Effect of High-Energy Milling on the Dissolution of Anti-HIV Drug Efavirenz in Different Solvents.

The anti-HIV drug efavirenz (EFV) displays low and variable bioavailability because of its poor aqueous solubility. Ball milling is a simple and cost-effective alternative to traditional micronization to improve the solubility and dissolution rate of EFV. A multibody dynamics model was employed to optimize the milling process parameters, while the motion of the balls in the mill jar was monitored in operando. This led to a better understanding of the milling dynamics for efficient comminution and enhancement of EFV dissolution. The variability of results for different EFV batches was also considered. Depending on the EFV batch, there were intrinsic differences in how the milling affected the dissolution behavior and inhibition of HIV-1 infection. High-energy grinding is more effective on EFV materials containing an amorphous fraction; it helps to remove agglomeration and enhances dissolution. Polyvinylpyrrolidone (PVP) addition improves the dissolution by forming a hydrophilic layer on the EFV surface, thereby increasing the drug wettability. Polymorphism also affects the quality, dosage, and effectiveness of the drug. The mechanical stress effect and PVP addition on the EFV polymorphic transformation were monitored by X-ray powder diffraction, while the residual of ground EFV was collected after dissolution, analyzed by scanning electron microscopy, and provided insights into the morphological changes.

SARS-CoV-2 can recruit a heme metabolite to evade antibody immunity.

The coronaviral spike is the dominant viral antigen and the target of neutralizing antibodies. We show that SARS-CoV-2 spike binds biliverdin and bilirubin, the tetrapyrrole products of heme metabolism, with nanomolar affinity. Using cryo-electron microscopy and x-ray crystallography, we mapped the tetrapyrrole interaction pocket to a deep cleft on the spike N-terminal domain (NTD). At physiological concentrations, biliverdin significantly dampened the reactivity of SARS-CoV-2 spike with immune sera and inhibited a subset of neutralizing antibodies. Access to the tetrapyrrole-sensitive epitope is gated by a flexible loop on the distal face of the NTD. Accompanied by profound conformational changes in the NTD, antibody binding requires relocation of the gating loop, which folds into the cleft vacated by the metabolite. Our results indicate that SARS-CoV-2 spike NTD harbors a dominant epitope, access to which can be controlled by an allosteric mechanism that is regulated through recruitment of a metabolite.

SARS-CoV-2 recruits a haem metabolite to evade antibody immunity.

The coronaviral spike is the dominant viral antigen and the target of neutralizing antibodies. We show that SARS-CoV-2 spike binds biliverdin and bilirubin, the tetrapyrrole products of haem metabolism, with nanomolar affinity. Using cryo-electron microscopy and X-ray crystallography we mapped the tetrapyrrole interaction pocket to a deep cleft on the spike N-terminal domain (NTD). At physiological concentrations, biliverdin significantly dampened the reactivity of SARS-CoV-2 spike with immune sera and inhibited a subset of neutralizing antibodies. Access to the tetrapyrrole-sensitive epitope is gated by a flexible loop on the distal face of the NTD. Accompanied by profound conformational changes in the NTD, antibody binding requires relocation of the gating loop, which folds into the cleft vacated by the metabolite. Our results indicate that the virus co-opts the haem metabolite for the evasion of humoral immunity via allosteric shielding of a sensitive epitope and demonstrate the remarkable structural plasticity of the NTD.