Findings Gladstone Institutes - Scientific Report for the Gladstone Institute of Cardiovascular Disease
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Gladstone Institutes Findings 2018 Scientific Report for the Gladstone Institute of Cardiovascular Disease
Mission Gladstone Institutes uses visionary science and technology to overcome major unsolved diseases. Vision Gladstone Institutes believes that life science research provides the most effective solutions to overcome major unsolved diseases and enables society to address health-related humanitarian issues worldwide. Based in San Francisco’s Mission Bay neighborhood, the Gladstone Institutes is an independent state-of-the-art biomedical research institution that empowers its world-class scientists to find new pathways to cures. It has a close academic affiliation with the University of California, San Francisco. Unified by a common vision, everyone at Gladstone believes that the best discoveries will come from bringing diverse thoughts, approaches, and people together to tackle scientific challenges in creative ways.
Gladstone Institutes
Contents Gladstone Leadership
Andrew S. Garb
Gladstone by the Numbers 2 Trustee
Message from the President 3 William S. Price III
Trustee
Institutes at Gladstone 4 Nicholas J. Simon
Trustee
Scientific Advisory Board 6
Deepak Srivastava, MD
President
Major Research Achievements 6
Scientific Report Staff
Laboratory Reports Megan McDevitt
Vice President of Communications
Thomas P. Bersot, MD, PhD 8
Thomas Becher
Producer for Web
Benoit G. Bruneau, PhD 10
Gary Howard, PhD
Bruce R. Conklin, MD 12 Editor
Julie Langelier
Sheng Ding, PhD 14 Editor
Saptarsi Haldar, MD 16 Giovanni Maki
Art Director
Todd C. McDevitt, PhD 18 Sarah Gardner
Graphic Designer
Deepak Srivastava, MD 20 Teresa Roberts
Design Assistant
Shinya Yamanaka, MD, PhD 22
Contributors
Programs and Initiatives 24 Martyna Ziemba-Martinez
Graphic Designer
Research Infrastructure 25 Diana Rothery
Photographer
Recent Discoveries 26
The reference period for this scientific
report is from January 1, 2016, to
December 31, 2017.
1650 Owens Street
San Francisco, CA 94158
415.734.2000
gladstone.org
@gladstoneinst
/gladstoneinstitutes
© 2018 Gladstone Institutes
Findings 2018
Gladstone by the Numbers
Science
292 24 9
Publications in scientific journals from January 1, 2016, Laboratories Core facilities and
to December 31, 2017. technology center
People Trainee Careers (since 2012)
472 After their Gladstone training, postdoctoral scholars have
moved on to:
199
35%
Scientific Staff 31%
103 Academia
(tenured and
Postdoctoral non-tenured Other
Scholars faculty positions) (government,
non-science
related,
further studies)
45
26 8%
Graduate
Investigators Students 26%
99 Science-related
(including a staff research
investigator and a visiting professions
Administration/ Industry (including policy and
investigator)
Support Staff business development)
Finances
$81M
$35.6M $12.5M $4.7M
Federal Grants Gladstone’s Endowment State Grants
$20.5M $7.7M
Private Support Commercial Revenue
The financial information presented above is unaudited and has been prepared in
accordance with accounting principles generally accepted in the United States.
2
Message from the President At the Gladstone Institutes, we believe that the key to making groundbreaking and paradigm-shifting discoveries lies in the power of interdisciplinary teams. When scientists from different fields come together to tackle a common problem, the combi- nation of their unique perspectives leads to more creative and comprehensive solutions. Ultimately, that’s how we can maximize the impact of biomedical research on improving human health. This successful model has spread throughout our organization. In the past few years, it led to the creation of several new research centers aiming to foster collaborations around broad thematic areas and exceed the potential of any single laboratory. Within the Gladstone Institute of Cardiovascular Disease (GICD), our team science approach has united basic biologists, chemists, computer scientists, and engineers, all working towards a shared goal of unraveling the most fundamental aspects of the cardio- vascular system. We also join forces with investigators through- out Gladstone and the San Francisco Bay Area, as well as the experts in our core facilities and technology centers. This breadth of knowledge and variety of viewpoints provide a rich learning environment for our trainees. A central part of Gladstone’s mission, mentoring is a priority for all our investigators as we recognize the importance of preparing graduate students and postdoctoral scholars for successful scientific careers. At GICD, our research focuses on understanding how the entire cardiovascular network develops and functions, both in health and disease. We explore the cellular and molecular mecha- nisms underlying pluripotency and cardiogenesis, as well as the basic concepts in gene regulation, particularly those disrupted in human disease. We combine stem cell biology, gene editing techniques, and chemical biology approaches, while also engi- neering 3D tissues and organoids, and developing novel tech- nologies. Our goal is to fill existing gaps in cardiac regeneration and genetics and to find better therapeutic strategies for patients with cardiovascular disease. Building on our discoveries, Gladstone launched a new biophar- maceutical company, Tenaya Therapeutics Inc., in 2016. This spin-off company was formed from the BioFulcrum initiative, developed to enhance Gladstone’s translational efforts by merging our basic science expertise with the resources and translational know-how of the biotechnology industry. We are very proud of Tenaya and continue to be closely involved in their work, which strives to create new therapeutics for cardiac regen- erative medicine and drug discovery for heart failure. As the new president of Gladstone, I am honored to represent my scientific colleagues and their outstanding research. I invite you to read this scientific report, which highlights the accomplish- ments of GICD’s eight investigators from 2016 and 2017.
Findings 2018 Institutes at Gladstone Gladstone houses four major institutes, each representing different yet interconnected areas of focus. Interwoven are multiple research centers that bring together common approaches or themes throughout the organization. Driven by their inquisitive nature, investigators have the freedom to follow their research wherever it leads, and work closely with their colleagues in all institutes to deeply probe important questions in biomedicine. Above all, they champion highly interactive, creative, and mold-breaking approaches to science as they seek prevention, treatments, and cures for major diseases. Supported by state-of-the-art core facilities and professionally trained staff, Gladstone scientists rely on the latest technologies to advance their work. And to deliver results to patients, as urgently as possible, they join forces with the Office of Corporate Ventures and Translation to develop fruitful collaborations with the San Francisco Bay Area biomedical industry. Gladstone investigators also actively invest in the future of research. They remain strongly committed to mentoring graduate students and postdoctoral scholars, who will have to overcome tomorrow’s scientific and medical challenges. 4
Gladstone Institutes
Gladstone Institute of Cardiovascular Disease Gladstone Institute of Neurological Disease
Cardiovascular disease remains the world’s leading cause of Diseases that affect the brain or other parts of the central
death, with heart failure alone afflicting over 26 million people nervous system raise fascinating neuroscientific questions
around the globe. Despite decades of work, patients and doctors and are among the most devastating and complex conditions
still need scientific and medical breakthroughs to combat these plaguing humankind. As populations around the world are living
devastating diseases, which include heart attacks, congenital longer, neurodegenerative disorders are rising in prevalence
heart defects, and other disorders. at an unprecedented pace. However, many of these diseases
remain without effective treatment options.
To address this critical situation, the Gladstone Institute of
Cardiovascular Disease (GICD) leverages important genetic, Consequently, the Gladstone Institute of Neurological Disease
developmental, chemical, biological systems, computational, and (GIND) maintains a strong focus on neurodegenerative and
engineering approaches to the study of heart disease and stem neuroinflammatory diseases, including Alzheimer’s disease, fron-
cell biology. totemporal dementia, Parkinson’s disease, Huntington’s disease,
amyotrophic lateral sclerosis, and multiple sclerosis. Given the
Recently, much of their work expands on two paradigm-shifting
overlap between these conditions and other challenging brain
discoveries: induced pluripotent stem cells and CRISPSR-Cas9
gene editing. Gladstone scientists uncovered new and more diseases, they also study epilepsy and neuropsychiatric disorders,
such as autism and depression.
efficient ways to use these technologies to study cardiovascular
disease and transform their research into therapies that help GIND investigators believe that pathogenic convergence points
repair damaged hearts. among these conditions will allow them to identify therapeutic
interventions that might benefit multiple disorders. They are
Gladstone Institute of Virology and Immunology using unconventional approaches to yield such multi-faceted
Since its foundation 25 years ago, the Gladstone Institute of solutions, while also expanding capabilities in regenerative and
Virology and Immunology (GIVI) has made significant contri- personalized medicine.
butions to fighting HIV/AIDS, which ranks among the deadliest
infectious epidemics ever recorded. Its investigators defined Gladstone Institute of Data Science and Biotechnology
the life cycle of HIV, paved the way for many medications In recent years, the deployment of advanced technologies has
currently in use, and led a global and groundbreaking effort in become crucial in driving novel scientific discovery. Researchers
HIV prevention. increasingly depend on creative data analysis and integration
to interrogate biological questions. To respond to this growing
Today, antiretroviral drugs can help prolong lifespan and improve
need, the Gladstone Institute of Data Science and Biotechnology
the quality of life for people with HIV. However, patients require
was launched in 2018.
lifelong treatment of daily medications, because the virus persists
in a latent form. Scientists in GIVI are uniquely positioned to Building on the success of the Convergence Zone, this new insti-
explore the biological basis for HIV latency, which represents the tute’s mission is to decode biomedical knowledge that is missed
greatest barrier to a cure. without rigorous statistical approaches. Their efforts will combine
multiple intellectual and physical assets, as well as new machine
The major successes in HIV research have provided Gladstone
learning and artificial intelligence approaches, that will impact
with an opportunity to shift this institute’s focus to a new field.
numerous disease areas. They will also develop new research
Investigators are working to identify a new area of research
technologies and platforms to propel groundbreaking science
that could be most impacted by capitalizing on the existing
throughout Gladstone and the San Francisco Bay Area.
strengths of GIVI.
5
Findings 2018
Scientific Advisory Board Major Research Achievements
Members of the 2018 Scientific Advisory Board
for the Gladstone Institute of Cardiovascular Disease Over the past 40 years, investigators at
Brian Black, PhD the Gladstone Institute of Cardiovascular
Cardiovascular Research Institute
University of California, San Francisco
Disease made important discoveries
George Q. Daley, MD, PhD that have significantly impacted
Children’s Hospital Boston
Harvard Medical School
the scientific community. These
Howard Hughes Medical Institute achievements became the foundation
Joseph Goldstein, MD for numerous subsequent research
Departments of Molecular Genetics and Internal Medicine
University of Texas Southwestern projects that continue to advance
Medical Center at Dallas
knowledge in the field of cardiovascular
Andrew R. Marks, MD
The Clyde and Helen Wu Center for Molecular Cardiology biology, cellular reprogramming,
Columbia University
and regenerative medicine.
Deborah Nickerson, PhD
Department of Genome Sciences
University of Washington School of Medicine
Eric N. Olson, PhD
Department of Molecular Biology
University of Texas Southwestern
Medical Center at Dallas
Marlene Rabinovitch, MD
Department of Pediatrics, Cardiology
Stanford Cardiovascular Institute
Stanford University School of Medicine
Janet Rossant, PhD
Developmental and Stem Cell Biology Program
The Hospital for Sick Children
University of Toronto
Christine Seidman, PhD
Department of Genetics and Medicine
Harvard Medical School
Irving L. Weissman, MD
Institute for Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Gladstone Institutes
Defined the sequence of molecular events in normal and Discovered genetic causes and underlying mechanisms
abnormal heart development and homeostasis of heart disease
For regenerative medicine to be successful in repairing human Since 1979, Gladstone investigators have been at the forefront
hearts, it is critical to understand normal and abnormal heart of characterizing heart disease. Early on, they made significant
development, as well as the morphogenetic and patterning contributions to the scientific community’s understanding of
processes that occur to assemble all of the heart’s components how cholesterol and apolipoproteins are involved in coronary
into a functional organ. Gladstone scientists achieved this by artery disease. Specifically, they showed how apolipoprotein E2
deciphering a basic blueprint for the development of the heart. contributes to type III hyperlipoproteinemia and premature heart
To do so, they systematically investigated the function of tran- disease. More recently, the scientists shifted their focus to better
scriptional and epigenetic regulators across the entire genome, understand early heart development and birth defects that affect
and gained a deeper understanding of how networks of genes the heart. They showed the mechanisms underlying human
are deployed for important patterning and morphogenetic cardiac septal defects and valve disease.
decisions in heart development. Recent recognition that broad
epigenetic dysregulation in heart failure underlies the reactiva-
Identified and defined apolipoprotein E and its role in
tion of fetal gene programming and activation of fibrotic path- cholesterol metabolism and heart disease
ways provides novel targets for treating cardiac dysfunction. Gladstone scientists identified and described the characteristics
of apolipoprotein E (apoE), one of the major lipoproteins involved
Reprogrammed resident cardiac fibroblasts to cardiomyocytes in cholesterol metabolism, heart disease, and neurological
in situ to regenerate damaged hearts disease. They determined the amino acid and gene sequences
When a heart attack occurs, blood flow is lost to a portion of of the three isoforms of apoE, their three-dimensional structures
the heart muscle. Without a steady supply of oxygen, the heart and their effects on function, and their involvement in cholesterol
muscle dies and cardiac fibroblasts—which make up about metabolism and heart disease. This influential research laid the
50 percent of the heart—move in to form non-beating scar tissue. basis for showing apoE4’s involvement in Alzheimer’s and other
In a seminal advance, Gladstone investigators reprogrammed the neurological diseases.
abundant fibroblasts in a mouse heart into beating heart muscle.
As a result, instead of forming scar tissue, the fibroblasts became
Identified and defined the function of lipid metabolism enzymes
cardiomyocytes, incorporated themselves into the heart tissue, Lipids, especially triglycerides, are the major energy storage
and began beating. The approach of harnessing resident cells molecules for animal and human cells. Excessive accumulation of
to regenerate the heart is now being developed toward clinical triglycerides, however, is associated with human diseases, such
application within the spin-out company, Tenaya Therapeutics. as obesity, diabetes, and steatohepatitis. Little was known about
the enzymes and synthetic pathways that make these fats, until
Developed cellular reprogramming process for multiple cell Gladstone scientists identified and characterized those enzymes,
types using chemicals including monoacylglycerol acyltransferases and diacylglycerol
The initial discovery of reprogramming adult cells into stem cells acyltransferases. Their studies helped define the possible roles
revolutionized biology and energized research into regenerative of these important enzymes in human health and diseases.
medicine. Gladstone scientists identified small molecules that
can replace the genetic material that was traditionally used to
reprogram cells. More recently, Gladstone scientists success-
fully reprogrammed fibroblasts to pluripotent stem cells using
CRISPR-Cas9 technology to activate enhancers and promoters of
pluripotency factors. They also identified discrete combinations
of small molecules that can reprogram fibroblasts directly into
heart, neural, liver, and pancreatic cells and control specific types
of T cells, simplifying the process of reprogramming to specific
cell types.
7
Laboratory Reports
Thomas P.
Bersot
MD, PhD
ASSOCIATE INVESTIGATOR
Highlights Since 1990, Thomas P. Bersot has directed the Gladstone
Lipid Disorders Training Center, helping educate health care
Conducted two courses to train
providers to better manage risk factors for cardiovascular
155 health care providers in
disease (CVD) in their patients, including overweight and
cardiovascular risk factors
obesity. The center has trained over 4,200 health care pro-
Oversaw the training of 68 residents viders in the Community Health Network of the San Francisco
in managing cardiovascular disease Department of Public Health and at San Francisco General
risk factors Hospital. In addition to physicians, the courses are offered to
Served as a member of the nurse practitioners, clinical pharmacists, dietitians, and ex-
institutional review board for ercise physiologists, given their substantial responsibility for
human research at San Francisco managing CVD risk factors in primary-care patients.
General Hospital Accomplishments
The Gladstone Lipid Disorders Training Center offers two types
of courses several times per year.
The basic 2-1/2-day course covers the physiology and patho-
physiology of plasma lipid metabolism, hypertension, and diabe-
tes mellitus. Bersot and his team review the evidence supporting
risk assessment tools and the use of diagnostic procedures and
therapies. They devote extensive time to diet, exercise, and
weight management, which are the cornerstones of CVD preven-
tion. They also review safe and appropriate use of medications.
In addition, attendees participate in a 1-day demonstration clinic
where they see actual patients, thus providing them with practi-
cal experience in patient management.
The other course is a 1-day update for previous students and
covers new diagnostic methods for assessing the risk of sustain-
ing a clinical vascular disease event and the treatment implica-
tions of recently completed clinical studies. Bersot reviews new
drug therapies and discusses significant developments in life-
style management.
Bersot and his team stress the value of a healthy lifestyle, which
can reduce vascular disease risk by 50 percent or more and
add to the benefits of drug therapy and invasive treatments
(angioplasty, stenting, and bypass surgery). The courses’ focusGladstone Institutes
Gladstone Lipid Disorders Training
The Center’s training courses are endorsed by the American
Heart Association. Since the founding of the center in
1990, these courses have educated over 4,200 health care
providers on ways to help patients reduce their risk of
cardiovascular disease.
on these issues is a uniformly popular feature, because encour- is expected to be reversed in the next 5 years. The good news
aging patients to comply with lifestyle change recommendations is that a significant proportion of the risk of heart disease is
is difficult. completely within our own hands.
Research Impact Future Direction
Coronary heart disease is still responsible for one in six deaths Bersot will continue his work to train physicians and other health
in the United States. If stroke and heart failure are included, CVD care providers using the latest methods of managing cardiovas-
caused about one in three deaths (≈690,000 deaths) in 2008. cular disease risk factors. He hopes that drawing attention to
the value of a healthy lifestyle will contribute to improving the
The number of annual deaths caused by CVD has significantly
management of these risk factors and, ultimately, reduce the
decreased since 1968, due to better treatments and improve-
number of deaths associated with CVD.
ments in the management of CVD risk factors. However, the
steadily increasing prevalence of overweight, obesity, and diabe-
tes has increased CVD death in adults by nearly 20 percent. As
of 2008, 68 percent of adults in the United States were over-
weight or obese, as were one in three children 12–19 years of
age, and nearly one-fifth of children 2–11 years of age.
If this trend of increasing overweight and obesity in young chil-
dren persists, the decline in CVD mortality that began in 1968
Top Five Overall Publications
Ling H et al. Genome-wide linkage and association analyses to
identify genes influencing adiponectin levels: the GEMS study.
Obesity (Silver Spring). 2
009.
Mahley RW et al. Low levels of high density lipoproteins in Turks,
a population with elevated hepatic lipase. High density lipoprotein
characterization and gender-specific effects of apolipoprotein E
genotype. Journal of Lipid Research. 2
000.
Rall SC Jr et al. Type III hyperlipoproteinemia associated with
apolipoprotein E phenotype E3/3. Structure and genetics of an
apolipoprotein E3 variant. Journal of Clinical Investment. 1989.
Bersot TP et al. Interaction of swine lipoproteins with the low-density
lipoprotein receptor in human fibroblasts. J ournal of Biological
Chemistry. 1976.
Mahley RW et al. Identity of very low density lipoprotein apo-proteins
of plasma and liver Golgi apparatus. Science. 1 970.
9Laboratory Reports
Benoit G.
Bruneau
PhD
ASSOCIATE DIRECTOR
AND SENIOR INVESTIGATOR
Highlights Benoit G. Bruneau and his team aim to understand how the
human genome is coordinately regulated to make specific cell
Defined the mechanism underlying
types, such as cardiomyocytes, and how this normally stable
3D genome organization
blueprint is misread in inherited and acquired heart disease.
Discovered how cardiac transcription
factors work together to form a heart Accomplishments
Bruneau’s laboratory demonstrated the interactions between
Discovered how chromatin-
three disease-related transcription factors—TBX5, NKX2-5, and
remodeling complexes change their
GATA4—at a genome scale. They found that these proteins
identity to shape the genome of a co-localize across the genome to regulate the cardiac gene
heart cell expression program, and elucidated some of the rules by which
they co-recruit one another to active cardiac enhancers. The
scientists also identified a protein-protein interaction that facili-
tates their shared binding, through the crystal structure of TBX5,
NKX2-5, and their shared DNA binding site.
In addition, the team studied the roles played by another tran-
scription factor, CTCF, in embryonic stem cells. Using a new
system that allows rapid and reversible depletion of CTCF, they
showed that the three-dimensional organization of chroma-
tin into structures called “topologically associated domains”
is highly dependent on CTCF. Through these studies, they
discovered new rules about chromatin organization and how it
impacts gene regulation.
They also examined the importance of a disease-related
Lab Members histone-modifying enzyme called KMT2D. The gene that encodes
* indicates current lab members this protein is often mutated in congenital heart disease. They
deleted KMT2D in mice and showed that it controls a set of
Laure Bernard Alexis Krup*
genes essential for embryonic cardiac function by depositing
Aaron Blotnick* Alejandra Lopez Delgado
Pervinder Choksi Luis Luna-Zurita
at regulatory elements in the genome a specific type of histone
Steven Cincotta Dario Miguel-Perez modification that helps genes become activated.
Walter Devine* Abigail Nagle*
Bayardo Garay Elphège-Pierre Nora*
Matthew George* Diego Quintero
Piyush Goyal* Kavitha Rao*
Swetansu Hota* Tanya Sukonnik
Vasumathi Kameswaran* Alec Uebersohn
Irfan Kathiriya* Sarah Wood*Gladstone Institutes
Molecular Players for Transcriptional Regulation
DNA Cis-regulatory elements containing DNA binding sites
Cardiomyocyte
are bound by transcription factors and modulate the
assembly of the Pre-Initiation Complex at promoters
through physical contacts driven by a three-dimensional
arrangement of chromatin, thereby acting as a molecular
platform between cellular signaling and gene activity.
Binding
Sites
Transcription
Factors
Co-Factor
Chromatin
Remodeling Mediator
Complex Complex
Repressed
Chromatin General
TFs
Pol II Pol II
Histone Modification
Reader/Writer mRNA
Research Impact Future Direction
Bruneau’s research is important for understanding basic This laboratory focuses on understanding how the earliest deci-
concepts in gene regulation, and how they are dysregulated sion by an embryo or a stem cell to become a heart precursor
in disease. The demonstrated interactions between cardiac is regulated, and how global gene regulation is coordinated in
transcription factors provided new insights into the tight regula- this process. The scientists are further exploring the cellular and
tion of gene cohorts, which has had immediate implications in molecular mechanisms by which discrete cell types contribute to
understanding how diseases in these transcription factor genes specific cardiac structures, such as the interventricular septum.
cause similar diseases. These findings are broadly impactful as They are also investigating how chromatin-remodeling complexes
they apply to any set of transcription factors, in any cell type. In establish a “go/no-go” switch in cardiac differentiation. Finally,
addition, his team’s work on CTCF and 3D genome organization they use human induced pluripotent stem cell models to under-
resolved several long-standing questions in various fields, and stand, at a single-cell resolution, how disease-causing mutations
raised new questions that now need answers. in TBX5 affect gene expression and chromatin states.
Selected Recent Publications Top Five Overall Publications
Sun X et al. Cardiac-enriched BAF chromatin-remodeling complex Nora E et al. Targeted degradation of CTCF decouples
subunit Baf60c regulates gene expression programs essential for local insulation of chromosome domains from genomic
heart development and function. Biology Open.2017. compartmentalization.Cell.2017.
Nora E et al. Targeted degradation of CTCF decouples Luna-Zurita L et al. Complex interdependence regulates heterotypic
local insulation of chromosome domains from genomic transcription factor distribution and coordinates cardiogenesis.
compartmentalization.Cell.2017. Cell.2016.
Hota S et al. ATP-dependent chromatin remodeling during Devine W et al. Early patterning and specification of cardiac
mammalian development.Development.2016. progenitors in gastrulating mesoderm.eLife.2014.
Ang S et al. KMT2D regulates specific programs in heart Wamstad J et al. Dynamic and coordinated epigenetic regulation of
development via histone H3 lysine 4 di-methylation. developmental transitions in the cardiac lineage.Cell.2012.
Development.2016.
Takeuchi J et al. Directed transdifferentiation of mouse mesoderm to
Luna-Zurita L et al. Complex interdependence regulates heterotypic heart tissue by defined factors.Nature.2009.
transcription factor distribution and coordinates cardiogenesis.
Cell.2016.
11Laboratory Reports
Bruce R.
Conklin
MD
SENIOR INVESTIGATOR
Highlights Decoding human genetic disease allows Bruce R. Conklin
and his team to develop models of pathology that can be
Established precise genome-editing
directly tested with gene correction or targeted drug therapy.
methods for disease modeling
Dominant negative mutations are particularly promising thera-
and therapy
peutic targets since they are resistant to traditional therapies,
Established an efficient and yet, precise excision of a disease-causing allele could
method to produce single-base provide a cure. This laboratory uses patient-derived induced
changes in iPSCs pluripotent stem cells (iPSCs) to model diseases in tissues that
Pioneered the use of CRISPRi are particularly susceptible to dominant negative mutations:
to epigenetically control gene cardiomyocytes, motor neurons, and retinal pigment epithelial
expression in iPSCs (RPE) cells. By developing CRISPR genome surgery in human
cells, they hope to devise improved cellular models and
human therapies.
Accomplishments
Conklin’s team has successfully created stem cell models of
genome surgery. By focusing on allele-specific gene excision,
they can select gene mutations that are highly penetrant, with
clear phenotypes in cell types that can be readily derived
from iPSCs. They use whole-genome sequencing to identify
common genetic polymorphisms, which can be used to selec-
tively inactivate the disease allele with CRISPR nucleases. The
diseased cell types allow them to decode the cellular signatures
of disease and determine if the excision of the disease allele
restores cellular functioning.
Lab Members
Genome surgery is a rapidly advancing field that uses state-of-
* indicates current lab members
the-art techniques to push the boundaries of cell and molecular
Amanda Jayne Carr Steven Mayerl biology. This laboratory uses advanced microscopy, tissue engi-
Carissa Feliciano Meghan McKenna*
neering, and single-cell genomics to optimize precise editing.
Vanessa Herrera* Michael Olvera*
Kristin Holmes Meiliang Pan* They are also developing computational methods to select
Nathaniel Huebsch* Juan Pérez-Bermejo* optimal CRISPR/Cas9 combinations in diverse populations. They
Olga Ivanova Edward Shin aim to produce therapies that are safe and cost effective so they
Christina Jensen Kenneth Tan* can benefit the maximal number of people. In collaboration with
Luke Judge* An Truong clinical scientists and the Innovative Genomics Institute, they are
Kathleen Keough* Hannah Watry* preparing large animal models and clinical-grade reagents for
Angela Ziqi Liu* Kenneth Wu*
human clinical trials.
Mohammadali Mandegar Perry WuGladstone Institutes
Genome Surgery for Dominant Negative Disease
Resulting Protein Complexes The CRISPR/Cas9 system can be used to selectively silence
the disease allele without altering the normal allele. In the
Diseased 97% lower panel, a dominant negative disease allele (orange)
Protein Diseased poisons the protein complex. CRISPR excision blocks the
disease allele to ensure all proteins remain healthy.
Normal 3%
Protein Healthy
Dominant
Negative
Target A Allele Target B
Target
Cas9 gRNA 100%
Healthy
Excision
Cleavage
Research Impact Future Direction
A major benefit of testing genome surgery in authentic cellular Genome engineering and stem cell biology have been the most
models is the mechanistic insights into the disease process and disruptive technologies of this millennia. Advances in iPSC
the potential for functional recovery. The reversion of a cellular differentiation and cell modeling will allow more cell types and
phenotype is proof that a dominant negative allele was causative sophisticated multicellular models of disease. These will provide
and that the disease process is reversible. molecular insights into many diseases, which are likely to lead
to improved drug therapy without gene correction. Conklin’s
Detailed cellular analysis often provides new insights into the
team aims to further enhance these sophisticated methods to
mechanism of the disease. Genomic deletions require detailed
intervene in genetic disease with epigenetic modification or
knowledge of the non-coding elements that are poorly under-
base editing that will expand the field of genome surgery. They
stood, such as enhancers, LncRNAs, and microRNAs. Each cell
continue to leverage these new advances to reach their goal of
type allows the researchers to probe the 3D architecture and
decoding and repairing genetic disease.
epigenetic state of the gene region, since distant DNA can be
in close proximity, allowing efficient excision of larger genomic
segments. Finally, as Conklin’s team learns to orchestrate precise
repair, they will better understand the DNA repair machinery of
each cell type.
Genome surgery is an emerging field of medicine that will drive
a new level of investigation into the molecular physiology of
diverse cell types, including cardiomyocytes, motor neurons, and
RPE cells. Only by understanding the basic cellular and molecular
physiology of these cells can scientists meet the challenges that
lie ahead.
Selected Recent Publications Top Five Overall Publications
Judge LM et al. BAG3 chaperone complex maintains cardiomyocyte Conklin BR et al. Engineering GPCR signaling pathways with RASSLs.
function during proteotoxic stress. JCI Insight. 2017. Nature Methods. 2008.
Liu SJ et al. CRISPRi-based genome-scale identification of functional Dahlquist KD et al. GenMAPP, a new tool for viewing and analyzing
long noncoding RNA loci in human cells.Science.2017. microarray data on biological pathways.Nature Genetics.2002.
Huebsch N et al. Miniaturized iPS-cell-derived cardiac muscles Redfern CH et al. Conditional expression and signaling of a
for physiologically relevant drug response analyses.Scientific specifically designed Gi-coupled receptor in transgenic mice.Nature
Reports.2016. Biotechnology.1999.
Mandegar MA et al. CRISPR interference efficiently induces gene Conklin BR et al. Substitution of three amino acids switches receptor
knockdown and models disease in iPSCs.Cell Stem Cell.2016. specificity of Gqα to that of Giα.Nature.1993.
Miyaoka Y et al. Systematic quantification of HDR and NHEJ reveals Federman AR et al. Hormonal stimulation of adenylyl cyclase through
effects of locus, nuclease, and cell type on genome-editing.Scientific Gi-protein βγ subunits.Nature.1992.
Reports.2016.
13Laboratory Reports
Sheng
Ding
PhD
SENIOR INVESTIGATOR
Highlights The team led by Sheng Ding develops new chemical biology
approaches to study stem cell biology and regeneration. Their
Reprogram human fibroblasts into
current work focuses on identifying and characterizing novel
cardiomyocytes with a cocktail of
small molecules that control cell fate and/or function in various
small molecules
systems, including maintenance of tissue-specific stem cells,
Reprogram mouse fibroblasts directed differentiation of pluripotent stem cells toward new
into neural progenitor cells with a cell lineages, reprogramming of lineage-restricted somatic
cocktail of small molecules cells to alternative cell fate, and regulation of cancer stem
Reprogram human Th17 cells into cells. The identified small molecules or generated cells are
regulatory T cells further characterized in vitro and in vivo. Furthermore, mech-
anistic studies of these small molecules provide new insights
underlying fundamental processes in cell fate regulation.
Accomplishments
The scientists recently developed a new paradigm in cellular
transdifferentiation using the cell-activation and signaling-directed
(CASD) lineage-conversion strategy. This method employs tran-
sient exposure of somatic cells with reprogramming molecules
(cell activation, CA) in conjunction with lineage-specific soluble
signals (signal-directed, SD) to reprogram somatic cells into
diverse lineage-specific cell types without entering the pluripotent
state. The strategy was demonstrated by directly converting fibro-
blasts into cardiac, neural, or definitive endoderm precursor cells
in mouse and human systems.
Importantly, Ding’s laboratory identified specific chemically
defined conditions that enable robust expansion of the repro-
grammed lineage-specific precursor cells, which could then
be further differentiated into mature functional cells in vitro.
Lab Members
Transplanting those CASD-reprogrammed cells rescued disease
* indicates current lab members phenotypes in corresponding mouse models, demonstrating
Nan Cao Shibing Tang potential utility in cell-based therapy. More significantly, they
Shengping Hou Haixia Wang identified chemical cocktails that enable CASD-based repro-
Ke Li Shaohua Xu gramming without genetic manipulations to generate cardiac and
Changsheng Lin* Tao Xu neural precursor cells from fibroblasts.
Kai Liu Chen Yu*
Peng Liu Mingliang Zhang
Tianhua MaGladstone Institutes
Small Molecule Inhibitor A Novel Path to Transdifferentiation
of Pluripotency Temporally restricted ectopic overexpression of reprogram-
iPSC Medium
and Extended ming factors in fibroblasts leads to the rapid generation of
Fibroblast
Expression of epigenetically “activated” cells, which can then be coaxed to
iPSC TFs iPSC “relax” back into various differentiated states, ultimately
giving rise to somatic cells entirely distinct from the
starting population. TF, transcription factor. iPSC, induced
Transient
Treatment with Cardiac pluripotent stem cell.
Molecules Induction
Medium Cardiac Cardiomyocyte
Precursor
Definitive
Endoderm
Induction Endoderm Pancreatic and
Medium Precursor Hepatocyte
Epigenetically Neural
“Activated” Induction
Medium Neural
Cell Population Precursor Neuron and Glial
Additional discoveries by the team include reprogramming Future Direction
pro-inflammatory Th17 cells into immune suppressive regulatory Ding will continue to pursue the research paradigm of identifying
T cells by a small molecule using a novel immunometabolism and further characterizing novel small molecules that control cell
mechanism, reprogramming white adipogenic cells into brown/ fate and/or function in vitro and in vivo, especially in the context
beige adipogenic cells, and showing in vivo reprogramming of of disease and injury.
those cell types by the small molecules and disease rescue in
relevant mouse models.
Research Impact
Ding and his team hope their continued studies will ultimately
facilitate therapeutic applications of stem cells and the devel-
opment of small-molecule drugs. These could be used to stim-
ulate the body’s own regenerative capabilities by promoting
survival, migration/homing, proliferation, differentiation, and
reprogramming of endogenous stem/progenitor cells or more
differentiated cells.
Selected Recent Publications Top Five Overall Publications
Xu T et al. Metabolic control of TH17 and induced Treg cell balance Li H et al. Versatile pathway-centric approach based on high-
by an epigenetic mechanism.Nature.2017. throughput sequencing to anticancer drug discovery.Proceedings of
the National Academy of Sciences.2012.
Nie B et al. Brown adipogenic reprogramming induced by a small
molecule.Cell Reports.2017. Li W et al. Rapid induction and long-term self-renewal of primitive
neural precursors from human embryonic stem cells by small
Cao N et al. Conversion of human fibroblasts into functional
molecule inhibitors.Proceedings of the National Academy of
cardiomyocytes by small molecules.Science.2016.
Sciences.2011.
Zhang M et al. Pharmacological reprogramming of fibroblasts into
Efe JA et al. Conversion of mouse fibroblasts into cardiomyocytes
neural stem cells by signaling-directed transcriptional activation.Cell
using a direct reprogramming strategy.Nature Cell Biology.2011.
Stem Cell.2016.
Shi Y et al. A combined chemical and genetic approach for the
Zhang Y et al. Expandable cardiovascular progenitor cells
generation of induced pluripotent stem cells.Cell Stem Cell.2008.
reprogrammed from fibroblasts.Cell Stem Cell.2016.
Chen S et al. Dedifferentiation of lineage-committed cells by a small
molecule.Journal of the American Chemical Society.2004.
15Laboratory Reports
Saptarsi
Haldar
MD
ASSOCIATE INVESTIGATOR
Highlights The goal of Saptarsi Haldar’s laboratory is to dissect the
molecular mechanisms used by cells to control gene expres-
Discovered several novel epigenetic
sion during cardiovascular and metabolic homeostasis.
regulators of cardiovascular
Furthermore, his team seeks to understand how these gene
homeostasis that may be druggable
regulatory mechanisms are dysregulated in disease and find
targets in heart failure
novel approaches to manipulate the epigenetic signaling
Leveraged deep epigenomic pathways for therapeutic gain.
interrogation to uncover novel core
regulatory circuitry in smooth muscle Accomplishments
cell phenotypic plasticity The research team discovered several novel epigenetic signaling
mechanisms that govern cardiovascular and metabolic homeo-
Discovered novel molecular pathway
stasis. For example, they found that BRD4, a member of the BET
in the liver that is essential for bromodomain family epigenetic reader proteins, is a critical regu-
systemic glucocorticoid hormone lator of cardiovascular stress responses and is important in heart
homeostasis failure pathogenesis and vascular remodeling. They showed
proof-of-concept that small-molecule inhibition of BRD4 has ther-
apeutic potential in cardiovascular disease.
Haldar’s team also found that another epigenetic protein, CDK7
(the core kinase in the TFIIH complex), is a novel druggable
target in heart failure pathogenesis. In addition, they identified
Salt-inducible kinases (SIKs) as novel effectors of cardiomyocyte
transcription control and pathological cardiac remodeling. They
also used deep epigenomic interrogation of smooth muscle cells
to discover novel core regulatory transcriptional circuitry that
drives smooth muscle phenotypic switching, findings which have
major implications for vascular diseases.
In the field of metabolism, they discovered that the transcription
factor KLF15 is a master regulator of hepatic cortisol binding
Lab Members globulin production and is an essential regulator of systemic
glucocorticoid bioactivity during physiology and disease. Using
* indicates current lab members CRISPR-Cas9–based genome editing, they epitope tagged the
Michael Alexanian* Austin Hsu* KLF15 allele in mice, and are now discovering the first endoge-
Priti Anand* Zhen Jiang* nous cistromes and interaction partners for this nodal metabolic
Rohan Bhardwaj Sarah McMahon*
transcription factor.
Anna Chen Arun Padmanabhan*
Qiming Duan* Sarah Wood*
Previn GanesanGladstone Institutes
Haldar’s team aims to understand how gene expression is
Diseased and
Committed Mature Dysfunctional controlled in the postnatal heart during physiology and
Cardiomyocyte Cardiomyocyte Cardiomyocyte disease at both the cellular and whole-organ levels. This
includes studying how DNA binding transcription factors
Cellular
Level (TFs) signal in the context of chromatin to fashion the
TF/ TF/ mature cardiomyocyte and adult heart during postnatal
Chromatin Chromatin growth and development, and how these mechanisms go
awry in the diseased heart. The same conceptual framework
is applied to understanding gene control mechanisms
Organ underlying cell state changes in blood vessels and key
Level metabolic organs, such as liver and skeletal muscle.
Newborn
Heart
Mature Heart
Diseased Heart
Research Impact Future Direction
The Haldar laboratory discovered new epigenetic signaling The team engineered a suite of genetically modified cells and
mechanisms used by cardiovascular and metabolic tissues to mice to deeply probe the molecular function of several epigen-
control gene expression. For a number of these gene regulatory etic regulators, including BET proteins, CDK7, SIKs, and KLF15,
pathways, they showed how mechanisms go awry in disease both in physiological homeostasis and disease. In a new discov-
and provided proof-of-concept that specific epigenetic signaling ery effort, they are leveraging deep epigenomic interrogation
effectors can be pharmacologically targeted in conditions, such to decipher the core transcription factor regulatory circuitry that
as heart failure, vascular dysfunction, and muscular dystrophy. controls postnatal cardiomyocyte maturation, which represents
Through a detailed mechanistic understanding of cardiovascular a major knowledge gap in the cardiac regeneration field. Finally,
and metabolic gene regulation, they ultimately hope to find new they are developing genetic screens to discover novel regulators
therapies for human disease. of cardiomyocyte homeostasis and plasticity.
Selected Recent Publications Top Five Overall Publications
Newman J et al. Ketogenic diet reduces midlife mortality and Duan Q et al. BET bromodomain inhibition suppresses innate
improves memory and aging in mice. C ell Metabolism.2017. inflammatory and profibrotic transcriptional programs in heart failure.
Science Translational Medicine.2017.
Duan Q et al. BET bromodomain inhibition suppresses innate
inflammatory and profibrotic transcriptional programs in heart failure. Morrison-Nozik A et al. Glucocorticoids enhance muscle endurance
Science Translational Medicine. 2017. and ameliorate Duchenne muscular dystrophy through a defined
metabolic program. P
roceedings of the National Academy of
Stratton M et al. Signal-dependent recruitment of BRD4 to
Sciences. 2015.
cardiomyocyte super-enhancers is suppressed by a microRNA. C
ell
Reports.2016. Anand P et al. BET bromodomains mediate transcriptional pause
release in heart failure. Cell. 2013.
Jeyaraj D et al. Circadian rhythms govern cardiac repolarization and
arrythmogenesis. N ature. 2012.
Haldar S et al. Klf15 deficiency is a molecular link between heart
failure and aortic aneurysm formation. Science Translational
Medicine. 2010.
17Laboratory Reports
Todd C.
McDevitt
PhD
SENIOR INVESTIGATOR
Highlights The overall goal of Todd C. McDevitt’s laboratory is to develop
novel technologies to enable the translation of stem cells for
Generated excitatory spinal
therapeutic and diagnostic applications. Much of their work
interneurons from human
focuses on engineering 3D microscale tissue constructs from
pluripotent stem cells capable of
human stem cells that recapitulate the phenotypic and func-
forming synaptic connections with
tional properties of native tissues. They employ scaffoldless
other neurons
tissue-engineering approaches to study the morphogenesis
Engineered heterotypic cardiac of pluripotent stem cells using a combination of biomateri-
microtissues that promote als-based approaches and cell-engineering techniques. They
the phenotypic and functional are also interested in characterizing and exploiting morpho-
maturation of human iPSC-derived genic factors produced by stem and progenitor cells that have
cardiomyocytes immunomodulatory and regenerative/rejuvenative effects on
Created predictable and robust somatic cells and tissues.
multicellular human iPSC patterns Accomplishments
via manipulation of intrinsic cell
Over the past decade, McDevitt and his team defined scalable
mechanisms
and robust technologies for enhanced control and consistency
of stem cell differentiation and microtissue engineering. Using
these platform technologies, they developed different heterotypic
models of cardiac, neural, and hepatic tissues from human stem
cell sources. One of their goals is to determine the specific effects
of 3D multicellular heterotypic interactions and cell-derived extra-
cellular matrix on individual cell phenotypes and the resulting
physiological properties of engineered microtissues.
The researchers have found that tissue-specific stromal cells (i.e.,
fibroblasts) significantly impact parenchymal cell (i.e., cardiomyo-
Lab Members cyte) phenotype through several different mechanisms, which
*indicates current lab members they continue to pursue in more depth. They also showed that
the phenotypic changes affect the relative maturity of the human
Jessica Butts* Oriane Matthys*
Ana De Andrade E Silva* Dylan McCreedy* induced pluripotent stem cell (iPSC)–derived cells.
Amy Foley Nik Mendoza-Camacho* The team was the first to report the differentiation of excitatory
Tracy Hookway Eszter Mihaly*
spinal interneurons from human pluripotent stem cells. They are
David Joy* Vaishaali Natarajan*
Michael Kang* Jessica Sepulveda
now examining the functional and potential therapeutic efficacy
Ariel Kauss* Diwakar Turaga* of using these cells to repair spinal cord injury and develop novel
Ashley Libby* Jenna Wilson organoid models of the central nervous system.
Ronald Manlapaz* Joshua ZimmermannGladstone Institutes
The McDevitt laboratory focuses on the creation of tissue
Stem Cells models and regenerative molecular therapies from stem
cells. They are developing cardiac, neural, and other tissues
to probe heterotypic impacts on development and disease,
while in parallel exploring and exploiting the unique and
complex cadre of molecules produced by stem cells.
Tissue Molecules
Cardiac Neural Matrix Secretome
They also devised a high-density perfusion bioreactor system for Future Direction
human iPSC culture that produces enhanced yields and concen- The team will continue to engineer robust methods to control
trations of morphogenic factors. They are identifying and examin- organoid development from human pluripotent and post-natal
ing the rejuvenative effects of human iPSC-factors on aged cells stem cells, and use these novel tissue models to study mecha-
and tissues in several mouse models. nisms of human development and disease ex vivo. Furthermore,
Research Impact they aim to innovate new microphysiological systems and func-
tional assays that enable the exploration of epigenetic effects of
Human tissue constructs and organoids derived from stem cells
environmental factors on human tissue structure and function.
offer novel model systems for probing mechanisms of embryonic
development. Furthermore, engineered human microtissues offer In addition, they will use the systematic and comprehensive anal-
tractable substrates to interrogate infectious and genetic diseases ysis of the molecules uniquely produced by stem cells to lead to
and the effects of exposure to other environmental factors. new molecular therapies and engineered materials that stimulate
McDevitt’s laboratory uses a bottom-up approach to understand regeneration and repair of somatic tissues.
how cells cooperatively interact to form complex tissues and
dictate multicellular organization and subsequent physiologic
function. They also seek to determine the relative contributions of
stromal cells to measurable physiological properties.
Selected Recent Publications Top Five Overall Publications
Khalil et al. Functionalization of microparticles with mineral coatings Sutha et al. Osteogenic embryoid body-derived material induces
enhances non-viral transfection of primary human cells. Scientific bone formation in vivo.Scientific Reports.2015.
Reports.2017.
Murphy et al. Materials as stem cell regulators.Nature
Jackson-Holmes EL et al. A microfluidic trap array for longitudinal Materials.2014.
monitoring and multi-modal phenotypic analysis of individual stem
McDevitt TC. Scalable culture of human pluripotent stem cells in 3D.
cell aggregates.Lab on a Chip.2017.
Proceedings of the National Academy of Sciences.2013.
Butts JC et al. Differentiation of V2a interneurons from human
Bratt-Leal AM et al. A microparticle approach to morphogen delivery
pluripotent stem cells.Proceedings of the National Academy of
within pluripotent stem cell aggregates.Biomaterials.2013.
Sciences.2017.
Singh A et al. Adhesion strength–based, label-free isolation of
Zimmerman JA et al. Enhanced immunosuppression of T cells
human pluripotent stem cells.Nature Methods.2013.
by sustained presentation of bioactive interferon-γ within three-
dimensional mesenchymal stem cell constructs.Stem Cells
Translational Medicine.2017.
Wang Y et al. Mineral particles modulate the osteo-chondrogenic
differentiation of embryonic stem cell aggregates.Acta
Biomaterialia.2016.
19Laboratory Reports
Deepak
Srivastava
MD
PRESIDENT AND
SENIOR INVESTIGATOR
Highlights Deepak Srivastava’s laboratory focuses on the fundamental
events involved in cell fate determination, differentiation,
Discovered combination of tran-
and organogenesis. Specifically, the team investigates the
scription factors and small molecule
molecular events regulating cardiogenesis. They focus on
inhibitors that optimally reprogram
signaling, transcriptional, and post-transcriptional networks
mouse and human cardiac fibroblasts
in this process. They have leveraged knowledge of key gene
to induced cardiomyocyte-like cells in
networks to reprogram resident fibroblasts directly into
vitro and in vivo and revealed mecha-
cardiomyocyte-like cells for regenerative purposes. They also
nisms underlying the transition
investigate the genetic causes of human cardiovascular disease
Identified a combination of genes and use human induced pluripotent stem cells (iPSCs) to reveal
that unlocks the proliferative poten- disruption of cardiogenic gene networks that lead to disease.
tial in cells that had permanently By using these approaches, they are discovering the biology
exited the cell cycle underlying cardiogenesis and cardiovascular disorders and
Showed a GATA4 missense muta- beginning to find novel therapeutic interventions.
tion disrupts a transcription factor Accomplishments
“code” at cardiac enhancers, leading
Srivastava’s team described complex signaling, transcriptional,
to abnormal cardiac septation and
and translational networks that guide early differentiation of
dysfunction
cardiac progenitors and later morphogenetic events during
Lab Members cardiogenesis. By leveraging these networks, they repro-
grammed disease-specific human cells to model human heart
*indicates current lab members
disease in patients with mutations in cardiac developmental
Yen Sin Ang Ethan Radzinsky genes. Deep epigenetic and transcriptome analyses revealed
Bonnie Cole* Sanjeev Ranade* perturbations in pivotal gene networks, which contribute to
Yvanka De Soysa* Janell Rivera
disease that could be corrected by altering dosage of nodal
Aryé Elfenbein* Gabriel Rubio
Giselle Galang Hazel Salunga* points in the network. These studies revealed mechanisms of
Laxmi Ghimire Ryan Samarakoon* NOTCH1 and GATA4 haploinsufficiency, and the researchers
Casey Gifford* Kaitlen Samse* showed the contribution of genetic variants inherited in an oligo-
Bárbara González Terán* Amelia Schricker* genic fashion in congenital heart disease.
Yu Huang* Nicole Stone*
Isabelle N. King Joke van Bemmel* They used a combination of cardiac regulatory factors and small
Wesley Kwong* Vasanth Vedantham molecules to directly reprogram resident cardiac fibroblasts into
Lei Liu* Pengzhi Yu* cardiomyocyte-like cells in vitro and in vivo to repair damaged
Elijah Martin Sarah Zambrano* hearts. Single-cell RNA-sequencing showed how heterogeneity
Kimberly R. Cordes Metzler Yu Zhang* of the reprogramming process may inhibit efficiency and could
Tamer Mohamed Ping Zhou*
be manipulated. The small molecules appear to increase acces-
Shanelle Nebre* Lili Zhu*
Karishma Pratt
sibility of reprogramming factors to their DNA-binding sites by
opening chromatin at those sites genome-wide.You can also read
