Forskolin Increases cAMP Levels and Enhances Recombinant Antibody Production in CHO Cell Cultures

Abstract

Cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger essential to many biological processes. Forskolin, a labdane diterpene, is widely used to increase cAMP levels in various cell types. When 5 µm of forskolin is added to recombinant Chinese hamster ovary (rCHO) cell cultures producing monoclonal antibody (mAb) (GSR cell line), it decreases specific growth rate (), but increases culture longevity and specific mAb productivity (q mAb). The beneficial effect of forskolin on culture longevity and q mAb outweighs its detrimental effect on , resulting in 1.5-fold increase in the maximum mAb concentration (MMC) without an adverse effect on mAb quality attributes such as aggregation, charge variation, and galactosylation. Forskolin induces cell cycle arrest at the G0/G1 phase via the LKB1/AMPK pathway and inhibits apoptosis via the CREB/Bcl-2 pathway. The beneficial effect of forskolin on mAb production is also demonstrated with another rCHO cell line (ABTAA). Addition of 5 µm forskolin to ABTAA cell cultures also results in 1.5-fold increase in the MMC. Taken together, the results obtained demonstrate that forskolin is an efficient chemical reagent to increase mAb production in rCHO cell cultures.

Pharmacological Evidence That Dictyostelium Differentiation-Inducing Factor 1 Promotes Glucose Uptake Partly via an Increase in Intracellular cAMP Content in Mouse 3T3-L1 Cells

“…At 20 µM, forskolin appeared to be slightly toxic to the cells as it changed cell morphology and made the boundaries between cells clearer after 20 h incubation. Since forskolin is expected to increase intracellular cAMP levels [33,34], our results suggest that an increase in intracellular cAMP levels can promote glucose uptake in confluent 3T3-L1 cells.…”

3D microfluidics-assisted modeling of glucose transport in placental malaria

“…As our model cannot reproduce these features of placenta and is a simplified model, in this case it represents a mixture of two types of relevant cells on the trophoblast side of the placenta, though not in multilayer arrangement. Third, and important, we grew two types of cells at the same time, and we did not want to treat EC with forskolin, as it significantly increases cAMP levels in treated cells 43 , which may produce some unexpected effects. Without this treatment we would have some level of syncytialization of BeWo cells and avoid potential effects of forskolin treatment on HUVEC.…”

Chemical and Genetic Modulation of Complex I of the Electron Transport Chain Enhances the Biotherapeutic Protein Production Capacity of CHO Cells

“…Of the 1280 small molecules tested, only two molecules, 3F3 (BrdU) and 7A8 (forskolin), enhanced the volumetric titer by a minimum of 1.4× in at least three of the eight conditions tested (4 clonal cell lines × 2 concentrations of each small molecule) (Figure 1B,C and Supplemental Table S1), which corresponds to a hit rate of 0.15% (2/1280 = 0.15%). Interestingly, forskolin has been previously reported to increase recombinant protein production in CHO cells via modulation of both cell cycle and apoptosis pathways [10]. BrdU, on the other hand, has not been previously reported to enhance the production of recombinant proteins, but rather is most recognized as a DNA intercalating agent that can alter DNA stability, prolong the cell cycle, and have effects on transcription and translation [24].…”

“…Interestingly, the benefits of supplementing BrdU into the media were observed in not only stably expressing cell lines, but also in a transient protein production system. While forskolin has been shown to induce cell cycle arrest via the LKB1/AMPK pathway, inhibit apoptosis via the CREB/Bcl-2 pathway, and ultimately increase cell-specific productivity of CHO cells expressing a monoclonal antibody, the effects of BrdU are not well-understood [10]. Since adding a pharmacologically active compound to a production process may not be a favorable path to enhance protein production, we performed differential gene expression analysis to determine which cellular pathways were being modulated by the addition of BrdU.…”