At 25 μM, EIPA inhibited albumin-supported growth of Ras-transformed cells under low-glutamine conditions. Added glutamine or dimethyl α-ketoglutarate suppressed this response, supporting a nutrient-dependent interpretation. [2]
EIPA also affected receptor-mediated albumin uptake in opossum kidney cells (extracellular endpoint IC50 ∼21 μmol l−1), while fluid-phase dextran uptake was preserved at 100 μmol l−1. Its endocytic effects therefore depend on the model and uptake pathway. [3]
EIPA (10 μM, 18–20 h) reduced basal and maximal oxygen consumption in both cancer cells and tissue-matched untransformed epithelial cells. This metabolic response was not restricted to cancer cells. [1]
EIPA suppresses cellular acid-load recovery
Cell-based study · BxPC3 pancreatic cancer and MDA-MB-157 breast cancer cells [1]
EIPA suppressed intracellular pH recovery after an acid load, used by the authors as an index of NHE1 activity. This is a cellular transport assay, not a purified-target potency measurement.
Literature-Reported Treatment Conditions
- Treatment concentration
- 10 μM
- Exposure duration
- 18–20 h treatment in the general pHi methods; S1A specifies 20 h. The S1B-C legend does not separately restate exposure duration.
- Method
- BCECF pHi recording in HEPES buffer after NH4Cl-induced acid load; untreated control comparison.
- Reported limits
- Vehicle and EIPA supplier are not specified in these methods; no biochemical IC50 is established.
Methods and source locations
Compound reported in the paper: 5-(N-ethyl-N-isopropyl)amiloride; Compound label in the paper: EIPA
Evidence concerns the named EIPA molecule under the reported conditions; it does not validate A20092 lots or establish salt equivalence.
Supplementary Figure S1B-C and S1 legend; Methods: Intracellular pH and NHE1 Activity; Results: EIPA Decreases Basal and Maximal Respiratory Capacity
pHi recovery after acid load: BCECF fluorescence calibrated to pHi; S1B-C traces — Supplementary Figure S1B-C and S1 legend; Methods: Intracellular pH and NHE1 Activity; Results: EIPA Decreases Basal and Maximal Respiratory Capacity
EIPA inhibits albumin-supported growth under low glutamine
Cell-based study · MIA PaCa-2, T24 and NIH 3T3 [K-RasV12] cells [2]
EIPA reduced albumin-supported growth under low glutamine. Additional glutamine or dimethyl α-ketoglutarate suppressed this response, limiting a simple interpretation as unconditional cytotoxicity.
Literature-Reported Treatment Conditions
- Treatment concentration
- 25 μM
- Exposure duration
- 6 days for Figure 4a; Figure 4b rescue duration is not separately restated in its caption.
- Method
- MTT assay; medium replaced every 24 h.
- Vehicle
- DMSO; vehicle-only controls
- Co-treatment
- 0.2 mM glutamine with 2% final albumin; rescue with increased glutamine or 7 mM dimethyl α-ketoglutarate.
- Reported limits
- EIPA salt/counterion not specified; MTT is a viability/metabolic proxy rather than direct cell counting.
Methods and source locations
Compound reported in the paper: 5-(N-ethyl-N-isopropyl)amiloride; Compound label in the paper: EIPA
Evidence concerns the named EIPA molecule under the reported conditions; it does not validate A20092 lots or establish salt equivalence.
Figure 4a-b and caption; Results paragraph beginning Heightened sensitivity; Methods: Glutamine Deprivation Assays
Albumin-supported viable-cell MTT signal: MTT after 6 days; Figure 4a normalized to 0.2Q, Figure 4b to 0.2Q+Alb; t-test significance shown in source — Figure 4a-b and caption; Results paragraph beginning Heightened sensitivity; Methods: Glutamine Deprivation Assays
EIPA affects receptor-mediated albumin uptake in renal cells
Cell-based study · Confluent opossum kidney (OK) proximal tubular cells [3]
EIPA inhibited receptor-mediated albumin uptake in this NHE3-expressing renal model. Fluid-phase FITC-dextran uptake was preserved, providing a model-specific limit to treating EIPA as a universal macropinocytosis marker.
Literature-Reported Treatment Conditions
- Treatment concentration
- 100 μmol l−1 for time-course and tracer controls; a separate extracellular concentration-response series was fitted.
- Exposure duration
- 10 min pretreatment and continued exposure during the Figure 1 uptake time course (up to 15 min).
- Method
- 10 mg l−1 FITC-albumin in Hepes Ringer; uptake at 37°C corrected for surface-bound material measured at 4°C.
- Reported limits
- Vehicle not identified in the read methods; tracer-control durations are not individually restated in Results.
Methods and source locations
Compound reported in the paper: 5-(N-ethyl-N-isopropyl)amiloride; Compound label in the paper: EIPA
Evidence concerns the named EIPA molecule under the reported conditions; it does not validate A20092 lots or establish salt equivalence.
Figure 1A-C and caption; Results: Time course of inhibition of endocytosis and Action profile; Methods: Materials, Uptake studies, Calculations and statistics
Receptor-mediated albumin uptake: Least-squares concentration-response fit; Figure 1C n=6–12 Petri dishes; endpoint IC50, not target binding affinity — Figure 1A-C and caption; Results: Time course of inhibition of endocytosis and Action profile; Methods: Materials, Uptake studies, Calculations and statistics
Radiolabeled albumin and fluid-phase dextran uptake: Results tracer controls; mean ± SEM; n denotes Petri dishes, with cells of at least three passages used per experimental series — Results: Time course of inhibition of endocytosis; Methods: Uptake studies, Calculations and statistics
EIPA reduces cellular oxygen consumption
Cell-based study · HPDE and BxPC3 pancreatic epithelial cells; MCF10A and MDA-MB-157 mammary epithelial cells [1]
EIPA decreased basal and maximal oxygen consumption in cancer and untransformed cells. These observations limit assumptions that the metabolic response is restricted to cancer cells; a direct mitochondrial target is not established.
Literature-Reported Treatment Conditions
- Treatment concentration
- 10 μM
- Exposure duration
- 18–20 h
- Method
- Seahorse XF24 mitochondrial stress assay; OCR normalized to cell number; untreated controls.
- Co-treatment
- Sequential mitochondrial stress-test reagents used as assay probes, not a therapeutic combination.
- Reported limits
- Vehicle and EIPA supplier not stated in the read methods. Figure 1G-H legend misnames pancreatic cells; plotted labels, Results and methods identify the mammary pair.
Methods and source locations
Compound reported in the paper: 5-(N-ethyl-N-isopropyl)amiloride; Compound label in the paper: EIPA
Evidence concerns the named EIPA molecule under the reported conditions; it does not validate A20092 lots or establish salt equivalence.
Figure 1A-H and caption; Results: EIPA Decreases Basal and Maximal Respiratory Capacity; Methods: Mitochondrial Stress and Fuel Assays, Data Presentation and Statistical Analysis
Basal and maximal mitochondrial oxygen consumption: Figure 1: n=5 separate preparations/experiments for HPDE/BxPC3; n=4 for MCF10A/MDA-MB-157 endpoint plots; Tukey–Kramer HSD. Kinetic-panel captions state five preparations. No curve-derived values adopted. — Figure 1A-H and caption; Results: EIPA Decreases Basal and Maximal Respiratory Capacity; Methods: Mitochondrial Stress and Fuel Assays, Data Presentation and Statistical Analysis
