Human M. tuberculosis infection/ disease: classical pathology and immunology (Slide -1) W. Henry Boom, M.D. Tuberculosis Research Unit (TBRU) Case Western Reserve University W. Henry Boom, M.D. Tuberculosis Research Unit (TBRU) Case Western Reserve University NIAID-DMID: -AI70022 Cattle Prod 1950

Route(s) of Infection & Natural Course (Slide 0) small vs. large droplet aerosol repeated exposure ?infectious dose (animals: 1-10 CFU) ?repeated infection small vs. large droplet aerosol repeated exposure ?infectious dose (animals: 1-10 CFU) ?repeated infection Bacterial Load I O PROGRESSIVE: PEDS.+IMMUNOCOMP. (5%) I O PROGRESSIVE: PEDS.+IMMUNOCOMP. (5%) REACTIVATION/ADULTS (5-10%) REACTIVATION/ADULTS (5-10%) INFECTION (90+%) INFECTION (90+%) Time (mos-yrs)

Pulmonary Tuberculosis (slide 1) Cough (+/-RBC), Wt. Loss, Night sweats CFU Diagnosis: Sputum Smear/Culture (<50% paucibacillary) Pathology: Caseating Granulomas, Necrosis, Cavitation (?Host or Microbe) Death: –Cachexia –Respiratory Failure –Dissemination (miliary, meningitis) –Massive Hemoptysis Cough (+/-RBC), Wt. Loss, Night sweats CFU Diagnosis: Sputum Smear/Culture (<50% paucibacillary) Pathology: Caseating Granulomas, Necrosis, Cavitation (?Host or Microbe) Death: –Cachexia –Respiratory Failure –Dissemination (miliary, meningitis) –Massive Hemoptysis

Immunology of M. tuberculosis infection and disease (slide 2) Bacterial Load REACTIVATION INFECTION InnateAdaptive Failure (Immunopathogenesis?) TLR’s Chemokines Cytokines Antigens T cell subsets Effector mech. Immune evasion TLR’s Chemokines Cytokines Antigens T cell subsets Effector mech. Immune evasion

“Known knowns, known unknowns, unknown unknowns” and dogma for immunology of human TB (slide 3, “adapted from Donald Rumsfeld ‘03”) Known: –Adaptive immunity –CD4+ T cell –TNF-alpha –IFN-gamma –IL-12 Unknown: –Genetics: which ones/stage (IFNgamma/IL12 pathway, NRAMP1, TNFalphaR, etc.) –TLRs: which ones/when –Chemokines: same (MCP1)? –What does IFN-gamma do? –Immunology of the lung: why so slow? –Antigens matter: which ones, when, where? Known: –Adaptive immunity –CD4+ T cell –TNF-alpha –IFN-gamma –IL-12 Unknown: –Genetics: which ones/stage (IFNgamma/IL12 pathway, NRAMP1, TNFalphaR, etc.) –TLRs: which ones/when –Chemokines: same (MCP1)? –What does IFN-gamma do? –Immunology of the lung: why so slow? –Antigens matter: which ones, when, where? Dogma: –“Immuno-pathogenesis” (HIV: cavitation related to CD4, but mortality still high) –CD8’s critical, cause of BCG failure –It is all about cytokines (cytokine interventions have failed) –Now it’s Tregs, Th17……… Unknown unknowns: –TLRs in last century –Why all T cell vaccines have failed so far (TB, HIV)? T MM TNF- , IL-12 IFN- 

Cytokines and M. tuberculosis IFN-  –IFN-  R deficient humans –IFN-  KO mice TNF-  –TNF-  and TNF-  R KO mice –anti-TNF-  antibodies in humans IL-12 –IL-12R deficient humans –IL-12 KO mice IL-10/TGF-  –Inhibit during active disease IFN-  –IFN-  R deficient humans –IFN-  KO mice TNF-  –TNF-  and TNF-  R KO mice –anti-TNF-  antibodies in humans IL-12 –IL-12R deficient humans –IL-12 KO mice IL-10/TGF-  –Inhibit during active disease T MM TNF- , IL-12 IFN- , IL-10

Immune Evasion by M. tuberculosis Innate: –Blocks Phagosomal Maturation Limited fusion with lysosomes Exclusion Na + Dep. Proton ATP-ase Retains Rab5 delays Rab7 acquisition ? Sticking TACO-tryptophane aspartate-containing coat protein (murine) Inhibition of Ca2+ rise prevents calmodulin/CaMKII recruitment of cathepsinD Roles for LAM, PIM –Inactivates Bactericidal Mechanisms oxygen radicals nitric oxide autophagy Adaptive: –Inhibition of IFN-gamma regulated genes –Inhibition of MHC II Antigen Processing (lipoproteins/TLR-2) –Direct modulation of CD4+ T cell function –Inhibitory Cytokines (IL-10, TGF-beta) –T cell Apoptosis Innate: –Blocks Phagosomal Maturation Limited fusion with lysosomes Exclusion Na + Dep. Proton ATP-ase Retains Rab5 delays Rab7 acquisition ? Sticking TACO-tryptophane aspartate-containing coat protein (murine) Inhibition of Ca2+ rise prevents calmodulin/CaMKII recruitment of cathepsinD Roles for LAM, PIM –Inactivates Bactericidal Mechanisms oxygen radicals nitric oxide autophagy Adaptive: –Inhibition of IFN-gamma regulated genes –Inhibition of MHC II Antigen Processing (lipoproteins/TLR-2) –Direct modulation of CD4+ T cell function –Inhibitory Cytokines (IL-10, TGF-beta) –T cell Apoptosis NIAID-DMID: -AI70022

Models for what aspect of TB? Pathology-granuloma, lung pathology, cavity, caseation (rabbit) Immunology-innate vs. adaptive T cell immunity (mouse, primate,?bovine) Genetics- risk for infection, progression, reactivation vs. relapse (?mouse, bovine) Drug Treatment-latent vs. active infection (efficacy, PK, ARV interactions) (primate) Vaccine- infection, dissemination vs. re-activation (mouse, guinea pig, primate) Co-pathogenesis-HIV or helminth co-infection (primate) Pathology-granuloma, lung pathology, cavity, caseation (rabbit) Immunology-innate vs. adaptive T cell immunity (mouse, primate,?bovine) Genetics- risk for infection, progression, reactivation vs. relapse (?mouse, bovine) Drug Treatment-latent vs. active infection (efficacy, PK, ARV interactions) (primate) Vaccine- infection, dissemination vs. re-activation (mouse, guinea pig, primate) Co-pathogenesis-HIV or helminth co-infection (primate)

Protective Adaptive Immunity to M. tuberculosis: Macrophages and T cells IL-12, TNF-alpha, IFN-gamma IL-2 for T cell expansion Cytotoxic Effector T Cells (CTL ) Growth inhibition of M. tuberculosis IL-12, TNF-alpha, IFN-gamma IL-2 for T cell expansion Cytotoxic Effector T Cells (CTL ) Growth inhibition of M. tuberculosis T cell MM CTL cytokines IL-12, IFN- , TNF-  (FasL/CD95L, granzymes, perforin, granulysin) Growth inhibition IL-2