Fighting Coccidioidomycosis Prevention: Vaccine Fighting Coccidioidomycosis Prevention: Vaccine Treatment: Nikkomycin Z Valley Fever Vaccine Project.

Fighting Coccidioidomycosis Fighting Coccidioidomycosis Prevention: VaccinePrevention: Vaccine

Treatment: Nikkomycin ZTreatment: Nikkomycin Z

Valley Fever Vaccine ProjectValley Fever Vaccine Project

A brief history…

Valley Fever Vaccine Project

• Epidemic of early 90’s spawned renewed interest in a vaccine

• Bakersfield locals created a group to organize project & secure financing

• With funds in hand, “exploratory phase” was initiated and research begun in ‘98

• VFVP formally began in 2000 with 5 yrs of funding for 5 research institutions

The Valley Fever Vaccine Project

Valley Fever Vaccine Project

University of Arizona– John Galgiani– Lisa Shubitz

UT San Antonio– Rebecca Cox– Mitch Magee (ASU)

Med. College of Ohio– Garry Cole– Chiung Yu

UC Davis– Demo Pappagianis– Susan Johnson

UC San Diego– Theo Kirkland

UC San Francisco– George Rutherford– Richard Hector

CSU Bakersfield

Supporting Organizations

Valley Fever Vaccine Project

• California HealthCare Foundation• State of California (Ashburn, Parra)• CDC (Thomas)• Rotary’s Valley Fever Project of the

Americas, Valley Fever Research Foundation, & many local contributors

• Kern County• NIH for sequencing of C. posadasii

Major AccomplishmentsMajor Accomplishments

• • Dozens of antigens identified, cloned Dozens of antigens identified, cloned

• • Attenuated mutant strains createdAttenuated mutant strains created

• • Genome of Genome of C. posadasiiC. posadasii sequenced sequenced

• • Expression systems & manufacturingExpression systems & manufacturing

evaluated for fusion proteinevaluated for fusion protein

• • Primate trial to evaluate fusion protein Primate trial to evaluate fusion protein

• • 10 patents filed; 4 patents issued10 patents filed; 4 patents issued

• • Canine incidence/prevalence trialCanine incidence/prevalence trial

• • Coccidioidin Phase 1 & 2 trialsCoccidioidin Phase 1 & 2 trialsValley Fever Vaccine Project

Possible Value Of A Vaccine

• Prevent ~3,500 disseminated cases/yr• Prevent ~10-30,000 primary illnesses/yr

– Public Health risk• Residents of California, Arizona, Nevada, Utah• Tourists to the Southwest• Relocated employees• Immunocompromised individuals

– Military risk• Desert training bases within Southwest• Biodefense

Valley Fever Vaccine Project

Cost-effectiveness of a vaccine

Valley Fever Vaccine Project

0

50,000

100,000

150,000

200,000

250,000

300,000

20% 30% 40% 50% 60% 70% 80% 90%

Vaccine Efficacy

Co

st-E

ffec

tive

nes

s R

atio

($/

QA

LY) Infants: vaccinate vs. no vaccination

Resident 15 yo: vaccinate vs. no vaccination

Immigrant 15 yo: vaccinate vs. no vaccination

Resident 35 yo: screen/vaccinate vs. no vaccination

Immigrant 35 yo: vaccinate vs. no vaccination

Base case

* Barnato et al. Cost-effectiveness of a potential vaccine for coccidioidomycosis. Emerg Infect Dis 2001; 7:797-806.

Vaccine CandidatesVaccine Candidates

• • Recombinant proteins:Recombinant proteins:Ag2/PRA106+CsaAg2/PRA106+CsaELI1ELI1

• • Live, attenuated mutants:Live, attenuated mutants: ∆∆cts2/∆ard1/ ∆cts3 straincts2/∆ard1/ ∆cts3 strain ∆ ∆chs5 strainchs5 strain

Valley Fever Vaccine Project

ValleyFever

Center forExcellence

Ag2/PRA106-CSA Chimeric Ag

Flag Ag2/PRA1-106 CSA

EcoRIGlu-Phe

BamHIGly-Ser

Patent issued 2006

Days Survived

0 10 20 30 40 50 60

Pro

port

ion

of M

ice

Sur

vivi

ng

0.0

0.2

0.4

0.6

0.8

1.0

Mouse Survival Studies with Fusion Protein*Mouse Survival Studies with Fusion Protein*

Adj. only

CSA

Ag2/PRA1-106

Both

*Intranasal infection with 80 arthroconidiaShubitz et al., Vaccine 24:5904, 2006

C57BL/6 mice251 and 218 spores IN, Silveira

Days Survived

0 10 20 30 40 50 60 70

Pro

port

ion

of M

ice

Sur

vivi

ng

0.0

0.2

0.4

0.6

0.8

1.0

PRA106+CS

106/CS Chimera

PRA106

CS

Adjuvant

Gehans Wilcoxon: Both > Ag2/PRA1-106 > CSA > Adjuvant

HigherInoculum

Mouse Survival Study with Higher InoculumMouse Survival Study with Higher Inoculum

Adding ELI-1 to Ag2/PRA+CSAAdding ELI-1 to Ag2/PRA+CSA

Days Postinfection

10 20 30 40 50 60 70 80 90 100

Per

cen

t Su

rviv

al

20

40

60

80

100 CPG/IFAAg2/CSAELI-Ag1Ag2/CSA + ELI-Ag1

P=0.04

Magee et al, 2006

Fusion Vaccine IssuesFusion Vaccine Issues

• Efficacy: may prevent dissemination, but Efficacy: may prevent dissemination, but clearly does not prevent infectionclearly does not prevent infection– Recombinant vaccines have not worked in Recombinant vaccines have not worked in

diseases like TBdiseases like TB– No data on duration of immunityNo data on duration of immunity

• Adjuvants: few CMI-promoting adjuvantsAdjuvants: few CMI-promoting adjuvants– MPL is not available to usMPL is not available to us– Alternatives are few & have safety issuesAlternatives are few & have safety issues

• Manufacturing issuesManufacturing issues– Expression is poor; aggregation is a problemExpression is poor; aggregation is a problem

Valley Fever Vaccine Project

Flagellin as an Adjuvant SubstituteFlagellin as an Adjuvant Substitute

• Flagellin binds to TLR 5 to stimulate DC activation and antigen presentation.

• Flagellin + antigen fusion proteins have been shown in mouse models to stimulate cell-mediated immunity

• This technology is being commercialized by Vaxinnate, Inc.; clinical trials underway

• Flagellin + Ag2/PRA+Csa experiment in progress in Galgiani lab

C. posadasiiC. posadasii ∆cts2/∆ard1/∆cts3 ∆cts2/∆ard1/∆cts3 Attenuated MutantAttenuated Mutant

Valley Fever Vaccine Project

Parental strain: C735 cts2/ard1/cts3

Cu

ltu

res

in C

onve

rse

med

ium

Infe

cted

mu

rin

e lu

ng

tiss

ue

Morphology of cts2/ard1/cts3 triple mutant in vitro vs in vivo

Sterile spherules

cts2/ard1/cts3 cts2/ard1/cts3

Days post-challenge0 5 10 15 20 25 30 35

% S

urv

ival

0

20

40

60

80

100

Neg Ct

cts2ard1cts3FKS

45 55

BALB/c mice80 spores i.n.

65 75

Evaluation of survival of BALB/c mice vaccinated with the live cts2/ard1/cts3

mutant or the FKS vaccine and i.n.-challenged

Data courtesy Garry Cole, MUO

Valley Fever Vaccine Project

Attenuated Mutant:Attenuated Mutant:Why it’s a “no go” for humansWhy it’s a “no go” for humans

Valley Fever Vaccine Project

• • Safety: possibility of reversionSafety: possibility of reversion

• • Restrictions on use in humansRestrictions on use in humans

• • Immunogenicity vs. ReactogenicityImmunogenicity vs. Reactogenicity

• • Manufacture of spore-formerManufacture of spore-former

Nikkomycin Z: Nikkomycin Z: The perfect drug for The perfect drug for

coccidioidomycosis?coccidioidomycosis?

History of Nikkomycin ZHistory of Nikkomycin Z

• 1970s: Evaluated by Bayer as agricultural fungicide

• 1980s: Cured mice with cocci (Hector/Pappagianis)

• 1990s: Clinical Development started by Shaman Pharmaceuticals: Phase Ia conducted

• 2001: Purchased at auction by CSUBF

• 2005: Rights transferred to University of Arizona

• 2006: Nik Z designated as orphan drug

• Oct 2007: Patients enrolled in Phase Ib/II @ UA

Nikkomycins Inhibit Chitin SynthaseNikkomycins Inhibit Chitin Synthase

• UDP-N-acetylglucosamine is a precursor to chitin

• Nikkomycin Z is a competitive inhibitor of chitin synthase

O

OHOH

HH

HH

O

N

NH

O

O

P

O

O-

OP

O

O

O-

O

HN

O

CH3

HO

OH

OHUDP-UDP-NN-Ac-glc-Ac-glc

O

OHOH

HN

N

NH

O

OOO-

O

NH2

CH3

OHN

HO

Nik ZNik Z

ValleyFever

Center forExcellence

Mature Mature C. immitisC. immitis stained with calcofluor white stained with calcofluor white

ValleyFever

Center forExcellence

A. fumigatusA. fumigatus stained with calcofluor white stained with calcofluor white

SUSCEPTIBILITY OF DIVERSE SUSCEPTIBILITY OF DIVERSE FUNGI TO NIKKOMYCIN ZFUNGI TO NIKKOMYCIN Z

OrganismOrganism

Coccidioides immitisCoccidioides immitisBlastomyces dermatitidisBlastomyces dermatitidisHistoplasma capsulatumHistoplasma capsulatum

Sporothrix schenkiiSporothrix schenkiiCandida albicansCandida albicans

Candida parapsilosisCandida parapsilosisCandida rugosaCandida rugosa

Candida tropicalisCandida tropicalisCandida kruseiCandida krusei

Candida lusitaniaeCandida lusitaniaeCryptococcus neoformansCryptococcus neoformans

Torulopsis glabrataTorulopsis glabrataAspergillus flavusAspergillus flavus

Aspergillus fumigatusAspergillus fumigatus

No.No.StrainsStrains

11101099

10105959101011775511

303021212222

Geometric MeanGeometric MeanMICMIC

100100 ( (g/ml)g/ml)

0.06250.06250.250.252.472.47

0.4070.4075.565.564.294.297.87.8

>500>500445445

>500>500144144

>500>500500500500500

Control vs. NZ-treatedControl vs. NZ-treated

SURVIVAL EXPERIMENT WITH SURVIVAL EXPERIMENT WITH COCCIDIOIDOMYCOSISCOCCIDIOIDOMYCOSIS

00

1100

2200

3300

4400

5500

6600

7700

8800

9900

110000

00 22 44 66 88 1100 1122 1144 1166 1188 2200 2222 2244 2266 2288

% S

UR

VIV

AL

--------THERAPY-----------------THERAPY---------

DAYS POST-INFECTIONDAYS POST-INFECTION

Nik Z Nik Z 50 mg/kg50 mg/kg

Nik ZNik Z20 mg/kg20 mg/kg

ControlControl

Nik Z Nik Z 5 mg/kg5 mg/kg

ANIMALS ANIMALS INFECTED withINFECTED with 9 x 109 x 1033 CFU I.N. CFU I.N.

ANIMALS ANIMALS INFECTED withINFECTED with 9 x 109 x 1033 CFU I.N. CFU I.N.

Hector et al. AAC, 1990

SHORT-TERM ORGAN LOADS (Lung) WITH COCCIDIOIDES IMMITIS

GROUPGROUP DOSEDOSE MEAN LOGMEAN LOG(mg/kg)(mg/kg) CFU ± S. E. M.CFU ± S. E. M.

CONTROLCONTROL

FLUCONAZOLEFLUCONAZOLE

““

NIK ZNIK Z

--

2.52.5

2525

5050

6.35 ± .066.35 ± .06

3.77 ± 1.123.77 ± 1.12

2.62 ± .822.62 ± .82

0.37 ± .37 0.37 ± .37

• Enrollment: 60 seropositive subjects with Enrollment: 60 seropositive subjects with uncomplicated cocci pneumoniauncomplicated cocci pneumonia

• Rising multiple doses: 100-2250 mg x 14 days Rising multiple doses: 100-2250 mg x 14 days vs. placebovs. placebo

• End of treatment response based on signs & End of treatment response based on signs & symptoms vs. pre-dosesymptoms vs. pre-dose

– Subjective questionnaire for 12 symptomsSubjective questionnaire for 12 symptoms

– Lab: ESR, C-reactive protein, procalcitonin, lung Lab: ESR, C-reactive protein, procalcitonin, lung lesion volume (CT)lesion volume (CT)

UA: Phase Ib/II DesignUA: Phase Ib/II Design

Valley Fever Vaccine ProjectValley Fever Vaccine Project