Cell engineering platform
The current subject matter provides a cell engineering platform of a diagnostic and clinical use scale for vector-free and/or viral delivery of payload/cargo compounds and compositions into non-adherent cells. The platform achieves delivery to a large number of cells quickly in a closed system. Related apparatus, systems, techniques, articles and compositions are also described.
1. A system comprising:
a housing configured to receive a filter plate comprising a well;
a differential pressure applicator configured to apply a differential pressure to the well;
a delivery solution applicator configured to deliver atomized delivery solution to the well;
a stop solution applicator configured to deliver a stop solution to the well; and
a culture medium applicator configured to deliver a culture medium to the well.
2. The system of claim 1 , wherein the housing comprises:
a chamber;
a lid disposed at a first end of the chamber; and
a base disposed at a second end of the chamber,
wherein the filter plate is disposed within the chamber.
3. The system of claim 2 , wherein the differential pressure applicator is connected to the lid of the housing and includes a shower head, which includes a plurality of apertures at an end portion thereof.
4. The system of claim 2 , wherein the stop solution applicator delivers the stop solution to the well through a septum provided in the lid of the housing.
5. The system of claim 2 , wherein the culture medium applicator delivers the culture medium to the well through a septum provided in the lid of the housing or through a port of the chamber.
6. The system of claim 1 , wherein the housing is configured to tilt the filter plate.
7. The system of claim 1 , wherein the delivery solution applicator includes a robotic arm and spray head, the robotic arm magnetically coupled to the spray head.
8. The system of claim 7 , further comprising a soft elastomeric barrier enclosing the filter plate and spray head, wherein the soft elastomeric barrier separates the robotic arm and the spray head.
9. The system of claim 7 , wherein the robotic arm is configured to carry the spray head to a plurality of locations on the filter plate.
10. The system of claim 1 , wherein the stop solution applicator and the culture medium applicator are formed integral within the housing as ports, and wherein the housing forms a vessel.
11. The system of claim 10 , wherein the vessel includes a waste media outlet for collecting waste media.
12. The system of claim 1 , further comprising a soft elastomeric barrier enclosing the filter plate and delivery solution applicator within the housing, the soft elastomeric barrier and the housing forming a bioreactor.
13. The system of claim 1 , wherein the housing includes a filter plate base configured to tilt, rotate, and/or vibrate the filter plate.
14. The system of claim 1 , wherein the delivery solution applicator includes a robotic arm and spray head, and the spray head is a single-use cartridge.
15. The system of claim 1 , wherein the filter plate is sized to hold greater than 1×10 7 T cells.
16. The system of claim 1 , wherein the system is configured to automatically:
provide to the filter plate cells in media;
remove culture media to form a cellular monolayer on top of the filter plate;
apply an atomized delivery solution to the cellular monolayer;
incubate the cells,
apply a stop solution to the incubated cellular monolayer; and
provide new media to the cellular monolayer.
17. The system of claim 16 , wherein the system is configured to automatically tilt, vibrate, and/or rotate the filter plate to re-suspend the cells in the new media.
18. The system of claim 16 , wherein the system is configured to repeat the application of the atomized delivery solution, the incubation, and the application of the stop solution.
19. The system of claim 1 , wherein the delivery solution applicator includes a nebulizer.
20. The system of claim 19 , wherein the delivery solution applicator further includes a mass flow controller or a volumetric flow controller to regulate a gas flow to operate the nebulizer.
21. The system of claim 1 , wherein the delivery solution applicator is configured to deliver 10-300 micro liters of the delivery solution per actuation.
22. The system of claim 1 , further comprising a temperature control system configured to control a temperature of the delivery solution and/or of the plate comprising the well.
23. The system of claim 1 , wherein the delivery solution includes an aqueous solution, the aqueous solution including the payload and an alcohol at greater than 2 percent (v/v) concentration.
24. The system of claim 23 , wherein said alcohol comprises ethanol.
25. The system of claim 23 , wherein said aqueous solution comprises greater than 5% ethanol.
26. The system of claim 23 , wherein said aqueous solution comprises between 5-30% ethanol.
27. The system of claim 23 , wherein said aqueous solution comprises 12% or 25% ethanol.
28. The system of claim 23 , wherein said aqueous solution comprises between 12.5-500 mM KCl.
29. The system of claim 23 , wherein said aqueous solution comprises 106 mM KCl.
30. The system of claim 1 , further comprising:
the filter plate, wherein the well is configured to contain a population of non-adherent cells.
31. The system of claim 30 , wherein said non-adherent cell comprises a peripheral blood mononuclear cell.
32. The system of claim 30 , wherein said non-adherent cell comprises an immune cell.
33. The system of claim 30 , wherein said non-adherent cell comprises a T lymphocyte.
34. The system of claim 23 , wherein said payload comprises a messenger ribonucleic acid (mRNA).
35. The system of claim 34 , wherein said mRNA encodes a gene-editing composition.
36. The system of claim 35 , wherein said gene editing composition reduces the expression of PD-1.
37. The system of claim 34 , wherein said mRNA encodes a chimeric antigen receptor.
38. The system of claim 1 for use to deliver a cargo compound or composition to a mammalian cell.
39. The system of claim 30 , wherein said population of non-adherent cells comprises a monolayer.