Live Attenuated ETEC Vaccine ACE527 With and Without dmLT Adjuvant in Adults

This study is a clinical trial in healthy adult volunteers to evaluate the safety,
immunogenicity and efficacy of a live attenuated ETEC vaccine, ACE527, with and without a
mucosal adjuvant, dmLT. This study was designed initially as a single site, Phase 1,
double-blind, randomized, placebo-controlled, clinical trial in healthy adult volunteers to
evaluate the safety and immunogenicity of a live attenuated ETEC vaccine, ACE527, with and
without a mucosal adjuvant, dmLT (Part A). The addition of a challenge step provides a unique
opportunity to evaluate the efficacy of the new lyophilized formulation of ACE527 vaccine,
given in a two or three dose regimen, with and without dmLT, against wild type ETEC strain
H10407 challenge (designated as Part B: Phase 2b Open Label Challenge Study). In addition,
challenging the volunteers may allow for the identification of immune correlates of
protection, taking advantage of newly available technologies (immune proteomics,
transcriptomics, etc.)

ACE527 comprises three genetically attenuated and engineered strains of E. coli, with antigen
profiles covering a wide range of ETEC surface colonization factor antigens (CFA/I, CFA/II
[CS1, CS2, CS3] and CFA/IV [CS5, CS6]) and also expressing LT-B, the inactive subunit of LT
(ETEC heat labile toxin). LT(R192G/L211A), or dmLT, is a derivative of wild-type
enterotoxigenic Escherichia coli heat-labile enterotoxin that has been genetically modified
by replacing the arginine at amino acid position 192 with glycine and the leucine at amino
acid position 211 with alanine.

Volunteers were enrolled in Part A into each of two separate Cohorts. Cohort 1 and 2
volunteers received 10^10 colony-forming units (cfu) total dose of ACE527, 10^10 cfu total
dose of ACE527 with 25 µg dmLT, or placebo at 0, 1, and 2 months. Enrollment and dosing of
Cohort 2 was dependent on an acceptable safety profile of the first dose of Cohort 1, based
on evaluation of data up until Day 3 by the Safety Review Committee (SRC). The first
immunization of each Cohort was administered in the Center for Immunization Research (CIR)
Inpatient Unit, followed by 72 hours of direct post-immunization observation. The SRC met
after the first dose of cohort to determine continuation of volunteer dosing on an outpatient
basis, and enrollment of Cohort 2. The SRC met again after the first dose of Cohort 2 after
concluding that the first dose appeared safe and well tolerated, subsequent doses would be
administered on an outpatient basis. Safety was assessed by solicited symptoms/subject memory
aid and laboratory evaluations. Adverse events (AE)s were graded according to standardized
criteria. The immunogenicity outcome measures of interest include serum immunoglobulin G
(IgG) and immunoglobulin A (IgA) antibodies by enzyme-linked immunosorbent assay (ELISA)
against all vaccine antigens, cytokine assays, B and T cell memory responses, shedding
profile of ACE527, and vaccine specific mucosal IgA responses.

Part B challenge cohorts were recruited among those participating in Part A; plus additional
unvaccinated control volunteers sufficient to enroll up to a total of 60 recruited, as
needed. Volunteers in the Phase 2b study were enrolled and challenged in 2-4 cohorts due to
the CIR Inpatient Unit capacity of 30 beds. A minimum of 8-10 controls were challenged with
each cohort of vaccinees to ensure comparability of attack rates across challenge cohorts.
Volunteers were admitted as inpatients and challenged, with approximately 2 x 10^7 cfu of the
fully virulent ETEC strain, H10407, followed by 5 days of direct observation. A Data Review
Committee (DRC) will be convened to review the clinical data for all challenged volunteers
and verify all outcomes per protocol definitions before any vaccine efficacy assessments were
made.

Inclusion Criteria:

1. Male and female healthy adults between 18 and 50 years of age at the time of
enrollment.

2. General good health, without clinically significant medical history, physical
examination findings or clinical laboratory abnormalities per clinical judgment of PI.

3. Negative pregnancy test at screening and before the first (V0), second (V28), and
third vaccinations (V56) for female volunteers of childbearing potential. Females of
childbearing potential must agree to use an efficacious hormonal or barrier method of
birth control during the study. Abstinence is acceptable. Female volunteers unable to
bear children must have this documented (e.g. tubal ligation or hysterectomy) or must
have negative pregnancy tests.

4. Willingness to participate in the study after all aspects of the protocol have been
explained and written informed consent obtained.

5. Completion of a training session and demonstrated comprehension of the protocol
procedures and knowledge of ETEC associated illness by passing a written examination
(70% pass score).

6. Availability for the study duration, including all planned follow-up visits.

7. Received at least 2 doses of ACE527 vaccine alone or in combination with 25 ug dmLT
4-6 months prior to challenge (Part B only)

Exclusion Criteria:

1. Presence of a significant medical or psychiatric condition which in the opinion of the
investigator precludes participation in the study. Some medical conditions which are
adequately treated and stable would not preclude entry into the study. These
conditions might include stable asthma controlled with inhalers or mild hypertension
stably controlled with a single agent.

2. Significant abnormalities in screening hematology, or serum chemistry as determined by
PI or PI in consultation with the Medical Officer and sponsor.

3. Presence in the serum of HIV antibody, HBsAg, or HCV antibody.

4. Evidence of IgA deficiency (serum IgA < 7 mg/dl or limit of detection of assay).

5. Evidence of current excessive alcohol consumption or drug dependence.

6. Volunteers whose Body Mass Index (BMI) is less than 19.0 or greater than 34.0 (kg/m2)

7. Recent vaccination or receipt of an investigational product (within 30 days before
vaccination).

8. Intention to donate blood or blood products within one month following the completion
of study participation (note: The Red Cross will not allow blood donations for 1 year
following participation in an investigational research study).

9. Any other criteria which, in the investigator's opinion, would compromise the ability
of the volunteer to participate in the study, the safety of the study, or the results
of the study

10. Working as a food handler, in child-care or as a healthcare worker with direct patient
contact.

11. Have household contacts who are <2 years old or >80 years old or infirm or
immunocompromised (for reasons including corticosteroid therapy, HIV infection, cancer
chemotherapy, or other chronic debilitating disease).

12. Abnormal stool pattern (fewer than 3 per week or more than 3 per day).

13. Regular (≥ weekly) use of laxatives, antacids, or other agents to lower stomach
acidity.

14. Use of any medication known to affect the immune function (e.g., corticosteroids and
others) within 30 days preceding the first vaccination or planned use during the
active study period.

15. Symptoms consistent with Traveler's Diarrhea concurrent with travel to countries where
ETEC infection is endemic (most of the developing world) within two years prior to
dosing, OR planned travel to endemic countries during the length of the study.

16. Vaccination for or ingestion of ETEC, cholera, or LT toxin within 3 years prior to
dosing.

17. Use of antibiotics during the 7 days before dosing or proton pump inhibitors, H2
blockers or antacids within 48hours prior to dosing.

18. History of diarrhea in the 7 days prior to vaccination (outpatient diarrhea is defined
as ≥ 3 unformed (grade 3 or greater) loose stools in 24 hours).

19. Known allergy to two of the three following antibiotics: Ciprofloxacin, amoxicillin,
and/or trimethoprim/sulfamethoxazole