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ADD (some) future directions

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Nathan Dwarshuis 2021-07-29 19:05:37 -04:00
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tex/thesis.tex
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@ -68,6 +68,7 @@
\newacronym{pbs}{PBS}{phosphate buffered saline}
\newacronym{bca}{BCA}{bicinchoninic acid assay}
\newacronym{bsa}{BSA}{bovine serum albumin}
\newacronym{hsa}{HSA}{human serum albumin}
\newacronym{stp}{STP}{streptavidin}
\newacronym{stppe}{STP-PE}{streptavidin-phycoerythrin}
\newacronym{snb}{SNB}{sulfo-nhs-biotin}
@ -3476,15 +3477,64 @@ In \cref{aim3} we determined that the \glspl{dms} expand T cells that also
performed better than beads \invivo{}. In the first experiment we performed, the
results were very clearly in favor of the \glspl{dms}. In the second experiment,
even the \gls{dms} group failed to fully control the tumor burden, but this is
not surprising given the low \ptcarp{} across all groups. Also, despite this, the
\gls{dms} group appeared to control the tumor better on average for early, mid,
and late T cell harvesting timepoints. It was not clear if this effect was due
to increased \cdp{} or overall increased fitness of the \gls{dms}-expanded T
not surprising given the low \ptcarp{} across all groups. Also, despite this,
the \gls{dms} group appeared to control the tumor better on average for early,
mid, and late T cell harvesting timepoints. It was not clear if this effect was
due to increased \cdp{} or overall increased fitness of the \gls{dms}-expanded T
cells given their higher expansion rate. The \ptmemp{} did not seem to be a
factor given that it was nearly the same in the first experiment between
\gls{dms} and bead groups despite the clear advantage seen in the \gls{dms} group.
\gls{dms} and bead groups despite the clear advantage seen in the \gls{dms}
group.
\section{future work}
\section{future directions}
There are several important next steps to perform with this work:
\subsection{Translation to GMP process}
While this work was done with translatability and \gls{qc} in mind, an important
feature that is missing from the process currently is the use of \gls{gmp}
methods and materials. The microcarriers themselves are made from
porcine-derived collagen, which itself is not \gls{gmp}-compliant due to its
non-human animal origins. However, using any other source of collagen should
work so long as the structure of the microcarriers remains relatively similar
and it has lysine groups that can react with the \gls{snb} to attach \gls{stp}
and \glspl{mab}. Obviously these would need to be tested and verified, but they
should not be insurmountable. Furthermore, the \gls{mab} binding step requires
\gls{bsa} to prevent adsorption to the non-polar polymer walls of the reaction
tubes. A human carrier protein such as \gls{hsa} could be used in its place to
eliminate the non-human animal origin material, but this could be much more
expensive. Alternatively, the use of protein could be replaced altogether by a
non-ionic detergent such as Tween-20 or Tween-80, which are already used for
commercial \gls{mab} formulations for precisely this purpose {\#}. Validating
the process with Tween would be the best next step to eliminate \gls{bsa} from
the process. The \gls{stp} and \glspl{mab} in this process were not
\gls{gmp}-grade; however, they are commonly used in clinical technology such as
dynabeads and thus the research-grade proteins used here could be easily
replaced. The \gls{snb} is a synthetic small molecule and thus does not have any
animal-origin concerns.
\subsection{Mechanistic investigation}
% why do the dms work?
% can we put anything on the dms to enhance their potency?
\subsection{Assessing performance using unhealthy donors}
All the work presented in this dissertation was performed using healthy donors.
This was mostly due to the fact that it was much easier to obtain healthy donor
cells and was much easier to control. However, it is indisputable that the most
relevant test cases of the \gls{dms} will be for unhealthy patient T cells, at
least in the case of autologous therapies. In particular, it will be interesting
to see how the \gls{dms} performs when assessed head-to-head with bead-based
expansion technology given that even in healthy donors, we observed the
\gls{dms} platform to work where the beads failed
(\cref{fig:dms_exp_fold_change}).
\subsection{translation to bioreactors}
% use dms in non-static bioreactors such as wave by first activating in a static
% environment
\onecolumn
\clearpage