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Article: Fibroblast Stromal Support Model for Predicting Human Papillomavirus-Associated Cancer Drug Responses.

James, Claire D / Lewis, Rachel L / Fakunmoju, Alexis L / Witt, Austin J / Youssef, Aya H / Wang, Xu / Rais, Nabiha M / Prabhakar, Apurva Tadimari / Otoa, Raymonde / Bristol, Molly L

bioRxiv : the preprint server for biology

2024

Abstract: Currently, there are no specific antiviral therapeutic approaches targeting Human papillomaviruses (HPVs), which cause around 5% of all human cancers. Specific antiviral reagents are particularly needed for HPV-related oropharyngeal cancers ( ... ...

Abstract	Currently, there are no specific antiviral therapeutic approaches targeting Human papillomaviruses (HPVs), which cause around 5% of all human cancers. Specific antiviral reagents are particularly needed for HPV-related oropharyngeal cancers (HPV
Language	English
Publishing date	2024-04-12
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.04.09.588680
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Article: Human Papillomavirus 16 replication converts SAMHD1 into a homologous recombination factor and promotes its recruitment to replicating viral DNA.

James, Claire D / Youssef, Aya / Prabhakar, Apurva T / Otoa, Raymonde / Witt, Austin / Lewis, Rachel L / Bristol, Molly L / Wang, Xu / Zhang, Kun / Li, Renfeng / Morgan, Iain M

bioRxiv : the preprint server for biology

2023

Abstract: We have demonstrated that SAMHD1 (sterile alpha motif and histidine-aspartic domain HD-containing protein 1) is a restriction factor for the HPV16 life cycle. Here we demonstrate that in HPV negative cervical cancer C33a cells and human foreskin ... ...

Abstract	We have demonstrated that SAMHD1 (sterile alpha motif and histidine-aspartic domain HD-containing protein 1) is a restriction factor for the HPV16 life cycle. Here we demonstrate that in HPV negative cervical cancer C33a cells and human foreskin keratinocytes immortalized by HPV16 (HFK+HPV16), SAMHD1 is recruited to E1-E2 replicating DNA. Homologous recombination (HR) factors are required for HPV16 replication and viral replication promotes phosphorylation of SAMHD1, which converts it from a dNTPase to an HR factor independent from E6/E7 expression. A SAMHD1 phosphor-mimic (SAMHD1 T592D) reduces E1-E2 mediated DNA replication in C33a cells and has enhanced recruitment to the replicating DNA. In HFK+HPV16 cells SAMHD1 T592D is recruited to the viral DNA and attenuates cellular growth, but does not attenuate growth in isogenic HFK cells immortalized by E6/E7 alone. SAMHD1 T592D also attenuates the development of viral replication foci following keratinocyte differentiation. The results indicated that enhanced SAMHD1 phosphorylation could be therapeutically beneficial in cells with HPV16 replicating genomes. Protein phosphatase 2A (PP2A) can dephosphorylate SAMHD1 and PP2A function can be inhibited by endothall. We demonstrate that endothall reduces E1-E2 replication and promotes SAMHD1 recruitment to E1-E2 replicating DNA, mimicking the SAMHD1 T592D phenotypes. Finally, we demonstrate that in head and neck cancer cell lines with HPV16 episomal genomes endothall attenuates their growth and promotes recruitment of SAMHD1 to the viral genome. The results suggest that targeting cellular phosphatases has therapeutic potential for the treatment of HPV infections and cancers. Importance: Human papillomaviruses are causative agents in around 5% of all human cancers. The development of anti-viral therapeutics depends upon an increased understanding of the viral life cycle. Here we demonstrate that HPV16 replication converts SAMHD1 into an HR factor via phosphorylation. This phosphorylation promotes recruitment of SAMHD1 to viral DNA to assist with replication. A SAMHD1 mutant that mimics phosphorylation is hyper-recruited to viral DNA and attenuates viral replication. Expression of this mutant in HPV16 immortalized cells attenuates the growth of these cells, but not cells immortalized by the viral oncogenes E6/E7 alone. Finally, we demonstrate that the phosphatase inhibitor endothall promotes hyper-recruitment of endogenous SAMHD1 to HPV16 replicating DNA and can attenuate the growth of both HPV16 immortalized human foreskin keratinocytes and HPV16 positive head and neck cancer cell lines. We propose that phosphatase inhibitors represent a novel tool for combating HPV infections and disease.
Language	English
Publishing date	2023-11-15
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.11.13.566899
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Article: Mouse and human embryonic stem cell models of hematopoiesis: past, present, and future.

Chen, Dong / Lewis, Rachel L / Kaufman, Dan S

BioTechniques

2003 Volume 35, Issue 6, Page(s) 1253–1261

Abstract: Human embryonic stem (ES) cells provide a unique model and an important resource to analyze early hematopoietic development. Other systems to study mammalian hematopoiesis include mouse ES cells, dissection of timed mouse embryos, or use of human ... ...

Abstract	Human embryonic stem (ES) cells provide a unique model and an important resource to analyze early hematopoietic development. Other systems to study mammalian hematopoiesis include mouse ES cells, dissection of timed mouse embryos, or use of human postnatal hematopoietic tissue typically isolated from bone marrow or umbilical cord blood. All these models have particular strengths and weaknesses. The extensive studies on murine hematopoiesis provide a basis for work on the human developmental system. Since there are likely some important species differences, use of human ES cells now provides an optimal means to evaluate basic cellular and molecular mechanisms that regulate the beginning stages of human blood development, prior to derivation of hematopoietic stem cells (HSCs). Eventually, research on human ES cells may provide an alternative source of HSCs and other blood products for hematopoietic cell transplantation or other cellular therapies.
MeSH term(s)	Animals ; Cell Culture Techniques/methods ; Cell Culture Techniques/trends ; Embryo, Mammalian/cytology ; Hematopoiesis/physiology ; Hematopoietic Stem Cells/classification ; Hematopoietic Stem Cells/cytology ; Hematopoietic Stem Cells/physiology ; Humans ; Models, Animal ; Rats ; Species Specificity ; Stem Cell Transplantation/methods ; Stem Cell Transplantation/trends
Language	English
Publishing date	2003-09-04
Publishing country	England
Document type	Journal Article ; Review
ZDB-ID	48453-2
ISSN	1940-9818 ; 0736-6205
ISSN (online)	1940-9818
ISSN	0736-6205
DOI	10.2144/03356ss05
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Article ; Online: Defined substrates for human embryonic stem cell growth identified from surface arrays.

Derda, Ratmir / Li, Lingyin / Orner, Brendan P / Lewis, Rachel L / Thomson, James A / Kiessling, Laura L

ACS chemical biology

2007 Volume 2, Issue 5, Page(s) 347–355

Abstract: Methods for the rapid identification of defined cell growth conditions are lacking. This deficiency is a major barrier to the investigation and application of human embryonic stem (ES) cells. To address this problem, we developed a method for generating ... ...

Abstract	Methods for the rapid identification of defined cell growth conditions are lacking. This deficiency is a major barrier to the investigation and application of human embryonic stem (ES) cells. To address this problem, we developed a method for generating arrays of self-assembled monolayers (SAMs) in which each element constitutes a defined surface. By screening surface arrays, we identified peptidic surfaces that support ES cell growth and self-renewal. The ability of the active surface array elements to support ES cell growth depends on their composition: both the density of the peptide presented and its sequence are critical. These findings support a role for specific surface-cell interactions. Moreover, the data from the surface arrays are portable. They can be used to design an effective 3D synthetic scaffold that supports the growth of undifferentiated human ES cells. Our results demonstrate that synthetic substrates for promoting and probing human ES cell self-renewal can be discovered through SAM surface arrays.
MeSH term(s)	Alkanes/chemistry ; Cell Adhesion/physiology ; Cell Culture Techniques ; Cell Differentiation/drug effects ; Cell Proliferation/drug effects ; Embryonic Stem Cells/cytology ; Embryonic Stem Cells/physiology ; Gold/chemistry ; Humans ; Ligands ; Peptide Fragments/chemistry ; Peptide Fragments/pharmacology ; Substrate Specificity ; Sulfhydryl Compounds/chemistry ; Surface Properties
Chemical Substances	Alkanes ; Ligands ; Peptide Fragments ; Sulfhydryl Compounds ; Gold (7440-57-5)
Language	English
Publishing date	2007-05-22
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S.
ISSN	1554-8937
ISSN (online)	1554-8937
DOI	10.1021/cb700032u
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: Arrays for the combinatorial exploration of cell adhesion.

Orner, Brendan P / Derda, Ratmir / Lewis, Rachel L / Thomson, James A / Kiessling, Laura L

Journal of the American Chemical Society

2004 Volume 126, Issue 35, Page(s) 10808–10809

Abstract: A new method for the fabrication of arrays of self-assembled monolayers (SAMs) of alkane thiols (ATs) on gold to combinatorially assay surfaces for cell adhesion is reported. A fluorous SAM, which is both cytophobic and solvophobic, was used as the ... ...

Abstract	A new method for the fabrication of arrays of self-assembled monolayers (SAMs) of alkane thiols (ATs) on gold to combinatorially assay surfaces for cell adhesion is reported. A fluorous SAM, which is both cytophobic and solvophobic, was used as the background between the array features. The resulting solvophobic background permits the application of an assembly after conjugation strategy for fabrication. SAMs containing mixtures of ATs and peptide-terminated ATs were generated. Multiple cell types demonstrated differential and specific binding to these surfaces. Additionally, pluripotent human embryonic stem cells proliferated on surfaces generated by this method.
MeSH term(s)	Animals ; Cell Adhesion/physiology ; Cell Line, Tumor ; Combinatorial Chemistry Techniques/methods ; Gold/chemistry ; Humans ; Mice ; Neuroblastoma/pathology ; Oligopeptides/chemistry ; Oligopeptides/metabolism ; Peptides/chemistry ; Peptides/metabolism ; Pluripotent Stem Cells/cytology ; Polyethylene Glycols/chemistry ; Polyethylene Glycols/metabolism ; Sulfhydryl Compounds/chemistry ; Surface Properties ; Swiss 3T3 Cells
Chemical Substances	Oligopeptides ; Peptides ; Sulfhydryl Compounds ; glycyl-tyrosyl-isoleucyl-glycyl-seryl-arginine (110590-63-1) ; Polyethylene Glycols (30IQX730WE) ; Gold (7440-57-5) ; arginyl-glycyl-aspartic acid (78VO7F77PN)
Language	English
Publishing date	2004-09-08
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S. ; Research Support, U.S. Gov't, P.H.S.
ZDB-ID	3155-0
ISSN	1520-5126 ; 0002-7863
ISSN (online)	1520-5126
ISSN	0002-7863
DOI	10.1021/ja0474291
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Article: Functional endothelial cells derived from rhesus monkey embryonic stem cells.

Kaufman, Dan S / Lewis, Rachel L / Hanson, Eric T / Auerbach, Robert / Plendl, Johanna / Thomson, James A

Blood

2003 Volume 103, Issue 4, Page(s) 1325–1332

Abstract: We have used rhesus monkey embryonic stem (ES) cells to study endothelial cell development. Rhesus ES cells (R366.4 cell line) exposed to medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like ... ...

Abstract	We have used rhesus monkey embryonic stem (ES) cells to study endothelial cell development. Rhesus ES cells (R366.4 cell line) exposed to medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF), and epidermal growth factor (EGF) assumed a relatively uniform endothelial cell morphology and could be propagated and expanded with a consistent phenotype and normal karyotype. When placed in Matrigel, these rhesus ES cell-derived endothelial cells (RESDECs) formed capillary-like structures characteristic of endothelial cells. Immunohistochemical and flow cytometric analysis of RESDECs showed that they take up acetylated low-density lipoprotein (LDL), express CD146, von Willebrand factor, and the integrin alpha v beta 3, and bind the lectin ulex europaeus agglutinin-1. These cells also express the VEGF receptor Flk-1 and secrete VEGF. When introduced in a Matrigel plug implanted subcutaneously in mice, RESDECs formed intact vessels and recruited new endothelial cell growth. In vivo function was demonstrated by coinjection of RESDECs with murine tumor cells subcutaneously into immunocompromised adult mice. RESDECs injected alone did not form measurable tumors. Tumor cells grew more rapidly and had increased vascularization when coinjected with the RESDECs. Immunohistochemical staining demonstrated that the RESDECs participated in forming the tumor neovasculature. RESDECs provide a novel means to examine the mechanisms of endothelial cell development, and may open up new therapeutic strategies.
MeSH term(s)	Animals ; Biocompatible Materials ; Cell Differentiation/drug effects ; Cell Differentiation/physiology ; Cell Line ; Collagen ; Drug Combinations ; Endothelium, Vascular/cytology ; Epidermal Growth Factor/pharmacology ; Fibroblast Growth Factor 2/pharmacology ; Immunophenotyping ; Insulin-Like Growth Factor I/pharmacology ; Laminin ; Macaca mulatta ; Mice ; Neovascularization, Physiologic/physiology ; Proteoglycans ; Stem Cells/cytology ; Umbilical Veins/cytology ; Vascular Endothelial Growth Factor A/pharmacology
Chemical Substances	Biocompatible Materials ; Drug Combinations ; Laminin ; Proteoglycans ; Vascular Endothelial Growth Factor A ; Fibroblast Growth Factor 2 (103107-01-3) ; matrigel (119978-18-6) ; Epidermal Growth Factor (62229-50-9) ; Insulin-Like Growth Factor I (67763-96-6) ; Collagen (9007-34-5)
Language	English
Publishing date	2003-10-16
Publishing country	United States
Document type	Journal Article
ZDB-ID	80069-7
ISSN	1528-0020 ; 0006-4971
ISSN (online)	1528-0020
ISSN	0006-4971
DOI	10.1182/blood-2003-03-0799
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Article: A microarray analysis of the emergence of embryonic definitive hematopoiesis.

Chen, Dong / Wang, Ping / Lewis, Rachel L / Daigh, Christine A / Ho, Coral / Chen, Xin / Thomson, James A / Kendziorski, Christina

Experimental hematology

2007 Volume 35, Issue 9, Page(s) 1344–1357

Abstract: Objective: Human embryonic stem (ES) cells provide a unique model for studying the development and function of human tissues and have proven utility in a number of areas. However, results from ES cell-based studies have been limited by the paucity of ... ...

Abstract	Objective: Human embryonic stem (ES) cells provide a unique model for studying the development and function of human tissues and have proven utility in a number of areas. However, results from ES cell-based studies have been limited by the paucity of information available about early human hematopoietic development. Methods: To better understand early development of the hematopoietic lineage, we use microarray analysis to examine the temporal patterns of gene expression in embryoid bodies derived from human ES cells, focusing around the time of the emergence of definitive hematopoiesis. We use an empirical Bayes hierarchical modeling approach, called EBarrays, to classify genes into each of the possible temporal patterns of gene expression for five different time points, and correlate those patterns with the emergence of hematopoiesis. Results: We find a distinct group of genes previously identified as important in adult hematopoietic self-renewal (such as PIK3R1, ABCB1/MDR-1, RGS18, IRS1, SENP6/SUMO-1, and Wnt5A, etc.) temporally correlates with the emergence of the definitive hematopoiesis. Microarray-based results are further supported via flow cytometry and reverse transcription-polymerase chain reaction studies. Conclusion: The novel genes demonstrating the same expression pattern as this group could further facilitate the understanding of the molecular mechanisms of embryonic hematopoiesis.
MeSH term(s)	Antigens, CD34/metabolism ; Base Sequence ; Cell Differentiation/genetics ; Cell Line ; Cell Lineage ; Cells, Cultured ; Embryonic Stem Cells/chemistry ; Embryonic Stem Cells/physiology ; Gene Expression Regulation, Developmental ; Hematopoiesis/genetics ; Humans ; Microarray Analysis ; Molecular Sequence Data ; Up-Regulation
Chemical Substances	Antigens, CD34
Language	English
Publishing date	2007-09
Publishing country	Netherlands
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	185107-x
ISSN	1873-2399 ; 0301-472X ; 0531-5573
ISSN (online)	1873-2399
ISSN	0301-472X ; 0531-5573
DOI	10.1016/j.exphem.2007.06.004
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Article: Human embryonic stem cell-derived hematopoietic cells are capable of engrafting primary as well as secondary fetal sheep recipients.

Narayan, A Daisy / Chase, Jessica L / Lewis, Rachel L / Tian, Xinghui / Kaufman, Dan S / Thomson, James A / Zanjani, Esmail D

Blood

2005 Volume 107, Issue 5, Page(s) 2180–2183

Abstract: The human/sheep xenograft model has proven valuable in assessing the in vivo hematopoietic activity of stem cells from a variety of fetal and postnatal human sources. CD34+/lineage- or CD34+/CD38- cells isolated from human embryonic stem cells (hESCs) ... ...

Abstract	The human/sheep xenograft model has proven valuable in assessing the in vivo hematopoietic activity of stem cells from a variety of fetal and postnatal human sources. CD34+/lineage- or CD34+/CD38- cells isolated from human embryonic stem cells (hESCs) differentiated on S17 feeder layer were transplanted by intraperitoneal injections into fetal sheep. Chimerism in primary transplants was established with polymerase chain reaction (PCR) and flow cytometry of bone marrow and peripheral blood samples. Whole bone marrow cells harvested from a primary recipient were transplanted into a secondary recipient. Chimerism was established as described before. This animal was stimulated with human GM-CSF, and an increase in human hematopoietic activity was noted by flow cytometry. Bone marrow aspirations cultured in methylcellulose generated colonies identified by PCR to be of human origin. We therefore conclude that hESCs are capable of generating hematopoietic cells that engraft primary recipients. These cells also fulfill the criteria for long-term engrafting hematopoietic stem cells as demonstrated by engraftment and differentiation in the secondary recipient.
MeSH term(s)	ADP-ribosyl Cyclase 1/metabolism ; Animals ; Antigens, CD34/metabolism ; Bone Marrow Cells/cytology ; Bone Marrow Cells/physiology ; Colony-Forming Units Assay/methods ; Embryo, Mammalian/cytology ; Embryo, Mammalian/physiology ; Fetus/physiology ; Flow Cytometry/methods ; Graft Survival/physiology ; Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells/cytology ; Hematopoietic Stem Cells/physiology ; Humans ; Polymerase Chain Reaction/methods ; Sheep/physiology ; Transplantation Chimera/physiology ; Transplantation, Heterologous
Chemical Substances	Antigens, CD34 ; ADP-ribosyl Cyclase 1 (EC 3.2.2.6)
Language	English
Publishing date	2005-11-08
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	80069-7
ISSN	1528-0020 ; 0006-4971
ISSN (online)	1528-0020
ISSN	0006-4971
DOI	10.1182/blood-2005-05-1922
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