Supercooling methods for preservation of biological samples
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Publication Number
US-11564388-B2
Publication Date
2023-01-31
Expiration Date
Abstract
The present invention provides methods for cryopreserving a population of cells with improved cell viability. In some aspects, the method comprises contacting a population of cells with a peptoid polymer comprising one or more polar peptoid monomers, e.g., formulated in a cryoprotectant solution, and cooling the population of cells at a temperature of from 0° C. to about −20° C. for a time period of at least about 3 hours to produce a population of supercooled cells. The supercooling methods of the present invention provide excellent post-thaw cell survival and recovery. In certain embodiments, the population of cells is present in a tissue or an organ that is cryopreserved by performing the supercooling methods of the present invention.
Core Innovation
The invention relates to preserving a population of cells by contacting the cells with a solution comprising a peptoid polymer or a salt thereof comprising one or more polar peptoid monomers. The peptoid polymer is present in an amount sufficient to reduce or inhibit ice crystal formation at the cooling temperature.
The solution is cooled to a temperature of from 0°C to about -20°C for a time period of at least about 3 hours to produce a population of preserved cells. The solution comprising the preserved cells is unfrozen at the temperature, and certain embodiments maintain a liquid, ice-free suspension at the specified temperature.
After the cooling period, at least about 50% of the population of preserved cells survive after warming to above 0°C, including warming to 37°C. The preservation is described in connection with improved viability and post-warming recovery, including proliferation.
Claims Coverage
The claim coverage includes one independent claim directed to preserving cells by contacting cells with a peptoid polymer or salt comprising polar peptoid monomers, cooling the solution under conditions intended to keep the solution unfrozen, and achieving a defined post-warm survival threshold. Dependent features further refine cooling conditions, peptoid monomer characteristics, allowable peptoid salt forms, and the post-warm survival endpoint.
Cell preservation with polar peptoid polymer solution and controlled unfrozen cooling
A method for preserving a population of cells by contacting the population of cells with a solution comprising a peptoid polymer or a salt thereof comprising one or more polar peptoid monomers, cooling the solution to a temperature of from 0°C to about -20°C for a time period of at least about 3 hours to produce a population of preserved cells, wherein the peptoid polymer is present in an amount sufficient to reduce or inhibit ice crystal formation at the temperature, wherein the solution comprising the preserved cells is unfrozen at the temperature, and wherein at least about 50% of the population of preserved cells survive after warming to above 0°C.
At least two polar monomers with hydroxyalkyl side chains
At least 2 of the polar peptoid monomers each have a side chain containing an independently selected, optionally substituted C1-18 hydroxyalkyl group.
Cooling temperature narrowed to about -225°C to about -220°C
The temperature is within the range of about -225°C to about -220°C.
Survival after warming to 37°C
At least about 50% of the population of preserved cells survive after being warmed to 37°C.
Selected peptoid polymer salt forms
The peptoid polymer salt is selected from the group consisting of hydrochloride salt, acetate salt, sulfate salt, phosphate salt, maleate salt, citrate salt, mesylate salt, nitrate salt, tartrate salt, gluconate salt, and a combination thereof.
Liquid, ice-free suspension at the specified temperature
A population of preserved cells is kept in a liquid, ice-free suspension at the specified temperature.
The claims center on preserving cells using a peptoid polymer or salt with polar peptoid monomers to reduce or inhibit ice crystal formation while cooling to keep the solution unfrozen, with at least about 50% cell survival after warming. Dependent features further specify hydroxyalkyl-containing polar monomers, narrower cooling ranges, a survival endpoint at 37°C, enumerated peptoid salt forms, and a liquid ice-free suspension condition.
Stated Advantages
Reduces or inhibits ice crystal formation at the cooling temperature.
Maintains the solution comprising the preserved cells unfrozen at the cooling temperature.
Supports survival of at least about 50% of the preserved cell population after warming to above 0°C.
Supports at least about 50% survival after warming to 37°C.
Supports liquid, ice-free suspension of preserved cells at the specified temperature.
Documented Applications
Preserving a population of cells by contacting the cells with a solution comprising a peptoid polymer or salt comprising polar peptoid monomers and cooling the solution so it remains unfrozen.
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