When LONG^®R³IGF-I is supplemented in serum-free medium it promotes cell proliferation, increased cell survival, increased productivity through greater proliferation and anti-apoptotic signaling. LONG^®R³IGF-I provides equivalent or better performance to recombinant insulin depending on the cell line and clone. LONG^®R³IGF-I has been used with numerous cell types including CHO, BHK, HEK 293, Vero, PER.C6^®, MDCK and fibroblasts (graphs 1-4). All cell types that have a growth response to insulin in cell culture have the potential to respond to LONG^®R³IGF-I.

LONG^®R³IGF-I is an analogue of human-like growth factor I (IGF-I) specifically engineered for use in research and industrial cell culture. Insulin, like IGF-I and their receptors (the insulin receptor (IR) and the type-I IGF receptor IGF-IR) have similar amino acid sequence and protein structure. As a consequence, insulin and IGF-I are able to bind to each other's receptor with relatively low affinity. It is widely accepted that in CHO cells the effects of insulin are mediated by the IGF-IR, due to the fact there are relatively few IR present on the CHO cells and that insulin must be present at a high, non-physiological concentration typically 1 -10 mg/L, to be effective. A more effective growth factor is one which targets and activates the IGF-IR directly, such as IGF-I or the analogue LONG^®R³IGF-I. LONG^®R³IGF-I has a distinct biological advantage over native IGF-I due to its low affinity for IGF Binding Proteins (IGFBPs). All mammalian cells secrete IGFBPs, which bind to and inhibit native IGF-I. The substitution of an arginine for glutamine acid at position three in LONG^®R³IGF-I, in conjunction with the 13 amino acid N-terminal extension peptide, results in > 1000-fold reduced affinity for IGFBPs enhancing bioavailability and effectiveness in comparison to native IGF-I.

Under bioreactor conditions, stress induced apoptosis is the major cause of loss of cell viability. Activation of the IGF-IR results in the stimulation of a number of signal transduction cascades that have been identified as important for cell survival and proliferation. LONG^®R³IGF-I not only results in greater activation of IGF-IR over insulin, but also results in greater activation of key anti-apoptotic and proliferative signaling molecules: Akt and MAPK. Increased activation of these signaling molecules prevents the loss of viability caused by different culture conditions.

Under normal cell culture conditions, LONG^®R³IGF-I is more stable than insulin, persisting up to two times longer. Insulin is degraded in cell culture by enzymes (insulinases) secreted by cells into culture media. In addition, insulin is more rapidly internalized and degraded compared with IGF-I and LONG^®R³IGF-I. The extended cell culture stability of LONG^®R³IGF-I results in prolonged activity and associated benefits to cell culture. (Graph 5).

LONG^®R³IGF-I is manufactured under cGMP compliance with the International Conference on Harmonization (ICH) Q7A guidelines. The manufacturing plant is regularly audited by major European, US and Japanese biopharmaceutical companies. LONG^®R³IGF-I is produced in a proprietary E. coli fermentation process that is completely animal free (AF). No animal-derived materials are used during the manufacture or storage of LONG^®R³IGF-I. A certificate of origin is available upon request. Manufactured by Novozymes Biopharma, LONG^®R³IGF-I has been supplied to the research and industrial cell culture market for over 15 years. In addition, LONG^®R³IGF-I is used in the manufacture of several FDA (US), EMEA (Europe) and MHLW (Japan) approved marketed products, with many more in late-phase clinical trials.