Cell Culture Media Supplement and Method of Molecular Stress Control

Abstract

A novel class of agents has been identified to serve as cell-guard agents and/or target-specific supplements to increase cell quality and yield, as well as select for target cell populations. Laboratory experiments have demonstrated the use of cell-guard agents and/or target-specific supplements in the bioprocessing of cells as well as in selecting out a desired cell population. Several potential additive agents (both natural and synthetic) have been identified during these studies, including Vitamin D 3 , NAC, resveratrol, salubrinal, AKT, and tunicamycin (among others) that hold promise for application in cell models. In one embodiment, hypothermic stress regimes are utilized. In another embodiment, normothermic conditions are utilized while other stressors are tested in the processing. The methods of maintaining mass cell cultures and/or selecting out particular cell populations for further research and clinical use represents an important step in therapeutic discovery. The cell-guard agents are process-matched and cell-matched to protect and/or select particular cells during laboratory and clinical manipulation. The components modulate cell stress and survival pathways.

1 . A method of modulating a molecular stress pathway to improve bioprocessing comprising: exposing a cell population to a stressful condition or environment which initiates a stress response within a cell population, inhibiting the stress response using a target-specific supplement, and then allowing the cell population to recover. 
     
     
         2 . The method of  claim 1 , further comprising a step of selecting a cell population while utilizing the stress response of the cell population in combination with the target-specific supplement. 
     
     
         3 . The method of  claim 2 , wherein the step of selecting a cell population comprises selecting stem cells. 
     
     
         4 . The method of  claim 1 , wherein the stress response is a stress-induced pathway including a UPR pathway, apoptotic pathway, or free-radical scavenging system. 
     
     
         5 . The method of  claim 1 , wherein the cell population comprises a eukaryotic cell system. 
     
     
         6 . The method of  claim 5 , wherein the eukaryotic cell system includes human cells, other mammalian cells, or a malignant form thereof in vitro or in vivo. 
     
     
         7 . The method of  claim 1 , wherein the stress response is a stress-induced pathway associated with apoptosis including membrane-mediated, mitochondrial mediated, or nuclear mediated pathways. 
     
     
         8 . The method of  claim 1 , wherein the method of modulating the stress response results in increased metabolic activity, membrane integrity, cellular attachment, cell function, or overall sample viability. 
     
     
         9 . The method of  claim 1 , wherein the target-specific supplement is salubrinal, resveratrol, free radical scavengers, caspase inhibitors, UPR inhibitors, apoptotic inhibitors, Vitamins D 3 , A, C, E, or B, or other derivatives thereof. 
     
     
         10 . The method of  claim 1 , wherein the method of modulating is based on a stress regime which is mechanical, thermal, hypoxic, or based on other stressed normothermic conditions and targets optimal cell survival. 
     
     
         11 . The method of  claim 1 , wherein the stress response is cell-type specific to enable stress based cell selection such that a molecular stress pathway is manipulated to selectively eliminate or selectively protect a targeted cell population. 
     
     
         12 . The method of  claim 1 , wherein the target-specific supplement modulates a cell response to the physical environment surrounding the cell population to reduce mitochondrial stress. 
     
     
         13 . The method of  claim 1 , wherein the target-specific supplement is a caspase inhibitor, salubrinal, tunicamycin, AKT, or ROCK. 
     
     
         14 . The method of  claim 1 , wherein the target-specific supplement is an antioxidant comprising one or more of resveratrol, free radical scavengers, NAC, NO, glutathione, DMSO, ubiquinol, oxalic acid, tannins, and phytic acid. 
     
     
         15 . The method of  claim 1 , wherein the target-specific supplement is a vitamin, protein, antibody, or nucleotide sequence including but not limited to DNA, RNA, RNAi, siRNA, and SNIPS. 
     
     
         16 . The method of  claim 1 , wherein the target-specific supplement comprises natural and synthetic reagents that reduce a cell stress response and promote cell survival. 
     
     
         17 . The method of  claim 1 , wherein the cell population is utilized in cell culture, bioprocessing, cell therapy, drug discovery, molecular characterization, basic research, medical diagnosis, pathology, treatment screening, regenerative medicine, cancer therapy, or other various therapeutic technologies. 
     
     
         18 . A process for controlling a molecular pathway comprising the steps of activating a response pathway in a cell population and modulating a cell stress response using a media supplement of UPR inhibitors or antioxidants to prevent spontaneous differentiation and inhibit cell death. 
     
     
         19 . The process of  claim 18 , further comprising a step of selecting stem cells including iPSCs or hESCs to purify the cell population prior to stem cell therapy. 
     
     
         20 . The process of  claim 18 , further comprising a step of up-regulating ER-sensors during a recovery phase to provide a more effective molecular repair response in the cell population. 
     
     
         21 . A composition for protecting and sustaining biological material during bioprocessing or cell selection, said composition comprising natural or synthetic components, wherein said composition modulates cell signaling pathways and sustains cell membrane integrity using selective antioxidants or inhibitors of stress-induced cell death. 
     
     
         22 . The composition of  claim 21 , wherein the biological material is selected from the group comprising cells, cell aggregates, tissues, organs, natural liposomes and synthetic liposomes.