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Article ; Online: Characterization of human induced pluripotent stems cells: Current approaches, challenges, and future solutions

Suresh Babu, Sahana / Duvvuru, Haritha / Baker, Jillian / Switalski, Stephanie / Shafa, Mehdi / Panchalingam, Krishna Morgan / Dadgar, Saedeh / Beller, Justin / Ahmadian Baghbaderani, Behnam

Biotechnology Reports. 2023 Mar., v. 37 p.e00784-

2023

Abstract: Human induced pluripotent stem cells (iPSC) have demonstrated massive potentials for use in regenerative and personalized medicine due to their ability to expand in culture and differentiate into specialized cells with therapeutic benefits. However, in ... ...

Abstract	Human induced pluripotent stem cells (iPSC) have demonstrated massive potentials for use in regenerative and personalized medicine due to their ability to expand in culture and differentiate into specialized cells with therapeutic benefits. However, in order to industrialize iPSC-derived therapies, it is necessary to address the existing challenges surrounding the analytics implemented in the manufacturing process to evaluate and monitor cell expansion, differentiation, and quality of the final products. Here, we review some of the key analytical methods used as part of identity, potency, or safety for in-process or final product release testing and highlighted the challenges and potential solutions for consideration in the Chemistry, Manufacturing and Controls (CMC) strategy for iPSC-based therapies. Some of the challenges associated with characterization and testing of iPSC-based products are related to the choice of analytical technology (to ensure fit-for-purpose), assay reliability and robustness. Automation of analytical methods may be required to reduce hands on time, and improve reliability of the methods through reducing assay variability. Indeed, we have shown that automation of analytical methods is feasible (evaluated using an ELISA based assay) and would result in more precise measurements (demonstrated by lower co-efficient of Variation and standard deviation), less hands-on time, and swift compared to a manually run assay. Therefore, in order to support commercialization of iPSC-based therapies we suggest a well-designed testing strategy to be established in the development phase while incorporating robust, reproducible, reliable, and potentially automated analytics in the manufacturing process.
Keywords	analytical methods ; automation ; biotechnology ; chemistry ; commercialization ; humans ; precision medicine ; standard deviation ; iPSCs ; Cell therapies ; Assay robustness
Language	English
Dates of publication	2023-03
Publishing place	Elsevier B.V.
Document type	Article ; Online
Note	Use and reproduction
ZDB-ID	2801018-8
ISSN	2215-017X
ISSN	2215-017X
DOI	10.1016/j.btre.2023.e00784
Database	NAL-Catalogue (AGRICOLA)

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Article: Characterization of human induced pluripotent stems cells: Current approaches, challenges, and future solutions.

Suresh Babu, Sahana / Duvvuru, Haritha / Baker, Jillian / Switalski, Stephanie / Shafa, Mehdi / Panchalingam, Krishna Morgan / Dadgar, Saedeh / Beller, Justin / Ahmadian Baghbaderani, Behnam

Biotechnology reports (Amsterdam, Netherlands)

2023 Volume 37, Page(s) e00784

Abstract	Human induced pluripotent stem cells (iPSC) have demonstrated massive potentials for use in regenerative and personalized medicine due to their ability to expand in culture and differentiate into specialized cells with therapeutic benefits. However, in order to industrialize iPSC-derived therapies, it is necessary to address the existing challenges surrounding the analytics implemented in the manufacturing process to evaluate and monitor cell expansion, differentiation, and quality of the final products. Here, we review some of the key analytical methods used as part of identity, potency, or safety for in-process or final product release testing and highlighted the challenges and potential solutions for consideration in the Chemistry, Manufacturing and Controls (CMC) strategy for iPSC-based therapies. Some of the challenges associated with characterization and testing of iPSC-based products are related to the choice of analytical technology (to ensure fit-for-purpose), assay reliability and robustness. Automation of analytical methods may be required to reduce hands on time, and improve reliability of the methods through reducing assay variability. Indeed, we have shown that automation of analytical methods is feasible (evaluated using an ELISA based assay) and would result in more precise measurements (demonstrated by lower co-efficient of Variation and standard deviation), less hands-on time, and swift compared to a manually run assay. Therefore, in order to support commercialization of iPSC-based therapies we suggest a well-designed testing strategy to be established in the development phase while incorporating robust, reproducible, reliable, and potentially automated analytics in the manufacturing process.
Language	English
Publishing date	2023-01-24
Publishing country	Netherlands
Document type	Journal Article ; Review
ZDB-ID	2801018-8
ISSN	2215-017X
ISSN	2215-017X
DOI	10.1016/j.btre.2023.e00784
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Low RNA stability signifies strong expression regulatability of tumor suppressors.

Gao, Xinlei / Yi, Yang / Lv, Jie / Li, Yanqiang / Arulsamy, Kulandaisamy / Babu, Sahana Suresh / Bruno, Ivone / Zhang, Lili / Cao, Qi / Chen, Kaifu

Nucleic acids research

2023 Volume 51, Issue 21, Page(s) 11534–11548

Abstract: RNA expression of a gene is determined by not only transcriptional regulation, but also post-transcriptional regulation of RNA decay. The precise regulation of RNA stability in the cell plays an important role in normal development. Dysregulation of RNA ... ...

Abstract	RNA expression of a gene is determined by not only transcriptional regulation, but also post-transcriptional regulation of RNA decay. The precise regulation of RNA stability in the cell plays an important role in normal development. Dysregulation of RNA stability can lead to diseases such as cancer. Here we found tumor suppressor RNAs tended to decay fast in normal cell types when compared with other RNAs. Consistent with a negative effect of m6A modification on RNA stability, we observed preferential deposition of m6A on tumor suppressor RNAs. Moreover, abundant m6A and fast decay of tumor suppressor RNAs both tended to be further enhanced in prostate cancer cells relative to normal prostate epithelial cells. Further, knockdown of m6A methyltransferase METTL3 and reader YTHDF2 in prostate cancer cells both posed stronger effect on tumor suppressor RNAs than on other RNAs. These results indicated a strong post transcriptional expression regulatability mediated by abundant m6A modification on tumor suppressor RNAs.
MeSH term(s)	Humans ; Male ; Methyltransferases/genetics ; Prostatic Neoplasms/chemistry ; Prostatic Neoplasms/genetics ; RNA/genetics ; RNA Stability ; Genes, Tumor Suppressor ; RNA, Messenger/chemistry
Chemical Substances	Methyltransferases (EC 2.1.1.-) ; METTL3 protein, human (EC 2.1.1.62) ; RNA (63231-63-0) ; RNA, Messenger
Language	English
Publishing date	2023-10-13
Publishing country	England
Document type	Journal Article
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad838
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Article ; Online: Low RNA stability signifies increased post-transcriptional regulation of cell identity genes.

Li, Yanqiang / Yi, Yang / Lv, Jie / Gao, Xinlei / Yu, Yang / Babu, Sahana Suresh / Bruno, Ivone / Zhao, Dongyu / Xia, Bo / Peng, Weiqun / Zhu, Jun / Chen, Hong / Zhang, Lili / Cao, Qi / Chen, Kaifu

Nucleic acids research

2023 Volume 51, Issue 12, Page(s) 6020–6038

Abstract: Cell identity genes are distinct from other genes with respect to the epigenetic mechanisms to activate their transcription, e.g. by super-enhancers and broad H3K4me3 domains. However, it remains unclear whether their post-transcriptional regulation is ... ...

Abstract	Cell identity genes are distinct from other genes with respect to the epigenetic mechanisms to activate their transcription, e.g. by super-enhancers and broad H3K4me3 domains. However, it remains unclear whether their post-transcriptional regulation is also unique. We performed a systematic analysis of transcriptome-wide RNA stability in nine cell types and found that unstable transcripts were enriched in cell identity-related pathways while stable transcripts were enriched in housekeeping pathways. Joint analyses of RNA stability and chromatin state revealed significant enrichment of super-enhancers and broad H3K4me3 domains at the gene loci of unstable transcripts. Intriguingly, the RNA m6A methyltransferase, METTL3, preferentially binds to chromatin at super-enhancers, broad H3K4me3 domains and their associated genes. METTL3 binding intensity is positively correlated with RNA m6A methylation and negatively correlated with RNA stability of cell identity genes, probably due to co-transcriptional m6A modifications promoting RNA decay. Nanopore direct RNA-sequencing showed that METTL3 knockdown has a stronger effect on RNA m6A and mRNA stability for cell identity genes. Our data suggest a run-and-brake model, where cell identity genes undergo both frequent transcription and fast RNA decay to achieve precise regulation of RNA expression.
MeSH term(s)	Chromatin/genetics ; Epigenesis, Genetic ; Gene Expression Regulation ; Methyltransferases/metabolism ; RNA/chemistry ; RNA Stability
Chemical Substances	Chromatin ; Methyltransferases (EC 2.1.1.-) ; RNA (63231-63-0)
Language	English
Publishing date	2023-04-25
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, U.S. Gov't, Non-P.H.S. ; Research Support, Non-U.S. Gov't
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad300
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Article ; Online: Enhancer-associated long noncoding RNAs: A novel frontier with new perception in cardiac regeneration.

Jeyabal, Prince / Thandavarayan, Rajarajan A / Babu, Sahana Suresh / Krishnamurthy, Prasanna

Trends in cardiovascular medicine

2015 Volume 25, Issue 7, Page(s) 603–605

MeSH term(s)	Animals ; Genetic Therapy/methods ; Heart Failure/therapy ; Humans ; Myocardium/metabolism ; Myocytes, Cardiac/transplantation ; RNA, Long Noncoding/metabolism ; Regeneration
Chemical Substances	RNA, Long Noncoding
Language	English
Publishing date	2015-10
Publishing country	United States
Document type	Comment ; Editorial ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	1097434-9
ISSN	1873-2615 ; 1050-1738
ISSN (online)	1873-2615
ISSN	1050-1738
DOI	10.1016/j.tcm.2015.02.010
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: The senescence accelerated mouse prone 8 (SAMP8): A novel murine model for cardiac aging.

Karuppagounder, Vengadeshprabhu / Arumugam, Somasundaram / Babu, Sahana Suresh / Palaniyandi, Suresh S / Watanabe, Kenichi / Cooke, John P / Thandavarayan, Rajarajan A

Ageing research reviews

2017 Volume 35, Page(s) 291–296

Abstract: Because cardiovascular disease remains the major cause of mortality and morbidity world-wide, there remains a compelling need for new insights and novel therapeutic avenues. In this regard, the senescence-accelerated mouse prone 8 (SAMP8) line is a ... ...

Abstract	Because cardiovascular disease remains the major cause of mortality and morbidity world-wide, there remains a compelling need for new insights and novel therapeutic avenues. In this regard, the senescence-accelerated mouse prone 8 (SAMP8) line is a particularly good model for studying the effects of aging on cardiovascular health. Accumulating evidence suggests that this model may shed light on age-associated cardiac and vascular dysfunction and disease. These animals manifest evidence of inflammation, oxidative stress and adverse cardiac remodeling that may recapitulate processes involved in human disease. Early alterations in oxidative damage promote endoplasmic reticulum stress to trigger apoptosis and cytokine production in this genetically susceptible mouse strain. Conversely, pharmacological treatments that reduce inflammation and oxidative stress improve cardiac function in these animals. Therefore, the SAMP8 mouse model provides an exciting opportunity to expand our knowledge of aging in cardiovascular disease and the potential identification of novel targets of treatment. Herein, we review the previous studies performed in SAMP8 mice that provide insight into age-related cardiovascular alterations.
MeSH term(s)	Aging, Premature/metabolism ; Aging, Premature/physiopathology ; Animals ; Apoptosis/physiology ; Cardiovascular System/metabolism ; Cardiovascular System/physiopathology ; Cellular Senescence/physiology ; Disease Models, Animal ; Inflammation/physiopathology ; Mice ; Oxidative Stress/physiology
Language	English
Publishing date	2017-05
Publishing country	England
Document type	Journal Article ; Review
ZDB-ID	2075672-0
ISSN	1872-9649 ; 1568-1637
ISSN (online)	1872-9649
ISSN	1568-1637
DOI	10.1016/j.arr.2016.10.006
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Small Engine, Big Power

Darukeshwara Joladarashi / Rajarajan A. Thandavarayan / Sahana Suresh Babu / Prasanna Krishnamurthy

International Journal of Molecular Sciences, Vol 15, Iss 9, Pp 15891-

MicroRNAs as Regulators of Cardiac Diseases and Regeneration

2014 Volume 15911

Abstract: Cardiac diseases are the predominant cause of human mortality in the United States and around the world. MicroRNAs (miRNAs) are small non-coding RNAs that have been shown to modulate a wide range of biological functions under various pathophysiological ... ...

Abstract	Cardiac diseases are the predominant cause of human mortality in the United States and around the world. MicroRNAs (miRNAs) are small non-coding RNAs that have been shown to modulate a wide range of biological functions under various pathophysiological conditions. miRNAs alter target expression by post-transcriptional regulation of gene expression. Numerous studies have implicated specific miRNAs in cardiovascular development, pathology, regeneration and repair. These observations suggest that miRNAs are potential therapeutic targets to prevent or treat cardiovascular diseases. This review focuses on the emerging role of miRNAs in cardiac development, pathogenesis of cardiovascular diseases, cardiac regeneration and stem cell-mediated cardiac repair. We also discuss the novel diagnostic and therapeutic potential of these miRNAs and their targets in patients with cardiac diseases.
Keywords	miRNA ; cardiac development ; cardiovascular diseases ; hypertrophy ; fibrosis ; arrhythmia ; cardiac regeneration ; stem cells ; Biology (General) ; QH301-705.5 ; Chemistry ; QD1-999
Subject code	610
Language	English
Publishing date	2014-09-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: Long-Term Stability and Differentiation Potential of Cryopreserved cGMP-Compliant Human Induced Pluripotent Stem Cells.

Shafa, Mehdi / Walsh, Tylor / Panchalingam, Krishna Morgan / Richardson, Thomas / Menendez, Laura / Tian, Xinghui / Suresh Babu, Sahana / Dadgar, Saedeh / Beller, Justin / Yang, Fan / Baghbaderani, Behnam Ahmadian

International journal of molecular sciences

2019 Volume 21, Issue 1

Abstract: The clinical effectiveness of human induced pluripotent stem cells (iPSCs) is highly dependent on a few key quality characteristics including the generation of high quality cell bank, long-term genomic stability, post-thaw viability, plating efficiency, ... ...

Abstract	The clinical effectiveness of human induced pluripotent stem cells (iPSCs) is highly dependent on a few key quality characteristics including the generation of high quality cell bank, long-term genomic stability, post-thaw viability, plating efficiency, retention of pluripotency, directed differentiation, purity, potency, and sterility. We have already reported the establishment of iPSC master cell banks (MCBs) and working cell banks (WCBs) under current good manufacturing procedure (cGMP)-compliant conditions. In this study, we assessed the cellular and genomic stability of the iPSC lines generated and cryopreserved five years ago under cGMP-compliant conditions. iPSC lines were thawed, characterized, and directly differentiated into cells from three germ layers including cardiomyocytes (CMs), neural stem cells (NSCs), and definitive endoderm (DE). The cells were also expanded in 2D and 3D spinner flasks to evaluate their long-term expansion potential in matrix-dependent and feeder-free culture environment. All three lines successfully thawed and attached to the L7
MeSH term(s)	Cell Differentiation ; Cell Line ; Cryopreservation ; Humans ; Induced Pluripotent Stem Cells/cytology ; Induced Pluripotent Stem Cells/metabolism ; Myocytes, Cardiac/cytology ; Myocytes, Cardiac/metabolism ; Neural Stem Cells/cytology ; Neural Stem Cells/metabolism ; Time Factors
Language	English
Publishing date	2019-12-23
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms21010108
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: RNA-stabilizing proteins as molecular targets in cardiovascular pathologies.

Suresh Babu, Sahana / Joladarashi, Darukeshwara / Jeyabal, Prince / Thandavarayan, Rajarajan A / Krishnamurthy, Prasanna

Trends in cardiovascular medicine

2015 Volume 25, Issue 8, Page(s) 676–683

Abstract: The stability of mRNA has emerged as a key step in the regulation of eukaryotic gene expression and function. RNA stabilizing proteins (RSPs) contain several RNA recognition motifs, and selectively bind to adenylate-uridylate-rich elements in the 3' ... ...

Abstract	The stability of mRNA has emerged as a key step in the regulation of eukaryotic gene expression and function. RNA stabilizing proteins (RSPs) contain several RNA recognition motifs, and selectively bind to adenylate-uridylate-rich elements in the 3' untranslated region of several mRNAs leading to altered processing, stability, and translation. These post-transcriptional gene regulations play a critical role in cellular homeostasis; therefore act as molecular switch between 'normal cell' and 'disease state.' Many mRNA binding proteins have been discovered to date, which either stabilize (HuR/HuA, HuB, HuC, HuD) or destabilize (AUF1, tristetraprolin, KSRP) the target transcripts. Although the function of RSPs has been widely studied in cancer biology, its role in cardiovascular pathologies is only beginning to evolve. The current review provides an overall understanding of the potential role of RSPs, specifically HuR-mediated mRNA stability in myocardial infarction, hypertension and hypertrophy. Also, the effect of RSPs on various cellular processes including inflammation, fibrosis, angiogenesis, cell-death, and proliferation and its relevance to cardiovascular pathophysiological processes is presented. We also discuss the potential clinical implications of RSPs as therapeutic targets in cardiovascular diseases.
MeSH term(s)	Cardiovascular Agents/therapeutic use ; Cardiovascular Diseases/genetics ; Cardiovascular Diseases/physiopathology ; Cell Death/genetics ; Cell Proliferation/genetics ; ELAV-Like Protein 1/drug effects ; ELAV-Like Protein 1/genetics ; Gene Expression Regulation ; Humans ; Molecular Targeted Therapy ; Neovascularization, Pathologic/genetics ; RNA Stability/genetics ; RNA, Messenger/drug effects ; RNA, Messenger/genetics ; Sensitivity and Specificity
Chemical Substances	Cardiovascular Agents ; ELAV-Like Protein 1 ; RNA, Messenger
Language	English
Publishing date	2015-11
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	1097434-9
ISSN	1873-2615 ; 1050-1738
ISSN (online)	1873-2615
ISSN	1050-1738
DOI	10.1016/j.tcm.2015.02.006
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Article ; Online: Small engine, big power: microRNAs as regulators of cardiac diseases and regeneration.

Joladarashi, Darukeshwara / Thandavarayan, Rajarajan Amirthalingam / Babu, Sahana Suresh / Krishnamurthy, Prasanna

International journal of molecular sciences

2014 Volume 15, Issue 9, Page(s) 15891–15911

Abstract	Cardiac diseases are the predominant cause of human mortality in the United States and around the world. MicroRNAs (miRNAs) are small non-coding RNAs that have been shown to modulate a wide range of biological functions under various pathophysiological conditions. miRNAs alter target expression by post-transcriptional regulation of gene expression. Numerous studies have implicated specific miRNAs in cardiovascular development, pathology, regeneration and repair. These observations suggest that miRNAs are potential therapeutic targets to prevent or treat cardiovascular diseases. This review focuses on the emerging role of miRNAs in cardiac development, pathogenesis of cardiovascular diseases, cardiac regeneration and stem cell-mediated cardiac repair. We also discuss the novel diagnostic and therapeutic potential of these miRNAs and their targets in patients with cardiac diseases.
MeSH term(s)	Arrhythmias, Cardiac/genetics ; Arrhythmias, Cardiac/metabolism ; Arrhythmias, Cardiac/pathology ; Cardiomegaly/genetics ; Cardiomegaly/metabolism ; Cardiomegaly/pathology ; Fibrosis/genetics ; Fibrosis/metabolism ; Fibrosis/pathology ; Heart/growth & development ; Heart/physiology ; Heart Diseases/genetics ; Heart Diseases/pathology ; Heart Diseases/therapy ; Humans ; MicroRNAs/metabolism ; Myocardium/metabolism ; Regeneration ; Stem Cells/metabolism
Chemical Substances	MicroRNAs
Language	English
Publishing date	2014-09-09
Publishing country	Switzerland
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms150915891
Database	MEDical Literature Analysis and Retrieval System OnLINE

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