Development of Brain Penetrant Pyridazine Pantothenate Kinase Activators

Abstract

Conversion of pantothenate to phosphopantothenate in humans is the first dedicated step in the coenzyme A (CoA) biosynthesis pathway and is mediated by four isoforms of pantothenate kinase. These enzymes are allosterically regulated by acyl-CoA levels, which control the rate of CoA biosynthesis. Small molecule activators of the PANK enzymes that overcome feedback suppression increase CoA levels in cultured cells and animals and have shown great potential for the treatment of pantothenate kinase-associated neurodegeneration and propionic acidemias. In this study, we detail the further optimization of PANK pyridazine activators using structure-guided design and focus on the cellular CoA activation potential, metabolic stability, and solubility as the primary drivers of the structure-activity relationship. These studies led to the prioritization of three late-stage preclinical lead PANK modulators with improved pharmacokinetic profiles and the ability to substantially increase brain CoA levels. Compound 22 (BBP-671) eventually advanced into clinical testing for the treatment of PKAN and propionic acidemia.

Figure 1

(A) Initial leads 1 and 2. (B) Functional group (FG) modifications explored in this study.

Figure 2

Crystal structures of representative compounds in the FG¹ series. (A) Overview of compound 1 bound across the dimer interface of the PANK3·AMPPNP·Mg²⁺·1 complex (PDB entry 6PE6). The locations of the substitutions at FG¹–FG³ are indicated. The interacting residues from the two PANK3 protomers are colored gold and cyan (carbons), and the compounds are colored purple. (B) FG¹ area of the PANK3·AMPPNP·Mg²⁺·2 structure (PDB entry 6PE6) illustrating the cyclopropyl replacement of the isopropyl in 1. (C) FG¹ area of the PANK3·AMPPNP·Mg²⁺·3 structure (PDB entry 7UE3) illustrating the location of the hydroxyl group and its hydrogen bond interactions with the AMPPNP γ-phosphate. (D) FG¹ area of the PANK3·AMPPNP·Mg²⁺·6 structure (PDB entry 7UE5) showing the two orientations of the CF₃ group. (E) FG¹ area of the PANK3·AMPPNP·Mg²⁺·14 structure (PDB entry 7UEQ) illustrating the location of the planar N,N-dimethylamine group compared to the tetrahedral isopropyl group.

Figure 3

Crystal structures of representative compounds in the FG² and FG³ series. The interacting residues from the two PANK3 protomers are colored gold and cyan (carbons), and the compounds are colored purple. (A) PANK3·AMPPNP·Mg²⁺·18 structure (PDB entry 7UE6) illustrating how the pyridine nitrogen is oriented with respect to the hydrogen bond to Wat1. (B) PANK3·AMPPNP·Mg²⁺·21 structure (PDB entry 7UE7) showing the two orientations of the fluorinated ring. (C) PANK3·AMPPNP·Mg²⁺·23 structure (PDB entry 7UEO). (D) PANK3·AMPPNP·Mg²⁺·29 structure (PDB entry 7UEP) showing the interaction of the CH₃ group with L263′.

Figure 4

Liver, forebrain, and hindbrain CoA concentrations in mice treated with members of the FG¹ compound series. CoA concentrations of compounds 3 and 5 were compared to those of 2 at a dose of 10 mg/kg. Compounds 8 and 9 were compared to 2 at a dose of 15 mg/kg: (A) liver CoA, (B) forebrain CoA, and (C) hindbrain CoA. The control mice were given the vehicle (30% Captisol). All mice were given five doses 12 h apart. An asterisk indicates mice were given three doses 24 h apart. All tissues were harvested 4 h after the last dose. The mean ± SEM is plotted, and the p values from a nonparametric t test are colored red.

Figure 5

Liver, forebrain, and hindbrain CoA concentrations in mice treated with members of the FG² and FG³ compound series. R is Cl or CN in that position. (A–C) CoA concentrations of FG² pyridine compounds 17 and 18 compared to those of 2 at a dose of 15 mg/kg. FG² fluorinated compounds 21, 22, and 24 compared to 2 at a dose of 10 mg/kg: (A) liver CoA, (B) forebrain CoA, and (C) hindbrain CoA. (D–F) CoA concentrations of the FG³ methyl substitutions in 27 and 29 compared to those of 2 at a dose of 10 mg/kg. FG³ methyl substitution combined with FG² fluorination in 31 and 33 compared to 2 and 5 at a dose of 10 mg/kg: (D) liver CoA, (E) forebrain CoA, and (F) hindbrain CoA. The control mice were given the vehicle (30% Captisol). All mice were given five doses 12 h apart. An asterisk indicates mice were given three doses 24 h apart. All tissues were harvested 4 h after the last dose. The mean ± SEM is plotted, and the p values from a nonparametric t test are colored red.