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REF update todos and comments for myself

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Nathan Dwarshuis 2021-07-31 18:37:14 -04:00
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tex/thesis.tex
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@ -415,8 +415,6 @@ better mimic these \invivo{} expansion conditions of T cells, can significantly
improve the quality and quantity of manufactured T cells and provide better improve the quality and quantity of manufactured T cells and provide better
control on the resulting T cell phenotype. control on the resulting T cell phenotype.
% TODO mention the Cloudz stuff that's in my presentation
A variety of solutions have been proposed to make the T cell expansion process A variety of solutions have been proposed to make the T cell expansion process
more physiological. One strategy is to use modified feeder cell cultures to more physiological. One strategy is to use modified feeder cell cultures to
provide activation signals similar to those of \glspl{dc}\cite{Forget2014}. provide activation signals similar to those of \glspl{dc}\cite{Forget2014}.
@ -499,8 +497,6 @@ The specific aims of this dissertation are outlined in
\subsection*{aim 1: develop and optimize a novel T cell expansion process that \subsection*{aim 1: develop and optimize a novel T cell expansion process that
mimics key components of the lymph nodes} mimics key components of the lymph nodes}
% TODO this might be easier to break apart in separate aims
In this first aim, we demonstrated the process for manufacturing \glspl{dms}, In this first aim, we demonstrated the process for manufacturing \glspl{dms},
including quality control steps that are necessary for translation of this including quality control steps that are necessary for translation of this
platform into a scalable manufacturing setting. We also demonstrate that the platform into a scalable manufacturing setting. We also demonstrate that the
@ -540,6 +536,8 @@ present our final conclusions in Chapter~\ref{conclusions}.
\chapter{background and significance}\label{background} \chapter{background and significance}\label{background}
\section*{background} \section*{background}
% TODO mention cloudz stuff
% TODO break this apart into mfg tech and T cell phenotypes/quality % TODO break this apart into mfg tech and T cell phenotypes/quality
% TODO consider adding a separate section on microcarriers and their use in % TODO consider adding a separate section on microcarriers and their use in
% bioprocess % bioprocess
@ -1284,7 +1282,6 @@ to the microcarrier suspension (which itself is in \gls{pbs}) this result
indicated that hydrolysis is not of concern when adding \gls{snb} within indicated that hydrolysis is not of concern when adding \gls{snb} within
minutes. minutes.
% TODO use the water vs pbs curve here
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -1369,7 +1366,7 @@ the \gls{mab} reaction should proceed in {\#}{mab curve}.
\subsection{DMSs can efficiently expand T cells compared to beads} \subsection{DMSs can efficiently expand T cells compared to beads}
% TODO add other subfigures here % RESULT add other subfigures here
We next sought to determine how our \glspl{dms} could expand T cells compared to We next sought to determine how our \glspl{dms} could expand T cells compared to
state-of-the-art methods used in industry. All bead expansions were performed as state-of-the-art methods used in industry. All bead expansions were performed as
per the manufacturer’s protocol, with the exception that the starting cell per the manufacturer’s protocol, with the exception that the starting cell
@ -1384,7 +1381,7 @@ also observed no T cell expansion using \glspl{dms} coated with an isotype
control mAb compared to \glspl{dms} coated with \acd{3}/\acd{28} \glspl{mab} control mAb compared to \glspl{dms} coated with \acd{3}/\acd{28} \glspl{mab}
{Figure X}, confirming specificity of the expansion method. {Figure X}, confirming specificity of the expansion method.
% TODO make sure the day on these is correct % FIGURE make sure the day on these is correct
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -1431,7 +1428,7 @@ control mAb compared to \glspl{dms} coated with \acd{3}/\acd{28} \glspl{mab}
\input{../tables/inside_fraction_regression.tex} \input{../tables/inside_fraction_regression.tex}
\end{table} \end{table}
% TODO state the CI of what are inside the carriers % RESULT state the CI of what are inside the carriers
We also asked how many cells were inside the \glspl{dms} vs. free-floating in We also asked how many cells were inside the \glspl{dms} vs. free-floating in
suspension and/or loosely attached to the surface. We qualitatively verified the suspension and/or loosely attached to the surface. We qualitatively verified the
presence of cells inside the \glspl{dms} using a \gls{mtt} stain to opaquely presence of cells inside the \glspl{dms} using a \gls{mtt} stain to opaquely
@ -1468,8 +1465,7 @@ cells after \SI{14}{\day} were on the interior surface of the \glspl{dms}
\label{fig:dms_flowchart} \label{fig:dms_flowchart}
\end{figure*} \end{figure*}
% TODO double check the timing of this experiment (it might not be day 14) % FIGURE double check the timing of this experiment (it might not be day 14)
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -1542,9 +1538,9 @@ observing the CD4+ and CD8+ fractions of the naïve/memory subset (CD62L+CCR7+)
\label{fig:dms_exp} \label{fig:dms_exp}
\end{figure*} \end{figure*}
% TODO add a paragraph for this figure % RESULT add a paragraph for this figure
% TODO this figure has weird proportions % FIGURE this figure has weird proportions
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -1633,9 +1629,7 @@ cells (albeit in this case at day 7 and with an undefined \gls{moi})
we also found that the number of \ptcar{} T cells was greater for \gls{dms} than we also found that the number of \ptcar{} T cells was greater for \gls{dms} than
for bead (\cref{fig:car_bcma_total}). for bead (\cref{fig:car_bcma_total}).
% TODO the right half if bigger than the left half % FIGURE the right half if bigger than the left half
% TODO add memory stuff to this since I have it (it wasn't the right size so I
% haven't included it yet)?
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -1653,7 +1647,6 @@ for bead (\cref{fig:car_bcma_total}).
\label{fig:car_bcma} \label{fig:car_bcma}
\end{figure*} \end{figure*}
\subsection{DMSs efficiently expand T cells in Grex bioreactors} \subsection{DMSs efficiently expand T cells in Grex bioreactors}
% RESULT update this in light of the grex data % RESULT update this in light of the grex data
@ -1666,7 +1659,7 @@ with past results, \glspl{dms}-expanded T cells had higher \pthp{} compared to
beads, but only had slightly higher \ptmemp{} compared to beads beads, but only had slightly higher \ptmemp{} compared to beads
(\cref{fig:grex_mem,fig:grex_cd4}). (\cref{fig:grex_mem,fig:grex_cd4}).
% TODO is this discussion stuff? % DISCUSSION is this discussion stuff?
These discrepancies might be explained in light of our other data as follows. These discrepancies might be explained in light of our other data as follows.
The Grex bioreactor has higher media capacity relative to its surface area, and The Grex bioreactor has higher media capacity relative to its surface area, and
we did not move the T cells to a larger bioreactor as they grew in contrast with we did not move the T cells to a larger bioreactor as they grew in contrast with
@ -1717,8 +1710,6 @@ demonstrates that \gls{dms} could lead to more robust activation and fitness.
\label{fig:grex_luminex} \label{fig:grex_luminex}
\end{figure*} \end{figure*}
% FIGURE grex + car (maybe, IDK if I actually have this data)
\subsection{DMSs do not leave antibodies attached to cell product} \subsection{DMSs do not leave antibodies attached to cell product}
We asked if \glspl{mab} from the \glspl{dms} detached from the \gls{dms} surface We asked if \glspl{mab} from the \glspl{dms} detached from the \gls{dms} surface
@ -1760,7 +1751,7 @@ include those experiments where the surface density of the CD3 and CD28
\glspl{dms}). This ultimately resulted in a dataset with 162 runs spanning 15 \glspl{dms}). This ultimately resulted in a dataset with 162 runs spanning 15
experiments between early 2017 and early 2021. experiments between early 2017 and early 2021.
% TODO add some correlation analysis to this % FIGURE add some correlation analysis to this
Since the aim of the analysis was to perform causal inference, we determined 6 Since the aim of the analysis was to perform causal inference, we determined 6
possible treatment variables which we controlled when designing the experiments possible treatment variables which we controlled when designing the experiments
@ -1807,7 +1798,7 @@ significant in all cases except for the \dpthp{}; however, we also observe that
relatively little of the variability is explained by these simple models ($R^2$ relatively little of the variability is explained by these simple models ($R^2$
between 0.17 and 0.44). between 0.17 and 0.44).
% TODO add the regression diagnostics to this % RESULT add the regression diagnostics to this
We then included all covariates and unbalanced treatment parameters and We then included all covariates and unbalanced treatment parameters and
performed linear regression again performed linear regression again
(\cref{tab:ci_controlled,fig:metaanalysis_fx}). We observe that after (\cref{tab:ci_controlled,fig:metaanalysis_fx}). We observe that after
@ -1830,7 +1821,7 @@ using a Grex entails changing the cell surface and feeding strategy for the T
cells, and any one of these ‘mediating variables’ might actually be the cause of cells, and any one of these ‘mediating variables’ might actually be the cause of
the responses. the responses.
% TODO these tables have extra crap in them that I don't need to show % TABLE these tables have extra crap in them that I don't need to show
\begin{table}[!h] \centering \begin{table}[!h] \centering
\caption{Causal Inference on treatment variables only} \caption{Causal Inference on treatment variables only}
\label{tab:ci_treat} \label{tab:ci_treat}
@ -1868,7 +1859,7 @@ the responses.
\section{discussion} \section{discussion}
% TODO this is fluffy % DISCUSSION this is fluffy
We have developed a T cell expansion system that recapitulates key features of We have developed a T cell expansion system that recapitulates key features of
the in vivo lymph node microenvironment using DMSs functionalized with the in vivo lymph node microenvironment using DMSs functionalized with
activating mAbs. This strategy provided superior expansion with higher number of activating mAbs. This strategy provided superior expansion with higher number of
@ -1904,7 +1895,7 @@ apoptosis\cite{Yang2017}. Despite evidence for the importance of CD4 T cells,
more work is required to determine the precise ratios of CD4 and CD8 T cell more work is required to determine the precise ratios of CD4 and CD8 T cell
subsets to be included in CAR T cell therapy given a disease state. subsets to be included in CAR T cell therapy given a disease state.
% TODO this mentions the DOE which is in the next aim % DISCUSSION this mentions the DOE which is in the next aim
When analyzing all our experiments comprehensively using causal inference, we When analyzing all our experiments comprehensively using causal inference, we
found that all three of our responses were significantly increased when found that all three of our responses were significantly increased when
controlling for covariates (Figure 3, Table 2). By extension, this implies that controlling for covariates (Figure 3, Table 2). By extension, this implies that
@ -1980,7 +1971,7 @@ subtype (\ptmem{}) in this study. Future work will focus on other memory
subtypes such as tissue resident memory and stem memory T cells, as well as the subtypes such as tissue resident memory and stem memory T cells, as well as the
impact of using the DMS system on the generation of these subtypes. impact of using the DMS system on the generation of these subtypes.
% TODO this sounds sketchy % DISCUSSION this sounds sketchy
Another advantage is that the DMS system appears to induce a faster growth rate Another advantage is that the DMS system appears to induce a faster growth rate
of T cells given the same IL2 concentration compared to beads (Supplemental of T cells given the same IL2 concentration compared to beads (Supplemental
Figure 8) along with retaining naïve and memory phenotype. This has benefits in Figure 8) along with retaining naïve and memory phenotype. This has benefits in
@ -1997,7 +1988,7 @@ economically advantageous to grow as many T cells as possible in one batch in
the allogeneic model (reduced start up and harvesting costs, fewer required cell the allogeneic model (reduced start up and harvesting costs, fewer required cell
donations), the DMSs offer an advantage over current technology. donations), the DMSs offer an advantage over current technology.
% TODO this is already stated in the innovation section % DISCUSSION this is already stated in the innovation section
It should be noted that while we demonstrate a method providing superior It should be noted that while we demonstrate a method providing superior
performance compared to bead-based expansion, the cell manufacturing field would performance compared to bead-based expansion, the cell manufacturing field would
tremendously benefit from simply having an alternative to state-of-the-art tremendously benefit from simply having an alternative to state-of-the-art
@ -2006,7 +1997,7 @@ licensed accordingly; having an alternative would provide more competition in
the market, reducing costs and improving access for academic researchers and the market, reducing costs and improving access for academic researchers and
manufacturing companies. manufacturing companies.
% TODO this isn't relevent to this aim but should be said somewhere % DISCUSSION this isn't relevent to this aim but should be said somewhere
Finally, while we have demonstrated the DMS system in the context of CAR T Finally, while we have demonstrated the DMS system in the context of CAR T
cells, this method can theoretically be applied to any T cell immunotherapy cells, this method can theoretically be applied to any T cell immunotherapy
which responds to anti-CD3/CD28 mAb and cytokine stimulation. These include which responds to anti-CD3/CD28 mAb and cytokine stimulation. These include
@ -2177,7 +2168,6 @@ suggested that some of these unknown features belonged to the same molecules
(not shown). Additional multidimensional NMR experiments will be required to (not shown). Additional multidimensional NMR experiments will be required to
determine their identity. determine their identity.
% TODO add footnote saying what I did in this
\subsection{machine learning and statistical analysis} \subsection{machine learning and statistical analysis}
Linear regression analysis of the \glspl{doe} was performed as described in Linear regression analysis of the \glspl{doe} was performed as described in
@ -2308,9 +2298,7 @@ between T cells. Since \gls{il2} is secreted by activated T cells themselves,
T cells in the \gls{dms} system may need less or no \gls{il2} if this hypothesis T cells in the \gls{dms} system may need less or no \gls{il2} if this hypothesis
were true. were true.
% TODO this plots proportions look dumb % FIGURE this plots proportions look dumb
% TODO explain what the NLS lines are in b
% TODO plot the differences in lower IL2 concentrations to better show this
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -2334,7 +2322,7 @@ were true.
\label{fig:il2_mod} \label{fig:il2_mod}
\end{figure*} \end{figure*}
% TODO the nls stuff is a bit iffy % RESULT the nls stuff is a bit iffy
We varied the concentration of \gls{il2} from \SIrange{0}{100}{\IU\per\ml} and We varied the concentration of \gls{il2} from \SIrange{0}{100}{\IU\per\ml} and
expanded T cells as described in \cref{sec:tcellculture}. T cells grown with expanded T cells as described in \cref{sec:tcellculture}. T cells grown with
either method expanded robustly as \gls{il2} concentration was increased either method expanded robustly as \gls{il2} concentration was increased
@ -2361,7 +2349,7 @@ at \SI{10}{\IU\per\ml} throughout the remainder of this aim.
% different % different
\subsection{DOE shows optimal conditions for expanded potent T cells} \subsection{DOE shows optimal conditions for expanded potent T cells}
% TODO not all of these were actually use, explain why by either adding columns % RESULT not all of these were actually used, explain why by either adding columns
% or marking with an asterisk % or marking with an asterisk
\begin{table}[!h] \centering \begin{table}[!h] \centering
\caption{DOE Runs} \caption{DOE Runs}
@ -2392,7 +2380,7 @@ the range of \SIrange{10}{30}{\IU\per\ml}, \pdms{} in the range of
\SIrange{500}{2500}{\dms\per\ml}, and \pmab{} in the range of \SIrange{500}{2500}{\dms\per\ml}, and \pmab{} in the range of
\SIrange{60}{100}{\percent}. \SIrange{60}{100}{\percent}.
% TODO explain why not all runs were used % RESULT explain why not all runs were used
After performing the first \gls{doe} we augmented the original design matrix After performing the first \gls{doe} we augmented the original design matrix
with an \gls{adoe} which was built with three goals in mind. Firstly we wished with an \gls{adoe} which was built with three goals in mind. Firstly we wished
to validate the first \gls{doe} by assessing the strength and responses of each to validate the first \gls{doe} by assessing the strength and responses of each
@ -2475,7 +2463,7 @@ confidence to the location of this second order feature. The remainder of the
responses showed mostly linear relationships in all parameter cases responses showed mostly linear relationships in all parameter cases
(\cref{fig:doe_responses_cd4,fig:doe_responses_mem4,fig:doe_responses_ratio}). (\cref{fig:doe_responses_cd4,fig:doe_responses_mem4,fig:doe_responses_ratio}).
% TODO it seems arbitrary that I went straight to a third order model, the real % RESULT it seems arbitrary that I went straight to a third order model, the real
% reason is because it seemed weird that a second order model didn't find % reason is because it seemed weird that a second order model didn't find
% anything to be significant % anything to be significant
We performed linear regression on the three input parameters as well as a binary We performed linear regression on the three input parameters as well as a binary
@ -2501,7 +2489,7 @@ that our data might be underpowered for a model this complex. Further
experiments beyond what was performed here may be needed to fully describe this experiments beyond what was performed here may be needed to fully describe this
response. response.
% TODO combine these tables into one % TABLE combine these tables into one
We performed linear regression on the other three responses, all of which We performed linear regression on the other three responses, all of which
performed much better than the \ptmem{} response as expected given the much performed much better than the \ptmem{} response as expected given the much
lower apparent complexity in the response plots lower apparent complexity in the response plots
@ -2562,7 +2550,7 @@ Symbolic Regression (SR), was implemented to molecularly characterize TN+TCM
cells and to extract predictive features of quality early on their expansion cells and to extract predictive features of quality early on their expansion
process (Fig.1d-e). process (Fig.1d-e).
% TODO this table looks like crap, break it up into smaller tables % TABLE this table looks like crap, break it up into smaller tables
\begin{table}[!h] \centering \begin{table}[!h] \centering
\caption{Results for data-driven modeling} \caption{Results for data-driven modeling}
\label{tab:mod_results} \label{tab:mod_results}
@ -2656,6 +2644,8 @@ IL13 and IL15 were found predictive in combination with these using SR
% optimization of process features % optimization of process features
% TODO this sounds like total fluff % TODO this sounds like total fluff
% DISCUSSION integrate figures
CPPs modeling and understanding are critical to new product development and in CPPs modeling and understanding are critical to new product development and in
cell therapy development, it can have life-saving implications. The challenges cell therapy development, it can have life-saving implications. The challenges
for effective modeling grow with the increasing complexity of processes due to for effective modeling grow with the increasing complexity of processes due to
@ -2805,7 +2795,7 @@ interest using \glspl{mab}.
\subsection{DMSs temporal modulation} \subsection{DMSs temporal modulation}
% TODO The concentration for the surface marker cleavage experiment was much % METHOD The concentration for the surface marker cleavage experiment was much
% higher, if that matters % higher, if that matters
\glspl{dms} were digested in active T cell cultures via addition of sterile \glspl{dms} were digested in active T cell cultures via addition of sterile
\product{\gls{colb}}{Sigma}{11088807001} or \product{\gls{colb}}{Sigma}{11088807001} or
@ -2866,7 +2856,7 @@ To block soluble \gls{il15}, we supplemented analogously with
\subsection{adding or removing DMSs alters expansion and phenotype} \subsection{adding or removing DMSs alters expansion and phenotype}
% TODO state what collagenase actually targets % RESULT state what collagenase actually targets
We hypothesized that adding or removing \gls{dms} in the middle of an active We hypothesized that adding or removing \gls{dms} in the middle of an active
culture would alter the activation signal and hence the growth trajectory and culture would alter the activation signal and hence the growth trajectory and
phenotype of T cells. While adding \glspl{dms} was simple, the easiest way to phenotype of T cells. While adding \glspl{dms} was simple, the easiest way to
@ -2882,7 +2872,7 @@ the \gls{cold} group were similar to that of the buffer group, while the
enzymatic cleavage (\cref{fig:collagenase_fx}). Based on this result, we used enzymatic cleavage (\cref{fig:collagenase_fx}). Based on this result, we used
\gls{cold} moving forward. \gls{cold} moving forward.
% TODO this figure is tall and skinny like me % FIGURE this figure is tall and skinny like me
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -3015,10 +3005,6 @@ domains present on the \gls{dms} group could be creating pro-survival and
pro-expansion signals to the T cells through \gls{a2b1} and \gls{a2b2}, causing pro-expansion signals to the T cells through \gls{a2b1} and \gls{a2b2}, causing
them to grow better in the \gls{dms} system. them to grow better in the \gls{dms} system.
% TODO perhaps these figs should be combined
% TODO actually make the captions for these
% TODO add some background into why integrins are important and the proposed mechanism
% TODO add an experimental timeline to these showing when I added the mabs
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -3103,9 +3089,7 @@ is not due to signaling through \gls{a2b1} or \gls{a2b2}.
\subsection{blocking IL15 signaling does not alter expansion or phenotype} \subsection{blocking IL15 signaling does not alter expansion or phenotype}
% BACKGROUND why is IL15 important? % RESULT cite the luminex data
% TODO cite the luminex data
\gls{il15} is a cytokine responsible for memory T cell survival and maintenance. \gls{il15} is a cytokine responsible for memory T cell survival and maintenance.
Furthermore, we observed in other experiments that it is secreted to a much Furthermore, we observed in other experiments that it is secreted to a much
greater extend in \gls{dms} compared to bead cultures. One of our driving greater extend in \gls{dms} compared to bead cultures. One of our driving
@ -3115,7 +3099,6 @@ many conditions, we hypothesized that \gls{il15} may be responsible for this,
and further that the unique \textit{cis/trans} activity of \gls{il15} may be and further that the unique \textit{cis/trans} activity of \gls{il15} may be
more active in the \gls{dms} system due to higher cell density. more active in the \gls{dms} system due to higher cell density.
% TODO add some background into why IL15 is important and the proposed mechanism
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -3139,8 +3122,8 @@ more active in the \gls{dms} system due to higher cell density.
\label{fig:il15_1} \label{fig:il15_1}
\end{figure*} \end{figure*}
% TODO how did I determine how much to add? % RESULT how did I determine how much to add?
% TODO just gate these as normal because this looks sketchy % FIGURE just gate these as normal because this looks sketchy
We first tested this hypothesis by blocking \gls{il15r} with either a specific We first tested this hypothesis by blocking \gls{il15r} with either a specific
\gls{mab} or an \gls{igg} isotype control. We observed no difference in the \gls{mab} or an \gls{igg} isotype control. We observed no difference in the
expansion rate of blocked or unblocked cells (this experiment also had expansion rate of blocked or unblocked cells (this experiment also had
@ -3180,7 +3163,7 @@ We next tried blocking soluble \gls{il15} itself using either a \gls{mab} or an
change, viability, or marker histograms between any of these markers, showing change, viability, or marker histograms between any of these markers, showing
that blocking \gls{il15} led to no difference in growth or phenotype. that blocking \gls{il15} led to no difference in growth or phenotype.
% TODO this can probably be worded more specifically in terms of the cis/trans % RESULT this can probably be worded more specifically in terms of the cis/trans
% action of IL15 % action of IL15
In summary, this data did not support the hypothesis that the \gls{dms} platform In summary, this data did not support the hypothesis that the \gls{dms} platform
gains its advantages via the \gls{il15} pathway. gains its advantages via the \gls{il15} pathway.
@ -3203,7 +3186,7 @@ are included in the \ptmem{} phenotype). Taken together, these imply that
temporally or spatially altering the \gls{dms} concentration, and thus the temporally or spatially altering the \gls{dms} concentration, and thus the
activation signal, has similar effects. activation signal, has similar effects.
% TODO this sounds like background? % BACKGROUND this sounds like background?
% There are several plausible explanations for the observed phenotypic differences % There are several plausible explanations for the observed phenotypic differences
% between beads and DMSs. First, the DMSs are composed of a collagen derivative % between beads and DMSs. First, the DMSs are composed of a collagen derivative
% (gelatin); collagen has been shown to costimulate activated T cells via % (gelatin); collagen has been shown to costimulate activated T cells via
@ -3247,7 +3230,7 @@ came back negative, we would be fairly confident that the \gls{a2b1} and
We also failed to uphold our hypothesis that the \gls{dms} system gains its We also failed to uphold our hypothesis that the \gls{dms} system gains its
advantage via \gls{il15} signaling. advantage via \gls{il15} signaling.
% TODO not sure if this belongs here, although it might make sense to offer % DISCUSSION not sure if this belongs here, although it might make sense to offer
% alternative explanations of why the DMSs "work" given this negative data % alternative explanations of why the DMSs "work" given this negative data
% Second, there is evidence that providing a larger % Second, there is evidence that providing a larger
% contact area for T cell activation provides greater stimulation16,43; the DMSs % contact area for T cell activation provides greater stimulation16,43; the DMSs
@ -3296,7 +3279,7 @@ potency\cite{Ghassemi2018}.
\subsection{\invivo{} therapeutic efficacy in NSG mice model} \subsection{\invivo{} therapeutic efficacy in NSG mice model}
% TODO use actual product numbers for mice % METHOD use actual product numbers for mice
All mice in this study were male \gls{nsg} mice from Jackson Laboratories. At All mice in this study were male \gls{nsg} mice from Jackson Laboratories. At
day 0 (-7 day relative to T cell injection), 1e6 firefly luciferase-expressing day 0 (-7 day relative to T cell injection), 1e6 firefly luciferase-expressing
\product{Nalm-6 cells}{ATCC}{CRL-3273} suspended in ice-cold PBS were injected \product{Nalm-6 cells}{ATCC}{CRL-3273} suspended in ice-cold PBS were injected
@ -3403,7 +3386,7 @@ case of beads (\cref{fig:mouse_dosing_qc_mem}).
\input{../tables/mouse_dose_car.tex} \input{../tables/mouse_dose_car.tex}
\end{table} \end{table}
% TODO put growth first in this figure % FIGURE put growth first in this figure
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -3424,7 +3407,7 @@ case of beads (\cref{fig:mouse_dosing_qc_mem}).
\label{fig:mouse_dosing_qc} \label{fig:mouse_dosing_qc}
\end{figure*} \end{figure*}
% TODO explain what statistical test was used here % FIGURE explain what statistical test was used here
\begin{figure*}[ht!] \begin{figure*}[ht!]
\begingroup \begingroup
@ -3479,7 +3462,7 @@ the same.
\label{fig:mouse_timecourse_overview} \label{fig:mouse_timecourse_overview}
\end{figure*} \end{figure*}
% TODO find literature saying that CAR T cells grow slower % RESULT find literature saying that CAR T cells grow slower
As was the case with the first \invivo{} experiment, T cells activated with As was the case with the first \invivo{} experiment, T cells activated with
\glspl{dms} expanded much more efficiently compared to those expanded with beads \glspl{dms} expanded much more efficiently compared to those expanded with beads
(\cref{fig:mouse_timecourse_qc_growth}). When we quantified the \ptcarp{} of T (\cref{fig:mouse_timecourse_qc_growth}). When we quantified the \ptcarp{} of T
@ -3591,7 +3574,7 @@ results suggest that the higher proportion of memory T cells in DMS groups
efficiently kill tumor cells as recently reported in efficiently kill tumor cells as recently reported in
literature\cite{Fraietta2018, Sommermeyer2015}. literature\cite{Fraietta2018, Sommermeyer2015}.
% TODO try and find literature explaining what the ideal ratio is % DISCUSSION try and find literature explaining what the ideal ratio is
When testing \gls{car} T cells at earlier timepoints relative to day 14 as used When testing \gls{car} T cells at earlier timepoints relative to day 14 as used
in the first \invivo{} experiment, we noted that none of the \gls{car} in the first \invivo{} experiment, we noted that none of the \gls{car}
treatments seemed to work as well as they did in the first experiment. However, treatments seemed to work as well as they did in the first experiment. However,