Aldolase triggers metabolic reprogramming in colorectal cancer in hypoxia and stiff desmoplastic microenvironments.

Colorectal cancer (CRC) progression is highly associated with desmoplasia. Aerobic glycolysis is another distinct feature that appears during the CRC phase of the adenoma-carcinoma sequence. However, the interconnections between the desmoplastic microenvironment and metabolic reprogramming remain largely unexplored. In our in vitro model, we investigated the compounding influences of hypoxia and substrate stiffness, two critical physical features of desmoplasia, on the CRC metabolic shift by using engineered polyacrylamide gels. Unexpectedly, we found that compared to cells on a soft gel (approximately 1.5 kPa, normal tissue), cells on a stiff gel (approximately 8.7 kPa, desmoplastic tissue) exhibited reduced glucose uptake and glycolysis under both normoxia and hypoxia. In addition, the increasing substrate stiffness activated focal adhesion kinase (FAK)/phosphoinositide 3-kinase signaling, but not the mitochondrial respiratory inhibitor HIF-1α. However, the presence of aldolase B (ALDOB) reversed the CRC metabolic response to mechanosignaling; enhanced glucose uptake (approximately 1.5-fold) and aerobic glycolysis (approximately 2- to 3--fold) with significantly decreased mitochondrial oxidative phosphorylation. ALDOB also changed the response of CRC traction force, which is related to tumor metastasis, under hypoxia/normoxia. In summary, our data suggest a counter influence of hypoxia and substrate stiffness on glucose uptake, and ALDOB upregulation can reverse this, which drives hypoxia and stiff substrate to enhance the CRC aerobic glycolysis synergistically. The results not only highlight the potential impacts on metabolic reprogramming led by physical alterations in the microenvironment, but also extend our understanding of the essential role of ALDOB in CRC progression from a biophysical perspective.