Abstract
Aim: Sustained cancer cell growth requires up-regulation of nutrient acquisition mechanisms. Novel approaches can identify "tumour fuel", the way it is obtained and then altered for cellular usage. We investigated the contribution of macropinocytosis and autophagy as the cellular mechanisms involved in protein uptake and degradation, respectively, to myeloma cells growth and survival.
Method: Macropinosomes were visualized using confocal microscopy in KRAS-mutated and WT human myeloma cells utilising TMR-dextran and 5-[N-ethyl-N-isopropyl] amiloride (EIPA) as a marker and specific inhibitor of macropinocytosis, respectively. Degradation of internalized albumin and its co-localization with TMR-dextran were evaluated. Cell sensitivity to glutamine deprivation was examined with an ATP-based viability test. To evaluate the possible role of macropinocytosis and autophagy under glutamine-deprived conditions, the effect of albumin supplementation on cell growth and viability was studied in the presence or absence of EIPA or chloroquine. The intracellular concentration of glutamine was directly measured after albumin supplementation.
Result: KRAS-mutated KMS28-PE, KMS18 and MM1S cells showed heightened EIPA-sensitive macropinocytosis compared to WT-KRAS KMS34, KMS12-BM and TK1 cells. Macropinocytosis led to internalization and subsequent degradation of albumin. The viability of WT-KRAS cells was reduced in sub-physiological concentrations of glutamine when compared with KRAS-mutated cells that remained viable and even continued to slowly proliferate over 7 days. Glutamine rescued the compromised growth of KRAS-mutated cells at low concentrations of albumin, which was abrogated by EIPA or chloroquine (p˂0.05). KRAS-mutated cells demonstrated an EIPA-sensitive increase in intracellular concentration of glutamine following albumin supplementation (p˂0.05) abrogated by chloroquine. In WT cells, however, intracellular glutamine level remained unchanged.
Conclusion: Autophagy and macropinocytosis maintain cellular growth and survival in the context of nutrient deprivation by provision of glutamine, and potentially other amino acids, in RAS-mutated myeloma. These data suggest that novel anti-cancer strategies interrupting these cellular mechanisms may represent a promising therapeutic approach to myeloma.
Method: Macropinosomes were visualized using confocal microscopy in KRAS-mutated and WT human myeloma cells utilising TMR-dextran and 5-[N-ethyl-N-isopropyl] amiloride (EIPA) as a marker and specific inhibitor of macropinocytosis, respectively. Degradation of internalized albumin and its co-localization with TMR-dextran were evaluated. Cell sensitivity to glutamine deprivation was examined with an ATP-based viability test. To evaluate the possible role of macropinocytosis and autophagy under glutamine-deprived conditions, the effect of albumin supplementation on cell growth and viability was studied in the presence or absence of EIPA or chloroquine. The intracellular concentration of glutamine was directly measured after albumin supplementation.
Result: KRAS-mutated KMS28-PE, KMS18 and MM1S cells showed heightened EIPA-sensitive macropinocytosis compared to WT-KRAS KMS34, KMS12-BM and TK1 cells. Macropinocytosis led to internalization and subsequent degradation of albumin. The viability of WT-KRAS cells was reduced in sub-physiological concentrations of glutamine when compared with KRAS-mutated cells that remained viable and even continued to slowly proliferate over 7 days. Glutamine rescued the compromised growth of KRAS-mutated cells at low concentrations of albumin, which was abrogated by EIPA or chloroquine (p˂0.05). KRAS-mutated cells demonstrated an EIPA-sensitive increase in intracellular concentration of glutamine following albumin supplementation (p˂0.05) abrogated by chloroquine. In WT cells, however, intracellular glutamine level remained unchanged.
Conclusion: Autophagy and macropinocytosis maintain cellular growth and survival in the context of nutrient deprivation by provision of glutamine, and potentially other amino acids, in RAS-mutated myeloma. These data suggest that novel anti-cancer strategies interrupting these cellular mechanisms may represent a promising therapeutic approach to myeloma.
| Original language | English |
|---|---|
| Number of pages | 1 |
| Publication status | Published - Oct 2018 |
| Event | Blood 2018 - Brisbane, Australia Duration: 21 Oct 2018 → 24 Oct 2018 http://www.blood2018.com |
Conference
| Conference | Blood 2018 |
|---|---|
| Country/Territory | Australia |
| City | Brisbane |
| Period | 21/10/18 → 24/10/18 |
| Other | The combined Annual Scientific Meeting of the Haematology Society of Australia and New Zealand, Australian and New Zealand Society of Blood Transfusion and Thrombosis and Haemostasis society of Australia and New Zealand |
| Internet address |
