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Chemoprevention of bilirubin encephalopathy with a nanoceutical agent

Abstract

Background

Targeted rapid degradation of bilirubin has the potential to thwart incipient bilirubin encephalopathy. We investigated a novel spinel-structured citrate-functionalized trimanganese tetroxide nanoparticle (C-Mn3O4 NP, the nanodrug) to degrade both systemic and neural bilirubin loads.

Method

Severe neonatal unconjugated hyperbilirubinemia (SNH) was induced in neonatal C57BL/6j mice model with phenylhydrazine (PHz) intoxication. Efficiency of the nanodrug on both in vivo bilirubin degradation and amelioration of bilirubin encephalopathy and associated neurobehavioral sequelae were evaluated.

Results

Single oral dose (0.25 mg kg−1 bodyweight) of the nanodrug reduced both total serum bilirubin (TSB) and unconjugated bilirubin (UCB) in SNH rodents. Significant (p < 0.0001) UCB and TSB-degradation rates were reported within 4–8 h at 1.84 ± 0.26 and 2.19 ± 0.31 mg dL−1 h−1, respectively. Neural bilirubin load was decreased by 5.6 nmol g−1 (p = 0.0002) along with improved measures of neurobehavior, neuromotor movements, learning, and memory. Histopathological studies confirm that the nanodrug prevented neural cell reduction in Purkinje and substantia nigra regions, eosinophilic neurons, spongiosis, and cell shrinkage in SNH brain parenchyma. Brain oxidative status was maintained in nanodrug-treated SNH cohort. Pharmacokinetic data corroborated the bilirubin degradation rate with plasma nanodrug concentrations.

Conclusion

This study demonstrates the in vivo capacity of this novel nanodrug to reduce systemic and neural bilirubin load and reverse bilirubin-induced neurotoxicity. Further compilation of a drug-safety-dossier is warranted to translate this novel therapeutic chemopreventive approach to clinical settings.

Impact

  • None of the current pharmacotherapeutics treat severe neonatal hyperbilirubinemia (SNH) to prevent risks of neurotoxicity.

  • In this preclinical study, a newly investigated nano-formulation, citrate-functionalized Mn3O4 nanoparticles (C-Mn3O4 NPs), exhibits bilirubin reduction properties in rodents.

  • Chemopreventive properties of this nano-formulation demonstrate an efficacious, efficient agent that appears to be safe in these early studies.

  • Translation of C-Mn3O4 NPs to prospective preclinical and clinical trials in appropriate in vivo models should be explored as a potential novel pharmacotherapy for SNH.

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Fig. 1: In vivo bilirubin degradation efficacy of C-Mn3O4 NPs nanodrug in SNH model (postnatal age 10 days).
Fig. 2: Effect of the nanodrug on prevention of SNH-associated neurotoxicity.
Fig. 3: Effect of the nanodrug on SNH-associated brain (cerebellar) oxidative distress.
Fig. 4: Effect of the nanodrug on SNH-associated neuromotor function.
Fig. 5: “Long-term” effects in learning and memory.
Fig. 6: Pharmacokinetics and pharmacodynamics of the nanodrug.

Data availability

All data that support the findings of this study are available within the published article. Study-related additional data will be available from S.K.P. upon legitimate request.

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Funding

This study has been supported by Abdul Kalam Technology Innovation National Fellowship (INAE/121/AKF), Indian National Academy of Engineering (INAE), Science and Engineering Research Board (SERB), Department of Science and Technology (DST), Govt. of India and financial grant under BOOST scheme (339/WBBDC/1P-2/2013), Department of Biotechnology (DBT-WB), Govt. of West Bengal. M.D. received Junior Research Fellowship (JRF) from University Grants Commission (UGC), Govt. of India.

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Contributions

A.A., V.K.B., A.K.M and S.K.P. conceived and planned the study. A.A., S.M., M.D., N.P., and S.D. performed animal experiments and biochemical studies. A.A. executed numerical calculations and statistical analysis. A.A., S.D., S.S.B., D.P., A.K.M., V.K.B, and S.K.P. contributed to the interpretation of the results. A.K.D. carried out and evaluated the histological studies. A.A. and V.K.B. took the lead in writing the manuscript. All authors provided critical feedback and helped in shaping the research, analysis, interpretation, and writing. All authors reviewed, edited, and approved the final version of the manuscript.

Corresponding author

Correspondence to Samir Kumar Pal.

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The authors declare no competing interests.

Ethics approval and consent to participate

This study does not involve human subject or clinical study. Thus, patient consent was not required. All animal studies and experimental procedures were performed at Central Animal Facility, Department of Zoology, Uluberia College, India (Reg. No.: 2057/GO/ReRcBi/S/19/CPCSEA) following the protocol approved by the Institutional Animal Ethics Committee (Ref: 02/S/UC-IAEC/01/2019) as per standard guideline of Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), Govt. of India.

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Adhikari, A., Bhutani, V.K., Mondal, S. et al. Chemoprevention of bilirubin encephalopathy with a nanoceutical agent. Pediatr Res (2022). https://doi.org/10.1038/s41390-022-02179-5

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