Academic Research & Preclinical Studies
The focus of academic or biopharma scientists who use collagenase products for enzyme-mediated cell isolation of their cell of interest to either:
Improve their understanding of a cell function
Manipulate or alter that function by exposing cells to other agents
Identify a biomarker that qualitatively or quantitively correlates with cell identification, function, or functional state
The focus for funding research has changed from research to increase knowledge to translational science to apply new knowledge and techniques to improve patient care. This shift happened in 2012 when NIH increased the funding of its Clinical and Translational Science awards. These and all NIH biomedical funding were re-directed to apply new knowledge and techniques to improve healthcare.
Traditional Collagenase Manufacturing Process
Traditional collagenase products are still commonly used to isolate cells when using enzyme mediated cell isolation methods. These crude or enriched collagenase products are minimally processed with the enzyme activities reflecting the variability of enzyme expression in each fermentation batch. Moreover, collagenase accounts for a small percentage of the product (< 8% w/w). End-users adapt these “off the shelf” products to meet their needs. Lot-to-lot variability requires most end-users pre-qualify new lots of collagenase products before use. The only adjustment that the end-user can make is adjusting the enzyme dose used in the isolation procedure.
Purified-Defined Collagenase Manufacturing Process
Purifying the collagenase and protease enzymes required for enzyme-mediated cell isolation appeared to overcome the need for lot pre-qualification. The development of Liberase HI Purified Enzyme Blend for isolating human islets showed that at least three enzymes were required for successful enzyme-mediated cell isolation; specifically, class I and class II C. histolyticum collagenases, and at least one neutral proteases from any source.
For many years, Liberase HI was the gold standard product for human islet isolation.However, a change in the manufacturing process of the first purified product, Liberase™ HI Purified Enzyme Blend, resulted in inconsistent recoveries of human islets that delayed the completion of a human clinical trial using human islets to manage adult, brittle, type 1 diabetic patients.
VitaCyte analyzed the enzymes that gave suboptimal islet yields and found that these results could be explained by proteolytic degradation of class I C. histolyticum collagenase. These results convinced VitaCyte to more rigorously characterize collagenase enzymes it supplies to customers.
VitaCyte’s Anticipating the Needs for Translational Research
As the understanding of the structure-function of collagenase enzymes improved, VitaCyte adopted more rigorous purification and characterization procedures creating the new category, “purified-defined” collagenase products. These enzymes offer significant advantages over other collagenase products in that the purification eliminates loss of collagen degradation activity. Consistent activity allows others to reproduce results or to transfer the process to a cell manufacturing facility.
Moreover, VitaCyte’s deep understanding of enzyme-mediated cell isolation led to the formulation of a mechanistic model to understand the roles collagenase and protease enzymes play in releasing cells from tissue. This knowledge is applied to optimize the composition and dose of enzymes used for cell isolation.
Purified-Defined Collagenase vs Traditional / Purified Collagenase
| Traditional | Purified | Purified-Defined | |
|---|---|---|---|
| Lot Qualification | Required | ± requirement depending on product consistency | Once dose/composition identified, no need for lot pre-qualification |
| Knowledge Gained from Lot Qualification | None: Enzyme composition unknown | Some: Dependent on rigor of collagenase or protease analyses | Extensive: Defined enzyme composition |
| Ability to Modify Enzyme Formulation | None | ± dependent on supplier’s offering | Yes, broad product offering |
| Technical Support | Little to none | Little to none | Experts in field |
| Relative Lifetime Costs | High: Expense of lot qualification | Moderate: Dependent on product consistency | Low: No need for lot qualification |
