Chikungunya viruses that escape monoclonal antibody therapy are clinically attenuated, stable, and not purified in mosquitoes.
Chikungunya virus (CHIKV) is a reemerging mosquito-transmitted alphavirus that causes epidemics of debilitating polyarthritis in humans. A prior study identified two anti-CHIKV monoclonal antibodies ([MAbs] CHK-152 and CHK-166) against the E2 and E1 structural proteins, which had therapeutic efficacy in immunocompetent and immunocompromised mice. Combination MAb therapy was required as administration of a single MAb resulted in the rapid selection of neutralization escape variants and treatment failure in mice. Here, we initially evaluated the efficacy of combination MAb therapy in a nonhuman primate model of CHIKV infection. Treatment of rhesus macaques with CHK-152 and CHK-166 reduced viral spread and infection in distant tissue sites and also neutralized reservoirs of infectious virus. Escape viruses were not detected in the residual viral RNA present in tissues and organs of rhesus macaques. To evaluate the possible significance of MAb resistance, we engineered neutralization escape variant viruses (E1-K61T, E2-D59N, and the double mutant E1-K61T E2-D59N) that conferred resistance to CHK-152 and CHK-166 and tested them for fitness in mosquito cells, mammalian cells, mice, and Aedes albopictus mosquitoes. In both cell culture and mosquitoes, the mutant viruses grew equivalently and did not revert to wild-type (WT) sequence. All escape variants showed evidence of mild clinical attenuation, with decreased musculoskeletal disease at early times after infection in WT mice and a prolonged survival time in immunocompromised Ifnar1(-/-) mice. Unexpectedly, this was not associated with decreased infectivity, and consensus sequencing from tissues revealed no evidence of reversion or compensatory mutations. Competition studies with CHIKV WT also revealed no fitness compromise of the double mutant (E1-K61T E2-D59N) neutralization escape variant in WT mice. Collectively, our study suggests that neutralization escape viruses selected during combination MAb therapy with CHK-152 plus CHK-166 retain fitness, cause less severe clinical disease, and likely would not be purified during the enzootic cycle.


OBJECTIVE
Chikungunya virus (CHIKV) causes explosive epidemics of acute and chronic arthritis in humans in Africa, the Indian subcontinent, and Southeast Asia and recently has spread to the New World. As there are no approved vaccines or therapies for human use, the possibility of CHIKV-induced debilitating disease is high in many parts of the world. To this end, our laboratory recently generated a combination monoclonal antibody therapy that aborted lethal and arthritogenic disease in wild-type and immunocompromised mice when administered as a single dose several days after infection. In this study, we show the efficacy of the antibody combination in nonhuman primates and also evaluate the significance of possible neutralization escape mutations in mosquito and mammalian cells, mice, and Aedes albopictus vector mosquitoes. Our experiments show that escape viruses from combination antibody therapy cause less severe CHIKV clinical disease, retain fitness, and likely would not be purified by mosquito vectors.
Department of Molecular Microbiology, Washington University School of Medicine, Saint Louis, Missouri, USA.
Pankaj
P
Pal
Department of Medicine, Washington University School of Medicine, Saint Louis, Missouri, USA.
Julie M
JM
Fox
Departments of Microbiology and Immunology, University of Colorado School of Medicine, Aurora, Colorado, USA.
David W
DW
Hawman
Biosecurity Research Institute and Department of Diagnostic Medicine and Pathobiology, Kansas State University, Manhattan, Kansas, USA.
Yan-Jang S
YJ
Huang
Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, Oregon, USA Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA Division of Pathobiology and Immunology, Oregon National Primate Research Center, Beaverton, Oregon, USA University of California-Riverside, Riverside, California, USA.
Ilhem
I
Messaoudi
Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, Oregon, USA.
Craig
C
Kreklywich
Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, Oregon, USA.
Michael
M
Denton
Division of Pathobiology and Immunology, Oregon National Primate Research Center, Beaverton, Oregon, USA.
Alfred W
AW
Legasse
Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, Oregon, USA.
Patricia P
PP
Smith
MacroGenics, Rockville, Maryland, USA.
Syd
S
Johnson
Division of Pathobiology and Immunology, Oregon National Primate Research Center, Beaverton, Oregon, USA.
Michael K
MK
Axthelm
Biosecurity Research Institute and Department of Diagnostic Medicine and Pathobiology, Kansas State University, Manhattan, Kansas, USA.
Dana L
DL
Vanlandingham
Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, Oregon, USA Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA Division of Pathobiology and Immunology, Oregon National Primate Research Center, Beaverton, Oregon, USA.
Daniel N
DN
Streblow
Biosecurity Research Institute and Department of Diagnostic Medicine and Pathobiology, Kansas State University, Manhattan, Kansas, USA.
Stephen
S
Higgs
Departments of Microbiology and Immunology, University of Colorado School of Medicine, Aurora, Colorado, USA.
Thomas E
TE
Morrison
Department of Molecular Microbiology, Washington University School of Medicine, Saint Louis, Missouri, USA Department of Medicine, Washington University School of Medicine, Saint Louis, Missouri, USA Department of Pathology and Immunology, Washington University School of Medicine, Saint Louis, Missouri, USA diamond@borcim.wustl.edu.
Michael S
MS
Diamond
D000330:Aedes / Q000821:virology*; D018354:Alphavirus Infections / Q000628:therapy / Q000821:virology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000627:therapeutic use*; D000914:Antibodies, Viral / Q000276:immunology / Q000627:therapeutic use*; D065632:Chikungunya Fever; D002646:Chikungunya virus / Q000276:immunology / Q000302:isolation & purification / Q000472:pathogenicity / Q000502:physiology*; D004195:Disease Models, Animal; D005260:Female; D018398:Homeodomain Proteins; D007167:Immunotherapy / Q000379:methods*; D008253:Macaca mulatta; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D016019:Survival Analysis; D014774:Virulence; D014779:Virus Replication
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D018398:Homeodomain Proteins; C489656:Not protein, mouse