pmid:28865429 pmcid:PMC5581930 fatcat:wlveqvoqr5byhfloqqxzorqyx4

Efficient Delivery of RNA Interference Effectors via in vitro-Packaged SV40 Pseudovirions

Chava Kimchi-Sarfaty, Scott Brittain, Susan Garfield, Natasha J. Caplen, Qingquan Tang, Michael M. Gottesman
2005 Human Gene Therapy  
; Kimchi-Sarfaty et al., 2003; Kimchi-Sarfaty et al., 2004a; Kimchi-Sarfaty and Gottesman, 2004) .  ...  Previous experiments (Kimchi-Sarfaty et al., 2002; Kimchi-Sarfaty et al., 2003) showed that .45 cells could be transduced with a maximum volume of one reaction as described in Materials and Methods (  ... 
doi:10.1089/hum.2005.16.1110 pmid:16149909 pmcid:PMC1618762 fatcat:oc7odk3mffasvow7fv3vbcfnge

Building better drugs: developing and regulating engineered therapeutic proteins

Chava Kimchi-Sarfaty, Tal Schiller, Nobuko Hamasaki-Katagiri, Mansoor A. Khan, Chen Yanover, Zuben E. Sauna
2013 TIPS - Trends in Pharmacological Sciences  
We thank Mr Ofer Kimchi, Mr Andrew Wu, and Ms Yini Qi for editorial assistance.  ... 
doi:10.1016/j.tips.2013.08.005 pmid:24060103 fatcat:af42iow4yvdi7ac6tqo2xxqupy

Sensitive measurement of single-nucleotide polymorphism-induced changes of RNA conformation: application to disease studies

Raheleh Salari, Chava Kimchi-Sarfaty, Michael M. Gottesman, Teresa M. Przytycka
2012 Nucleic Acids Research  
Single-nucleotide polymorphisms (SNPs) are often linked to critical phenotypes such as diseases or responses to vaccines, medications and environmental factors. However, the specific molecular mechanisms by which a causal SNP acts is usually not obvious. Changes in RNA secondary structure emerge as a possible explanation necessitating the development of methods to measure the impact of single-nucleotide variation on RNA structure. Despite the recognition of the importance of considering the
more » ... ges in Boltzmann ensemble of RNA conformers in this context, a formal method to perform directly such comparison was lacking. Here, we solved this problem and designed an efficient method to compute the relative entropy between the Boltzmann ensembles of the native and a mutant structure. On the basis of this theoretical progress, we developed a software tool, remuRNA, and investigated examples of its application. Comparing the impact of common SNPs naturally occurring in populations with the impact of random point mutations, we found that structural changes introduced by common SNPs are smaller than those introduced by random point mutations. This suggests a natural selection against mutations that significantly change RNA structure and demonstrates, surprisingly, that randomly inserted point mutations provide inadequate estimation of random mutations effects. Subsequently, we applied remuRNA to determine which of the disease-associated non-coding SNPs are potentially related to RNA structural changes.
doi:10.1093/nar/gks1009 pmid:23125360 pmcid:PMC3592397 fatcat:tzuucmawjfbbfktbv2f62q7qgm

Splicing dysregulation contributes to the pathogenicity of several F9 exonic point variants

Upendra K. Katneni, Aaron Liss, David Holcomb, Nobuko H. Katagiri, Ryan Hunt, Haim Bar, Amra Ismail, Anton A. Komar, Chava Kimchi-Sarfaty
2019 Molecular Genetics & Genomic Medicine  
Komar, and Chava Kimchi-Sarfaty This article is available at EngagedScholarship@CSU: https://engagedscholarship.csuohio.edu/scibges_facpub/97 machinery primarily consists of five small nuclear ribonucleoproteins  ...  Synonymous variants do not alter the underlying aa sequence and are considered to primarily cause disease by altering splicing of pre-mRNA transcript (Hunt, Simhadri, Iandoli, Sauna, & Kimchi-Sarfaty,  ...  ORCID Chava Kimchi-Sarfaty https://orcid.org/0000-0002-9355-8585 SUPPORTING INFORMATION Additional supporting information may be found online in the Supporting Information section at the end of the  ... 
doi:10.1002/mgg3.840 pmid:31257730 pmcid:PMC6687662 fatcat:3mfm474sufg5nchqoios3voh4a

Detection of intracellular ADAMTS13, a secreted zinc-metalloprotease, via flow cytometry

S. Geetha, Courtni E. Allen, Ryan C. Hunt, Elizabeth Plum, Susan Garfield, Scott L. Friedman, Kenji Soejima, Zuben E. Sauna, Chava Kimchi-Sarfaty
2009 Cytometry Part A  
Background-ADAMTS13 is a secreted metalloprotease that cleaves von Willebrand Factor multimers and maintains proper homeostasis. A severe deficiency in ADAMTS13 triggers a disorder known as thrombotic thrombocytopenic purpura (TTP). At present, ADAMTS13 expression levels are determined by immunoblotting. Methods-We established a flow cytometry methodology to detect intracellular ADAMTS13 in liver and kidney cells using a polyclonal antibody, BL154G, and several monoclonal antibodies previously
more » ... sed to detect ADAMTS13 by immunoblotting. Results were validated using confocal microscopy, immunoblotting and an activity assay (FRETS-VWF73). Results-We show that labeling ADAMTS13 with specific antibodies and detection by flow cytometry yields results that are comparable to previously established methods for ADAMTS13 detection. Specifically, we compared the endogenous expression levels of ADAMTS13 in various liver cell lines using flow cytometry and obtained results that parallel immunoblot analysis. Knock-down of ADAMTS13 expression via targeted siRNA resulted in significantly reduced median signal, displaying the sensitivity of this detection method. A further analysis of reliability and specificity was achieved through plasmid DNA and transfection reagent dose response studies. Conclusions-The flow cytometry method described here is useful in determining the expression of both endogenous and recombinant forms of intracellular ADAMTS13. Flow cytometry is a convenient, efficient and cost effective way to measure the expression levels of ADAMTS13.
doi:10.1002/cyto.a.20748 pmid:19526483 pmcid:PMC2778596 fatcat:t6xdd4r7ijdvhno7fycuf2kesi

Synonymous Mutations and Ribosome Stalling Can Lead to Altered Folding Pathways and Distinct Minima

Chung-Jung Tsai, Zuben E. Sauna, Chava Kimchi-Sarfaty, Suresh V. Ambudkar, Michael M. Gottesman, Ruth Nussinov
2008 Journal of Molecular Biology  
Which of the two states is the "native" and which is the "mutant" is unclear: the haplotype (1236C>T/2677T>A/3435C>T) described in the study by Kimchi-Sarfaty et al 1 is fully functional as a drug transporter  ... 
doi:10.1016/j.jmb.2008.08.012 pmid:18722384 pmcid:PMC2628389 fatcat:olhidegojraoxptqmog4adquby

An Optimized Purification Design for Extracting Active ADAMTS13 from Conditioned Media

Katarzyna I. Jankowska, Upendra Katneni, Brian C. Lin, Randilu Amarasinghe, Je-Nie Phue, Wells W. Wu, Nobuko Hamasaki-Katagiri, Wojciech Jankowski, Rong-Fong Shen, Chava Kimchi-Sarfaty
2022 Processes  
ADAMTS13 is a hemostatic enzyme that breaks down pro-thrombotic ultra-large multimers of von Willebrand factor (VWF). The deficiency of ADAMTS13 increases VWF-mediated thrombogenic potential and may lead to thrombotic thrombocytopenic purpura (TTP). Recently, clinical studies have shown the development of acquired TTP after COVID-19 infection and a correlation between low ADAMTS13 plasma levels and increased mortality. As a result, investigating ADAMTS13 as a potential recombinant therapeutic
more » ... of broad interest in the field of hematology. ADAMTS13 is considered challenging to purify in its biologically active state. Current purification methods utilize immobilized metal ions, which can interfere with ADAMTS13 metalloprotease activity. For this reason, we optimized an alternative strategy to isolate milligram quantities of highly active recombinant ADAMTS13 (rADAMTS13) from conditioned media after exogenous expression in human cell line, HEK293. HEK293 cells stably expressing C-terminal V5-His-tagged ADAMTS13 were grown in two parallel systems, culture bottles and flasks, for identifying an optimal cultivation strategy. Subsequently, we employed anion exchange followed by anti-V5-tag affinity chromatography to purify rADAMTS13, and extracted rADAMTS13 of high specific activity while preserving its native post-translational modifications. In addition, this process has been optimized and scaled up to produce active rADAMTS13 at levels sufficient for laboratory-scale structural, enzymatic, and biochemical studies.
doi:10.3390/pr10020322 fatcat:6ewuxc5gljhw5m4vzedxw2bccq

Potential impact on coagulopathy of gene variants of coagulation related proteins that interact with SARS-CoV-2 [article]

David Holcomb, Aikaterini Alexaki, Nancy Hernandez, Kyle Laurie, Jacob Kames, Nobuko Hamasaki-Katagiri, Anton A. Komar, Michael DiCuccio, Chava Kimchi-Sarfaty
2020 biorxiv/medrxiv  
Thrombosis has been one of the complications of the Coronavirus disease of 2019 (COVID-19), often associated with poor prognosis. There is a well-recognized link between coagulation and inflammation, however, the extent of thrombotic events associated with COVID-19 warrants further investigation. Poly(A) Binding Protein Cytoplasmic 4 (PABPC4), Serine/Cysteine Proteinase Inhibitor Clade G Member 1 (SERPING1) and Vitamin K epOxide Reductase Complex subunit 1 (VKORC1), which are all proteins
more » ... to coagulation, have been shown to interact with SARS proteins. We computationally examined the interaction of these with SARS-CoV-2 proteins and, in the case of VKORC1, we describe its binding to ORF7a in detail. We examined the occurrence of variants of each of these proteins across populations and interrogated their potential contribution to COVID-19 severity. Potential mechanisms by which some of these variants may contribute to disease are proposed. Some of these variants are prevalent in minority groups that are disproportionally affected by severe COVID-19. Therefore, we are proposing that further investigation around these variants may lead to better understanding of disease pathogenesis in minority groups and more informed therapeutic approaches.
doi:10.1101/2020.09.08.272328 pmid:32935103 pmcid:PMC7491517 fatcat:urjrkwvc3jbzba6ohl5rcpo7xa

Transduction of multiple cell types using improved conditions for gene delivery and expression of SV40 pseudovirions packaged in vitro

Chava Kimchi-Sarfaty, Nathan S. Alexander, Scott Brittain, Saadia Ali, Michael M. Gottesman
2004 BioTechniques  
Kimchi-Sarfaty, Nathan S.  ...  maintained in Iscove's modified essential medium with 55 μM β-mer-Transduction of multiple cell types using improved conditions for gene delivery and expression of SV40 pseudovirions packaged in vitro Chava  ... 
doi:10.2144/04372rr04 pmid:15335219 fatcat:slrksvw7nrazfpsvioc3ciuptm
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