authors@Fred Hutch

Frost, Sophia

April 20, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 4 | Fred Hutchinson Cancer Research Center 
 
A Vascular Niche for Boosted Blood Stem Cell 
Production 
April 20, 2015 
    SHL Frost 
Many patients with hematopoietic disease rely on transplantation of hematopoietic stem/progenitor 
cells (HSPC) for a cure. Regardless of transplant source—healthy donors or the patients' own 
genetically modified cells—treatment success depends on proper engraftment. Allogeneic 
transplantation has a good track record, but is limited by donor availability and adverse effects such 
as graft-versus-host disease. The advent of autologous stem cell gene therapies introduced a 
promising solution to both issues; however, the production of long-term engrafting HSPCs must be 
improved for treatments to be clinically realistic. 
How can this be achieved? First of all, different kinds of HSPCs have varied self-renewal, 
differentiation, and engraftment potential. Pluripotent stem cells (PSC) can propagate indefinitely 
and may be directed to make many cell types in the human body, and could therefore be used as an 
unlimited HSPC source. Hematopoietic multipotent progenitor cells (MPP) are derived from PSCs 
and share the basic blood stem cell features. However, clinical translation of blood therapeutics 
utilizing human PSC-derived MPPs has been hampered by inadequate engraftment and poor 
proliferation in mouse models, prompting exploration of the underlying mechanisms behind blood 
cell development. Findings indicate that endothelial cells (EC) are key players for development and 
maintenance of HSPCs, by forming specialized microenvironments within the bone marrow and 
other organs, referred to as "vascular niches"—units with both anatomic and functional dimensions 
that determine the various interactions that influence blood stem cell formation. More specifically, 
studies of signaling pathways and gene expression in the vascular endothelium suggest an 
important role for the Notch pathway for boosting the proliferation of human long-term MPPs—a 
possible lead to improved HSPC production. 
These findings inspired Drs. Jennifer Gori and Hans-Peter Kiem from the Clinical Research Division 
to hypothesize that EC-induced Notch ligands may stimulate blood stem cell specification, and that 
an artificial ex vivo vascular niche could increase the production of long-term blood MPPs from PSC 
precursors. The idea was tested by generating induced PSCs directly from mature macaque cells, 
allowing evaluation of PSC-derived MPP engraftment and proliferation in mice. Published in Journal 
of Clinical Investigation, the results showed a clear benefit of EC-induced PSC-MPPs over cytokine-
induced control cells for initial engraftment and retainment over time. In vivo selection using low-
April 20, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 4 | Fred Hutchinson Cancer Research Center 
 
dose chemotherapy of gene-modified PSC-MPPs—genetically modified to be chemotherapy-
resistant—further increased the levels, tripling the bone marrow engraftment of EC-induced 
progenitor cells compared with hematopoietic cells that were exposed to cytokines alone. In addition, 
cells produced through the artificial vascular niche exhibited desired stem cell-associated traits such 
as multipotency and self-renewal. 
To shed further light on the vascular niche functionality the investigators also studied differentiation 
and engraftment of human embryonic stem cells (hESC) with and without EC-mediated activation of 
the Notch pathway. The comprehensive evidence indicated a key role for the EC-associated Notch 
ligands JAG1 and DLL4 in the differentiation of PSCs, and that PSC-MPPs generated in the context 
of a vascular niche were significantly more active, both ex vivo and in vivo. "It has been well 
established that the vascular niche is the initial site of blood development in the embryo. Here, we 
show that EC-mediated activation of the Notch pathway in primitive blood stem cells facilitates the 
maturation and possibly the long-term engraftment of stem cells produced from PSCs," explained Dr. 
Gori. 
The studies demonstrate the potential for scaled production of functional HSPCs via reprogramming 
of patient cells, followed by growth factor-mediated induction of blood stem cell precursor formation, 
and culture of these precursors on endothelial cells for their maturation to an HSPC-like state—an 
important technical advance. "Being able to generate large numbers of hematopoietic stem cells has 
significant implications for autologous transplantation for patients with a number of hematologic 
malignancies," Dr. Kiem explained, adding that the discoveries apply also to gene therapy. "Now we 
can generate stem cells, correct them ex vivo, eg. for hemoglobinopathies, then expand corrected 
autologous stem cells and infuse them back into the patient." What's more, transplantation of larger 
amounts of corrected cells should require less pre-infusion chemo- or radiation therapy for 
successful engraftment. 
Dr. Kiem and colleagues are now adapting the methodology for generation of sufficient amounts of 
cells for clinical applications, and companies are working on scaling up the technology. Some 
questions remain, for instance whether the ECs need to be eliminated before infusion or if they can 
be irradiated and die off in the patient after ensuring engraftment of the expanded stem cells. Drs. 
Gori and Kiem do not hold all the answers yet, but the investigators are one important step closer to 
generating long-term HSPCs—a goal that could benefit many who suffer from hematologic disease. 
Gori JL, Butler JM, Chan Y-Y, Chandrasekaran D, Poulos MG, Ginsberg M, Nolan DJ, Elemento O, 
Wood BL, Adair JE, Rafii S, Kiem H-P. 2015. Vascular niche promotes hematopoietic multipotent 
progenitor formation from pluripotent stem cells. J Clin Invest. 125(3):1243-54. 
April 20, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 4 | Fred Hutchinson Cancer Research Center 
 
 
 
Image provided by Dr. Hans-Peter Kiem 
Expansion of hematopoietic stem cells using an artificial ex vivo vascular 
niche. Patient cells (fibroblasts or other somatic cells including 
lymphocytes) are reprogrammed to generate induced pluripotent stem 
cells, which differentiate into CD34-expressing progenitors (with or 
without additional gene therapy). The CD34+ cells are then cultured on 
endothelial cells that express JAG1 and DLL4, boosting the expansion 
to sufficient amounts for autologous transplantation back into the patient, 
to replace diseased blood cells. 
 


English

A Vascular Niche for Boosted Blood Stem Cell Production

Kocarnik, Jonathan

May 18, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 5 | Fred Hutchinson Cancer Research Center 
 
Risk of Gene-Fusion Positive Prostate Cancer is 
Lower Among Obese 
May 18, 2015 
    JM Kocarnik 
Approximately 50% of prostate cancers harbor a T2E gene fusion. As an early driver of prostate 
carcinogenesis, this gene fusion may characterize distinct molecular subtypes of prostate cancer 
with different etiologies.  It is not known, however, whether lifestyle factors such as obesity have 
different associations with prostate cancer risk depending on this molecular characterization.  In a 
recent study in theAmerican Journal of Epidemiology, Ms. Lieke Egbers and Dr. Janet Stanford and 
colleagues in the Public Health Sciences Division provide evidence that obesity is associated with a 
reduced risk of androgen-regulated T2E fusion-positive tumors, but not with T2E fusion-negative 
tumors. 
The T2E gene fusion consists of the fusion of the transmembrane protease, serine 2 (TMPRSS2) 
and erythroblast transformation-specific-related (ERG) genes. TMPRSS2 harbors androgen-
responsive elements in the promoter region, while ERG encodes a nuclear protein that acts as a 
transcription factor.  Androgens have been implicated in the induction of these fusion events, which 
once fused leads to overexpression of ERG and disruption of androgen receptor signaling.  As 
obesity alters the circulating levels of sex hormones compared to normal-weight men, including 
lowering androgen levels, the authors hypothesized that obesity may decrease the risk of T2E gene 
fusion-positive prostate tumors. 
The authors measured T2E gene fusion status in 563 prostate cancer tumors, using tumor tissue 
from two population-based case-control studies conducted in the King County, Washington 
area.  "The ability to obtain formalin-fixed paraffin embedded (FFPE) tumor tissue blocks and utilize 
the latest technologies for molecular profiling of such tissues is key for this type of molecular 
epidemiologic research," said lead author Ms. Egbers.  The association between quartiles of body 
mass index (BMI) and prostate cancer risk was then evaluated according to fusion status.  "This 
ability to stratify prostate cancer cases according to tumor molecular subtype (such as T2E fusion 
status) now allows us to study etiologic heterogeneity, with different molecular subtypes having 
distinct risk factor profiles," said Ms. Egbers.  
 
 
May 18, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 5 | Fred Hutchinson Cancer Research Center 
 
 
Comparing the highest BMI quartile with the lowest, the authors found an inverse association with 
the risk of T2E-positive prostate cancer, but no association was seen with risk of T2E-negative 
prostate cancer (see figure).  Other characterizations of obesity were consistent with this finding, 
such as evaluating standard WHO BMI categories (obese vs. normal) or BMI as a continuous 
variable (in 5-unit increments).  "These data provide new evidence that these different molecular 
subtypes of prostate cancer may have different etiologies," said Ms. Egbers.  
"At present the biological mechanism by which obesity decreases risk for T2E positive prostate 
cancer, but not for T2E negative tumors, is not known," said Ms. Egbers.  "Since the T2E fusion is 
responsive to androgens, we hypothesize that the altered steroid hormone profile in obese men 
(e.g., lower circulating levels of androgens) may contribute to the reduction in risk for T2Epositive 
prostate cancer. Further research will be needed to explore the biological mechanism behind this 
finding."  Understanding these etiological differences may lead to better understanding of risk factors 
for particular subtypes of prostate cancer, which should eventually lead to improvements in 
prevention, screening, and treatment. 
Other PHS researchers contributing to this project were Drs. Jonathan Wright and Marian 
Neuhouser, as well as Ms. Suzanne Kolb. 
 
Citation:  
Egbers L, Luedeke M, Rinckleb A, Kolb S, Wright JL, Maier C, Neuhouser ML, Stanford JL. 2015. 
Obesity and Prostate Cancer Risk According to Tumor TMPRSS2:ERG Gene Fusion Status. Am J 
Epidemiol. 181(9):706-13. doi: 10.1093/aje/kwu344. 
 
 
 
 
 
 
 
May 18, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 5 | Fred Hutchinson Cancer Research Center 
 
 
 
Image provided by Ms. Lieke Egbers 
Association of quartiles of body mass index (compared to the lowest 
quartile) with risk of T2E negative (top) or positive (bottom) prostate 
cancer. 
 


Risk of Gene-Fusion Positive Prostate Cancer is Lower Among Obese

June 15, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
CIVO: Drug Development in the Fast Lane 
June 15, 2015 
     SHL Frost 
Drug development is a slow and cumbersome process that includes assessment of new 
pharmaceutical compounds in countless pre-clinical studies and several phases of clinical trials. A 
critical stage is the transition from laboratory experiments to patient studies, whereupon the results 
all too often are disappointing in terms of therapeutic effect. The reason is as simple as it is 
challenging: cell-based experimental models are not always proper representations of human 
disease. Consequently, very few of the numerous tested anticancer drugs perform well enough to 
warrant regulatory approval, despite promising preclinical results. Huge efforts made at research 
centers and pharmaceutical companies are thus not only excruciatingly time consuming and 
expensive, but also - in most cases - futile. Meanwhile, patients in need are waiting for improved 
treatments. 
To address this fundamental problem, Dr. Jim Olson and colleagues in Fred Hutch's Clinical 
Research Division developed a device that enables more accurate and efficient testing of new 
pharmaceutical compounds, together with automated, image-based analysis. The technology 
platform, called CIVO, allows simultaneous testing of multiple drugs and drug combinations directly 
in human tumors, thereby bypassing issues with in vivo distribution, metabolism, excretion etc., 
allowing full focus on the interaction between drugs and cancer cells in an authentic biological 
microenvironment. The CIVO platform is licensed to Presage Biosciences, a Seattle-based company 
that sprung from Dr. Olson's laboratory in 2008. 
The technology and its benefits were demonstrated in the April 2015 issue of Science Translational 
Medicine. In the article, Dr. Olson and associates describe the principle of the handheld device, 
which enables localized administration of microliter volumes of up to eight different drugs per tumor, 
using an array of needles connected to drug reservoirs. The investigators showed results from 
several tumor models, including xenografted mice and canine patients, as well as four human 
lymphoma patients. 
A key factor for allowing simultaneous analysis of multiple compounds is separation between the 
infused agents, also over time. It was demonstrated that more than 96 % of the various injectates 
remained within 2 mm of their respective injection sites, at least up to 24 hours, without crossover 
between the different regions. Besides comparing various drugs and drug combinations, CIVO 
explores yet another dimension: tumor depth. During injection, the needles are pushed into the in-
June 15, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
patient tumor and then slowly pulled out as the drug solution is being infused at a constant flow rate. 
The result is uniform tracks of drugs through the z-axis of the tumor, in a column-like fashion. After 
tumor resection, samples from different depths into the tissue can be used to evaluate both the 
uniformity of response and the effect of different microenvironments on drug efficacy. For instance, 
deeper-lying tumor regions may not be sufficiently vascularized and thereby contain less oxygen. 
Will these hypoxic regions respond differently to treatments? This is one of many questions that may 
be answered using the CIVO platform. 
The investigators found that the system offered safe and efficient drug administration, with very mild 
adverse effects in human patients. To assess clinical predictive ability, comparisons were made in 
mice between microinjection-induced local tumor responses and responses after systemic drug 
delivery. CIVO-injection resulted in tumor growth inhibition that was comparable to that of systemic 
drug delivery, and the investigators successfully predicted both resistance and increased sensitivity 
in tumors of different contexts using the new platform. In addition, an unexpected sensitivity to the 
drug mafosfamide was discovered in otherwise chemotherapy-resistant lymphoma xenografts in 
mice. Importantly, this effect was not revealed using standard in vitrocell assays, underlining their 
relative inadequacy compared with in vivo screening methods. 
According to Dr. Olson, CIVO represents a major step forward towards improved cancer treatments 
by enabling evaluation of resistance or sensitivity to drugs in the context of a patient's own immune 
system. "In some cases, we can identify the specific mechanism of resistance to therapeutic agents," 
he added. 
The long term goal is clear: "To avoid treating patients with chemotherapy that will cause harm but 
provide no benefit - and to find the most effective and least toxic combinations of drugs," Dr. Olson 
said. The immediate plan is to advance the platform and broaden its range of clinical applications. 
Dr. Olson's enthusiasm is contagious: "Wouldn't it be interesting if we could accurately predict which 
patients will fail standard of care agents so that novel therapeutics could be evaluated in first or 
second line regimens instead of third or fourth?" he asked. It sure would! 
Klinghoffer RA, Bahrami SB, Hatton BA, Frazier JP, Moreno-Gonzalez A, Strand AD, Kerwin WS, 
Casalini JR, Thirstrup DJ, You S, Morris SM, Watts KL, Veiseh M, Grenley MO, Tretyak I, Dey J, 
Carleton M, Beirne E, Pedro KD, Ditzler SH, Girard EJ, Deckwerth TL, Bertout JA, Meleo KA, 
Filvaroff EH, Chopra R, Press OW, Olson JM. 2015. A technology platform to assess multiple cancer 
agents simultaneously within a patient's tumor. Sci Transl Med 7(284), 284ra58. 
June 15, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
 
Images provided by Presage Biosciences 
Presage Biosciences' CIVO patented 
arrayed microinjection drug delivery 
device (top) enables simultaneous 
assessment of multiple drugs or drug 
combinations directly in a single solid 
tumor while still in a patient's body. 
Presage analysis shows local response to 
microinjections of eight different cancer 
drugs within a canine patient's sarcoma 
tumor, as visualized by cleaved caspase 
staining in red for regions of apoptotic 
cell death due to drug response 
(bottom). Live tumor cells are shown in 
blue, the green is the injection site dye, 
and the red dots are dead tumor cells 
responding to the injected drugs. 
June 15, 2015 SCIENCE SPOTLIGHT 
 
4 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
 
 


CIVO: Drug Development in the Fast Lane

Zentner, Gabriel

June 15, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
Synaptojanin and Endophilin: Partners in 
Synaptic Transmission 
June 15, 2015 
   GE Zentner 
Endocytosis is a process by which cells uptake molecules by engulfing them with their 
membranes.  It is a critical process for all eukaryotic cells, being involved in numerous processes 
including plasma membrane homeostasis, signal transduction, nutrient uptake, and pathogen entry. 
At neuronal synapses, endocytosis is specialized to sustain efficient neurotransmission by recycling 
neurotransmitter storage compartments called synaptic vesicles. Defects in synaptic endocytosis 
can lead to severe consequences, ranging from neuronal degeneration to lethality.  While this 
indicates that vesicle recycling at synapses is critical for normal neural function, how it occurs is 
largely unknown.  Two of the major players are the synaptojanin and endophilin proteins, and it is 
thought that the synaptojanin proline-rich domain (PRD) guides synaptojanin to endocytic sites 
through high-affinity interactions with the endocytic protein endophilin's src homology 3 (SH3) 
domain. 
To investigate how synaptojanin and endophilin exert their positive effects on neurotransmission, 
postdoctoral fellow Dr. Yongming Dong and colleagues in the lab of Dr. Jihong Bai (Basic Sciences 
Division) studied mutant forms of synaptojanin in the nematode Caenorhabditis elegans.  "The PRD-
recruitment model has received a significant amount of attention over the years. However, we find 
that synaptojanin PRD is not required for synaptic activity in vivo. In fact, both the PRD and its 
binding partner (the endophilin SH3 domain) are dispensable for synaptic function," said Dr. Bai. 
The authors first tested the effect of deleting the synaptojanin PRD, proposed to mediate its 
interaction with endophilin, on synaptic transmission.  Strikingly, worms expressing PRD-deficient 
synaptojanin in the absence of wild-type synaptojanin showed no defects in synaptic vesicle 
endocytosis, neurotransmission, or behavior.  Simultaneous expression of PRD-deficient 
synaptojanin and endophilin lacking its SH3 domain in the absence of wild-type versions of either 
protein fully restored synaptic transmission, strongly arguing that the current model of synaptojanin 
PRD-endophilin SH3 interaction requires revision. 
In addition to the PRD, synaptojanin contains two catalytic domains that remove phosphates from 
phosphoinositide membrane lipids: Sac1 and a central phosphoinositide phosphatase  
 
June 15, 2015 SCIENCE SPOTLIGHT 
 
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domain.  Expression of synaptojanin carrying a mutation in the central phosphatase domain that 
abolishes its phosphatase activity completely abrogated its ability to rescue neurotransmission and 
behavioral defects in worms lacking wild-type synaptojanin.  In contrast, mutation of the Sac1 
domain had no such effect, consistent with the observation that humans with mutations in the 
synaptojanin Sac1 domain show no severe symptoms for the first few decades of life. 
While the authors found that the phosphatase activity of the Sac1 domain was dispensable for 
synaptic transmission, they wondered if it might be playing another role.  They thus expressed Sac1-
deleted synaptojanin in the absence of wild-type protein and found that this form of the protein did 
not rescue synaptic and behavioral deficits.  By analyzing synapses microscopically, they found that 
the Sac1 domain was required for targeting of synaptojanin to synapses.  To test this further, the 
authors tethered Sac1-deleted synaptojanin to endophilin.  Indeed, this bypassed the requirement 
for the Sac1 domain, rescuing the defects in behavior and neurotransmission of synaptojanin mutant 
worms.  This effect was dependent on the endophilin BAR domain. 
"Our study reveals a novel mechanism that coordinates the action of two essential proteins 
(synaptojanin and endophilin) for synaptic vesicle endocytosis, which significantly changes the 
current thinking in the field," said Dr. Bai.  "The functional connection between synaptojanin Sac1 
and endophilin BAR is highly intriguing because BAR proteins bend membranes during endocytosis. 
It suggests that the activity of synaptojanin is linked to the transient morphology of endocytic 
membranes. Together, these surprising findings revise our understanding of how synaptojanin and 
endophilin support brain activity, and have important ramifications for the field."  The roles of 
synaptojanin and endophilin also appear to extend to other tissues.  For instance, an isoform of 
synaptojanin increases cell migration and invasion in cell culture as well as metastasis of breast 
tumor xenografts in mice.  Thus, much remains to be learned about these intriguing proteins. 
 
Dong Y, Gou Y, Li Y, Liu Y, Bai J. 2015. Synaptojanin cooperates in vivo with endophilin through an 
unexpected mechanism. eLife10.7554/eLife.05660. 
 
 
 
June 15, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 6 | Fred Hutchinson Cancer Research Center 
 
 
 
Image provided by Dr. Jihong Bai 
Schematic showing cooperation between synaptojanin Sac1 and 
endophilin BAR at membranes to support synaptic vesicle recycling. 
 


Synaptojanin and Endophilin: Partners in Synaptic Transmission

Kocarnik, Jonathan M

August 17, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 8 | Fred Hutchinson Cancer Research Center 
 
Identifying barriers to disseminating genetic 
research results to Latinos 
Aug. 17, 2015 
     JM Kocarnik  
 
 
 
 
 
Family history information, often depicted in a 
pedigree (top), can provide useful information 
regarding current and future health risks for family 
members across generations (bottom). 
Image provided by Ms. Rachel Malen 
 
 
 
 
 
Increasingly, researchers working in community settings are seeking to engage the group they are 
working with in the research process.  These efforts not only help to translate the research into 
practice, but also show reciprocity with study participants by involving them in the research 
process.  Such community-based participatory research is common for work with underserved 
populations, and for research addressing health disparities and other social problems.  One 
important aspect of these research methods is seeking to understand the traditions and needs of the 
target community, so that research results can be disseminated in ways that are respectful, 
understandable, and beneficial to study participants.  A first step for any research program, then, is 
beginning a dialogue with the target population so as to identify potential barriers and 
opportunities.  In a recent report in the Journal of Community Genetics, Ms. Rachel Malen and Dr. 
Beti Thompson in the Public Health Sciences Division describe the results of a pilot study in which 
August 17, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 8 | Fred Hutchinson Cancer Research Center 
 
they interviewed Mexican-Americans living in rural Washington.  These interviews yielded insights 
into how this population perceives family health history, genetics, and disease risk.  
One area of rapidly growing interest is examining whether family history and genetic ancestry can 
partly explain observed health disparities for various diseases.  One such project is the Partnership 
for Understanding and Eliminating Disparate Outcomes (PUEDO) for Latinas, a transdisciplinary 
Center for Population Health and Health Disparities (CPHHD) at the Fred Hutch.  One of the goals of 
PUEDO is to examine the association of ancestral genetic background with breast cancer tumor 
biology and metabolic response to diet.  As this research requires discussing family health history 
and genetic information, the investigators felt it was important to first identify how this potentially 
sensitive information is understood and perceived by study participants and their communities.  
At the same time, the researchers also hoped to eventually be able to return their findings back to 
research participants, as well as to other Latinos around the state.  Said lead author Ms. Malen, 
"dissemination of research results to participants is becoming more and more common, but we were 
unsure how our Hispanic participants would receive them, especially any genetic findings.  Limited 
research has been done in this area.  Our goal was to gauge the basic understandings of family 
health history, genetics, and disease risk and how these concepts interact." 
To do so, the investigators worked with culturally-matched community health workers (promotores) 
to conduct pilot interviews among Spanish speaking Mexican-Americans living in rural agricultural 
communities in the lower Yakima Valley in Washington State.  Key themes identified from these 
interviews could be grouped into three major content areas, which provided useful insights into how 
family health history, genetics, and disease risk were perceived by these participants.   While 
participants were familiar with the concept of family health history and its relationship with disease 
risk, family discussions did not usually contain the type of specific information (age at onset, etc.) 
that would be important for clinical and public health applications.   
Participants expressed great interest in receiving genetic information about future risk of disease, as 
well as a strong desire for any health information that could help them live healthier lives.  Genetic 
literacy in this group was low, however, emphasizing the need for careful communication of results 
so that they are not misinterpreted or overhyped.  At the same time, genetic factors did not seem of 
great relevance to participants’ views of health or disease, and participants did not connect an 
increased risk of disease from family history to the concepts of biological inheritance or 
genetics.  Identifying such gaps in genetic knowledge and perceived relevance is the first step to 
addressing and overcoming these potential barriers, which will be necessary for effectively 
translating research results into interventions that can help reduce health disparities.  
August 17, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 8 | Fred Hutchinson Cancer Research Center 
 
Overall, the results of these pilot interviews highlight the importance of understanding the target 
audience before beginning the dissemination of research findings.  Said senior author Dr. 
Thompson, "this study helps us understand how Hispanics think heredity, health and genetics are 
related.  This is important information for this underserved population." Moving forward, the authors 
plan to go back to former participants and Latino community members to continue the dialogue 
around this line of research.  Said Dr. Thompson, "this work also launched a larger study examining 
the dissemination of research results to Hispanic women." 
Other Fred Hutch investigators contributing to this project were Ms. Sarah Knerr and Dr. Malia 
Fullerton. 
Citation:  
Malen R, Knerr S, Delgado F, Fullerton SM, Thompson B. 2015. Rural Mexican-Americans' 
perceptions of family health history, genetics, and disease risk: implications for disparities-focused 
research dissemination. J Community Genet. doi: 10.1007/s12687-015-0245-1 [Epub ahead of print] 
Support for this study was provided by the National Cancer Institute (NCI) and the American College 
of Medical Genetics Foundation. 
 
 


Identifying barriers to disseminating genetic research results to Latinos

In this monograph, Paul Neiman, M.D. (Member and Director Emeritus, Division of Basic Science, Fred Hutchinson Cancer Research Center), describes alternative organizing paradigms for developing research activity at biomedical research institutions.  He then contrasts the implications of each of these choices across a spectrum of elements that comprise the operating structure of the enterprise.  The philosophy and a selection of supporting policies used in developing the faculty in basic science at Fred Hutch, contrasted with a widely employed program-driven approach to both basic and applied research development in academic institutions, are used as examples that might inform decision making at such institutions

Neiman, Paul E

The biomedical research institute: a perspective on alternative paradigms for research development.

Herman, Jake

October 19, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 10 | Fred Hutchinson Cancer Research Center 
 
Picture perfect: alpha imaging confirms efficient 
targeting 
October 19, 2015 
     JA Herman 
 
 
A hematoxylin and eosin (H&E)-stained 
section of a canine lymph node 
showing the At-211 dose rate 
distribution in red/yellow, quantified 
through alpha imaging. The lymph node 
was biopsied and immediately frozen 
two hours after the radioactive injection, 
at which point the accumulation of At-
211 was high in the T cell-rich 
paracortex. 
Image provided by Dr. Tom Bäck 
 
 
Hematopoietic cell transplantation can successfully cure many hematological diseases including 
cancers. The success of this therapy depends on efficiently removing malignant cells, as well as 
suppressing the immune system of the patient. Chemotherapy and whole body gamma irradiation 
are standard conditioning regimens, yet these aggressive approaches have extensive side effects. 
Radiolabeled monoclonal antibodies are being explored as approaches to increase the specificity 
and efficacy for targeting host hematopoietic cells. Monoclonal antibodies (MAbs) targeting the cell 
surface protein CD45 are known to recognize nearly all hematopoietic cells and serve as an 
excellent targeting agent; however, to date studies have mainly paired anti-CD45-expressing MAbs 
with beta-emitting isotopes. Compared to beta emitters, alpha-emitting isotopes release higher-
energy particles with shorter range, which is likely to increase both the specificity and the efficacy of 
the treatment. Dr. Sofia Frost and colleagues from the Clinical Research Division demonstrate the 
feasibility of alpha-radioimmunotherapy in The Journal of Nuclear Medicine. 
The high energy and short range of alpha radiation is ideal for antibody-guided targeted therapies of 
hematological malignancies, however, the limited particle range makes calculation of absorbed 
radiation doses in tissues difficult. In this study, Dr. Frost and investigators overcame this challenge 
October 19, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 10 | Fred Hutchinson Cancer Research Center 
 
using ex vivo alpha imaging in a canine model. Anti-CD45-expressing MAbs were radiolabeled with 
the alpha emitter astatine-211 (
211
At) and infused into animals. Using alpha imaging and flow 
cytometry of biopsied bone marrow and lymph node samples the researchers demonstrated that this 
targeted approach was comparable to current pre-treatment approaches utilizing whole body gamma 
radiation, “ We are showing that the absorbed doses that are achieved in this model compare well 
with, or exceed, those used in standard external beam total body irradiation regimens” , said Dr. 
Frost. The high spatial resolution of the alpha images makes this techinque sensitive enough to 
efficiently reveal the specificity of 
211
At-conjugated anti-CD45 MAbs. Different regions within lymph 
nodes contain varying amounts of CD45-positive immune cells, and by alpha imaging these were 
clearly distinguished. The T cell-rich paracortex, for instance, was the region that contained the most 
211
At-anti-CD45. Thus the authors conclude that 
211
At-anti-CD45 radioimmunotherapy efficiently 
targets hematopoietic cells for transplant pre-treatment without severe toxicity, and that ex vivo 
alpha imaging enables better prediction and understanding of the therapeutic outcome. 
These initial safety and biodistribution studies will allow for test treatments of canine patients with 
spontaneous lymphoma. Excitingly, human applications are also underway, “ Clinical trials using 
alpha emitters like astatine-211 are currently being developed at Fred Hutch…and we anticipate they 
will open in the next year.”  said Dr. Frost. 
 
Frost SHL, Miller BW, Bäck TA, Santos EB, Hamlin DK, Knoblaugh SE, Frayo SL, Kenoyer AL, 
Strorb R, Press OW, Wilbur DS, Pagel JM, Sandmaier BM. 2015. Alpha imaging confirmed efficient 
targeting of CD45-positive cells after Astatine-211 (
211
At)-radioimmunotherapy for hematopoietic cell 
transplantation. J Nucl Med. 56(11):1766-73. doi: 10.2967/jnumed.115.162388. 
Funding for this research was provided by the NIH, the Giuliani Family Foundation, and a Walker 
Immunotherapy fellowship. 
 
 
 


Picture perfect: alpha imaging confirms efficient targeting.

Neves, Alexandre

January 19, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
Fbw7 Eludes the Two-Hit Hypothesis 
January 19, 2015 
    A Neves 
Ever since Alfred G. Knudson Jr. published his seminal statistical analysis of retinoblastoma, 
scientists have embraced the two-hit hypothesis, which suggested that both copies of tumor 
suppressor genes have to be mutated in order to drive cancer initiation. However, in some cases, 
tumor suppressors do not follow the two-hit rule. A recent Fred Hutch review article, co-authored by 
graduate student Ryan Davis, Staff scientist Dr. Markus Welcker and Principal Investigator Dr. Bruce 
Clurman (Human Biology Division), uses the ubiquitin ligase component and tumor suppressor Fbw7 
as a model to illustrate this point. This review concisely presents the most current research on Fbw7 
and entices us with potential therapeutic opportunities provided by cancers in which Fbw7 is 
mutated. 
Ubiquitin-mediated proteolysis has been implicated in a wide array of biological processes and 
components of the ubiquitin-proteasome system are mutated in many cancers. Large-scale 
sequencing of cancer genomes and mouse modeling experiments have shown that the F-box 
protein Fbw7 is a potent tumor suppressor. "The Fbw7 tumor suppressor is part of a ubiquitin ligase 
that degrades key oncoproteins, including c-Myc, c-Jun, Notch, and cyclin E. Fbw7 binds to 
substrates after they become phosphorylated, which results in their ubiquitylation and subsequent 
destruction by the proteasome", explained Dr. Clurman. Fbw7 is a part of the SCF
Fbw7
 ubiquitin 
ligase (see figure). Within SCF
Fbw7
, Fbw7 functions as an adaptor, providing substrate specificity to 
the SCF
Fbw7
 complex. Substrates that are phosphorylated on a motif known as the Cdc4 
phosphodegron (CPD) bind Fbw7, and three key arginine residues (R465, 479 and R505; hereafter 
Fbw7
ARG
) on Fbw7 that bind CPD are mutational hotspots in cancer. "The inactivating Fbw7 
mutations commonly found in cancers allow oncogenic substrates to accumulate and drives 
carcinogenesis," added Dr. Clurman. 
Many of the well-characterized Fbw7 oncoprotein substrates are transcription factors with 
established or emerging roles in development and tumorigenesis, which highlights Fbw7’s broad 
reach. For instance, c-Myc proteins are known to have profound effects on cancer at least in part 
through its transcriptional control of proliferation, cell growth, metabolism and protein synthesis. 
Importantly, the CPD on c-Myc is mutated in some human Burkitt’s lymphomas, highlighting a 
potential pathological consequence of impaired c-Myc degradation. The authors also point out that  
 
January 19, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
mutations in the Notch CPD are mutually exclusive with Fbw7 mutations that occur in T-cell acute 
lymphoblastic leukemia (T-ALL), which emphasizes the impact of Fbw7-dependent Notch 
degradation in T-ALL pathogenesis. 
Because the prognostic value of Fbw7 mutations had remained unclear despite extensive studies, 
the authors performed a meta-analysis (a computational study of previous research) of The Cancer 
Genome Atlas (TCGA), a widely used public database of cancer biology. This analysis confirmed a 
high prevalence of Fbw7
ARG
 mutations, particularly in cancer types where Fbw7 mutations are found 
in more than 10% of cases, such as T-ALL. Notably, the second Fbw7 allele often remained intact, 
contrasting with the classical two-hit scenario. The phenotypes of Fbw7
ARG
 mice are consistent with 
Fbw7
ARG
 acting in a dominant-negative fashion, but why there has been a strong biological selection 
for Fbw7
ARG
mutations, compared to nonsense or null mutations, is still unclear. Because Fbw7 
functions as a dimer in vivo, the authors propose that Fbw7
ARG
 dominantly inhibits the protein 
encoded by the wilde-type allele (Fbw7
WT
) and that Fbw7 
ARG
- Fbw7
WT
 dimers may have substrate-
specific effects. 
The investigators conclude the review by proposing therapeutic strategies to target the Fbw7 
pathway in cancer. The first approach would be to develop small molecule agonists to increase 
substrate affinity towards Fbw7
ARG
. Second, the oncoprotein substrates themselves could be 
targeted, and a previous study showed that pharmacologic inhibition of c-Myc in T-ALL to be a 
potentially viable strategy. Third, a synthetic lethal approach could be employed to identify pathways 
that are essential for the survival of Fbw7-defective, but not normal cells. Finally, a more general 
strategy would be to reduce, but not eliminate Fbw7 function, in such a way to eradicate cancer-
initiating cells of the hematopoietic system, while preserving Fbw7’s tumor-suppressive function in 
solid tissues. Overall, this review showed that our understanding of Fbw7’s tumor suppressive 
function has progressed tremendously over the past 5 years, through identification of substrates, 
mouse modeling, and studying the unique biological and oncogenic properties of 
Fbw7
ARG
 mutations. The network of oncogenic pathways regulated by Fbw7 offers a glimpse into 
how to target this pathway in the clinic. 
 
Davis RJ, Welcker M, Clurman BE. 2014. Tumor Suppression by the Fbw7 Ubiquitin Ligase: 
Mechanisms and Opportunities. Cancer Cell, 26(4), 455-64. 
 
 
January 19, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
 
 
 
Image provided by Mr. Ryan Davis. 
The Fbw7 protein binds both to the remainder of the SCF complex 
(Skp-1, Cullin-1, Rbx1 and an E2 enzyme) and its oncogenic 
substrates (purple ovals). This binding leads to substrate 
ubiquitylation (light blue hexagons) and degradation by the 
proteasome (purple hexagons). This degradation is impaired in many 
Fbw7-mutated cancers. 
 


Fbw7 Eludes the Two-Hit Hypothesis

Pattacini, Laura

January 19, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
A Mathematical Approach Lights up The Way to 
End Cholera Transmission 
January 19, 2015 
   L Pattacini 
In Western countries, the word cholera might sound like something belonging to the past. On the 
contrary, cholera still affects millions of people every year, with an estimated 120,000 deaths in 
2014. The most affected area is the Bay of Bengal, where Vibrio cholerae, the etiologic agent of 
cholera, is an autochthonous resident of the Ganges river delta. Although the final solution to the 
epidemic is universal access to clean water, hygiene, and sanitation, oral vaccines represent a 
temporary solution to limit cholera cases and ultimately deaths.  Although oral cholera vaccines have 
been proven safe and effective, their widespread use is difficult to justify economically, given the low 
mortality rate and incidence of the disease in most settings, even in endemic areas. However, large 
trials demonstrated that sufficient vaccine coverage leads to protection not only directly, but also 
through indirect mechanisms that benefit unvaccinated individuals. This aspect complicates attempts 
at cost-benefit analyses. Mathematical models are a powerful tool that can account for direct and 
indirect effects of vaccination campaigns. 
Dr. Dobromir Dimitrov and colleagues from the Vaccine and Infectious Disease Division (VIDD), in 
collaboration with co-investigators from the University of Florida, published a study on PLoS 
Neglected Tropical Diseases in which they present their estimates on the effects of different 
strategies for vaccination against cholera in Bangladesh. "A key feature of the model is that it 
integrates two transmission cycles", explained Dr. Dimitrov. "It allows estimating for transmission by 
contact with infected individuals (short cycle transmission) or by exposure to cholera in the 
environment (long cycle transmission). It makes this modeling approach applicable when other 
diseases such as typhoid are investigated. The current version of the model reproduces well the 
dynamics of endemic cholera from Matlab, Bangladesh. We plan to extend the analysis to non-
endemic settings in Africa using data provided by the African Cholera Surveillance Network 
(AFRICHOL). We are also working on evaluating the cost-effectiveness of different vaccination 
strategies in medium- and high-incidence states in Bangladesh to help planning for widespread 
cholera vaccination in Bangladesh and other countries." 
To identify the best strategy, the study focused on two main aspects: age of the target population 
and frequency of the vaccination campaign. Given that cholera affects primarily children, the model 
was applied to the total population, children 1-14, and children 1-4. The study showed that 
January 19, 2015 SCIENCE SPOTLIGHT 
 
2 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
vaccinating the whole population averts the greatest number and fraction of cholera cases while 
vaccinating children is more efficient measured by the number of vaccinations per cholera case 
averted. 
The study also compared the effectiveness of continuous compared to campaign vaccination 
strategies. The administration of the vaccine in a routine, continuous strategy maintains a high level 
of immunity and stops transmission of cholera at about 70% coverage. In contrast, the campaign 
strategy every five years averts a significant fraction of cholera cases but allows outbreaks due to 
waning of the immune response over time and the introduction of new susceptible individuals, 
through new births and migratory flux. 
The proposed approach, while maintaining the limits of a mathematical model, offers a solid platform 
to plan an effective and efficient strategy for vaccination. First, a once a year vaccination campaign 
would offer the same level of protection of a continuous vaccination strategy, while making its 
implementation more feasible. Second, while targeting the pre-school aged children could build upon 
pre-existing delivery programs, it would likely not suffice to eliminate cholera transmission. Finally, 
the provided data allow for an estimation of the cost to benefit ratio that could be used by policy 
makers to decide which strategy to use. 
"Mathematical modeling already plays an important role in assessing the potential impact of 
vaccination programs and other prevention interventions taking into account the existing 
epidemiological and prevention context." confirmed Dr. Dimitrov. "It is a valuable tool which allows 
interpreting the efficacy results from completed clinical trials and translating them into realistic 
projections of population level effectiveness and cost-effectiveness. As a member of the HPTN 
Modelling Center and the Center for Statistics and Quantitative Infectious Diseases, I witness the 
increasing interest in utilizing math modelling analyses at different stages of the infectious disease 
prevention process: to inform product development, to identify optimal combination prevention, to 
design and conduct of large community-based clinical trials, and help generalize results. Most 
importantly, modeling results often provoke debates among researchers, funders and policy makers 
and lead to shift in thinking about optimal resource allocation." 
In this case, we hope that the efforts made by Dr. Dimitrov and his co-investigators at the Fred Hutch 
can drive the efforts needed to eliminate cholera. 
Dimitrov DT, Troeger C, Halloran ME, Longini IM, Chao DL. 2014. Comparative effectiveness of 
different strategies of oral cholera vaccination in bangladesh: a modeling study. PLoS Negl Trop Dis, 
8(12), e3343. 
January 19, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 1 | Fred Hutchinson Cancer Research Center 
 
 
 
Image provided by Dr. Dobromir Dimitrov 
Modeling 5-year campaign and continuous vaccination strategies 
for different coverage levels. The overall effectiveness is the 
prevented fraction of cholera cases over 20 years. The vaccine was 
assumed to protect for an average of 5 years and is less effective 
among children from ages 1 to 4 years (40% efficacy) than among 
older individuals (65% efficacy). The bars represent the median 
values, the boxes represent the interquartile range and the whiskers 
cover 90% of the results from 100 simulations per scenario with 
parameters of seasonal environmental exposure sampled from 
ranges based on data from Matlab, Bangladesh. 
 


A Mathematical Approach Lights up The Way to End Cholera Transmission

February 16, 2015 SCIENCE SPOTLIGHT 
 
1 Volume 5, Issue 2 | Fred Hutchinson Cancer Research Center 
 
Nailing Down the Debate on Antibiotic Resistance 
in Post-Transplant Patients 
February 16, 2015 
     L Pattacini 
Following hematopoietic cell transplantion (HCT), the extreme neutropenia (decrease of neutrophil 
number) and the impairment of the immune response make the body more susceptible to bacterial 
infections. Gram-negative rod (GNR) bacteremias affect 5-10% of HCT patients and are associated 
with a higher mortality rate than gram-positive infections. To reduce the probability of such 
infections, the use of prophylactic antibiotics have been recommended by international guidelines. 
Among available antibiotic treatments, fluoroquinolones have been shown to reduce the rate of post-
transplant infections and mortality rates. Although these agents were thought to be less prone to 
resistance due to their unique molecular structure, excessive use in medicine and agriculture has led 
to increasing resistance throughout the world. 
Studies from investigators at the Fred Hutch Vaccine and Infectious Disease Division (VIDD) 
suggested an increase in gram-negative bacteremia events in allogeneic HCT recipients in recent 
years, raising the question of whether this increase was related to increased resistance to 
fluoroquinolones. In order to address this question, Epidemiology PhD student Arianna Miles-Jay 
from Dr. Steven Pergam's group in VIDD analyzed GNR bacteremia data from patients undergoing 
HCT at Seattle Cancer Care Alliance (SCCA). The group conducted a longitudinal retrospective 
study on data from patients that underwent HCT during an era in which levofloxacin (a member of 
the fluoroquinolone family) prophylaxis was standard practice for HCT recipients: January 2003 to 
December 2012. Results from the study were published in December in Biology of Blood and 
Marrow Transplantation. 
The study had two main purposes: first, to look at trends over time in the incidence of all GNR and 
FQ-resistant GNR bacteremia events in the first 100 days after HCT; and second, to compare the 
mortality rates following infection with FQ-sensitive versus resistant strains. "While antibiotics are a 
lifesaving necessity, especially in this high risk population, their widespread use does not come 
without consequences. Evaluating the local epidemiology of GNR infections, including antibiotic 
susceptibilities, is important for optimizing the standard of care for SCCA patients." said researcher 
Miles-Jay to introduce the reason for her interest. The study includes data from 2306 patients that 
underwent allogeneic HCT, of whom 12.1% experienced GNR bacteremia following transplantation.  
February 16, 2015 SCIENCE SPOTLIGHT 
 
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The bacteremias increased in the period of time between 2003 and 2009, and then declined in the 
following years. Although the study was not set to explain why such patterns occurred, the authors 
speculated that several infection control interventions implemented at SCCA between 2009 and 
2010 played a role in the post-2009 decrease. The incidence rate of FQ-resistant GNR followed a 
similar trend as total GNR, implying that the increased rate of GNR bacteremias were not due major 
increases in resistant strains. However, survival analyses showed that patients with a FQ resistant 
GNR bacteriemia had a significantly higher post-event cumulative mortality than patients 
experiencing a FQ-sensitive event, a finding that persisted after adjusting the analyses for various 
parameters, such as underlying illness, age at transplantation and conditioning regimen. This higher 
rate could be possibly be explained by delays in appropriate antibiotic therapy or that the bacteria 
most likely to acquire a resistant phenotype were ones known to be associated with higher mortality.  
Even though this study was a retrospective analysis, the large sample size and the fact that it was 
conducted from the center's robust long term longitudinal data make this publication an important 
step in addressing evidence-based decisions on the use of fluoroquinolones prophylaxis at the 
center. 
Arianna Miles-Jay concludes: "Since there is no evidence of overall increasing rates of 
fluoroquinolone resistance among gram-negative bacteremia events, levofloxacin is still appears to 
be a viable option for neutropenic prophylaxis for HCT recipients at the SCCA. However, the 
mortality difference we observed between FQ-resistant and sensitive infections highlights the 
importance of continued vigilance when it comes to antibiotic resistant infections." 
 
Miles-Jay A, Butler-Wu S, Rowhani-Rahbar A, Pergam SA. 2014. Incidence rate of fluoroquinolone 
resistant gram-negative rod bacteremia among allogeneic hematopoietic cell transplant patients 
during an era of levofloxacin prophylaxis. Biol Blood Marrow Transplant. S1083-8791(14):01399-8. 
 
 
 
 
 
February 16, 2015 SCIENCE SPOTLIGHT 
 
3 Volume 5, Issue 2 | Fred Hutchinson Cancer Research Center 
 
 
 
 
Image provided by Arianna Miles-Jay 
Incidence rate of first gram-negative bacteremia event by transplant 
year, 2003-2012. 
 


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