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Lactobacillus Crispatus WHAT 2nd Microbiome workshop WHEN 17 & 18 November 2016 WHERE Masur Auditorium at NIH Campus, Bethesda MD Modulating the Vaginal Microbiome to Prevent HIV Infection Laurel Lagenaur For more information: www.virology-education.com Conflict of Interest I work for Osel Inc., a microbiome company developing live biotherapeutic products to prevent diseases in women The Vaginal Microbiome and HIV Infection What’s normal /healthy/ optimal and what’s not? • Ravel Community Groups • Lactobacillus dominant vs. dysbiosis Why are Lactobacilli important for HIV prevention? • Dysbiosis = Inflammation = Increased risk of HIV acquisition • Efficacy of Pre-Exposure Prophylaxis decreased Modulation of the vaginal microbiome • LACTIN-V, live biotherapeutic-Lactobacillus crispatus • Ongoing clinical trials How we can use Lactobacilli and go a step further • Genetically modified Lactobacillus to prevent HIV acquisition HIV is Transmitted Across Mucosal Surfaces • HIV infection in women occurs in the mucosa of the vagina and cervix vagina cervix • Infection of underlying target cells All mucosal surfaces are continuously exposed to a Herrera and Shattock, community of microorganisms Curr Top Microbiol Immunol 2013 Vaginal Microbiome: Ravel Community Groups Vaginal microbiomes clustered into 5 groups: Group V L. jensenii Group II L. gasseri 4 were dominated by Group I L. crispatus Lactobacillus, Group III L. iners whereas the 5th had lower proportions of lactic acid bacteria and Group 4 Diversity higher proportions of Prevotella, strictly anaerobic Sneathia, Megasphaera, organisms Atopobium Shannon Diversity: Ravel et al. PNAS 2011 low species high species richness/evenness richness/evenness Vaginal Microbiome Lactobacilli-dominated Healthy Vagina Dysbiosis • Low diversity, low vaginal pH • Antagonize pathogens Low Diversity High Diversity • Non-inflammatory Low Nugent High Nugent pH Bacterial vaginosis >5.5 • Decrease H O -producing Lactobacilli 2 2 pH • Increased bacterial diversity 3.6-4.5 • High pH, odor, discharge • Increased risk of STI and HIV Bacterial communities in O + Adapted from Srinivasan 2012 Vaginal Microbiota vs. Host Inflammatory Responses Inflammatory Host vs. Microbiota Vaginal • CT 1 CT Community types Community CT 4 CT Alpha diversity Alpha IFN-γ (pg/ml) IL-1α (pg/ml) 1β, TNF 1β, cytokines of proinflammatory levels Increased IL- IL-\) IL- IL-\) - \) IL-\) \) IL-\) IFN α, Anahtar - γ , IL , et al et - IL-10 (pg/ml) IL-1β (pg/ml) IL and 10 . Immunity . 2015 Immunity - IL- 8 IL- IL-\) IL-\) \) IL-\) \) IL-\) - IL-8 (pg/ml) TNF-α (pg/ml) IL - 1α , IL IL- IL- - IL-\) IL-\) \) IL-\) \) IL-\) Genital Inflammation is Linked to HIV infection Abundance% Abundance % Abundance % Abundance0 >50 % Abundance 0 0 0 0 >50>50 >50 >50 Lactobacillus crispatus Lactobacillus crispatus Lactobacillus crispatus Lactobacillus iners Lactobacillus iners South African womenLactobacillus iners in the Prevotella bivia Prevotella bivia Prevotella bivia LactobacilluSneathias crispatus sp 99 Sneathia sp 99 Sneathia sp 99 Shuttleworthia sp 9 [BVAB-1] Shuttleworthia sp 9 [BVAB-1] Shuttleworthia sp 9 [BVAB-1] LactobacilluAerococcuss iners christensenii Aerococcus CAPRISAchristensenii study withAerococcus christensenii P. bivia were Veillonella montpellierensis Veillonella montpellierensis Veillonella montpellierensis PrevotellaPrevotella bivia amnii Prevotella amnii Prevotella amnii Atopobium vaginae Atopobium vaginae Atopobium vaginae SneathiaMegasphaera sp 99 sp 9 Megasphaera sp 9 19 times moreMegasphaera likely sp 9 to have Lactobacillus gasseri Lactobacillus gasseri Lactobacillus gasseri ShuttleworthiaMycoplasma-like sp 9 [BVAB-1] sp 9 Mycoplasma-like sp 9 Mycoplasma-like sp 9 AerococcusMycoplasma christensenii hominis Mycoplasma hominis genital inflammationMycoplasma hominis and 13 Atopobium sp 6 Atopobium sp 6 Atopobium sp 6 Shuttleworthia sp 58 [BVAB-1] Shuttleworthia sp 58 [BVAB-1] Shuttleworthia sp 58 [BVAB-1] VeilloStreptococcusnella montpellierensis agalactiae Streptococcus agalactiae Streptococcus agalactiae Lactobacillus sp 91 Lactobacillus sptimes 91 more likelyLactobacillu tos sp 91acquire HIV PrevotellaStaphylococcus amnii epidermidis Staphylococcus epidermidis Staphylococcus epidermidis AtopobiumSneathia vaginae sanguinegens Sneathia sanguinegens Sneathia sanguinegens Prevotella timonensis Prevotella timonensis Prevotella timonensis MegasphaeraCoriobacteriaceae sp 9 sp 9 Coriobacteriaceae sp 9 Coriobacteriaceae sp 9 Prevotella sp 101 Prevotella sp 101 Prevotella sp 101 LactobacilluUreaplasmas gasseri sp 1 Ureaplasma sp 1 Ureaplasma sp 1 Granulicatella elegans Granulicatella elegans Granulicatella elegans Mycoplasma-likeWillia sp 9 msia muralis Williamsia muralis Williamsia muralis MycoplasmaHIV hominis status HIV status HIV status Cytokine group Cytokine group Cytokine group 150 150 150 Trichomonas vaginalis Trichomonas vaginalis Trichomonas vaginalis d d d Atopobium sp 6 s s s e e e e e e v v v i i i r r 100 100 r 100 Chlamydia trachomatis Chlamydia trachomatis Chlamydia trachomatis c c c e e e e e e s s s p p p b b b Neisseria gonorrhoeae Neisseria gonorrhoeae Neisseria gonorrhoeae s s 50 50 s 50 Shuttleworthia sp 58 [BVAB-1] O O O 0 0 0 HSV-2 HSV-2 HSV-2 StreptococcusHSV agalactiae -1 HSV-1 HSV-1 Lactobacillus sp 91 Staphylococcus epidermidis Sneathia sanguinegens Prevotella timonensis Coriobacteriaceae sp 9 Prevotella sp 101 Ureaplasma sp 1 Granulicatella elegans Williamsia muralis HIV status Cytokine group 150 Trichomonas vaginalis d s e e v i r 100 Chlamydia trachomatis c e e s p b Neisseria gonorrhoeae s 50 O 0 HSV-2 HSV-1 Vaginal Microbiota May Influence Tenofovir Efficacy • Tenofovir disoproxil - Nucleotide analog reverse-transcriptase inhibitor used in Pre-Exposure Prophylaxis • Delivered as a 1% intravaginal gel (before and after sex) Tenofovir gel effective against HIV with Lactobacillus- dominance Lactobacilli-dominated Non-Lactobacillus dominant 0.40 0.40 A. LaA.ct obaLacctobaillusc idllousm indoamntinant 0.40 0.40 B. NoB.n No-Lanc-Latobactobacilluscil ldusom dinomanintant 0.35 0.35 0.35 0.35 n 61% efficacy n 18% efficacy n EfficaEfcyf,i c a6c1y%, 61% n EfficEfacfyic,a 1c8y,% 18% o o 0.30o 0.30 0.30o i 0.30 i i i t t t t c 95% CI, 11 to 84% c 95% CI, -77 to 63% c 95% CI, 11 to 84% c 95% CI, -77 to 63% e e e e f f f f n n 0.25n 0.25n i 0.25 i 0.25 i i P=0.644 V V P=0.644 V I V I I I H H H H f 0.20 f 0.20 f 0.20 f 0.20 o o o o y y t y t y i t i t l i l i i l i 0.15 0.15l i 0.15 i 0.15 b b b b a a a a b b b b o o o r 0.10 P=0.013 o r 0.10 r 0.10 P=0.013 r 0.10 P P P P 0.05 0.05 0.05 0.05 HR = 0.39 (95% CI: 0.20; 0.83) HR = 0.82 (95% CI: 0.40; 1.65) 0.00 HR = 0.39 (95% CI: 0.20; 0.83) HR = 0.82 (95% CI: 0.40; 1.65) 0.00 0.00 0.00 0.0 0.5 1.0 1.5 2.0 2.5 0.0 0.5 1.0 1.5 2.0 2.5 0.0 0.0 0.5 0.5 1.0 1.0 1.5 1.5 2.0 2.0 2.5 2.5 Years in Study Years in Study Years in Study Years in Study Tenofovir Placebo Tenofovir Placebo Tenofovir Placebo Tenofovir Placebo Lactobacillus dominant women at risk (Cumulative number of infections) Non-Lactobacillus dominant women at risk (Cumulative number of infections) Lactobacillus dominant women at risk (Cumulative number of infections) Non-Lactobacillus dominant women at risk (Cumulative number of infections) Tenofovir 205 (0) 204 (1) 183 (3) 129 (7) 46 (9) 0 (9) Tenofovir 140 (0) 137 (2) 123 (6) 87 (10) 32 (13) 0 (14) Tenofovir 205 (0) 204 (1) 183 (3) 129 (7) 46 (9) 0 (9) Tenofovir 140 (0) 137 (2) 123 (6) 87 (10) 32 (13) 0 (14) Placebo 202 (0) 196 (4) 173 (12) 123 (19) 51 (22) 0 (22) Placebo 141 (0) 137 (4) 116 (12) 84 (17) 28 (17) 0 (17) Placebo 202 (0) 196 (4) 173 (12) 123 (19) 51 (22) 0 (22) Placebo 141 (0) 137 (4) 116 (12) 84 (17) 28 (17) 0 (17) Tenofovir was rapidly depleted by Gardnerella but not Lactobacillus Tenofovir (supernatant) Tenofovir (cell) 1.5 Abiotic e g 0.003 L. iners L. iners n a ) h G. vaginalis G. vaginalis L C 1.0 m / d l g 0.002 o m F ( r i r i v 0.5 v o o f f o o 0.001 n n e e T T 0.0 0 10 20 30 0.000 Time (hours) 0 10 20 30 Time (hours) 4 hours: G. vag vs. L. iners: P=0.002 4 hours: G. vag vs Abiotic: P=0.005 G. vag vs. L. iners: P<0.001 24 hours: 24 hours: G. vag vs. L. iners: P<0.001 G. vag vs. L. iners: P<0.001 G. vag vs Abiotic: P<0.001 What Can We Do to Decrease HIV Infection in Women? • Treat their Bacterial vaginosis- Metronidazole Lactobacillus dominant Dysbiosis Abx Post-Abx Restoration ↓ H O + Lactobacillus 2 2 Live Biotherapeutic Product (LBP) Growth of diverse commensal and pathogenic bacteria LACTIN-V: Lactobacillus crispatus CTV-05 • Give women exogenous Lactobacillus Osel Product LACTIN-V (Lactobacillus crispatus) for Treatment of Bacterial Vaginosis LACTIN-V Clinical Studies Phase 1 • Safe and well-tolerated at all dose levels* * Hemmerling et al., S Trans Dis 2009 Phase 2a • Following Metronidazole treatment • Colonization up to 78% of women treated for BV** ** Hemmerling et al., S Trans Dis 2010 • Colonization correlated with reduced BV recurrence Phase 2b (sponsor NIAID/DMID ) • Prevention of recurrent BV • 1st patient enrolled Q2 2016 • Locations: University of California, San Diego, San Francisco General Hospital, Cook County Health and Hospitals System, Chicago, Washington University School of Medicine in St.
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