Safety, Tolerability, and PK Parameters of Sodium Nitrite Inhalation Solution in Healthy Subjects

Pulmonary hypertension (PH) is an increase in the blood pressure (BP) in the small pulmonary
vessels, arteries, veins or capillaries that results in progressive increases in right
ventricular afterload, often leading to right ventricular failure and death. Pulmonary
hypertension can result from a multitude of pathologies, including arterial etiologies,
venous, chronic hypoxia related, thromboembolic, and other miscellaneous etiologies. In the
recent Dana Point Classification, pulmonary arterial hypertension (PAH) was classified as
Group 1. In Group 1 PAH, the pathologic lesion is localized to the small muscular pulmonary
arteries, resulting in luminal narrowing and resistance to blood flow. Group 1 PAH includes
idiopathic or sporadic PAH and heritable PAH which includes those with a family history of
PAH. Associated PAH includes disease associated with connective tissue disease as well as
PAH associated with congenital systemic to pulmonary shunts, portal hypertension, and human
immunodeficiency virus (HIV) infection. Drug and toxin induced PAH is also in Group 1.
Untreated, patients with PAH have an average life expectancy of 3 years, which declines to
approximately 1 year if right heart failure is present.

Inhaled nitric oxide (NO) is a potent acute vasodilator. Its acute administration results in
improved hemodynamics in the 10 to 15% of patients with PAH who demonstrate acute
vasoreactivity. However, the usefulness of NO for chronic therapy is limited by the need for
continuous inhalation. In patients who demonstrate vasoreactivity during an acute challenge
with inhaled NO or a prostanoid challenge, therapy with calcium channel blockers has been
demonstrated to result in improved symptoms and survival. In patients who do not demonstrate
an acute vasodilator response, available therapies for PAH include prostanoids, endothelin
receptor antagonists (ERA), and phosphodiesterase type-5 (PDE-5) inhibitors. However, the
route and frequency of administration of the prostanoids, the hepatotoxicity of the ERAs,
and concerns about the sustained efficacy of both the ERAs and PDE-5 inhibitors suggests
that many patients with PAH could benefit from an effective therapy which offers ease of
administration and a favorable toxicity profile.

Aires Pharmaceuticals, Inc. is developing a novel therapeutic, AIR001, for the treatment of
PAH. The active ingredient in AIR001 is sodium nitrite, formulated in a buffered,
pH-adjusted solution for nebulization. Preclinical data suggest that under the hypoxic,
acidotic conditions present in the pulmonary hypertensive lung, inhaled sodium nitrite
serves as a sustained release source of NO which will act as an acute pulmonary vasodilator.
In addition, because decreased levels of NO have been shown to stimulate vascular
remodeling, the increased NO resulting from nitrite inhalation is postulated to attenuate or
reverse the pulmonary arterial remodeling process, resulting in both symptomatic improvement
and pulmonary hemodynamic improvement in patients with PAH.

Preclinical experiments have shown that AIR001 by oral, IV, and inhalation routes of
administration is effective in treating PAH induced by hypoxia or monocrotaline when
administered as infrequently as once weekly at doses that generate plasma concentrations of
approximately 1 to 10 uM and above. While the precise mechanism by which AIR001 exerts
anti-hypertensive actions in PAH models remains to be elucidated, nitrite itself has been
demonstrated to be metabolized to NO in animals and humans.

Single doses of AIR001 have been shown to be well tolerated in humans at dose levels that
generate peak plasma concentrations (Cmax) in the target range established in preclinical
models and no safety issues have been identified. AIR001 reduced PH induced by hypoxic gas
inhalation in healthy volunteers at doses that are well tolerated. Preclinical and clinical
data support further clinical investigation of inhaled AIR001 for the treatment of PAH.
Before progressing to studies in patients with PAH, the safety and tolerability of multiple
dose administration of AIR001 will be evaluated in Part A of the current study.

The PDE-5 inhibitor, sildenafil, is approved for use in patients with PAH and commonly used
in the initial treatment for PAH. AIR001 is converted to NO, which increases cGMP dependent
vasodilatation. Because PDE-5 inhibitors prevent the catabolism of cGMP by
phosphodiesterase, it is possible that an exaggerated drug effect could be observed when
AIR001 is administered to patients being treated with sildenafil. Therefore, combination
safety and tolerability (in particular orthostatic effects) of escalating single doses of
AIR001 will also be evaluated in combination with steady-state sildenafil in Part B of the
study.

Inclusion Criteria:

- Be informed of the nature of the study and is able to understand and has provided
written informed voluntary consent

- Be healthy males or females, of any race, at least 18 years of age or the legal age
of consent (whichever is greater) and less than 56 years of age at the time of the
first dose of study drug (or sildenafil)

- Have a body mass index (BMI) >=18.0 and <32.0 kg/m2 and weigh at least 50 kg

- Be in good general health with no clinically relevant abnormalities based on the
medical history, physical examination, clinical laboratory evaluations (hematology,
clinical chemistry, urinalysis, methemoglobin), and 12-lead ECG that, in the opinion
of the Investigator, would affect subject safety

- Agree to comply with the study procedures and restrictions

Exclusion Criteria:

- Any disease or condition that might affect drug absorption, metabolism, or excretion,
or clinically significant cardiovascular, ocular, hematological, renal, hepatic,
pulmonary, endocrine, gastrointestinal, immunological, dermatological, neurological,
or psychiatric disease

- Clinically significant illness (including lower respiratory tract infection) or
clinically significant surgery within 4 weeks before the administration of study drug
(or sildenafil)

- Use of any commercially marketed mouthwash or oral rinse with agents other than tap
water as well as tongue brushing or scraping from screening onwards through
completion of study

- Currently a smoker or has a past history of smoking (of >10 pack years)

- History of bronchial asthma or sleep apnea

- Evidence of restrictive or obstructive lung disease (Forced Expiratory Volume in 1
second (FEV1)/Forced Vital Capacity (FVC) <70%, FEV1 <70% predicted, and/or FVC <70%
predicted)

- Family history of primary PH

- History of pulmonary embolism

- Evidence of supine hypertension or hypotension (systolic BP >180 mmHg or <90 mmHg
and/or diastolic BP> 100 mmHg or <50 mmHg) pre-dose [NOTE: BP measurements may be
repeated twice, at least 10 minutes apart]

- Orthostatic hypotension defined as a drop in systolic BP by >=20 mmHg or diastolic BP
of >=10 mmHg at screening or predose or the development of significant postural
symptoms (dizziness, lightheadedness, vertigo) when going from the supine to the
standing position

- Personal or family history of congenital or acquired methemoglobinemia

- Personal or family history of RBC CYP B5 reductase deficiency

- Personal or family history or any evidence of hemoglobinopathy

- Known or suspected hypersensitivity or allergic reaction to sodium nitrite, sodium
nitrate, or saccharin

- Glucose-6-phosphate dehydrogenase (G6PD) deficiency or any contraindication to
receiving methylene blue

- History of hypersensitivity or idiosyncratic reaction to drugs from multiple drug
classes

- If female, is pregnant or breast feeding, or has a positive pregnancy test result
pre-dose

- If a sexually active female, is not surgically sterile (defined as having had a
hysterectomy, bilateral oophorectomy, or tubal ligation at least 6 months prior to
screening) or post-menopausal (defined as amenorrhea for the past 2 years if <50
years of age or for the past 1 year if <=50 years or, if on hormone replacement
therapy [HRT], documented follicle stimulating hormone [FSH] >30 IU/L before starting
HRT), or does not agree to utilize two effective methods of contraception
consistently and as intended from screening until at least 4 weeks after the last
dose of study drug. Subjects must use a barrier method (diaphragm with intravaginal
spermicide, cervical cap with intravaginal spermicide, or partner using condoms plus
use of intravaginal spermicide) in combination with at least one of the following
methods of contraception:

1. systemic hormonal contraceptive (oral, implant, injection, or patch)

2. intrauterine device

3. or male partner who has undergone a vasectomy at least 6 months prior to
screening.

- Unless approved by the Sponsor, chronic use of any systemic medications (with the
exception of systemic hormonal contraceptives and vitamins taken at standard
supplement doses); use of a drug therapy (including herbal preparations, e.g., St.
John's wort) known to induce or inhibit hepatic drug metabolism within 30 days before
the first dose of study drug (or sildenafil); use of prescription medication within
14 days before administration of study drug (or sildenafil) or over-the-counter [OTC]
products (including natural products, vitamins) within 7 days before administration
of study drug (or sildenafil). By exception, topical products without systemic
absorption will be allowed. (NOTE: in particular, use of any PDE-5 inhibitor [e.g.,
sildenafil, tadalafil, vardenafil] is prohibited)

- Current history or evidence of drug abuse, history of regular alcohol consumption
exceeding 21 drinks/week for men and 14 drinks/week for women (1 drink = 5 ounces
[150 mL] of wine or 12 ounces [360 mL] of beer or 1.5 ounces [45 mL] of hard liquor)
within 6 months of screening or a positive screen for substances of abuse or alcohol
at screening or pre-dose

- Positive screen for hepatitis B surface antigen (HBsAg), hepatitis C antibody, or HIV
antibody

- Receipt of an investigational product or device, or participation in a drug research
study within a period of 30 days (or 5 half-lives of the drug, whichever is longer)
before the first dose of study drug (sildenafil)

- Blood loss or blood donation of >550 mL within 90 days or plasma donation >500 mL
within 14 days before administration of study drug (or sildenafil)

- Any food allergy, intolerance, restriction, or special diet that, in the opinion of
the Investigator, should preclude the subject's participation in this study

Part B Only:

- Known or suspected hypersensitivity or allergic reaction to sildenafil or other
therapeutics of similar chemical structure or any of the components of Revatio®
tablets

- Any medical condition that constitutes a contraindication or risk to taking
sildenafil, including a history of hereditary degenerative retinal disorders such as
retinitis pigmentosa

- Ingestion of grapefruit or grapefruit juice within 72 hours before sildenafil
administration