ADD a bunch of future work fluff
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@ -2583,6 +2583,19 @@ CONCLUSIONS: We developed a simplified, semi-closed system for the initial selec
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publisher = {The American Association of Immunologists},
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publisher = {The American Association of Immunologists},
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}
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}
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@Article{Stephan2014,
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author = {Sirkka B Stephan and Alexandria M Taber and Ilona Jileaeva and Ericka P Pegues and Charles L Sentman and Matthias T Stephan},
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journal = {Nature Biotechnology},
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title = {Biopolymer implants enhance the efficacy of adoptive T-cell therapy},
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year = {2014},
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month = {dec},
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number = {1},
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pages = {97--101},
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volume = {33},
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doi = {10.1038/nbt.3104},
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publisher = {Springer Science and Business Media {LLC}},
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}
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@Comment{jabref-meta: databaseType:bibtex;}
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@Comment{jabref-meta: databaseType:bibtex;}
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@Comment{jabref-meta: grouping:
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@Comment{jabref-meta: grouping:
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@ -163,6 +163,7 @@
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\newacronym{qbd}{QbD}{quality-by-design}
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\newacronym{qbd}{QbD}{quality-by-design}
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\newacronym{aws}{AWS}{amazon web services}
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\newacronym{aws}{AWS}{amazon web services}
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\newacronym{qpcr}{qPCR}{quantitative polymerase chain reaction}
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\newacronym{qpcr}{qPCR}{quantitative polymerase chain reaction}
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\newacronym{cstr}{CSTR}{continuously stirred tank bioreactor}
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%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
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%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
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% SI units for uber nerds
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% SI units for uber nerds
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@ -733,7 +734,7 @@ much higher surface area than a traditional flask when matched for volume.
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Consequently, this means that microcarrier-based cultures can operate with much
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Consequently, this means that microcarrier-based cultures can operate with much
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lower footprints than flask-like systems. Microcarriers also allow cell culture
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lower footprints than flask-like systems. Microcarriers also allow cell culture
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to operate more like a traditional chemical engineering process, wherein a
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to operate more like a traditional chemical engineering process, wherein a
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stirred tank bioreactor may be employed to enhance oxygen transfer, maintain pH,
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\gls{cstr} may be employed to enhance oxygen transfer, maintain pH,
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and continuously supply nutrients\cite{Derakhti2019}.
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and continuously supply nutrients\cite{Derakhti2019}.
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A variety of microcarriers have been designed, primarily differing in their
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A variety of microcarriers have been designed, primarily differing in their
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@ -4315,7 +4316,8 @@ group.
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\section{future directions}
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\section{future directions}
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There are several important next steps to perform with this work:
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There are several important next steps to perform with this work, many of which
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will be relevent to using this technology in a clinical trial:
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\subsection{Translation to GMP process}
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\subsection{Translation to GMP process}
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@ -4341,12 +4343,33 @@ as dynabeads and thus the research-grade proteins used here could be easily
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replaced. The \gls{snb} is a synthetic small molecule and thus does not have any
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replaced. The \gls{snb} is a synthetic small molecule and thus does not have any
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animal-origin concerns.
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animal-origin concerns.
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\subsection{Mechanistic investigation}
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\subsection{mechanistic investigation}
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% why do the dms work?
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Despite the improved outcomes in terms of expansion and phenotype relative to
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% can we put anything on the dms to enhance their potency?
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beads, we don't have a good understanding of why they \gls{dms} platform works
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as well as it does. Several broad areas remain to be investigated, including the
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role of the increased cytokine output (including \il{15} which was explored to
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some extent in this work), the role of cells on the interior of the \gls{dms}
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relative to those outside the \gls{dms}, and the role of the physical surface
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properties of the \gls{dms} (including the morphology and the stiffness).
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\subsection{Assessing performance using unhealthy donors}
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\subsection{additional ligands and signals on the DMSs}
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In this work we only explored the use of \acd{3} and \acd{28} \glspl{mab} coated
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on the surface of the \gls{dms}. The chemistry used for the \gls{dms} is very
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general, and any molecule or protein that could be engineered with a biotin
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ligand could be attached without any further modification. There are many other
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ligands that could have profound effects on the expansion and quality of T cells
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which may be utilized. The simplest next step is to simply vary the ratio of
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\acd{3} and \acd{28} signal. Another obvious example is to attach
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\il{15}/\il{15R$\upalpha$} complexes to the surface to mimic \textit{trans}
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presentation from other cell types\cite{Stonier2010}. Other adhesion ligands or
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peptides such as GFOGER could be used to stimulate T cells and provide more
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motility on the \glspl{dms}\cite{Stephan2014}. Finally, viral delivery systems
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could theoretically be attached to the \gls{dms}, greatly simplifying the
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transduction step.
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\subsection{assessing performance using unhealthy donors}
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All the work presented in this dissertation was performed using healthy donors.
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All the work presented in this dissertation was performed using healthy donors.
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This was mostly due to the fact that it was much easier to obtain healthy donor
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This was mostly due to the fact that it was much easier to obtain healthy donor
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@ -4360,8 +4383,23 @@ expansion technology given that even in healthy donors, we observed the
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\subsection{translation to bioreactors}
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\subsection{translation to bioreactors}
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% use dms in non-static bioreactors such as wave by first activating in a static
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In this work we performed some preliminary experiments demonstrating that the
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% environment
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\gls{dms} platform can work in a Grex bioreactor. While an important first step,
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more work needs to be done to optimize how this system will or can work in a
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scalable environment using bioreactors. There are several paths to explore.
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Firstly, the Grex itself has additional automation accessories which could be
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tested, which would allow continuous media exchange and cytokine
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administration. While this is an improvement from the work done here, it is
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still a Grex and has all the disadvantages of an open system. Secondly, other
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static bioreactors such as the Quantum hollow fiber bioreactor (Terumo) could be
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explored. Essentially the \gls{dms} would be an additional matrix that could be
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supplied to this system which would enhance its compatibility with T cells.
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Finally, suspension bioreactors such as the classic \gls{cstr} or WAVE
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bioreactors could be tried. The caveat with these is that the T cells only seem
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to be loosely attached to the \gls{dms} throughout culture, so an initial
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activation/transduction step in static culture might be necessary before moving
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to a suspension system (alternatively the \gls{dms} could be coated with
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additional adhesion ligands to make the T cells attach more strongly).
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\onecolumn
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\onecolumn
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\clearpage
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\clearpage
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