title: New Verapamil Analogs Inhibit Intracellular Mycobacteria without Affecting the Functions of Mycobacterium-Specific T Cells.
abstract: There is a growing interest in repurposing mycobacterial efflux pump inhibitors, such as verapamil, for tuberculosis (TB) treatment. To aid in the design of better analogs, we studied the effects of verapamil on macrophages and Mycobacterium tuberculosis-specific T cells. Macrophage activation was evaluated by measuring levels of nitric oxide, tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), and gamma interferon (IFN-γ). Since verapamil is a known autophagy inducer, the roles of autophagy induction in the antimycobacterial activities of verapamil and norverapamil were studied using bone marrow-derived macrophages from ATG5(flox/flox) (control) and ATG5(flox/flox) Lyz-Cre mice. Our results showed that despite the well-recognized effects of verapamil on calcium channels and autophagy, its action on intracellular M. tuberculosis does not involve macrophage activation or autophagy induction. Next, the effects of verapamil and norverapamil on M. tuberculosis-specific T cells were assessed using flow cytometry following the stimulation of peripheral blood mononuclear cells from TB-skin-test-positive donors with M. tuberculosis whole-cell lysate for 7 days in the presence or absence of drugs. We found that verapamil and norverapamil inhibit the expansion of M. tuberculosis-specific T cells. Additionally, three new verapamil analogs were found to inhibit intracellular Mycobacterium bovis BCG, and one of the three analogs (KSV21) inhibited intracellular M. tuberculosis replication at concentrations that did not inhibit M. tuberculosis-specific T cell expansion. KSV21 also inhibited mycobacterial efflux pumps to the same degree as verapamil. More interestingly, the new analog enhances the inhibitory activities of isoniazid and rifampin on intracellular M. tuberculosis. In conclusion, KSV21 is a promising verapamil analog on which to base structure-activity relationship studies aimed at identifying more effective analogs.
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Allergy and Immunology, Saint Louis University, St. Louis, Missouri, USA abateg@slu.edu.
authors.forename: Getahun
authors.initials: G
authors.lastname: Abate
authors.affiliation: Centre for World Health and Medicine, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Peter G
authors.initials: PG
authors.lastname: Ruminiski
authors.affiliation: Department of Chemistry and South African Medical Research Council Drug Discovery and Development Research Unit, University of Cape Town, Rondebosch, South Africa.
authors.forename: Malkeet
authors.initials: M
authors.lastname: Kumar
authors.affiliation: Department of Chemistry and South African Medical Research Council Drug Discovery and Development Research Unit, University of Cape Town, Rondebosch, South Africa.
authors.forename: Kawaljit
authors.initials: K
authors.lastname: Singh
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Allergy and Immunology, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Fahreta
authors.initials: F
authors.lastname: Hamzabegovic
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Allergy and Immunology, Saint Louis University, St. Louis, Missouri, USA Department of Molecular Biology, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Daniel F
authors.initials: DF
authors.lastname: Hoft
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Allergy and Immunology, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Eickhoff
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, Allergy and Immunology, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Asmir
authors.initials: A
authors.lastname: Selimovic
authors.affiliation: Centre for World Health and Medicine, Saint Louis University, St. Louis, Missouri, USA.
authors.forename: Mary
authors.initials: M
authors.lastname: Campbell
authors.affiliation: Department of Chemistry and South African Medical Research Council Drug Discovery and Development Research Unit, University of Cape Town, Rondebosch, South Africa Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Rondebosch, South Africa.
authors.forename: Kelly
authors.initials: K
authors.lastname: Chibale
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects; D006801:Humans; D007538:Isoniazid / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000382:microbiology; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects / Q000382:microbiology; D008822:Mice, Transgenic; D009163:Mycobacterium bovis / Q000187:drug effects; D009169:Mycobacterium tuberculosis / Q000187:drug effects*; D012293:Rifampin / Q000031:analogs & derivatives / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects* / Q000382:microbiology; D014700:Verapamil / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: C000612283:KSV21 compound; C000612284:MKV4 compound; C016904:norverapamil; D014700:Verapamil; D007538:Isoniazid; D012293:Rifampin