A second generation leishmanization vaccine with a markerless attenuated Leishmania major strain using CRISPR gene editing

Wen-Wei Zhang; Subir Karmakar; Sreenivas Gannavaram; Ranadhir Dey; Patrick Lypaczewski; Nevien Ismail; Abid Siddiqui; Vahan Simonyan; Fabiano Oliveira; Iliano V Coutinho-Abreu; Thiago DeSouza-Vieira; Claudio Meneses; James Oristian; Tiago D Serafim; Abu Musa; Risa Nakamura; Noushin Saljoughian; Greta Volpedo; Monika Satoskar; Sanika Satoskar; Pradeep K Dagur; J Philip McCoy; Shaden Kamhawi; Jesus G Valenzuela; Shinjiro Hamano; Abhay R Satoskar; Greg Matlashewski; Hira L Nakhasi

doi:10.1038/s41467-020-17154-z

A second generation leishmanization vaccine with a markerless attenuated Leishmania major strain using CRISPR gene editing

Nat Commun. 2020 Jul 10;11(1):3461. doi: 10.1038/s41467-020-17154-z.

Authors

Wen-Wei Zhang^#¹, Subir Karmakar^#², Sreenivas Gannavaram^#², Ranadhir Dey^#², Patrick Lypaczewski¹, Nevien Ismail², Abid Siddiqui², Vahan Simonyan², Fabiano Oliveira³, Iliano V Coutinho-Abreu³, Thiago DeSouza-Vieira³, Claudio Meneses³, James Oristian³, Tiago D Serafim³, Abu Musa⁴, Risa Nakamura⁴, Noushin Saljoughian⁵, Greta Volpedo⁵, Monika Satoskar^{2

6}, Sanika Satoskar^{2

6}, Pradeep K Dagur⁷, J Philip McCoy⁷, Shaden Kamhawi³, Jesus G Valenzuela³, Shinjiro Hamano⁴, Abhay R Satoskar⁸, Greg Matlashewski⁹, Hira L Nakhasi¹⁰

Affiliations

¹ Department of Microbiology and Immunology, McGill University, Montreal, QC, H3A 2B4, Canada.
² Division of Emerging and Transfusion Transmitted Diseases, CBER, FDA, Silver Spring, MD, 20993, USA.
³ Vector Molecular Biology Section, Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, MD, 20852, USA.
⁴ Department of Parasitology, Institute of Tropical Medicine (NEKKEN), The Joint Usage/Research Center on Tropical Disease, Nagasaki University, Nagasaki, Japan and Nagasaki University Graduate School of Biomedical Sciences Doctoral Leadership Program, Nagasaki, Japan.
⁵ Department of Pathology and Microbiology, Ohio State University, Columbus, OH, 43210, USA.
⁶ Northeast Ohio Medical University, Rootstown, Ohio, 44272, USA.
⁷ National Institute of Heart, Lung and Blood Institute, NIH, Bethesda, MD, 20852, USA.
⁸ Department of Pathology and Microbiology, Ohio State University, Columbus, OH, 43210, USA. Abhay.Satoskar@osumc.edu.
⁹ Department of Microbiology and Immunology, McGill University, Montreal, QC, H3A 2B4, Canada. greg.matlashewski@mcgill.ca.
¹⁰ Division of Emerging and Transfusion Transmitted Diseases, CBER, FDA, Silver Spring, MD, 20993, USA. Hira.Nakhasi@fda.hhs.gov.

^# Contributed equally.

Abstract

Leishmaniasis is a neglected tropical disease caused by Leishmania protozoa transmitted by infected sand flies. Vaccination through leishmanization with live Leishmania major has been used successfully but is no longer practiced because it resulted in occasional skin lesions. A second generation leishmanization is described here using a CRISPR genome edited L. major strain (LmCen^-/-). Notably, LmCen^-/- is a genetically engineered centrin gene knock-out mutant strain that is antibiotic resistant marker free and does not have detectable off-target mutations. Mice immunized with LmCen^-/- have no visible lesions following challenge with L. major-infected sand flies, while non-immunized animals develop large and progressive lesions with a 2-log fold higher parasite burden. LmCen^-/- immunization results in protection and an immune response comparable to leishmanization. LmCen^-/- is safe since it is unable to cause disease in immunocompromised mice, induces robust host protection against vector sand fly challenge and because it is marker free, can be advanced to human vaccine trials.

Publication types

Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

MeSH terms

Animals
CD4-Positive T-Lymphocytes / drug effects
CD4-Positive T-Lymphocytes / metabolism
Clustered Regularly Interspaced Short Palindromic Repeats / genetics*
Dexamethasone / pharmacology
Female
Flow Cytometry
Gene Editing
Genetic Engineering
Humans
Immunosuppression Therapy
Leishmania major / genetics*
Leishmania major / pathogenicity*
Macrophages / metabolism
Mice
Mice, Inbred BALB C
Mice, Inbred C57BL
Psychodidae / parasitology
Reverse Transcriptase Polymerase Chain Reaction
Vaccines, Attenuated / therapeutic use*

Substances

Vaccines, Attenuated
Dexamethasone

Grants and funding

CIHR/Canada