Imipramine Inhibits Chikungunya Virus Replication in Human Skin Fibroblasts through Interference with Intracellular Cholesterol Trafficking

Size: px

Start display at page:

Download "Imipramine Inhibits Chikungunya Virus Replication in Human Skin Fibroblasts through Interference with Intracellular Cholesterol Trafficking"

Adrian Stewart
5 years ago
Views:

WICHIT Faculty of Medical Technology Mahidol University JOINT INTERNATIONAL

1 Imipramine Inhibits Chikungunya Virus Replication in Human Skin Fibroblasts through Interference with Intracellular Cholesterol Trafficking SINEEWANLAYA WICHIT Faculty of Medical Technology Mahidol University JOINT INTERNATIONAL TROPICAL MEDICINE MEETING 2017 Amari Watergate Bangkok, Thailand 7 th December 2017

2 ARBOVIRUS: (Arthropod-Borne Virus) Viruses that are transmitted to vertebrate hosts by hematophagous arthropod vectors Mosquito Sand fly Midges Tick Flaviviridae: Flavivirus Togaviridae: Alphavirus Dengue and Zika virus Chikungunya virus 2

ARBOVIRUS: Mosquito Vector Aedes aegypti

Americas, Africa and Europe beginning in 1985 3

3 ARBOVIRUS: Mosquito Vector Aedes aegypti Originated in sub-saharan Africa, spread throughout the tropics centuries ago Aedes albopictus Originated in Asia, spread to the Americas, Africa and Europe beginning in billion people living in Aedes-infested regions (Kraemer et al., 2015) 3

ARBOVIRUS: Geographic distribution 2006 VV 2016 Dengue Virus Isolated in Nagasaki (Human, 1943), Japan 154 countries distributed Furuya-Kanamori et al.

4 ARBOVIRUS: Geographic distribution 2006 VV 2016 Dengue Virus Isolated in Nagasaki (Human, 1943), Japan 154 countries distributed Furuya-Kanamori et al. (2016) Zika Virus V Isolated in Uganda (Monkey, 1947) in Nigeria (Human, 1952) V 62 countries distributed WHO (2016) Chikungunya Virus Isolated in Tanzania (Human, 1952) 99 countries distributed VV Adapted from (Cao-Lormeau, 2016, CDC, 2016, 2017b, Kindhauser et al., 2016, WHO, 2017) Furuya-Kanamori et al. (2016) 4

ARBOVIRUS: Clinical manifestation Fatality rate 0.5-3.5 % No Data 0.

Polyarthralgia (joints pain) Headache Backache, Headache Maculopapular rash Hemorrhage

Maculopapular rash Shock Microcephaly Incidence 0.

5 ARBOVIRUS: Clinical manifestation Fatality rate % No Data 0.1% Common Dengue Zika Chikungunya Two stages of disease Acute illness High fever Polyarthralgia (joints pain) Headache Backache, Headache Maculopapular rash Hemorrhage Conjunctivitis Late stage of illness Polyarthralgia (joints pain) more than 3 years Fever Maculopapular rash Shock Microcephaly Incidence 0.024% Arthralgia Up to 3 years Impact on quality of life leading to economic and community consequences Furuya-Kanamori et al. (2016), Ioos et al. (2014), Patterson et al. (2016) 5

6 CHIKUNGUNYA VIRUS: Geographic distribution CHIKUNGUNYA becomes one of the most important arboviral diseases Italy in 2007 St. Martin in 2013 Thailand in 2013 Tanzania in 1952 La Réunion in 2005 (Silva and Dermody, 2017) 6

CHIKUNGUNYA VIRUS: Geographic distribution Case 100000 10000 1000 100 10 1 First

2012 2013 2014 2015 2016 2017 year Re-emerged Surin Khon Kean Outbreak Loei and

North-Eastern part 1958 4 years 14 years 12 years 13 years 4 years 1962 1976

7 CHIKUNGUNYA VIRUS: Geographic distribution Case First case report Bangkok Outbreak Bangkok Re-emerged Prachin Buri year Re-emerged Surin Khon Kean Outbreak Loei and Phayao Southern part Nong Khai and Nakhon Si Thammarat (October) Outbreak North-Eastern part years 14 years 12 years 13 years 4 years Bureau of Epidemiology, Department of Disease Control, Ministry of Public Health 7

(2017) Two rounds of translation occur to produce 4 non-structural proteins (nsp1, 2, 3 and

8 CHIKUNGUNYA VIRUS: Replication cycle E1-E2 heterodimer Virus enters host cell by endocytosis (Viral Replication complex) Johan et al. (2017) Two rounds of translation occur to produce 4 non-structural proteins (nsp1, 2, 3 and 4) 5 structural proteins (C, E3, E2, 6k, E1) Assembly occurs at the cell surface New viral particles ready to buds from host cell membrane 8

9 Human Skin Fibroblasts Human skin fibroblasts are permissive to CHIKV and ZIKV (Ekchariyawat et al., 2015 and Hamel et al., 2015) Epidermis Fibroblasts V Dermis Hypodermis Missé et al. (2014) Nestle et al. (2009) 9

Cholesterol and Niemann-Pick type C Protein (NPC) V 2008 2009

activity causes NPC disease type C - a rare, autosomal

10 Cholesterol and Niemann-Pick type C Protein (NPC) V Free Cholesterol Kwon et al. (2009) Abnormalities in cholesterol metabolism Inhibits CHIKV infection 2010 Niemann-Pick C (NPC) disease Dysfunctional NPC activity causes NPC disease type C - a rare, autosomal recessive, neurodegenerative disorder Composes of NPC1 and NPC2 Causes cholesterol accumulation in LE/Ls 10

U18666A and IMIPRAMINE A class II amphiphilic hydrophobic amine Inhibits of cholesterol transport from late endosomes/lysosomes to ER Induces a Neimmann-Pick disease type C or NPC-like phenotype

11 U18666A and IMIPRAMINE A class II amphiphilic hydrophobic amine Inhibits of cholesterol transport from late endosomes/lysosomes to ER Induces a Neimmann-Pick disease type C or NPC-like phenotype U18666A inhibits the replication of EBOV (Carette et al., 2011, Shoemaker et al., 2013) DENV (Poh et al., 2012) HIV-1 (Tang, et al., 2009) IMIPRAMINE (Tricyclic antidepressant,tca) Inhibits the production of EBOV in human umbilical vein epithelial cells (Herbert et al., 2015) 11

12 U18666A and CHIKV replication qrt-pcr Vehicle U18666A (μm) Filipin LAMP-1 Overlay U18666A (μm) Vehicle Plaque assay U18666A *** U18666A causes accumulation of cholesterol in the LE/Ls and inhibits CHIKV replication U18666A (μm) 12

13 Imipramine and CHIKV replication Vehicle Imipramine (μm) % infection Flow Cytometry Imipramine (μm) qrt-pcr Imipramine (μm) nsp1 nsp2 nsp3 β-actin CHIKV Imipramine (μm) *** Imipramine causes accumulation of cholesterol in the LE/Ls and inhibits CHIKV replication 13

qrt-pcr Vehicle 10 25 50 75 100 VSV-G CHIKV (E)-VLP Imipramine (μm) Imipramine (μm) *** V

14 Imipramine and CHIKV replication cycle Fusion/entry step Post-entry step (HUH-7 CHIKV replicon) Transduction efficiency * * 85% Rluc activity * 24 h 48 h qrt-pcr Vehicle VSV-G CHIKV (E)-VLP Imipramine (μm) Imipramine (μm) *** V Imipramine exerts its inhibitory effects at least two different stages of the CHIKV infection cycle 14

NPC proteins are crucial for CHIKV replication

fusion Flow cytometry FSC Healthy NPC1 NPC2 50.52 16.

Healthy NPC1 NPC2 qrt-pcr Viral RNA copy number / ng

NPC1 NPC2 *** NPC1 or NPC2 deficiency has a strong

15 NPC proteins are crucial for CHIKV replication Healthy NPC1 NPC2 NPCs-deficient cells impaired CHIKV fusion Flow cytometry FSC Healthy NPC1 NPC ** ** Anti-CHIKV CHIKV (E)-VLP Healthy NPC1 NPC2 qrt-pcr Viral RNA copy number / ng ** Healthy NPC1 NPC2 Healthy NPC1 NPC2 *** NPC1 or NPC2 deficiency has a strong impact on the entry and/or fusion step of the CHIKV life cycle 15

16 Imipramine exerts antiviral activity against Flaviviruses DENV WNV Imipramine (μm) Imipramine (μm) ZIKV *** V Imipramine strongly inhibited, in a dose-dependent manner, the replication of DENV, WNV and ZIKV Imipramine (μm) 16

, Sci Rep. 2017 Jun 9;7(1):3145. https://www.ncbi.nlm.nih.

17 CONCLUSIONS IMIPRAMINE I. Causes accumulation of cholesterol in the LE/Ls II. Exerts its inhibitory effects at least two different stages of the CHIKV infection cycle Wichit S. et al., Sci Rep Jun 9;7(1): III. Exerts broad-spectrum antiviral activity against Alphavirus and Flavivirus 17

18 Dorothée MISSÉ s Team: Dr. Dorothée MISSÉ Dr. Florian LIEGOIS Dr. Julien POMPON Dr. Rodolphe HAMEL Dr. Fodé DIOP Dr. Pauline FERRARIS Dr. Peeraya EKCHARIYAWAT Dr. Manuel PERRERA Dr. Pornapat SURASOMBATPATTANA Loïc TALIGNANI Deborah GARCIA All internship students Dr. Frederic SIMARD All members in MIVEGEC unit 18

Public Health Image Library. CDC/ Cynthia Goldsmith. Image #

Public Health Image Library. CDC/ Cynthia Goldsmith. Image # Zika Virus Fredrick M. Abrahamian, D.O., FACEP, FIDSA Clinical Professor of Medicine UCLA School of Medicine Director of Education Department of Emergency Medicine Olive View-UCLA Medical Center Sylmar,