ADD (some) future directions
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@ -68,6 +68,7 @@
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\newacronym{pbs}{PBS}{phosphate buffered saline}
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\newacronym{pbs}{PBS}{phosphate buffered saline}
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\newacronym{bca}{BCA}{bicinchoninic acid assay}
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\newacronym{bca}{BCA}{bicinchoninic acid assay}
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\newacronym{bsa}{BSA}{bovine serum albumin}
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\newacronym{bsa}{BSA}{bovine serum albumin}
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\newacronym{hsa}{HSA}{human serum albumin}
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\newacronym{stp}{STP}{streptavidin}
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\newacronym{stp}{STP}{streptavidin}
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\newacronym{stppe}{STP-PE}{streptavidin-phycoerythrin}
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\newacronym{stppe}{STP-PE}{streptavidin-phycoerythrin}
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\newacronym{snb}{SNB}{sulfo-nhs-biotin}
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\newacronym{snb}{SNB}{sulfo-nhs-biotin}
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@ -3476,15 +3477,64 @@ In \cref{aim3} we determined that the \glspl{dms} expand T cells that also
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performed better than beads \invivo{}. In the first experiment we performed, the
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performed better than beads \invivo{}. In the first experiment we performed, the
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results were very clearly in favor of the \glspl{dms}. In the second experiment,
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results were very clearly in favor of the \glspl{dms}. In the second experiment,
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even the \gls{dms} group failed to fully control the tumor burden, but this is
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even the \gls{dms} group failed to fully control the tumor burden, but this is
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not surprising given the low \ptcarp{} across all groups. Also, despite this, the
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not surprising given the low \ptcarp{} across all groups. Also, despite this,
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\gls{dms} group appeared to control the tumor better on average for early, mid,
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the \gls{dms} group appeared to control the tumor better on average for early,
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and late T cell harvesting timepoints. It was not clear if this effect was due
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mid, and late T cell harvesting timepoints. It was not clear if this effect was
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to increased \cdp{} or overall increased fitness of the \gls{dms}-expanded T
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due to increased \cdp{} or overall increased fitness of the \gls{dms}-expanded T
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cells given their higher expansion rate. The \ptmemp{} did not seem to be a
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cells given their higher expansion rate. The \ptmemp{} did not seem to be a
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factor given that it was nearly the same in the first experiment between
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factor given that it was nearly the same in the first experiment between
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\gls{dms} and bead groups despite the clear advantage seen in the \gls{dms} group.
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\gls{dms} and bead groups despite the clear advantage seen in the \gls{dms}
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group.
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\section{future work}
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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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\subsection{Translation to GMP process}
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While this work was done with translatability and \gls{qc} in mind, an important
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feature that is missing from the process currently is the use of \gls{gmp}
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methods and materials. The microcarriers themselves are made from
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porcine-derived collagen, which itself is not \gls{gmp}-compliant due to its
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non-human animal origins. However, using any other source of collagen should
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work so long as the structure of the microcarriers remains relatively similar
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and it has lysine groups that can react with the \gls{snb} to attach \gls{stp}
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and \glspl{mab}. Obviously these would need to be tested and verified, but they
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should not be insurmountable. Furthermore, the \gls{mab} binding step requires
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\gls{bsa} to prevent adsorption to the non-polar polymer walls of the reaction
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tubes. A human carrier protein such as \gls{hsa} could be used in its place to
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eliminate the non-human animal origin material, but this could be much more
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expensive. Alternatively, the use of protein could be replaced altogether by a
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non-ionic detergent such as Tween-20 or Tween-80, which are already used for
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commercial \gls{mab} formulations for precisely this purpose {\#}. Validating
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the process with Tween would be the best next step to eliminate \gls{bsa} from
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the process. The \gls{stp} and \glspl{mab} in this process were not
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\gls{gmp}-grade; however, they are commonly used in clinical technology such as
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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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animal-origin concerns.
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\subsection{Mechanistic investigation}
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% why do the dms work?
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% can we put anything on the dms to enhance their potency?
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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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This was mostly due to the fact that it was much easier to obtain healthy donor
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cells and was much easier to control. However, it is indisputable that the most
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relevant test cases of the \gls{dms} will be for unhealthy patient T cells, at
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least in the case of autologous therapies. In particular, it will be interesting
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to see how the \gls{dms} performs when assessed head-to-head with bead-based
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expansion technology given that even in healthy donors, we observed the
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\gls{dms} platform to work where the beads failed
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(\cref{fig:dms_exp_fold_change}).
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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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% environment
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\onecolumn
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\onecolumn
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\clearpage
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\clearpage
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