In vitro neonatal biomimetic (nMIMIC) model and methods of using same
Inventors
GOMES, Evan • Kamala, Tirumalai • Mosquera, Luis • Wittman, Vaughan • Warren, William • Moser, Janice • Drake, Donald
Assignees
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Publication Number
US-11156601-B2
Publication Date
2021-10-26
Expiration Date
Abstract
In vitro biomimetic models of the neonatal immune system are provided along with methods of using the models in pre-clinical assessment of infant immune cell-mediated and humoral responses to immunogenic stimulation, such as vaccination. The models include one comprising cord blood-derived T follicular helper cells and B cells, and one comprising cord blood-derived dendritic cells and CD4+ T cells. The models can be used, for example, to assess candidate vaccines via analysis of cellular responses to antigen and vaccine exposure.
Core Innovation
The invention provides neonatal modular immune in vitro constructs (nMIMIC) for assessing early-life immune responses to neonatal vaccines. The constructs include a first in vitro cell culture comprising cord blood CD4+ T cells maintained under conditions promoting development of vaccine-specific T follicular helper (T_FH) cells, with neonatal vaccine and one or more cellular activators including an anti-CD28 antibody and cytokines selected from IL-2, IL-6, IL-12, IL-21, and IL-23.
Irradiated feeder cells and additional vaccine are added after initial vaccine addition at defined day windows to support the development of vaccine-specific T_FH cells. A second in vitro cell culture is formed that comprises the developed T_FH cells and cord blood B cells, with the ratio of T_FH cells to cord blood B cells ranging from about 1:1 to about 1:100.
In some embodiments, the second cell culture comprises autologous cord blood B cells in a three-dimensional matrix, including a gel matrix. A vaccine-specific antigen is added and the culture is maintained for conditions promoting a cellular response, with IL-21 added upon addition of the antigen and at defined day windows after preparation of the second cell culture.
Cellular response readouts are obtained by analyzing supernatant and/or cells from the antigen-stimulated culture to assess outcomes. The response includes production of an antibody with binding specificity for the vaccine-specific antigen by the B cells, thereby screening neonatal vaccines for activity and assessing efficacy, and detection of a vaccine-specific cellular response and/or an antigen-binding antibody is used to predict in vivo efficacy.
Claims Coverage
The independent claims include 5 inventive claim scopes: screening for activity, assessing efficacy, and predicting in vivo efficacy. Across the independent claims, the inventive features consistently include induction of vaccine-specific T_FH cells from cord blood CD4+ T cells using defined cellular activators and staged addition of irradiated feeder cells and additional vaccine, followed by co-culture with cord blood B cells at a defined T_FH-to-B ratio and downstream antigen restimulation with defined readouts.
T_Fh induction from cord blood CD4+ T cells with defined cellular activators
Adding a neonatal vaccine and one or more cellular activator to a first in vitro cell culture comprising cord blood CD4+ T cells and maintaining the cell culture under conditions promoting development of vaccine-specific T follicular helper (T_FH) cells, wherein the cellular activators are one or more of an anti-CD28 antibody, IL-2, IL-6, IL-12, IL-21, and IL-23.
Staged addition of irradiated feeder cells and additional vaccine during T_Fh development
Adding irradiated feeder cells and additional vaccine on one of day 2, 3 or 4 after the initial vaccine addition, and on one of day 5, 6 or 7 after the initial vaccine addition.
Co-culture of T_Fh cells with cord blood B cells at defined T_Fh-to-B ratio
Forming a second in vitro cell culture comprising T_FH cells developed in the first culture and cord blood B cells, wherein the ratio of T_FH cells to cord blood B cells in the second cell culture ranges from about 1:1 to about 1:100.
Three-dimensional matrix co-culture with autologous cord blood B cells
Forming a second cell culture comprising T_FH cells developed in the first culture and autologous cord blood B cells in a three-dimensional matrix, wherein the ratio of T_FH cells to cord blood B cells in the second cell culture ranges from about 1:1 to about 1:100.
Antigen restimulation with IL-21 added upon antigen addition and during defined windows
Adding a vaccine-specific antigen to the cell culture and maintaining the culture under conditions promoting a cellular response, wherein IL-21 is added to the cell culture upon addition of the antigen and at defined day windows after preparation of the second cell culture.
Screening by analyzing supernatant and/or cells for a cellular response and antigen-binding antibody production
Analyzing supernatant and/or cells from the cell culture for a cellular response thereby screening a neonatal vaccine for activity, wherein production of an antibody with binding specificity for the vaccine-specific antigen by the B cells is used to screen neonatal vaccine for activity.
Assessing neonatal vaccine efficacy using the same in vitro workflow and cellular response readouts
Analyzing supernatant and/or cells from the cell culture for a cellular response, thereby assessing efficacy of a neonatal vaccine.
Predicting in vivo efficacy from detection of a vaccine-specific cellular response or antigen-binding antibody
Analyzing supernatant and/or cells from the cell culture for a cellular response, wherein when a cellular response is found the neonatal vaccine is predicted to have in vivo efficacy, or analyzing for production of an antibody with binding specificity for the vaccine-specific antigen by the B cells, wherein when such antibody is found the neonatal vaccine is predicted to have in vivo efficacy.
Across the independent claims, the core claim coverage centers on an in vitro neonatal vaccine testing workflow that induces vaccine-specific T_FH cells from cord blood CD4+ T cells using specified cellular activators, applies staged irradiated feeder cells and additional vaccine during T_FH development, co-cultures T_FH cells with cord blood B cells at a defined T_FH-to-B ratio, and then restimulates with a vaccine-specific antigen with IL-21 added at defined time windows. The resulting supernatant and/or cells are analyzed to screen for activity, assess efficacy, and predict in vivo efficacy using cellular responses and/or antigen-binding antibody production by B cells.
Stated Advantages
Documented Applications
Screening a neonatal vaccine for activity using cord blood-based in vitro cell culture to produce and analyze vaccine-specific cellular responses and/or antibody binding specificity.
Assessing efficacy of a neonatal vaccine using the described in vitro workflow and analyzing supernatant and/or cells for a cellular response.
Predicting in vivo efficacy of a neonatal vaccine using the described in vitro workflow, where detection of a cellular response and/or antigen-binding antibody indicates predicted in vivo efficacy.
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