Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation

One of the main hallmarks of tuberculosis (TB) is the ability of the causative agent to transform into a stage of dormancy and the capability of long persistence in the host phagocytes. It is believed that approximately one-third of the population of the world is latently infected with Mycobacterium...

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Autores principales:	Kata Horváti, Kinga Fodor, Bernadett Pályi, Judit Henczkó, Gyula Balka, Gergő Gyulai, Éva Kiss, Beáta Biri-Kovács, Zsuzsanna Senoner, Szilvia Bősze
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	tuberculosis cell penetrating peptides isoniazid Dhvar4 Transwell spheroid Immunologic diseases. Allergy RC581-607
Acceso en línea:	https://doaj.org/article/b74306c2370f46539db589adcc8c01a1
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spelling	oai:doaj.org-article:b74306c2370f46539db589adcc8c01a12021-11-12T04:42:56ZNovel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation1664-322410.3389/fimmu.2021.750496https://doaj.org/article/b74306c2370f46539db589adcc8c01a12021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fimmu.2021.750496/fullhttps://doaj.org/toc/1664-3224One of the main hallmarks of tuberculosis (TB) is the ability of the causative agent to transform into a stage of dormancy and the capability of long persistence in the host phagocytes. It is believed that approximately one-third of the population of the world is latently infected with Mycobacterium tuberculosis (Mtb), and 5%–10% of these individuals can develop clinical manifestations of active TB even decades after the initial infection. In this latent, intracellular form, the bacillus is shielded by an extremely robust cell wall and becomes phenotypically resistant to most antituberculars. Therefore, there is a clear rationale to develop novel compounds or carrier-conjugated constructs of existing drugs that are effective against the intracellular form of the bacilli. In this paper, we describe an experimental road map to define optimal candidates against intracellular Mtb and potential compounds effective in the therapy of latent TB. To validate our approach, isoniazid, a first-line antitubercular drug was employed, which is active against extracellular Mtb in the submicromolar range, but ineffective against the intracellular form of the bacteria. Cationic peptide conjugates of isoniazid were synthesized and employed to study the host-directed drug delivery. To measure the intracellular killing activity of the compounds, Mtb-infected MonoMac-6 human monocytic cells were utilized. We have assessed the antitubercular activity, cytotoxicity, membrane interactions in combination with internalization efficacy, localization, and penetration ability on interface and tissue-mimicking 3D models. Based on these in vitro data, most active compounds were further evaluated in vivo in a murine model of TB. Intraperitoneal infectious route was employed to induce a course of slowly progressive and systemic disease. The well-being of the animals, monitored by the body weight, allows a prolonged experimental setup and provides a great opportunity to test the long-term activity of the drug candidates. Having shown the great potency of this simple and suitable experimental design for antimicrobial research, the proposed novel assay platform could be used in the future to develop further innovative and highly effective antituberculars.Kata HorvátiKata HorvátiKinga FodorBernadett PályiJudit HenczkóGyula BalkaGergő GyulaiÉva KissBeáta Biri-KovácsZsuzsanna SenonerSzilvia BőszeSzilvia BőszeFrontiers Media S.A.articletuberculosiscell penetrating peptidesisoniazidDhvar4TranswellspheroidImmunologic diseases. AllergyRC581-607ENFrontiers in Immunology, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	tuberculosis cell penetrating peptides isoniazid Dhvar4 Transwell spheroid Immunologic diseases. Allergy RC581-607
spellingShingle	tuberculosis cell penetrating peptides isoniazid Dhvar4 Transwell spheroid Immunologic diseases. Allergy RC581-607 Kata Horváti Kata Horváti Kinga Fodor Bernadett Pályi Judit Henczkó Gyula Balka Gergő Gyulai Éva Kiss Beáta Biri-Kovács Zsuzsanna Senoner Szilvia Bősze Szilvia Bősze Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
description	One of the main hallmarks of tuberculosis (TB) is the ability of the causative agent to transform into a stage of dormancy and the capability of long persistence in the host phagocytes. It is believed that approximately one-third of the population of the world is latently infected with Mycobacterium tuberculosis (Mtb), and 5%–10% of these individuals can develop clinical manifestations of active TB even decades after the initial infection. In this latent, intracellular form, the bacillus is shielded by an extremely robust cell wall and becomes phenotypically resistant to most antituberculars. Therefore, there is a clear rationale to develop novel compounds or carrier-conjugated constructs of existing drugs that are effective against the intracellular form of the bacilli. In this paper, we describe an experimental road map to define optimal candidates against intracellular Mtb and potential compounds effective in the therapy of latent TB. To validate our approach, isoniazid, a first-line antitubercular drug was employed, which is active against extracellular Mtb in the submicromolar range, but ineffective against the intracellular form of the bacteria. Cationic peptide conjugates of isoniazid were synthesized and employed to study the host-directed drug delivery. To measure the intracellular killing activity of the compounds, Mtb-infected MonoMac-6 human monocytic cells were utilized. We have assessed the antitubercular activity, cytotoxicity, membrane interactions in combination with internalization efficacy, localization, and penetration ability on interface and tissue-mimicking 3D models. Based on these in vitro data, most active compounds were further evaluated in vivo in a murine model of TB. Intraperitoneal infectious route was employed to induce a course of slowly progressive and systemic disease. The well-being of the animals, monitored by the body weight, allows a prolonged experimental setup and provides a great opportunity to test the long-term activity of the drug candidates. Having shown the great potency of this simple and suitable experimental design for antimicrobial research, the proposed novel assay platform could be used in the future to develop further innovative and highly effective antituberculars.
format	article
author	Kata Horváti Kata Horváti Kinga Fodor Bernadett Pályi Judit Henczkó Gyula Balka Gergő Gyulai Éva Kiss Beáta Biri-Kovács Zsuzsanna Senoner Szilvia Bősze Szilvia Bősze
author_facet	Kata Horváti Kata Horváti Kinga Fodor Bernadett Pályi Judit Henczkó Gyula Balka Gergő Gyulai Éva Kiss Beáta Biri-Kovács Zsuzsanna Senoner Szilvia Bősze Szilvia Bősze
author_sort	Kata Horváti
title	Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
title_short	Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
title_full	Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
title_fullStr	Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
title_full_unstemmed	Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation
title_sort	novel assay platform to evaluate intracellular killing of mycobacterium tuberculosis: in vitro and in vivo validation
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/b74306c2370f46539db589adcc8c01a1
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Novel Assay Platform to Evaluate Intracellular Killing of Mycobacterium tuberculosis: In Vitro and In Vivo Validation

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