GBT1118, a potent allosteric modifier of hemoglobin O2 affinity, increases tolerance to severe hypoxia in mice

Abstract

Adaptation to hypoxia requires compensatory mechanisms that affect O₂ transport and utilization. Decreased hemoglobin (Hb) O₂ affinity is considered part of the physiological adaptive process to chronic hypoxia. However, this study explores the hypothesis that increased Hb O₂ affinity can complement acute physiological responses to hypoxia by increasing O₂ uptake and delivery compared with normal Hb O₂ affinity during acute severe hypoxia. To test this hypothesis, Hb O₂ affinity in mice was increased by oral administration of 2-hydroxy-6-{[(2S)-1-(pyridine-3-carbonyl)piperidin-2yl] methoxy}benzaldehyde (GBT1118; 70 or 140 mg/kg). Systemic and microcirculatory hemodynamics and oxygenation parameters were studied during hypoxia in awake-instrumented mice. GBT1118 increased Hb O₂ affinity and decreased the Po₂ at which 50% of Hb is saturated with O₂ (P₅₀) from 43 ± 1.1 to 18.3 ± 0.9 mmHg (70 mg/kg) and 7.7 ± 0.2 mmHg (140 mg/kg). In a dose-dependent fashion, GBT1118 increased arterial O₂ saturation by 16% (70 mg/kg) and 40% (140 mg/kg) relative to the control group during 5% O₂ hypoxia. In addition, a GBT1118-induced increase in Hb O₂ affinity reduced hypoxia-induced hypotension compared with the control group. Moreover, microvascular blood flow was higher during hypoxia in GBT1118-treated groups than the control group. The increased O₂ saturation and improved blood flow in GBT1118-treated groups preserved higher interstitial tissue Po₂ than in the control group during 5% O₂ hypoxia. In conclusion, increased Hb O₂ affinity enhanced physiological tolerance to hypoxia, as evidenced by improved hemodynamics and tissue oxygenation. Therefore, pharmacologically induced increases in Hb O₂ affinity become a potential therapeutic approach to improve tissue oxygenation in pulmonary diseases characterized by severe hypoxemia.NEW & NOTEWORTHY This study establishes that pharmacological modification of hemoglobin O₂ affinity can be a promising and novel therapeutic strategy for the treatment of hypoxic hypoxia and paves the way for the clinical development of molecules that prevent hypoxemia.

Keywords: hemoglobin; hypoxia; microcirculation; oxygen; oxygen delivery; partial pressure of oxygen at which hemoglobin is 50% saturated with oxygen; tissue partial pressure of oxygen.

Fig. 1.

A: hypoxic challenge design. Mice were dosed orally with GBT1118 (70 or 140 mg/kg) or vehicle only. At 1 h after GBT1118 or vehicle administration, mice were exposed to hypoxia by stepwise decreases in O₂ concentration to 15%, 10%, and 5%. Animals were kept at each hypoxic level for 0.5 h. To assess tolerance to hypoxia, mice were kept at 5% O₂ hypoxia for an additional 1.5 h. B: changes in O₂ affinity after GBT1118 administration, shown as representative O₂ equilibrium curves (OECs) of blood (40% hematocrit) from vehicle- and GBT1118-treated mice (n = 6 animals/group). GBT1118 at 70 or 140 mg/kg decreased the Po₂ at which Hb is 50% saturated with O₂ (P₅₀) of blood from 43 ± 1.1 to 18.3 ± 0.9 and 7.7 ± 0.2 mmHg, respectively. A leftward shift in the OEC, representing a decrease in P₅₀, indicates a GBT1118 dose-dependent increase in Hb O₂ affinity relative to vehicle-only control.

Fig. 2.

A: GBT1118 pharmacokinetics in male mice after intravenous (IV) dosing at 10 mg/kg and oral (PO) dosing at 100 mg/kg (n = 3 mice/time point). Blood samples were collected up to 8 h (intravenous) and 24 h (oral) after administration of GBT1118. Mean blood and plasma concentration-time profiles are shown. B: GBT1118 pharmacodynamics, shown as the effect of GBT1118 on OECs after intravenous and oral administration. Blood samples were collected for hemoximetry at different time points after single intravenous (10 mg/kg) and oral (100 mg/kg) GBT1118 administration. Both doses/routes of administration elicited a leftward shift in the blood OEC and a subsequent decrease in P₅₀ relative to vehicle, indicating an increase in Hb O₂ affinity (P₅₀ = 38, 31.6, and 13.6 mmHg for vehicle control, intravenous GBT1118, and oral GBT1118, respectively). C: GBT1118 pharmacokinetics and pharmacodynamics, shown as pharmacokinetics-pharmacodynamics correlation after single intravenous and oral doses of GBT1118. Blood samples were collected at maximum concentration after a single intravenous (10 mg/kg) or oral (70, 100 or 140 mg/kg) dose of GBT1118. Blood samples were analyzed for GBT1118 blood concentrations as well as for effects on Hb O₂ affinity using OECs (n = 18 total mice for the various indicated doses).

Fig. 3.

A: arterial Po₂ ($\mathrm{P}{\mathrm{a}}_{\mathrm{O}}{}_{{}_2}$) decreased during hypoxia. B: GBT1118 increased arterial O₂ saturation ($\mathrm{S}{\mathrm{a}}_{{\mathrm{O}}_2}$) during hypoxia. Values are means ± SD of 6 animals/group. *P < 0.05 vs. control at each percent O₂; ξP < 0.05 vs. GBT1118 (70 mg/kg); †P < 0.05 vs. baseline (BL).

Fig. 4.

Blood gases, respiratory rates, and interstitial tissue Po₂. GBT1118 prevented acidosis and improved O₂ delivery to tissues during hypoxia. A–F: changes in arterial blood pH, lactate concentration, respiration rate (in breaths/min), arterial blood Pco₂ ($\mathrm{P}{\mathrm{a}}_{\mathrm{C}\mathrm{O}}{}_{{}_2}$), arterial blood $\mathrm{H}\mathrm{C}{\mathrm{O}}_3^{-}$, and interstitial tissue Po₂ during hypoxia. Values are means ± SD of 6 animals/group. *P < 0.05 vs. control at each percent O₂; ξP < 0.05 vs. GBT1118 (70 mg/kg); †P < 0.05 vs. 21% O₂. Dashed line, BL values (pH 7.3, lactate = 1.17 mmol/l, respiration rate = 173.5 breaths/min, $\mathrm{P}{\mathrm{a}}_{\mathrm{C}\mathrm{O}}{}_{{}_2}$ = 38.4 mmHg, and $\mathrm{H}\mathrm{C}{\mathrm{O}}_3^{-}$ = 20 meq/l).

Fig. 5.

Systemic response during hypoxia. A and B: change in mean arterial pressure and heart rate (in beats/min) during hypoxia. Values are means ± SD of 6 animals/group. *P < 0.05 vs. control at each percent O₂; ξP < 0.05 vs. GBT1118 (70 mg/kg); †P < 0.05 vs. BL.

Fig. 6.

GBT1118 preserves microvascular blood flow during hypoxia. A and B: changes in arteriolar diameter and blood flow during hypoxia. Values are means ± SD of 6 animals/group. †P < 0.05 vs. 21% O₂.

Fig. 7.

GBT1118 reduces tissue hypoxia and improves tolerance to hypoxia in mice. A: hypoxic tissues (brain, heart, kidney, intestine, and liver) positively stained by pimonidazole relative to control during extreme hypoxia. B: tolerance to hypoxia in mice. Values are means ± SD of 6 animals/group.