Safety Study of CZ10 in Healthy Volunteers

A randomized, double-blind, placebo-controlled, parallel group, dose escalation study in healthy subjects to evaluate the safety of intravenous CZ10 medical imaging contrast material.

Development of a new intravenous clinical CT contrast agent would provide diagnostic value for CT, particularly for patients unable to tolerate existing agents. The current study will be the first-in-human test of a novel CZ10 contrast material with excellent signal and safety in preclinical testing.

Supporting Preclinical Safety Data

CZ10 has been the subject of substantial in vivo and safety research. CZ10 undergos renal elimination with low retention in other organs, comparable to those of current CT contrast agents. CZ10 is well tolerated, having been administered to rats at 3000 mg/kg, and repeatedly in rabbits and pigs at 400 - 540 mg/kg. CZ10 showed minimal, transient increase in only one urinary kidney stress marker (neutrophil gelatinase-associated lipocalin, NGAL) after 1500 mg/kg IV injection in rats, but no histopathological evidence of kidney injury was seen. Blood counts and clinical chemistry of rats given CZ10 at 1500 and 3000 mg/kg were unchanged from controls. Blood counts and clinical chemistry measurements from rabbits and swine also showed no adverse effects, although these studies were imaging-focused where safety was not an endpoint. Peritoneal injection of CZ10 at 500 mg/kg in rats showed rapid clearance at CT imaging on day 2 with no observable toxicity. Good laboratory practice testing of CZ10 in rat and dog showed no toxicity referable to CZ10.

INVESTIGATIONAL PLAN

OVERALL STUDY DESIGN

This is a randomized, double-blind, placebo-controlled, parallel group, dose escalation study in healthy subjects to evaluate the safety of intravenous CZ10 CT contrast material.

This first-in-human clinical phase 1 trial will test the safety of escalating doses of CZ10 that has shown high safety and efficacy as an intravenous contrast agent in extensive preclinical testing.

The subjects for this study will be healthy adult volunteers, including obese but otherwise healthy subjects up to 450 lbs. Subjects will be recruited and randomly assigned to cohort and test article.

A total of 24 subjects will be evaluated with 18 subjects randomized to receive active drug and 6 subjects randomized to receive placebo control in a double blind manner. A total of three cohorts of 8 subjects will be enrolled. The first cohort subjects will receive a low dose. A review of safety data from all 8 subjects will be performed. If less than 2 subjects show severe adverse events, then the second cohort will be enrolled. A review of safety data from these 8 subjects will then be reviewed, and if less than 2 subjects show severe adverse events, the last cohort will be enrolled. This last cohort will receive a high dose of the intravenous test article. For each cohort, six subjects will be randomized to receive CZ10 and two to placebo.

The study will be performed at a dedicated Clinical Phase 1 center. A dedicated Clinical Phase 1 center is desirable due to the specialized testing and subject recruitment required and the need for strict adherence to the procedures in the study manual. Good clinical practice will be emphasized and documented for all aspects of this trial.

INVESTIGATIONAL PRODUCT - INJECTABLE CONTRAST

Investigational Product and Placebo Description CZ10 will be administered as an injectable formulation. The aqueous formulation will have been prepared as a sterile bottled transparent fluid. A similar appearing saline placebo will be prepared.

Investigational Product Preparation The bottle of the agent will be opened in the CT suite and loaded manually into the CT injector syringe in the same manner as conventional CT contrast material, with care to remove gas bubbles. The volume of injection is likely to be 2 mL per kg body weight, but may be increased to 5 mL per kg of body weight in the high dose group. In some obese subjects, a dose volume of 2 or 4 mL per kg may not be achievable due to limitations of the contrast power injector. In such cases, the maximum feasible dose will be used (the limit of the power injector, which is 220 mL). This adjustment and maximum dose volume reflects standard practice in clinical radiology.

Investigational Product Storage and Accountability Investigational product will be stored at room temperature in a secure, locked area, under the responsibility of the clinical research unit pharmacy or its designee. The Investigator or Investigational Pharmacy must maintain accurate records of the receipt of all study drug, including date received, lot number, amount received, condition of the package and the disposition. Dispensing records will be maintained including the date and amount of study drug dispensed and the identity of the subject receiving the study drug.

Following dosing, all empty bottles will be retained until reconciliation with the dispensing and shipping records is complete. Empty bottles may then be destroyed per investigative pharmacy policy.

Following final reconciliation (at the end of the study), unused study drug may either destroyed according to site guidelines or returned to Nextrast, Inc.

Investigational Product Packaging and Labeling Labels will be accordance with all applicable regulatory requirements for the labeling of active pharmaceutical ingredients. Labels will contain the drug name, protocol number, date of manufacture, storage conditions and a caution statement that the drug is for clinical investigational use only.

Methods of Assigning Subjects to Dose Groups:

The subject treatment assignment will be based on a computer generated randomization scheme with a ratio of 3:1 (active:placebo) for each of the three drug level cohorts. The study drug blind will be maintained through a randomization schedule held by the Pharmacist randomization personnel in the CRU pharmacy.

SUBJECT POPULATION

Inclusion Criteria

To be eligible for enrollment, a subject must meet the following criteria:

1. Understands the requirements of the study and provides written informed consent prior to undergoing any study-related procedures 2. Subject is between the ages of 18 to 85 years old, inclusive 3. Has good venous access as determined by the Investigator at screening 4. Is able to lie flat with arms above head for 15 mintues and hold breath for 15 seconds 5. Weighs up to 450 lbs

Exclusion Criteria

Subjects will be excluded from the study if they meet any of the following criteria:

1. Is pregnant (as determined by a urine pregnancy test at the time of consent) 2. Has significant cardiovascular, respiratory, hepatic, gastrointestinal, hematological, neurological, psychiatric or other disease that may interfere with the objectives of the study and/or pose safety concerns; 3. Has evidence of moderate to severe renal insufficiency or renal failure (defined as an estimated glomerular filtration rate \<60 ml/minute) 4. Has an allergy to iodinated CT or other medical intravenous contrast material 5. Weighs more than 450 pounds which would result in inability to be scanned on CT 6. Is breastfeeding 7. Is a woman of childbearing age who is not using double protection birth control

Replacement of Subjects This study plans to enroll 24 evaluable subjects. For the purpose of this study "evaluable" subjects are those that are successfully injected with the full dose of intravenous test article. Non-evaluable subjects may be replaced following discussion between the Principal Investigator and Nextrast.

SCHEDULE OF PROCEDURES

Study Visits There are 4 planned in-clinic visits and 1 planned follow-up phone call for this study. Additional visits may be needed if follow up for an adverse event is needed or if screening cannot take place in a single visit. Investigators should schedule visits within the protocol-specified timelines unless they receive permission from Nextrast, or unforeseeable events prevent the subject from complying.

All study procedures must be recorded in source documentation (primary occurrence of a record) maintained by the clinical site.

Informed Consent Prior to performing any study-related activities under this protocol, including screening tests and assessments, written informed consent with the IRB-approved ICF must be obtained from the subject in accordance with local practice and regulations.

A copy of the ICF, signed and dated by the subject, must be given to the subject and documented in the subject's source documentation. The original signed consent form will be retained with the study records

Screening Visit (Days -30 to 0):

Once informed consent is obtained, the following assessments will be done to establish a subject's eligibility for the study. Note: in cases where subjects have given consent for general pre-screening at the site, some of these assessments may be done via chart review.

When possible, the assessments should be done in the order listed here to avoid the demands of sample collection and ECG prior to the subject passing basic entry criteria requirements.

* Review of eligibility requirements - inclusion and exclusion criteria * Collection of demographic information and medical history, including review of prior and ongoing medications (taken in previous 30 days) * Limited Physical Exam * Vital signs measurement (blood pressure, pulse rate, respiratory rate and oral temperature) * In women of child-bearing potential, urine sample for pregnancy test Once eligibility is established, subjects will be scheduled for their Day 1 visit

Day 1 Visit: Dosing and Imaging

Subjects will be instructed to arrive early in the morning to the clincal research unit. The following assessments must be done in the order noted:

1. Pre-dose: May be done up to 2 hours before start of dosing