POLY (BETA-AMINO ESTER) MEDIATED GENE DELIVERY TO
HUMAN ADIPOSE DERIVED MESENCHYMAL STEM CELLS (hAMSCs) &
ROUTES OF hAMSCs ADMINISTRATION FOR BRAIN TUMOR TREATMENT
by
Akhila Jyothy Denduluri
A thesis submitted to Johns Hopkins University in conformity with the
requirements for the degree of Master of Science in Engineering
Baltimore, Maryland
May, 2015
© 2015 Akhila Jyothy Denduluri
All Rights Reserved
Abstract
Glioblastoma (GB) is a severe type of brain tumor with 26,000 adult, 5,000
pediatric cases diagnosed every year. In addition to primary tumors, ~170,000 new
metastatic cancers to brain are diagnosed annually1,2. The median patient survival
remains a mere 14.6 months despite an expensive treatment combining surgical
resection, chemotherapy and radiation. There is an immediate need to develop
alternative therapies for GB that can be easily administered at an affordable cost. Stem
cells have provided an attractive platform to develop therapies in this regard due to their
inherent tropism towards inflammation or site of injury and their proliferation capacity.
Having the ability to non-virally modify stem cells to deliver therapeutic gene(s) of
interest will change the way treatment is strategized and administered for GB. The first
step towards developing this treatment involves efficiently engineering patient-derived
human adipose derived mesenchymal stem cells (hAMSCs) to express gene(s) of
interest and characterizing the modified hAMSCs. This thesis involves identifying an
optimal poly-beta-amino-ester (PBAE) polymer formulation to transfect patient-derived
hAMSCs and characterizing the migration, proliferation and stemness of hAMSCs
modified to express single or multiple genes of interest.
The thesis consists of two parts. The first half comprises of identification of a
polymer
formulation for
optimal
transfection
of
patient-derived
hAMSCs
and
characterization of nanoparticle modified hAMSCs. We were able to screen for optimal
gene expression using a library of PBAE polymers previously created in our laboratory.
We showed that primary patient-derived hAMSCs can be transfected using PBAE
nanoparticles and the gene expression can be seen even five days after transfection.
Though there is not just one single polymer formulation to obtain optimal expression, a
combination of polymer structure, polymer to DNA weights ratio and DNA dosage
resulted in transfection efficacy of up to 60%. We also observed variability in transfection
ii
efficacy among hAMSCs obtained from individual patients, suggesting that every
hAMSC culture does not result in similar gene expression for a given formulation.
However, we observed a transfection efficacy of at least 30-40% for any given patientderived hAMSC culture, with some cultures having transfection efficacy as high as 60%.
Multiple cultures of primary hAMSCs obtained from patients have been transfected to
express individual gene or a combination of genes. We observed that hAMSCs modified
to express receptor protein GFP before and after transfection maintained their migration
ability, proliferation capacity and stemness. In addition, hAMSCs expressing two genes
maintained their stemness as well. These results suggest that we have the ability to
transfect multiple plasmids within a single PBAE nanoparticle resulting in an effective
and efficient multiple gene delivery, without significant cytotoxicity.
The latter half of the thesis evaluates the routes of administration of hAMSCs in a
mouse model of glioblastoma in vivo. Virally transduced bioluminescent hAMSCs have
been administered to mice through intracardiac, nasal, intrathecal or intravenous
delivery. The distribution of hAMSCs was studied over a period of 5 days. The total flux
of the fluorescence signal for each mouse (for days 0-4) was qualitatively compared for
different routes. We observed the most diffused distribution of hAMSCs in for
intracardiac administration.
These findings indicate the PBAE polymers can be used to effectively transfect
primary patient-derived hAMSCs to express gene(s) of interest. Further, changes to
polymer structure, polymer to DNA weights ratio and DNA dosage influence the
transfection efficacy. Moreover, hAMSCs can be used as delivery vehicles in a brain
tumor model, with different routes of administration resulting in varied distribution of the
injected cells. In addition, patient-derived hAMSCs non-virally engineered to express
gene(s) of interest maintain their migration ability, proliferative capacity and stemness,
thus making hAMSCs an attractive gene delivery vehicle for brain tumor patients.
iii
Thesis Committee:
Alfredo Quinones-Hinojosa, M.D. (primary advisor, reader)
Professor, Neurological Surgery
Johns Hopkins Medicine
Jordan Green, Ph.D. (co-advisor, reader)
Associate Professor, Biomedical Engineering
Johns Hopkins University
Jennifer H. Elisseeff, Ph.D. (reader)
Professor, Biomedical Engineering
Johns Hopkins University
Honorary Readers:
Alessandro Olivi, M.D.
Professor, Neurological Surgery
Johns Hopkins Medicine
John J. Laterra, M.D.
Professor, Neurology
Johns Hopkins Medicine
Honggang Cui, Ph.D.
Assistant Professor, Materials Science and Engineering
Johns Hopkins University
iv
Acknowledgements
It is indeed a privilege to have an opportunity to express my sincerest gratitude to
so many incredible people in my life, who’ve helped me, taught me and inspired me to
lead a meaningful life as a person, and an ever so curious a life as a student. The
completion of this thesis is another new lesson learnt, and my grateful thanks to
everyone who made this possible.
The two most important people who’ve helped me with research and life, within
the lab and outside of lab are Dr. Alfredo Quiñones-Hinojosa (Dr. Q) and Dr. Jordan
Green. I thank them both for their guidance and support, and for their faith in me. It is a
great honor to work for Dr. Q, who has taught me the power of persistence, his passion
for medicine and the importance of responsibility. It’s a dream come true to work with Dr.
Green, and I cannot thank him enough for his patience, for inspiring me to be a creative
scientist and for all the incredible conversations. For providing funding for this research, I
would like to thank NIH, HHMI, MSCRF, JHU-Coulter Translational Partnership and
Microscopy and Imaging Core Module of the Wilmer Core Grant.
I especially thank Dr. Jennifer Elisseeff for taking time to review my
thesis. I would also like to thank Dr. Alessandro Olivi, Dr. John Laterra and Dr.
Honggang Cui for taking time to read this thesis. For all the suggestions, answers, and
time, I thank Samuel Bourne. I would also like to thank Dr. Hugo-Guerrero Cazares for
being nothing but patient with me, and answering all my questions, helping in
experimental design and explaining the science of logic, every day in the lab.
Collaborative work involves a lot many conversations, learning new concepts, clarifying
silly questions and having long discussions. For all these, and for their time and most
importantly, for their friendship, I thank Stephany Tzeng and Kristen Kozielski very
much. I knew nothing about cell culture and mouse models when I started at Hopkins,
and I have to thank Olindi Wijesekera for teaching me all that and helping start my thesis
v
project. I would like to thank Young Lee for being a hard-working teammate and a friend.
I also thank Dr. Kaisorn Chaichana for all his support and guidance. Scientific research
is an interdisciplinary effort, and I like to thank Dr. Sudipto Ganduly and Dr. Hao Zhang
for all their time and contributions.
Being a research student means having a chance to talk to your friends about
science and learning from each other. I couldn’t have asked for better friends and
inquisitive scientific minds than my labmates in the Q lab – Keila Alvarado, Paula
Schiapparelli, Gabi Drummond, Emily Lavell, Sagar Shah, Alejandro Ruiz-Valls, Juan
Carlos, Bhavika Patel, Sara Abbadi, Onur Kilic, Yun Feng, Vivian Gonzalez, Sean
Dangelmajer, Ernest Scalabrin, Kevin Stanko, Adela Wu, Jennifer Rios, Christopher
Smith, Roxana Magana-Madonado, Mingxin Zhu, Qian Li and friends from the Green lab
– Jayoung Kim, Ron Shmueli, Corey Bishop, Randall Meyer, David Wilson, Camilla
Zamboni.
I’ve been extremely fortunate in having friends at Hopkins and afar, who’ve been
with me through thick and thin, supported me and helped me, in more ways than I could
count. Thank you for your everlasting friendship, and omnipresent life lessons
Neeharika, Abhinit, Suboth, Padmaja, Carmen, Mithra, Hee Kyung-Baek, Sona, Minmin,
Colin, Luke, Nicole, Ani, Prathima, Spandana, Ruhban and Shiva-Spurthi.
Most importantly, I have to express my most heartfelt gratitude to my parents and
my sister, who have been so loving and supporting, and an endless source of strength. I
would like to thank my uncles, Dr. Subhash Pasumarthy and Dr. Sury Vepa and their
families, for being my family in a foreign land.
vi
“By believing passionately in something that still does not exist, we create it. The
nonexistent is whatever we have not sufficiently desired.”
-
vii
Franz Kafka
For my parents, and my sister,
viii
Table of Contents
Abstract ……………………………………………………………………………………….. ii
Acknowledgements …………………………………………………………………………. v
Table of contents ……………………………………………………………………………. ix
List of figures ………………………………………………………………………………… xi
Chapter 1: Introduction to the thesis ………………………………………………………1
1.1 Glioblastoma Multiforme ……………………………………………………………….......1
1.2 Mesenchymal Stem Cells ………………………………………………………………….2
1.3 Polymeric Gene Delivery ………………………………………………………………......3
1.4 References ……………………………………………………………………………….....5
1.5 Figures ……………………………………………………………………………………....9
Chapter 2: Specific aims ………………………………………………………………........11
2.1 Overview ……………………………………………………………………………….......11
2.2 Specific aims ……………………………………………………………………………….11
Chapter 3: Optimization of Poly (beta-amino-ester) (PBAE) nanoparticles for
effective gene expression of human adipose-derived Mesenchymal Stem Cells
(hAMSCs) …………………………………………………………………………………….. 13
3.1 Introduction ………………………………………………………………………………...13
3.2 Methods & Materials ……………………………………………………………………...14
3.3 Results & Discussion …………………………………………………………………......19
3.4 Conclusion ……………………………………………………………………………........25
3.5 References ………………………………………………………………………………...26
3.6 Figures ……………………………………………………………………………………...32
Chapter 4: Evaluating routes of administration of hAMSCs in a murine model of
glioblastoma …………………………………………………………………………………..44
4.1 Introduction …………………………………………………………………………….......44
ix
4.2 Methods & Materials …………………………………………………………….………..45
4.3 Results & Discussion …………………………………………………………….……….47
4.4 Conclusion ………………………………………………………………………….……...52
4.5 References ………………………………………………………………………….……..53
4.6 Figures ……………………………………………………………………………….…….57
Chapter 5: Conclusion & Future directions ……………………………………….…….63
Chapter 6: Curriculum Vitae.………………………………………………………….…....65
x
List of figures
Figure 1.1. Glioblastoma (GBM) is an aggressively infiltrating tumor and is
recurrent. Images of brain MRI sections (Axial view) of a glioblastoma patient. Preoperative, post-operative and most recent images are shown. The bright regions of the
MRI image represent tumor…………………………………………………………………….9
Figure 1.2. MSCs as cancer drug delivery vehicles. Transgene strategies potentiating
MSCs for tumor therapy. Stem cells can be designed to achieve different anti-tumor
effects. Image reproduced/reprinted, with copyrights license from: Shah, Khalid.
"Mesenchymal stem cells engineered for cancer therapy."Advanced drug delivery
reviews 64.8 (2012): 739-748. ……………………………………………………………….10
Figure 3.1. hAMSCs can be used to target BTICs for GBM treatment. (3.1a)Scheme
showing the brain tumor initiating cell (BTIC) population, known to be responsible for
aggressive tumor recurrence. (3.1b)Scheme showing the modification of AMSCs for
therapeutic applications………………………………………………………………………..32
Figure 3.2. Biodegrabdable PBAE polymer based gene delivery. Reprinted/Included
in this thesis with permission from: Green, Jordan J., Robert Langer, and Daniel G.
Anderson. "A combinatorial polymer library approach yields insight into nonviral gene
delivery." Accounts of chemical research 41.6 (2008): 749-759. General scheme
showing the critical steps involved in polymer-based gene delivery and expression in a
cell. ………………………………………………………………………………………………33
Figure 3.3. Monomers used to synthesize PBAE polymers. (3.3a) Figure showing the
synthesis of poly-beta-amino-ester (PBAE) polymer. Structures of the base, side chain
and end capping monomers used to create PBAE library for screening are also shown.
Reproduced/Included in this thesis, with permission from: Tzeng, Stephany Y., et al.
Biomaterials 32.23 (2011): 5402-5410. (3.3b) Gel permeation chromatography (GPC)
results of the leading polymers indicating number average (MN) and weight average
(MW) molecular weight and polydispersity (PDI) of each polymer. Reproduced/Included
in this thesis, with permission from: Mangraviti, Antonella, et al. "Polymeric Nanoparticles
for Nonviral Gene Therapy Extend Brain Tumor Survival in Vivo." ACS nano 9.2 (2015):
1236-1249. ……………………………………………………………………………………..34
Figure 3.4. Scheme showing nanoparticle formation and transfection of cells. …….….35
Figure 3.5. Scheme elaborating the process of obtaining adipose tissue from patient,
culturing hAMSCs and transfection using PBAE nanoparticles…………………….……..35
Figure 3.6. Effect of polymer structure on patient-derived hAMSCs transfection.
Patient-derived hAMSCs (1082) t = 24 hrs. after transfection. Transfection for different
xi
polymer structures is shown (for the same dose and w/w ratio). Images above show
phase contrast and images below show GFP fluorescence (merged images). ………..36
Figure 3.7. Duration of gene expression post-transfetction Bright field images (top)
and GFP fluorescence images (bottom) of GFP/PBAE nanoparticle modified patientderived hAMSCs (1082) at different time points t = 24 hrs, 3 days, 5 days posttransfection.. ………………………………………………………………………………….. 36
Figure 3.8. Effect of PBAE/DNA nanoparticle concentration on patient-derived
hAMSCs transfection. Patient-derived 1082 hAMSCs four days after transfection with
PBAE/GFP nanoparticles. Images above show phase contrast and images below show
GFP fluorescence………………………………………………………………………………37
Figure 3.9. Effect of DNA dosage on patient-derived hAMSCs transfection. Patientderived hAMSCs (1123) t = 24 hrs. after transfection. For a given PBAE polymer, 536 at
40 w/w, different doses of DNA plasmid were given to the cells. Images above show
phase contrast and images below show GFP fluorescence. ……………………………..37
Figure 3.10. Effect of polymer to DNA w/w ratio on patient-derived hAMSCs
transfection. Patient-derived hAMSCs (1082) t = 24 hrs. after transfection. PBAE
polymer 447 with DNA dosage of 600 ng/well (5 ng/uL concentration) with varying w/w
ratio was used for transfection. Images above show phase contrast and images below
show GFP fluorescence (merged images). …………………………………………………38
Figure 3.11. Graph showing the effect of PBAE polymer structure, polymer to DNA
weights ratio and DNA dosage on (3.11a) Percentage transfection of patient-derived
hAMSCs. (3.11b) Normalized Geometric Mean GFP expression. ……………………… 39
Figure 3.12. Characterizing the hAMSCs after modification with PBAE/DNA
nanoparticles. (3.12a) MTT assay was used to measure the proliferation of modified
and unmodified (control) hAMSCs (n = 4). (3.12b) Scheme showing the Boyden
Chamber migration assay. (3.12 c-d) 3D nanopattern migration assay was used to
measure the distance and speed of modified and unmodified (control) hAMSCs (n = 2).
(3.12e) 2D migration assay showing the number of cells migrated per membrane for
modified and unmodified (control) hAMSCs (n = 2). ……………………………………… 40
Figure 3.13. Flow characterization of patient-derived hAMSCs (1199) for stemness
surface markers (CD 31-/45- and CD 73+/90+/105+) for three groups (3.13a)
unmodified (control) (3.13b) PBAE/BMP4+ & GFP+ nanoparticle modified hAMSCs and
(3.13c) PBAE/CXC4+ & GFP+ nanoparticle modified hAMSCs. The unmodified and
modified hAMSCs have similar surface marker expression, indicating no significant
difference in stemness as a result of PBAE nanoparticle modification………………….. 41
xii
Figure 3.14. Flow characterization of unmodified (control) and PBAE nanoparticle
modified patient-derived hAMSCs (1082). The cells have been characterized for
stemness surface markers (CD 31-/34-/45- and CD 73+/90+/105+). The unmodified and
modified hAMSCs have similar surface marker expression, indicating no significant
difference in stemness as a result of PBAE nanoparticle modification………………….. 42
Figure 3.15. PBAE/DNA nanoparticle modification of five patient-derived hAMSCs
cultures. Graph showing the percentage of GFP expression and normalized geometric
mean t = 2 d. after transfection with GFP/PBAE nanoparticles for five different patientderived hAMSCs. ………………………………………………………………………………43
Figure 4.1. Comparison of commercially available hAMSCs distribution among
four different routes of administration over a period of 5 days. hAMSCs distribution
is qualitatively measured by the total flux (p/s) per group on a given day. (4.1a)
Intracardiac administration (4.1b) Intravenous (Tail vein) administration (4.1c) Nasal
administration 4.1d) Intrathecal (Subarachnoid) administration. Significance: p<0.05
……………………………………………………………………………………………………57
Figure 4.2. Distribution of commercially available hAMSCs in vivo in mice with
tumor after intracardiac administration. The distribution over duration of five days is
shown through small animal IVIS representative images on days 0,1,3,5……………….58
Figure 4.3. Distribution of commercially available hAMSCs in vivo in mice with
tumor after nasal administration. The distribution over duration of five days is shown
through small animal IVIS representative images on days 0,1,3,5.
……………………………………………………………………………………………………59
Figure 4.4. Distribution of commercially available hAMSCs in vivo in mice with
tumor after intravenous (tail vein) administration. The distribution over duration of
five days is shown through small animal IVIS representative images on days 0,1,3,5.
……………………………………………………………………………………………………60
Figure 4.5. Distribution of commercially available hAMSCs in vivo in mice with
tumor after intrathecal (subarachnoid) administration. The distribution over duration
of five days is shown through small animal IVIS representative images on days 0,1,3,5.
……………………………………………………………………………………………………61
Figure 4.6. Qualitative comparison of distribution of commercial hAMSCs between days
0 and 5, administered via four different routes in vivo in a murine tumor model………...62
Figure 5.1. Freshly extracted adipose tissue (F.A.T) transfection using GFP/PBAE
nanoparticles.…………………………………………………………………………………...64
xiii
Chapter 1
Introduction to thesis
1.1 Glioblastoma Multiforme
Gliomas are a collection of tumors arising from glia or their precursors within the
Central Nervous System. Clinically, gliomas are divided into four grades. Glioblastoma
(GBM) is the most aggressive of these gliomas3. The World Health Organization
classifies GBM as a grade IV, malignant neuroepithelial intracranial cancer of the central
nervous system. GBM is the most common malignant primary brain tumor in adults, with
an occurrence of 25,000 new cases and 16,000 deaths per annum. The median survival
remains at a dismal 14.6 months post-diagnosis despite combinatorial treatment strategy
consisting of surgical resection, chemotherapy and radiation. Implied in the name,
glioblastoma is a multiforme. Glioblastoma is a multiforme grossly with regions of
necrosis ad hemorrhage; microscopically with microvascular proliferation, pleomorphic
nuclei and cells; and genetically with deletions, amplifications and point mutations in
signal transduction pathways associate with angiogenesis, cell cycle checkpoint
regulation, survival and cell migration3. Glioblastoma is an aggressively diffusive in
nature (figure 1.1). The glioma cells infiltrate through the normal tissue, surround
neurons and vessels, and migrate through the white matter tracts. These finger-like
protrusions form the infiltrative patterns of glioblastoma preventing a complete surgical
resection of the tumor, and tumor recurrence3-7.
From within a population of CD 133+ cells from human glioblastoma samples,
scientists were able to identify and isolate a subpopulation of cells capable of forming
human brain tumor in vitro and in vivo, called the Brain Tumor Initiating Cells (BTICs)8.
1
Later on, several studies have shown that BTICs are resistant to radiotherapy and
chemotherapy (temozolomide) treatments9-15. Considering that a majority of the antitumor therapy for recurrent tumors comprises of chemo and radiotherapy, it is evident
that BTICs evade these therapeutic strategies and play a role in aggressive tumor
recurrence. Hence, there is a clear need to develop therapeutic strategies with the ability
to target this specific subpopulation of BTIC cells to reduce the invasion, delay the tumor
recurrence and prolong survival.
1.2 Mesenchymal Stem Cells
Mesenchymal stem cells (MSC) are a population of multi-potent self-renewing
adherent cells that have the ability to form colonies, proliferate and differentiate into
mesenchymal lineages, including osteoblasts, chondrocytes, adipocytes, fibroblasts,
myoblasts and in some cases, dopaminergic neurons and oligodendrocytes16,17. The
International Society for Cellular Therapy has provided with certain minimum criteria for
defining multipotent human mesenchymal stromal cells, which include plastic adherence
under standard culture conditions; positive for expression of CD73, CD90 and CD105,
and negative for expression of CD31, CD45, CD11a, CD19 and HLA-DR; differentiation
ability into osteocytes, adipocytes and chondrocytes in vitro under specific stimulus18.
There are many sources to isolate MSCs, with the most common being bone-marrow,
adipose tissue and umbilical cord19. However, the exact role of MSCs has not been
defined, though studies have shown that MSCs participate in formation and maintenance
of hematopoietic microenvironment, immune response control, wound healing, tissue
repair and regeneration and angiogenesis16.
Though Bone Marrow derived mesenchymal stem cells (BM-MSCs) have been
extensively studied, in comparison, adipose-derived mesenchymal stem cells (AMSCs)
have some advantages. AMSCs are genetically and morphologically more stable in long2
term culture, have a higher proliferative capacity and lower senescence ratio, and are
much easier to obtain due to the abundance of adipose tissue and easy isolation from
subcutaneous regions of the patient. Moreover, AMSCs have been shown to have an
endogenous tropism to tumors and sites of inflammation and injury, which makes them
an attractive option for therapeutic delivery vehicles19-22 AMSCs can be especially used
cases where systemic administration of drug causes unwanted, harmful side effects and
in scenarios where a certain population of cells or tissues is being targeted, including
cancer therapy. An overview of the various therapeutic molecules, agents and drugs is
shown (figure 1.2). In this thesis, we make use of patient-derived hAMSCs as delivery
vehicles for nanoparticles to deliver DNA to the brain tumor site.
1.3 Polymeric Gene Delivery
Gene therapy is a form of molecular medicine, where genetic material is
introduced into a cell, tissue or whole organ to either cure a disease or reduce the
progression of the disease. A critical factor in gene therapy is an effective delivery
system for efficient gene transfer, with minimal pathogenic or harmful side effects. There
are two main delivery vehicles or vectors used for DNA delivery: (1) viral vectors and (2)
non-viral (synthetic) vectors23. Viral vectors, though result in long term gene expression,
do not provide as much control in therapy design, in addition to potentially causing
insertional mutagenesis, adverse immune responses and a lack of specificity24.
However, non-viral synthetic vectors and delivery vehicles have not been able to match
the efficiency of viral vectors. Any synthetic delivery vehicle for DNA delivery should be
able to overcome extra- and intracellular barriers, maintain stability until nuclear import
occurs but have weak enough interactions with the DNA to release once in the nucleus,
and have the ability for endocytosis and endosomal escape25-27. Rather then developing
a single synthetic vector to modify the cells or deliver the genes, an effective approach
3
with broader applications would be to design and develop a library of polymeric vectors
and evaluate their efficacy. Such an approach is described and discussed in a study by
Green et al., where a library of over 2000 structurally unique poly (β-amino esters)
(PBAEs) were developed and tested for effective gene delivery28. In addition to being
comparable to adenovirus for in vitro and in vivo gene delivery to primary human cells,
these biodegradable PBAE polymers show polymer structure to cell type specificity29-31.
In this context, optimizing gene delivery for a certain cell type begins with a large screen
of polymeric nanoparticles, which will result in leading polymer choices for a given cell
type. Though the cause for this specificity remains unknown, it is a powerful tool in
optimizing gene expression and reducing non-specific delivery and related side effects.
In this thesis, the first half focuses on optimizing DNA delivery and gene expression of
patient-derived human Adipose-derived Mesenchymal Stem Cells (hAMSCs) using
PBAE-based nanoparticles, with a specific application in glioblatoma therapy.
4
1.4 References
1
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practice of oncology-advances in oncology. 9th edn, Vol. 1 1700-49 (Lippincott
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Hutter, A., Schwetye, K. E., Bierhals, A. J. & McKinstry, R. C. Brain neoplasms:
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3
Holland, E. C. Glioblastoma multiforme: The terminator. Proceedings of the
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(2000).
4
Stupp, R. et al. Radiotherapy plus concomitant and adjuvant temozolomide for
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Erpolat, O. P. et al. Outcome of newly diagnosed glioblastoma patients treated
by radiotherapy plus concomitant and adjuvant temozolomide: a long-term
analysis. Tumori 95, 191-197 (2009).
6
Chaichana, K. L. et al. Supratentorial glioblastoma multiforme: the role of surgical
resection versus biopsy among older patients. Annals of surgical oncology 18,
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Chaichana, K. L. et al. Establishing percent resection and residual volume
thresholds affecting survival and recurrence for patients with newly diagnosed
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8
Singh, S. K. et al. Identification of human brain tumour initiating cells. Nature 432,
396-401,
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9
Bao, S. et al. Glioma stem cells promote radioresistance by preferential
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Bleau, A. M. et al. PTEN/PI3K/Akt pathway regulates the side population
phenotype and ABCG2 activity in glioma tumor stem-like cells. Cell stem cell 4,
226-235, doi:10.1016/j.stem.2009.01.007 (2009).
11
Eramo, A. et al. Chemotherapy resistance of glioblastoma stem cells. Cell death
and differentiation 13, 1238-1241, doi:10.1038/sj.cdd.4401872 (2006).
12
Murat, A. et al. Stem cell-related "self-renewal" signature and high epidermal
growth factor receptor expression associated with resistance to concomitant
chemoradiotherapy in glioblastoma. Journal of clinical oncology : official journal
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Sakariassen, P. O., Immervoll, H. & Chekenya, M. Cancer stem cells as
mediators of treatment resistance in brain tumors: status and controversies.
Neoplasia 9, 882-892 (2007).
14
Wei,
J.
et
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Glioma-associated
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Association for Cancer Research 16, 461-473, doi:10.1158/1078-0432.CCR-091983 (2010).
15
Xie, T. X. et al. Aberrant NF-kappaB activity is critical in focal necrosis formation
of human glioblastoma by regulation of the expression of tissue factor.
International journal of oncology 33, 5-15 (2008).
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16
Strioga, M., Viswanathan, S., Darinskas, A., Slaby, O. & Michalek, J. Same or
not the same? Comparison of adipose tissue-derived versus bone marrowderived mesenchymal stem and stromal cells. Stem cells and development 21,
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17
Eftimov, P. B., Guy Wouters. "The Mesenchymal Stem Cells Therapy - New
Challenges and Opportunities. Clinical Case Report.". Animal Studies &
Veterinary Medicine III, 6-10.
18
Dominici, M. et al. Minimal criteria for defining multipotent mesenchymal stromal
cells. The International Society for Cellular Therapy position statement.
Cytotherapy 8, 315-317, doi:10.1080/14653240600855905 (2006).
19
Shah, K. Mesenchymal stem cells engineered for cancer therapy. Advanced drug
delivery reviews 64, 739-748 (2012).
20
Aboody, K. S. et al. Neural stem cells display extensive tropism for pathology in
adult brain: evidence from intracranial gliomas. Proceedings of the National
Academy of Sciences 97, 12846-12851 (2000).
21
Aboody, K. S. et al. Neural stem cell-mediated enzyme/prodrug therapy for
glioma: preclinical studies. Science translational medicine 5, 184ra159,
doi:10.1126/scitranslmed.3005365 (2013).
22
Frank, R. T., Najbauer, J. & Aboody, K. S. Concise review: stem cells as an
emerging platform for antibody therapy of cancer. Stem Cells 28, 2084-2087,
doi:10.1002/stem.513 (2010).
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8
1.5 Figures
1.1c.
Post-operative MRI
After 2 months
(10-October-2014)
Figure 1.1. Images of brain MRI sections (Axial view) of a
glioblastoma patient. Pre-operative, post-operative and most recent
images are shown. The bright regions of the MRI image represent
tumor.
1.1a.
Pre-operative MRI
1.1b.
Post-operative MRI
Immediate
9
Figure 1.2. Transgene strategies potentiating MSCs
for tumor therapy. Stem cells can be designed to
achieve
different
anti-tumor
effects.
Image
reproduced/reprinted, with copyrights license from:
Shah, Khalid. "Mesenchymal stem cells engineered for
cancer therapy."Advanced drug delivery reviews 64.8
(2012): 739-748.
10