A Bioinformatic Study Of The Effect Of Lactic Acidosis On 4T1 Cancer Cells Adaption To Glucose Deprivation

Background:The adaption of cancer cells in solid tumor to glucose deprivation, while its mechanism remains unknown, is critically important for tumor progressive growth. While the growth of solid tumor requires a large amount of nutrients, nutrient supply in solid tumor is very limited, largely due to the disorganized vasculature and dysfunctional capillary. Constant or intermittent glucose is a common feature of solid tumor, particularly the poorly vascularized solid tumors. Glucose is an essential nutrient for tumor growth. Cancer cells in vitro can neither grow nor survival under glucose deprivation. The fact that cancer cells in solid tumor can tolerate glucose deprivation suggests an unknown adaptive mechanism may exist to help cancer cell survive. Because cancer cells generate excess lactate anion and proton via aerobic glycolysis or Warburg effect, even in the presence of ample oxygen, lactic acidosis (lactate anion+proton) is a common feature of tumor microenvironment. High level of lactate is associated with increase metastasis, poor disease-free and overall survival. Decreasing lactate fermentation by displacing PKM2 with PKM1 reduced cancer cells' ability to form tumor in nude mice.In addition, our preliminary study showed that lactic acidosis significantly extended the cell survival rate. After glucose was completely consumed, without additional exogenous lactic acid, over 90% cells died within a single day, whereas supplemented with 20mM lactic acid,90% cells died in a period of 10 days, with viable cells identified even 65 days after glucose was depleted, in contrast to only 4 days for control cells. These cells resumed proliferation upon nutritional restoration. Lactate was barely consumed by cells after glucose was exhausted, indicating that lactate was not used as a fuel by cells under glucose deprivation. Lactic acidosis also significantly extended the life span of other cancer cells (human breast cancer cell line Bcap37, human colon cancer cell line RKO, human gastric cancer cell line SGC7901) under glucose starvation. We propose here that lactic acidosis may play an important role in role in the adaption of cancer cells to glucose deprivation.Methods and Results:Murine breast cancer cells 4T1 were cultured in RPMI-1640 medium containing 3 mM glucose with 20 mM lactic acid (lactic acidosis). The cells under cultured with 0,1, 3,4,5,7,9,11 days were collected, and RNA was extracted, then hydride to Mouse Gene 1.0 ST microarrays. Using transcriptional profiling technology and data analysis, we uncovered that lactic acidosis is a key factor that renders cancer cells adaptive to glucose deprivation:(1)Lactic acidosis regulates cell cycle checkpoints of cell cycle by loosening G2/M transition and spindle assembly checkpoint but tightening G1/S transition. While the former may permit cells to progress through cell cycle and to survive by avoiding checkpoint-induce apoptosis, the latter can arrest cells at "quiescent" status, which fits to survival under metabolic stress. (2) Lactic acidosis globally shuts down anabolic branch which is energy consuming, increases expression of enzymes for fatty acid?-oxidation but reduces expression of glycolytic enzymes, actively adapting to glucose starvation by shifting energy generating from glycolysis to fatty acid catabolism. (3) Lactic acidosis attenuates glucose deprivation-induced ER and oxidative stress. (4) Lactic acidosis activates a restrained autophagy, which not only recycles cellular components for essential metabolism but also prevents cells from metabolic stress-induced apoptosis. (5) Lactic acidosis upregulates endocytosis-lysosome pathway that permits cells to acquire extracellular nutrients. (6) Lactic acidosis inhibits apoptosis. (7) Lactic acidosis activates unfolded protein response under glucose deprivation.Conclusion:Lactic acidosis rescued cancer cells under glucose deprivation via regulating many cellular processes including cell cycle, autophagy, apoptosis, stress response, and metabolism, which orchestrate to attenuate numerous death signals emanating from glucose deprivation. Lactic acidosis, a common feature of solid tumor, potentially provides a general mechanism whereby cancer cells survive under glucose deprivation.