Pre-formulation development of highly potent nontuberculous mycobacterial agents

Nontuberculous mycobacteria (NTM) are pathogens that particularly infect patients with structural lung diseases, such as chronic obstructive pulmonary disease or cystic fibrosis. Several novel indole-2-carboxamides (ICs) that were synthesized in our laboratory have demonstrated potent antimicrobial activity against a wide panel of NTM pathogens with in vivo efficacy against Mycobacterium tuberculosis and Mycobacterium abscessus-infected mouse models. Lead ICs suffer poor aqueous solubility and permeability leading to high dosing regimens to achieve efficacy. The objective of this work is to conduct preformulation studies on two of the lead ICs (N2 and N21, US Patent 20180036283A1). Scanning electron microscopy (SEM), thermal analyses [differential scanning calorimetry (DSC), thermogravimetric analysis (TGA)], hot-stage microscopy (HSM)], and X-ray powder diffraction (XRD) were used for their solid-state characterization. Octanol/water partition coefficient (LogP), pKa, solubility, pH stability, intrinsic dissolution and Caco-2 cell monolayer permeability were determined. Thermal analyses and XRD indicated that both ICs are crystalline solids. N21 exists at least in two polymorphic forms and the polymorphic transition was monotropic in nature. However, N2 did not show any polymorphism. These molecules with poor aqueous solubility (<0.2 μg/mL), and high Caco-2 cell monolayer permeability (>18 × 10⁻⁶ cm/s), suggested that both ICs belong to Biopharmaceutics Classification System (BCS) class II and may pose absorption and bioavailability challenges in vivo, primarily due to the poor aqueous solubility. The BCS class II designation is defined by marketed doses, therefore, since N2 and N21 are currently under development, our class II designation is defined by the poor aqueous solubility and high permeability. These molecules are weakly acidic and most stable at pH 6.8 with pKa values between 6.7 and 7.9. Results from the preformulation studies set the stage for the formulation of these drug candidates for clinical investigation and new drug application in the future for the treatment of NTM infections.