Entolimod, an Adjuvant for Vaccine Augmentation

This is a single-blind (participants are blinded), pilot study with no therapeutic or
diagnostic intent and limited drug exposure. The study will be conducted at Baylor College of
Medicine in normal, healthy, and non-drug using male volunteers who have not received the Td
vaccination in the past 5 years.

The purpose of this study is to evaluate the safety and surrogate efficacy of low dose
entolimod in normal, healthy, non-patient subjects by comparing anti-tetanus (TT) antibody
levels between patients getting entolimod vs no entolimod combined with tetanus-diphtheria
(Td) vaccine. Subjects will be randomly assigned to one of two arms in a 3:5 ratio (for every
8 subjects, 3 will be randomly assigned to the control group and 5 to the treatment group) to
receive a single dose of:

Arm 1: Td (4 ug) (n=15), Arm 2: Td (4 ug) + Entolimod (1 ug) (n=25)

We propose to give the normal dose of Td given routinely to humans (i.e. TT at 2μg and
diphtheria toxoid at 2 ug) for a total protein dose of 4 ug. The proposed dose of entolimod
at 1 ug is 30-40 times lesser than that previously given safely to humans. We expect that Td
+ entolimod will double the anti-tetanus toxoid antibody (anti-TT AB) and anti-diphtheria AB
levels over Td alone. We predict that this low dosing of entolimod (1μg) produces no
difference in adverse effects from giving Td vaccine alone and that any adverse events will
be no different than what is typical as reported in the Physicians' Desk Reference for Td
vaccine in humans.

For purposes of this trial, study population age cut-off will be 40 years of age. The reason
for this cut-off is that the body produces higher antibody levels in response to vaccines in
younger age groups compared to older groups. Further, this study is restricted to males only
as a safety measure so that women of child-bearing age are not exposed to entolimod, which
has not been given to women of childbearing potential.

We consider our strategy adequate for human safety for several reasons related to risks for
this new combination of an existing vaccine and new adjuvant. First, our proposed
Td-Entolimod starting dose is the usual approved clinical dose of Td (4 μg containing 2 ug TT
+ 2 ug diphtheria toxoid). Second, the proposed dose for the adjuvant entolimod (1 μg) is
supported by both animal toxicology studies and human safety/PK/PD studies cross-referenced
from IND 100480. The results of these safety studies, which have given humans up to 30 μg and
animals up to 100μg, strongly support that entolimod at 1μg is a safe human dose. The other
adjuvant is aluminum phosphate, which is contained within the standard Td vaccine in humans.
Third, further support for safety comes from the recent recommendations to boost pregnant
women with Td at each pregnancy regardless whether or not they have recently received a Td
shot. This recommendation of acceptable enough safety for ACIP to recommend it for pregnant
women, supports that this study's single dosing with Td even at less than a 5 year interval
would be relatively low risk. Fourth, other FDA approved conjugate vaccines using TT have
contained 8 to 10 times greater doses of TT than in our planned dose. Fifth, we will have a
randomization of 25 subjects to the Td-Entolimod combination vaccine, which will provide
sufficient numbers of subjects to detect common adverse events such as fever and local site
reactions in more than one human subject in our vaccinated group compared to the Td alone
control group. Local injection site and systemic inflammatory changes also will be monitored
in planned clinical study trials with later TT conjugate anti-addiction vaccines.

The close monitoring of objective and subjective drug effects by direct observation, blood
work (CBC and Comprehensive Metabolic Panel), weekly telephone check-ins over a 6 week period
from point of vaccination to close-out visit to monitor for potential adverse reactions, and
a close-out physical will allow objective evaluation of the effects of the vaccines in these
normal subjects. We will also have post treatment follow up period with monthly telephone
visits up to 1 year from vaccination to further monitor for adverse reactions.

At 6-weeks post-vaccination, we will analyze level of anti-TT AB and anti-diphtheria AB and
compare both study arms compared to baseline AB levels to determine the immune potentiating
capability of entolimod. In addition to not expecting any difference in adverse events (AE)
from giving Td alone compared to those AE listed in the Physicians' Desk Reference for Td in
normals, we also expect the AB levels at 6 weeks post vaccination and the ratio of 6-week to
baseline AB levels from Td+entolimod (n=25) to be at least double the levels from Td alone
(n=15).

Inclusion Criteria:

1. Healthy, non-patient normal males between 18 and 40 years of age

2. No baseline anti-TT AB (tetanus toxoid antibody) levels greater than 5 µg/ml

3. Able to read/write/understand English and provide signed written informed consent
prior to any study specific procedures.

Exclusion Criteria:

1. Inability to read/write English

2. Inability to provide written informed consent.

3. Any major cardiovascular, renal, endocrine, immunological (including positive test
result for human immunodeficiency virus types 1 or 2 antibodies) or hepatic disorder
(including positive test result for hepatitis B, hepatitis C virus antibodies, or
liver function tests (LFT's)(SGOT, SGPT) that are 20% greater than normal levels or
history of severe alcohol use disorder (defined as a score of 5 of greater on the
MINI, based on DSM criteria).

4. Medical contraindication to treatment with vaccine as indicated by a history of
autoimmune disease, immune deficiency, or hypersensitivity to other vaccines. Specific
disorders include a prior Arthus-type hypersensitivity reaction to tetanus vaccine or
any of its components, prior Guillain-Barre Syndrome, Ongoing active infection of any
kind with clinical signs of febrile illness (temperature >99.5ºF).

5. Use of psychoactive or hepatotoxic medications or medications that might interact with
the tetanus toxoid based vaccine including steroids within 30 days of screening,
immunosuppressive or immunostimulant therapy.

6. Received other vaccines, including flu vaccine, within 30 days prior to screening or
Tetanus vaccine within 5 years of this study (Advisory Committee on Immunization
Practices (ACIP) guidelines.

7. Blood or blood products given in the 3 months prior to vaccination.

8. Participation in another investigational pharmaceutical trial within 30 days prior to
screening.

9. Greater than Grade 1 abnormality in baseline safety labs (e.g, CBC and CMP), as
defined using the FDA Guidance Document entitled "Toxicity Grading Scale for Healthy
Adult and Adolescent Volunteers Enrolled in Preventative Vaccine Clinical Trials"

10. Greater than Grade 1 abnormality in baseline vital signs

11. Women, as a standard safety measure, will be excluded from the study due to
childbearing potential and to minimize the effects of differences in hormonal levels
during the menstrual cycle and of sex differences in antibody titer responses to
vaccines. Furthermore, our sample size is too small to control for these effects in
women.

12. Other concerns that in the PI's judgment will be a potential safety issue for the
subject including cognitive impairment that precludes the ability to provide informed
consent, and individuals with behavioral issues evidenced during screening or through
the course of the trial that are disruptive and/or pose a safety concern to self or
other subjects.