065 Designing potent inhibitors of Ice recrystallization - from C-linked antifreeze glycoproteins (AFGPs) to small molecules

Abstract

Cell-based therapeutics has emerged as a critical aspect of modern health care and has shaped new models of care in stem cell and regenerative therapy. Dimethyl sulfoxide and glycerol are the two most common cryoprotectants. Unfortunately, many of the current cryopreservation protocols are suboptimal resulting in poor cell recoveries and/or compromised cellular function post cryopreservation thus limiting applications. One of the most significant causes of decreased cell viability and impaired function during cryopreservation is direct mechanical damage from ice crystals. Preventing this damage using ice recrystallization inhibitors may greatly reduce cellular injury during cryopreservation and enhance the recovery of specialized cellular functions. Antifreeze glycoproteins (AFGPs) are peptide-based structures found in many deep-sea Teleost fish inhabiting sub-zero environments. These compounds prevent the seeding of ice crystals in vivo and ultimately protect these organisms against cryoinjury and death. For the past decade our laboratory has been actively pursuing the rational design and synthesis of functional carbon-linked (C-linked) AFGP analogues possessing custom-tailored antifreeze activity. These compounds are potent inhibitors of ice recrystallization but do not exhibit thermal hysteresis properties making them attractive candidates as cryoprotectants for the preservation of various biological materials. Structure-function studies of these C-linked glycoconjugates have indicated that the most active ice recrystallization inhibitors contain highly hydrated carbohydrate residues. This discovery has lead to the development of several classes of small molecule carbohydrate derivatives that exhibit potent ice recrystallization inhibition activity and may constitute a novel class of cryoprotectants. To date, several different classes of small molecule ice recrystallization inhibitors (IRIs) have been identified including amino acid-based organogelators, non-ionic carbohydrate-based amphiphiles (aryl and alkyl alditols) and O-linked alkyl and aryl glycoside derivatives. The rational design of these compounds, assessment and quantification of their IRI activity will be presented and the key structural features necessary for potent ice recrystallization inhibition activity will be discussed. Finally, the ability of both C-linked AFGP and the small molecule IRIs to function as cryoprotectants will be described. To properly assess this, several cell lines and primary cells have been examined. In addition, we have investigated the cryopreservation of oncolytic viruses (both encapsulated non-encapsulated). Source of funding: NSERC, CIHR, Canadian Blood Services, Green Centre Canada. Conflict of interest: None declared. rben@uottawa.ca