(mRNA and CRISPER are powerful tools, here is another (sic) expression of that)
Milestone in diabetes treatment: Transgenic cow produces human insulin in milk
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"In the old days, we used to just slam DNA in and hope it got expressed where you wanted it to," Wheeler said. "We can be much more strategic and targeted these days. Using a DNA construct specific to mammary tissue means there's no human insulin circulating in the cow's blood or other tissues. It also takes advantage of the mammary gland's capabilities for producing large quantities of protein."
When the cow reached maturity, the team unsuccessfully attempted to impregnate her using standard artificial insemination techniques. Instead, they stimulated her first lactation using hormones. The lactation yielded milk, but a smaller quantity than would occur after a successful pregnancy. Still, human proinsulin and, surprisingly, insulin were detectable in the milk.
"Our goal was to make proinsulin, purify it out to insulin, and go from there. But the cow basically processed it herself. She makes about three to one biologically active insulin to proinsulin," Wheeler said. "The mammary gland is a magical thing."
The insulin and proinsulin, which would need to be extracted and purified for use, were expressed at a few grams per liter in the milk. But because the lactation was induced hormonally and the milk volume was smaller than expected, the team can't say exactly how much insulin would be made in a typical lactation.
Conservatively, Wheeler says if a cow could make 1 gram of insulin per liter and a typical Holstein makes 40 to 50 liters per day, that's a lot of insulin. Especially since the typical unit of insulin equals 0.0347 milligrams.
"That means each gram is equivalent to 28,818 units of insulin," Wheeler said. "And that's just one liter; Holsteins can produce 50 liters per day. You can do the math."
The team plans to re-clone the cow, and is optimistic they'll achieve greater success with pregnancy and full lactation cycles in the next generation. Eventually, they hope to create transgenic bulls to mate with the females, creating transgenic offspring that can be used to establish a purpose-built herd. Wheeler says even a small herd could quickly outcompete existing methods -; transgenic yeast and bacteria -; for producing insulin, and could do so without having to create highly technical facilities or infrastructure.
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https://www.news-medical.net/news/20240314/Milestone-in-diabetes-treatment-Transgenic-cow-produces-human-insulin-in-milk.aspxHuman proinsulin production in the milk of transgenic cattle
Pseudo-lentivirus containing the bovine β-casein promoter and human insulin sequences was used to produce modified adult fibroblasts, and the cells were used for nuclear transfer. Transgenic embryos were transferred to recipient cows, and one pregnancy was produced. Recombinant protein in milk was evaluated using western blotting and mass spectrometry. One transgenic cow was generated, and in milk analysis, two bands were observed in western blotting with a molecular mass corresponding to the proinsulin and insulin. The mass spectrometry analysis showed the presence of human insulin more than proinsulin in the milk, and it identified proteases in the transgenic milk that could convert proinsulin into insulin and insulin-degrading enzyme that could degrade the recombinant protein
https://analyticalsciencejournals.onlinelibrary.wiley.com/doi/10.1002/biot.202300307