Safety and Tolerability of SYNB1618 in Healthy Adult Volunteers and Adult Subjects With Phenylketonuria

This Phase 1/2a, first-in-human, oral single and multiple dose-escalation, randomized,
double-blinded, placebo-controlled study of SYNB1618 in healthy adult volunteers and adult
subjects with phenylketonuria will evaluate safety, tolerability, kinetics, and
pharmacodynamics within the following 2 study parts:

Part 1 comprises a single-ascending dose (SAD) study conducted (3 treated:1 placebo) over 4
days in HV male and female subjects evaluated in up to 6 dose cohorts to identify maximum
tolerated dose (MTD) within the single dose-dose range studied. Up to 24 healthy subjects may
be enrolled in this part of the study. Following attainment of the MTD in HV, a SAD cohort of
up to 4 subjects (male and female, >18 years old) previously diagnosed with PKU will be
enrolled (3 treated:1 placebo).

Part 2 comprises a multiple-ascending dose (MAD) study conducted in an inpatient setting (6
treated:2 placebo) over 10 days in HV male and female subjects evaluated in up to 4 cohorts
at doses that will not exceed the MTD from the SAD part of the study to identify the MTD of
SYNB1618 within the multiple-dose range studied. Up to 32 healthy subjects may be enrolled in
this part of the study. Once the highest MAD cohort and the SAD PKU cohort have been
completed, a multiple-dose cohort of male and female subjects (>18 years old) previously
diagnosed with PKU will be evaluated. Up to 20 subjects with PKU may be enrolled in the MAD
PKU cohort (12 treated: 8 placebo).

Inclusion Criteria:

1. Age > 18 to ≤ 64 years.

2. Able and willing to voluntarily complete the informed consent process (subject or
subject's representative).

3. Available for and agree to all study procedures, including feces, urine, and blood
collection and adherence to diet control, inpatient monitoring, follow-up visits, and
IP ingestion compliance.

4. Male subjects who are sexually abstinent or surgically sterilized (vasectomy), or
those who are sexually active with a female partner(s) and agree to use an acceptable
method of contraception (such as condom with spermicide) combined with an acceptable
method of contraception for their non-pregnant female partner(s) (as defined in
Inclusion Criterion #5) after informed consent, throughout the study, and for a
minimum of 90 days after the last dose of IP, and who do not intend to donate sperm in
the period from screening until 3 months following administration of the
investigational medical product.

5. Female subjects that meet one of the following:

1. Woman of childbearing potential (WOCBP) must have a negative serum pregnancy test
(HCG) at screening and at baseline prior to the start of IP and must agree to use
acceptable method(s) of contraception, combined with an acceptable method of
contraception for their male partner(s) (as defined in Inclusion Criterion #4)
after informed consent, throughout the study and for a minimum of 90 days after
the last dose of IP. Acceptable methods of contraception include hormonal
contraception, hormonal or non-hormonal intrauterine device, bilateral tubal
occlusion, complete abstinence, vasectomized partner with documented azoospermia
90 days after procedure, diaphragm with spermicide, cervical cap with spermicide,
vaginal sponge with spermicide, or male or female condom with or without
spermicide.

2. Premenopausal woman with one of the following:

i. Documented hysterectomy

ii. Documented bilateral salpingectomy

iii. Documented bilateral oophorectomy

iv. Documented tubal ligation/occlusion

v. Sexual abstinence is preferred or usual lifestyle of the subject

c. Postmenopausal woman (12 months or more amenorrhea verified by follicle stimulating
hormone [FSH] assessment and over 45 years of age in the absence of other biological
or physiological causes).

6. Screening laboratory evaluations (e.g., chemistry panel, CBC with differential,
prothrombin time [PT]/activated partial thromboplastin time [aPTT], urinalysis, C
reactive protein [CRP], creatinine clearance) and ECG must be within normal limits or
judged to be not clinically significant by the Investigator.

7. Stable diet including protein intake for at least 60 days prior to screening
assessments.

8. Able to produce at least 2 bowel movements per week on average without the assistance
of laxatives.

In addition to the above criteria for HV, inclusion criteria for PKU subjects are as
noted below.

9. Diagnosis of classic PKU by either medical history of blood Phe concentration of >1200
µmol/L at any time OR genetic diagnosis.

10. Blood Phe concentration of ≥ 600 µmol/L at Screening.

11. Stable diet including stable medical formula regimen (if used) for 60 days prior to
screening assessments.

Exclusion Criteria:

1. Acute or chronic medical, surgical, psychiatric, or social condition or laboratory
abnormality that may increase subject risk associated with study participation,
compromise adherence to study procedures and requirements, or may confound
interpretation of study safety or PD results and, in the judgment of the investigator,
would make the subject inappropriate for enrollment.

2. Body mass index < 18.5 or ≥ 30 kg/m2 (> 40 kg/m2 for PKU subjects).

3. History of or current immunodeficiency disorder including autoimmune disorders and
human immunodeficiency virus (HIV) antibody positivity.

4. Hepatitis B surface antigen positivity (subjects with hepatitis B surface antibody
positivity and hepatitis B core antibody positivity are not excluded, provided that
the hepatitis B surface antigen is negative).

5. Hepatitis C antibody positivity, unless a hepatitis C virus ribonucleic acid test is
performed and the result is negative.

6. History of febrile illness, confirmed bacteremia, or other active infection within 30
days prior to the anticipated first dose of IP.

7. History of active or chronic passage of 3 or more loose stools per day.

8. Active laxative use within 30 days prior to the anticipated first dose of
investigational product.

9. Active inflammatory or irritable bowel disorder of any grade.

10. Active or past history of gastrointestinal (GI) bleeding within 60 days prior to the
Screening Visit as confirmed via hospitalization-related event(s) or medical history
of hematemesis or hematochezia.

11. Intolerance of or allergic reaction to E. coli Nissle or any of the ingredients in
SYNB1618 or placebo formulations.

12. Any condition, prescription medication, or over-the-counter product that may possibly
affect absorption of medications or nutrients (e.g., celiac disease, gastrectomy,
bypass surgery, ileostomy).

13. Currently taking or plans to take any type of systemic (e.g., oral or intravenous)
antibiotic within 28 days prior to the anticipated first dose of investigational
product through the final outpatient follow-up. Exception: topical antibiotics are
allowed.

14. Major surgery (an operation upon an organ within the cranium, chest, abdomen, or
pelvic cavity) or inpatient hospital stay within the 3 months prior to the anticipated
first dose of investigational product.

15. Planned surgery, hospitalizations, dental, or interventional studies between screening
and last anticipated visit that might require antibiotics.

16. Taking or planning to take probiotic supplements (enriched foods excluded) within 28
days prior to the anticipated first dose of investigational product and for the
duration of participation and follow-up.

17. Dependence on drugs of abuse.

18. Regular alcohol consumption in excess of 14 standard drinks/week for men and in excess
of 7 standard drinks/week for women and/or any evidence of binge or heavy drinking
(according to National Institute on Alcohol Abuse and Alcoholism guidelines). One
drink is equivalent to 12 g of alcohol: 12 ounces (360 mL) of beer, 5 ounces (150 mL)
of wine or 1.5 ounces (45 mL) of 80 proof distilled spirits.

19. Administration or ingestion of an investigational drug within 60 days or 5 half-lives,
whichever is longer, prior to the Screening Visit or current enrollment in an
investigational study. (PKU subjects who participated in the PKU SAD cohort may be
eligible for screening for the PKU MAD cohort > 30 days after the last dose of IP.)

20. Evidence or history of clinically significant hematological, renal, endocrine,
pulmonary, GI, cardiovascular (including stable coronary artery disease/angina or
prior cardiac stent), hepatic, neurologic, or allergic disease including drug
allergies.

21. Screening laboratory parameters within the acceptable range.

In addition to the above criteria for HV, exclusion criteria for PKU subjects are as
noted below.

22. Currently taking (within 1 week prior to screening) sapropterin (KUVAN®).

23. Currently taking (within 6 months prior to screening) pegylated recombinant
phenylalanine ammonia lyase (PALYNZIQ™).

24. History of a severe immune reaction based on National Cancer Institute (NCI) Common
Terminology Criteria for Adverse Events (CTCAE) after administration of pegylated
recombinant phenylalanine ammonia lyase (PALYNZIQ).