Vitamin C Infusion for TReatment In Sepsis Associated Acute Lung Injury

CITRIS-ALI

Protocol Version 9

1. BACKGROUND ..... 12
   1.1. Introduction ..... 12
   1.2. Preliminary Progress - Vitamin C Intervention in Experimental Septic Lung Injury ..... 14
   1.3 Phase I Human Trial: Vitamin C Infusion In Human Sepsis: Preliminary Results From A Safety Trial ..... 17
   1.4. Potential Mechanisms of Action of Ascorbic Acid in Sepsis ..... 19
   1.5. Ascorbic Acid Dose Selection ..... 19
   1.6. Study Rationale ..... 20
2. OBJECTIVES ..... 20
   2.1. Primary Objectives ..... 20
   2.2. Hypothesis ..... 21
3. END-POINTS ..... 21
   3.1. Primary Endpoints ..... 21
   3.2. Secondary End Points ..... 21
   3.3. Focused Safety Analysis: ..... 22
   3.3.1 Renal Monitoring Plan ..... 22
   3.3.2 Glucose Monitoring Plan ..... 22
4. STUDY POPULATION AND ENROLLMENT ..... 23
   4.1. Number/Source/Screening ..... 23
   4.2. Inclusion Criteria ..... 23
   4.2.1 CITRIS-ALI Inclusion Criteria: ..... 23
   4.3. Enrollment, Randomization, and Study Initiation Time Window ..... 25
   4.4. Informed Consent ..... 25
   4.5. Randomization ..... 25
   4.7. Minorities and Women ..... 25
5. STUDY PROCEDURES ..... 25
   5.1. Vitamin C or Placebo Administration ..... 26
   5.2. Drug Level Specimens (venous blood) ..... 26
   5.3. Completion of Study Drug Administration ..... 26
   5.4. Premature Withdrawal from Treatment ..... 27
   5.5. Ventilator Procedures ..... 27
   5.6. On-Study Fluid Management ..... 27
6. DATA COLLECTION ..... 28
   6.1. Background Assessments ..... 28
   6.2. Baseline/Hour 0 Assessments and Procedures ..... 29
   6.3. Assessments after Enrollment ..... 29
7. STATISTICAL CONSIDERATIONS ..... 30
   7.1. Statistical Methods ..... 30
8. DATA COLLECTION AND SITE MONITORING ..... 31
   8.1. Data Collection ..... 31
   8.2. Site Monitoring ..... 31
9. RISK ASSESSMENT ..... 32
   9.1. Risks of Active Study Drug ..... 32
   9.2. Risks of Blood Draws ..... 32
   9.3. Minimization of Risks ..... 32
   9.4. Potential Benefits ..... 33
   9.5. Risks versus Benefits ..... 33
10. HUMAN SUBJECTS ..... 33
    10.1. Selection of Subjects ..... 33
    10.1.1. Equitable Selection of Subjects ..... 33
    10.1.2. Vulnerable Subjects ..... 33
    10.2. Informed Consent ..... 34
    10.3. Identification of Surrogates ..... 34
    10.4. Justification of Surrogate Consent ..... 34
    10.5. Additional Safeguards for Vulnerable Subjects. ..... 35
    10.6. Confidentiality ..... 35
11. ADVERSE EVENT REPORTING/SAFETY REPORTING ..... 35
12. APPENDICES ..... 38
    APPENDIX A: Guidelines for evidence of infection ..... 38
    APPENDIX B: Pleiotropic Effects of Vitamin C ..... 40
    APPENDIX C: Ventilator Procedures ..... 42
    C.1. Ventilator Management. ..... 42
    C.2. Weaning ..... 43
    C.3. Definition of Unassisted Breathing ..... 44
    C.4. Definition of Extubation ..... 44
    C.5. Completion of Ventilator Procedures ..... 44
    C.6. Removal from the Ventilator Management Protocol ..... 45
    APPENDIX D: FACTT LITE Conservative Fluid Management Approach ..... 46
    APPENDIX E: Time Events Schedule ..... 47
    APPENDIX F: Adverse Events ..... 48
    APPENDIX G: Modified SOFA Score Calculator ..... 53
    APPENDIX H: P/F from S/F Calculator ..... 54
    REFERENCES CITED: ..... 55

ABBREVIATIONS

DEFINITIONS

Part I: Study Summary

(CITRIS-ALI)

Hypothesis:

Study Design:

1. Multi-center, prospective, randomized, placebo-controlled clinical trial
2. A maximum of 170 patients will be enrolled
3. Participants will be randomized to receive either intravenous Vitamin C (mixed in dextrose in water) or placebo (5% dextrose in water)
4. Active treatment will continue for 96 hours, discharge from study hospital, discharge from the ICU, study withdrawal, or death, whichever comes first.
5. All participants will be followed for a total of 60 days.

Analysis/Interim Monitoring:

1. The principal analysis will be on the basis of the intention-to-treat.
2. Protocol compliance will be monitored by the study team by presentation of two enrolled subjects per site to the team. This will take place via investigator conference call and will address challenges encountered. Trial progress will be monitored by an independent Data and Safety Monitoring Board to determine if the study should stop for safety reasons. As an early phase study it is important to collect as much data as possible. For this reason the study will only be halted by the DSMB for reasons of patient safety concerns. The first scheduled analysis will occur after the enrollment of 80 patients or semi-annually, whichever happens first. The next review will occur after enrollment of the last enrolled subject, or semi-annually, whichever comes first. In the event that safety concerns arise prior to the scheduled analysis, the DSMB may request an unscheduled review at any time. The Data and Safety Monitoring Board (DSMB) will also monitor trial quality.
3. Regulatory compliance, GCP, and risk-based monitoring will be provided by an independent CRO.

CITRIS-ALI Inclusion and Exclusion Criteria:

CITRIS-ALI Inclusion Criteria:

1. Suspected or proven infection: (e.g., thorax, urinary tract, abdomen, skin, sinuses, central venous catheters, and central nervous system, see Appendix A).
2. The presence of a systemic inflammatory response: Defined as: fever: (any route) or hypothermia: (core temp only), tachycardia: heart rate beats/min or receiving medications that slow heart rate or paced rhythm, leukocytosis: or leukopenia: or band forms. Respiratory rate breaths per minute or or invasive mechanical ventilation.
3. The presence of sepsis associated organ dysfunction: (any of the following thought to be due to infection)
   a. Sepsis associated hypotension (systolic blood pressure (SBP) or an SBP decrease > 40 mm Hg unexplained by other causes or use of vasopressors for blood pressure support (epinephrine, norepinephrine, dopamine , phenylephrine)
   b. Arterial hypoxemia or supplemental .
   c. Lactate > upper limits of normal laboratory results
   d. Urine output hour for two hours despite adequate fluid resuscitation
   e. Platelet count per mcL
   f. Coagulopathy (INR > 1.5)
   g. Bilirubin
   h. Glasgow Coma Scale < 11 or a positive CAM ICU score
4. ARDS characterized by all the following criteria
   a. Lung injury of acute onset, within 1 week of an apparent clinical insult and with progression of respiratory symptoms
   b. Bilateral opacities on chest imaging not explained by other pulmonary pathology (e.g. pleural effusions, lung collapse, or nodules)
   c. Respiratory failure not explained by heart failure or volume overload
   d. Decreased arterial ratio
   e. Minimum PEEP of

CITRIS-ALI Exclusion Criteria:

1. Known allergy to Vitamin C
2. inability to obtain consent;
3. age years;
4. Not on a ventilator
5. more than 48 hrs since meeting ARDS criteria;
6. No indwelling venous or arterial catheter in patients requiring insulin in a manner that requires glucose being checked more than twice daily (e.g. continuous infusion, sliding scale)
7. Presence of diabetic ketoacidosis
8. patient or surrogate or physician not committed to full support (not excluded if patient would receive all supportive care except for cardiac resuscitation);
9. pregnancy or breast feeding,
10. moribund patient not expected to survive 24 hours;
11. home mechanical ventilation (via tracheotomy or noninvasive) except for CPAP/BIPAP used only for sleep-disordered breathing;
12. on home , except for with CPAP/BIPAP
13. diffuse alveolar hemorrhage (vasculitis);
14. interstitial lung disease requiring continuous home oxygen therapy;
15. Active kidney stone
16. Non English speaking;
17. ward of the state (inmate, other).

Primary Objective:

Endpoints:

Secondary Endpoints:

• Oxygenation Index ( Mean Airway Pressure/ ) at study hour 0, 48, 96, 168 if still intubated in ascorbate infused patient compared to placebo.
• VE-40 (Vent RR x TV/Weight) x ( ) at study hour if still intubated, in ascorbate infused patient compared to placebo
• SOFA scores at hours 48, 96, 168
• SOFA Score Components at hours 48,96, 168
• SpO2/FiO2
• Platelets
• Total Bilirubin
• Vasopressor status
• GCS
• Creatinine or Urine Output
• Procalcitonin, Receptor for Advanced Glycation End Products, Tissue Factor Pathway Inhibitor at study hour 0, 48, 96, 168
• Ascorbate level at hour 0,48,96,168
• Ventilator Free Days to day 28
• ICU-free days at day 28
• All cause mortality to day 28
• Hospital-free days at day 60

Study Drug Dosing:

1. First study drug dose ( L -ascorbic acid or placebo) will be considered "Dose 1 " and will be administered within 6 hours of randomization or the earliest available time post any clinically indicated procedure which requires the patient to be off the unit. All doses will be administered in the ICU. Patients receiving vitamin C will receive of the total daily calculated dosing ( hours) and will be infused over 30 minutes for this first dosing.
2. Subsequent doses which represent of the day's total dose will be infused every six hours through 96 hours ( hours).
   a. Timing of Dose 2 will be triggered by the physician order for q 6 hour administration and will therefore be listed on the bedside MAR. As such, timing of Dose 2 may be out of the +/- 3 hour window and will not trigger a protocol deviation.
   b. If for any reason any other maintenance dose is not administered within window, the dose will be skipped and the next scheduled dose will be given and documented in the data collection tool.

1. Final drug dose at 96 hours
2. Discharge from study hospital
3. Loss of indwelling venous or arterial catheter with no intent to replace the line, making it impossible to monitor glucose levels via central laboratory without multiple peripheral sticks.
4. Discharge from the ICU
5. Withdrawal from study
6. Death

Part II: Study DescriptionVitamin C Infusion for Treatment In Sepsis Associated Acute Lung Injury CITRIS-ALI

1. Background

1.1. Introduction

1.2. Preliminary Progress - Vitamin C Intervention in Experimental Septic Lung Injury

Figure 1

Figure 2

Figure 3

Figure 4 shows H&E stains of sections of lungs removed at 16 hours following onset of feces induced peritonitis. As seen in this figure, intense cellular sequestration and septal edema is present in the lung of unprotected FIP-treated mice. In contradistinction, AscA treatment significantly attenuated the histological findings of murine sepsis.

Figure 7

DIC

Figure 8

1.3 Phase I Human Trial: Vitamin C Infusion In Human Sepsis: Preliminary Results From A Safety Trial

Figure 9

Figure 10

Sequential Organ Failure Assessment (SOFA) Scores SOFA

Figure 11

Figure 12

1.4. Potential Mechanisms of Action of Ascorbic Acid in Sepsis

• Our experimental data in two murine models of sepsis induced lung injury suggest that vitamin C when infused acts in a pleotropic manner, attenuating NF%B inducible genes (chemokines, tissue factor) while boosting expression of genes leading to active fibrinolysis (i.e., tissue factor pathway inhibitor).
• Humans lack L-gulono- -lactone oxidase, the final enzyme in vitamin C biosynthesis.
• Sodium-dependent vitamin C transporters move vitamin C into cells in reduced form or via facilitative glucose transporters in oxidized form as dehydroascorbic acid (DHA). DHA is rapidly reduced and trapped intracellular as reduced vitamin C or L-ascorbic acid.
• Though vitamin C circulates in normal human plasma at , it accumulates normally in millimolar concentrations in host defense cells (i.e., neutrophils, platelets, macrophages) and endothelium. Together with glutathione, vitamin C constitutes a primary line of defense against ROS and promotes recycling of other antioxidants (e.g., vitamin E).
• Subnormal plasma vitamin C concentrations in septic patients correlate inversely with multiple organ failure and directly with survival. Vitamin C depletion in sepsis results from: 1) ascorbate consumption by reduction of plasma free iron, 2) ascorbate consumption by the scavenging of aqueous free radicals, and 3) by destruction of DHA. Sepsis associated vitamin C destruction permits uncontrolled oxidant activity.
• Clinical protocols currently in use for hospitalized septic patients are inadequate to normalize plasma vitamin C levels.
• Ascorbate infusion into septic animals: 1) improves survival, corrects hypotension, improves capillary blood flow, protects endothelial barrier function, attenuates peroxynitrite formation, attenuates ALI, and disrupts lung capillary microvascular thrombosis (see preliminary murine data above).

1.5. Ascorbic Acid Dose Selection

1.6. Study Rationale

2. Objectives

2.1. Primary Objectives

• VS (Body weight, blood pressure, heart rate, Temperature, mean arterial pressure, oxygen saturation, central venous pressure, glasgow coma score)
• Vasopressor use (amount and type) (mcg/kg/min)
• Ventilator data - Is the patient on or off vent?
  ✓ If on mechanical ventilation: Mean Airway Pressure, tidal volume, Peak Inspiratory Pressure, FiO2, Respiratory Rate, Plateau Pressure, Positive End Expiratory Pressure (data recorded from chart at the 8am time point or closest time point to 8am available)
• Arterial Blood Gases: - for as long as subject has arterial line
• Laboratory (Sodium, Potassium, Chloride, Metabolic Glucose, Hemoglobin, Hematocrit, Platelets, White Blood Cell Count, Creatinine, Blood Urea Nitrogen, Bilirubin (data recorded from chart at the 8am time point or closest time point to 8am available).
• Biomarker blood sample
• Sequential Organ Failure Assessment Score (SOFA): (if intubated use ratio for calculation. If not intubated see Appendix H for caluculating FiO 2 and from SpO 2 . Calculate using the worst value in the 24 hours preceding the Score time point for each component. See Appendix G for Modified SOFA Score Calculator.

2.2. Hypothesis

3. End-Points

3.1. Primary Endpoints

3.2. Secondary End Points

• Oxygenation Index ( Mean Airway Pressure/ ) at study hour 0, 48, 96, 168 if still intubated in ascorbate infused patient compared to placebo.
• VE-40 (Vent RR x TV/Weight) at study hour if still intubated, in ascorbate infused patient compared to placebo
• SOFA scores at hours 48, 96, 168
• SOFA Score Components at hours 48,96, 168
• Platelets
• Total Bilirubin
• Vasopressor status
• GCS

- Creatinine or Urine Output

• Angiopoietin-2, Procalcitonin, Receptor for Advanced Glycation End Products, Tissue Factor Pathway Inhibitor at study hour 0, 48, 96, 168
• Ascorbate level at hour 0, 48, 96, 168
• Ventilator Free Days to day 28
• ICU-free days at day 28
• All cause mortality to day 28
• Hospital-free days at day 60

3.3. Focused Safety Analysis:

3.3.1 Renal Monitoring Plan

3.3.2 Glucose Monitoring Plan

• Critical care Nursing and Physician leadership at all study sites must be informed of vitamin C's effect on point of care (glucometer) blood glucose and arterial blood gas glucose point of care values.
• In-service training will be documented in the Study Training Log
• Bold signage will be displayed on all study instructions, data collection forms, and at the patient's head of bed, stating:
• STOP! Do not use Accuchek or other Point of Care devices to measure glucose on this patient
• Use only metabolic or gas lab glucose screening methods
• This patient is enrolled in a study with Vitamin C, which artefactually increases POC glucose testing
• Do Not Initiate or Utilize Sliding Scale, Scheduled Insulin, or Continuous Insulin Infusion Without Laboratory Confirmation of Blood Glucose
• Those receiving insulin infusion or sliding scale insulin will have metabolic glucose screening on the schedule determined by the primary physician and paid for by the study
• Blood glucose monitoring for insulin administration guidance should only be by a metabolic or blood gas laboratory measured blood glucose results, whether or not the study patient is receiving insulin
• Study personnel will follow each study patient closely to monitor insulin use to ensure that point of care glucose screening is suspended for the research subject.
• If subject loses central venous access (PICC line and arterial line acceptable), Vitamin C infusions are to stop but subject not withdrawn. Data collected through end of study.
• Point of care glucose testing may resume 36 hours after the last infusion of study drug.

4. Study Population and Enrollment

4.1. Number/Source/Screening

4.2. Inclusion Criteria

4.2.1 CITRIS-ALI Inclusion Criteria:

CITRIS-ALI Inclusion Criteria:

1. Suspected or proven infection: (e.g., thorax, urinary tract, abdomen, skin, sinuses, central venous catheters, and central nervous system, see Appendix A).
2. The presence of a systemic inflammatory response: Defined as: fever: (any route) or hypothermia: (core temp only), tachycardia: heart rate beats/min or receiving medications that slow heart rate or paced rhythm, leukocytosis: or leukopenia: or band forms. Respiratory rate breaths per minute or or invasive mechanical ventilation.
3. The presence of sepsis associated organ dysfunction: (any of the following thought to be due to infection)
   a. Sepsis associated hypotension (systolic blood pressure (SBP) or an SBP decrease > 40 mm Hg unexplained by other causes or use of vasopressors for blood pressure support (epinephrine, norepinephrine, dopamine , phenylephrine, vasopressin)
   b. Arterial hypoxemia or supplemental .
   c. Lactate > upper limits of normal laboratory results
   d. Urine output hour for two hours despite adequate fluid resuscitation
   e. Platelet count per mcL
   f. Coagulopathy (INR )
   g. Bilirubin
   h. Glasgow Coma Scale < 11 or a positive CAM ICU score
4. ARDS characterized by all the following criteria
   a. Lung injury of acute onset, within 1 week of an apparent clinical insult and with progression of respiratory symptoms
   b. Bilateral opacities on chest imaging not explained by other pulmonary pathology (e.g. pleural effusions, lung collapse, or nodules)
   c. Respiratory failure not explained by heart failure or volume overload
   d. Decreased arterial ratio
   e. Minimum PEEP of

CITRIS-ALI Exclusion Criteria:

1. Known allergy to Vitamin C
2. inability to obtain consent;
3. age years;
4. Not on a ventilator
5. No indwelling venous or arterial catheter in patients requiring insulin in a manner that requires glucose being checked more than twice daily (e.g. continuous infusion, sliding scale)
6. Presence of diabetic ketoacidosis
7. more than 48 hrs since meeting ARDS criteria;
8. patient or surrogate or physician not committed to full support (not excluded if patient would receive all supportive care except for cardiac resuscitation);
9. pregnancy or breast feeding,
10. moribund patient not expected to survive 24 hours;
11. home mechanical ventilation (via tracheotomy or noninvasive) except for CPAP/BIPAP used only for sleep-disordered breathing;
12. on home , except for with CPAP/BIPAP
13. diffuse alveolar hemorrhage (vasculitis);
14. interstitial lung disease requiring continuous home oxygen therapy;
15. Active kidney stone
16. Non English speaking;
17. Ward of the state (inmate, other)

4.3. Enrollment, Randomization, and Study Initiation Time Window

4.4. Informed Consent

4.5. Randomization

4.7. Minorities and Women

5. Study Procedures

5.1. Vitamin C or Placebo Administration

1. First study drug dose ( L -ascorbic acid or placebo) will be considered "Dose 1" and will be administered within 6 hours of randomization or the earliest available time post any clinically indicated procedure which requires the patient to be off the unit. All doses will be administered in the ICU. Patients receiving vitamin C will receive of the total daily calculated dosing ( hours) and will be infused over 30 minutes for this first dosing.
2. Subsequent doses which represent of the day's total dose will be infused every six hours through 96 hours (+/- 3 hours).
   a. Timing of Dose 2 will be triggered by the physician order for q 6 hour administration and will therefore be listed on the bedside MAR. As such, timing of Dose 2 may be out of the +/- 3 hour window and will not trigger a protocol deviation.
   b. If for any reason any other maintenance dose is not administered within window, the dose will be skipped and the next scheduled dose will be given and documented in the data collection tool.

5.2. Drug Level Specimens (venous blood)

5.3. Completion of Study Drug Administration

1. Final drug dose at 96 hours or discharge from ICU, whichever comes first.
2. Discharge from study hospital
3. Loss of indwelling venous or arterial catheter with no intent to replace the line, making it impossible to monitor glucose levels via central laboratory without multiple peripheral sticks.
4. Withdrawal from study
5. Death

5.4. Premature Withdrawal from Treatment

5.5. Ventilator Procedures

5.6. On-Study Fluid Management

6. Data Collection

6.1. Background Assessments

1. Demographic:
   a. Gender
   b. Age
   c. Race/Ethnicity
2. Insurance status
   a. Privately insured
   b. Medicaid
   c. Medicare
   d. Other public
   e. uninsured
3. Pertinent Medical History and Physical Examination
   a. Etiology of Sepsis
   b. diabetic status - Hx of Diabetes □ Yes □ No
   i. Insulin received? □ Yes □ No
   c. patient place of residence: Home independently, home with help (supervision, direction, personal assistance), home with professional help (nursing/nursing service), intermediate care or rehab facility, skilled nursing facility, other (specify)
   d. Patient admitted directly from: OR, Recovery Room, ER, Floor, another special care unit, another hospital, direct admit, step-down unit
   e. Hx of alcohol use via the AUDIT-C Questionnaire:
   i. How often do you have a drink containing alcohol?
   □ Never
   □ Monthly or less
   □ 2-4 times per month
   □ 2-3 times per week
   □ 4 or more times per week
   ii. How many standard drinks containing alcohol do have on a typical day?
   □ 1 or 2
   □ 3 or 4
   □ 5 or 6
   □ 7 to 9
   □ 10 or more
   iii. How often do you have six or more drinks on one occasion?
   □ Never
   □ Less then monthly
   □ Monthly
   □ Weekly

6.2. Baseline/Hour 0 Assessments and Procedures

6.3. Assessments after Enrollment

Reference Measurements

1. Ventilator Data - The following conditions will be ensured prior to measurements: no endobronchial suctioning for 10 minutes; no invasive procedures or ventilator changes for 30 minutes.
2. Arterial Blood Gases when Arterial line is in place for clinical reasons
3. Vital Signs
4. Labs
5. Vasopressor
6. Use of Steroids
7. Blood samples for biomarkers.

7. Statistical Considerations

7.1. Statistical Methods

8. Data Collection and Site Monitoring

8.1. Data Collection

8.2. Site Monitoring

9. Risk Assessment

9.1. Risks of Active Study Drug

9.2. Risks of Blood Draws

9.3. Minimization of Risks

9.4. Potential Benefits

9.5. Risks versus Benefits

10. Human Subjects

10.1. Selection of Subjects

10.1.1. Equitable Selection of Subjects

10.1.2. Vulnerable Subjects

10.2. Informed Consent

10.3. Identification of Surrogates

10.4. Justification of Surrogate Consent

10.5. Additional Safeguards for Vulnerable Subjects

10.6. Confidentiality

11. Adverse Event Reporting/Safety Reporting

For this trial, a reportable adverse event is defined as:

1. Any clinically important untoward medical occurrence in a patient receiving study drug or undergoing study procedures which is different from what is expected in the clinical course of a patient with severe sepsis associated ALI, or,
2. Any clinically important, untoward medical occurrence that is thought to be associated with the study drug or procedures, regardless of the "expectedness" of the event for the course of a patient with severe sepsis associated ALI.
3. Investigators will report all serious, unexpected, AND study-related adverse events from the time of informed consent through study hour 168 that are considered to be harmful and unintended responses to the investigational product and/or study related procedures in the participants' case report forms. 'Responses to investigational product' means that the causal relationship between an investigational product and an adverse event cannot be ruled out.

Expected Events For ALI considered unreportable: refer to Appendix F Table

1. Defined as: Untoward clinical occurrences perceived by the investigator to occur with reasonable frequency in the day to day care of patients with ALI treated in an intensive care unit with mechanical ventilation.
2. Examples of untoward clinical occurrences that are expected in the course of ALI include: 1) transient hypoxemia, 2) agitation, 3) delirium, 4) nosocomial infections, 5) skin breakdown, and 6) gastrointestinal bleeding. Such events, which are often the focus of prevention efforts as part of usual ICU care, will not be considered reportable adverse events unless the event is considered by the investigator to be associated with the study drug or procedures, or unexpectedly severe or frequent for an individual patient with ALI. Examples of unexpectedly frequent untoward clinical occurrences would be repeated episodes of unexplained hypoxemia. This would be in contrast to an isolated episode of transient hypoxemia (e.g., Sp02 ), related to positioning or suctioning. This latter event would not be considered unexpected by nature, severity or frequency.
3. Adverse events occurring from the time of informed consent through study hour 168 or until discharged from the hospital, withdrawal from the study or death, will not be considered 'reportable' Adverse Events (AE)/Serious Adverse Events (SAE) unless the Investigator has a reasonable doubt regarding the relatedness of the event to the investigational product.

• Unexpected, in terms of nature, severity, or frequency, given the research procedures that are described in the protocol-related documents, such as the IRB-approved research protocol and informed consent document; and the characteristics of the subject population being studied;
• Related or possibly related to participation in the research, in this guidance document, possibly related means there is a reasonable possibility that the incident, experience, or outcome may have been caused by the procedures involved in the research;
• Suggests that the research places subjects or others at a greater risk of harm (including physical, psychological, economic, or social harm) than was previously known or recognized.

1. Safety Reporting:

12. APPENDICES

APPENDIX A: Guidelines for evidence of infection

1. Infections of the thorax:
   a) Chest x -ray or CT scan showing a new or progressive infiltrate, consolidation, cavitation, collection, or pleural effusion, and a clinical presentation consistent with pneumonia or empyema
   b) Pneumonia can be defined as the presence of new infiltrate (s), absence of a noninfectious explanation and either signs of SIRS as per protocol or purulent sputum production with an identifiable pathogen.
   c) Aspiration Pneumonitis in the acute phase is not considered an infection.
   d) However, if SIRS persists > 24 hours after aspiration, then an infectious etiology can be presumed.
2. Abdominal infection:
   a) Perforated viscus or ischemic bowel with either localized peritonitis
   b) Peritoneal fluid with > 250 PMNs
   c) Clinical signs of cholangitis or appendicitis
   d) Clostridium difficile toxin positive with evidence of colon dilation
   e) Suspicion of peritonitis by clinical examination only
3. Skin or soft tissue infection: Acute onset infection of the skin, such as erysipelas, or infection involving deeper soft tissue
4. Bacterial meningitis: cerebrospinal fluid analyses if available and a clinical presentation consistent with bacterial meningitis
5. Urinary Tract:
   a) Positive test for granulocyte esterase or nitrate in urine, or a positive culture (defined as CFU/mL)
   b) Urinalysis with increased WBC count or positive Gram stain
6. Central Line infections:
   a) Catheter-related bloodstream infections (CR-BSIs) are defined as bacteremia/fungemia in a patient with an intravascular catheter with at least one positive blood culture obtained from a peripheral vein, clinical manifestations of infection (i.e., fever, chills, and/or hypotension), and no apparent source for the bloodstream infection except the catheter. The catheter must be in place for at least 48 hours prior to development of the bloodstream infection.

7. Sinusitis

The following are not considered evidence of infection:

APPENDIX B: Pleiotropic Effects of Vitamin C

1. Cell culture/in vitro studies

• Reduced human neutrophil adhesion to endothelial cells
• Protects neutrophils against intracellular effects of superoxide generation.
• Protects monocytes against oxidative damage
• Reduced PMA induction of NF-
• Protects against oxidized-LDL-induced expression of MCP-1 in cultured human umbilical vein endothelial cells
• Modulates the inhibition of platelet aggregation by neutrophils
• Inhibits expression of platelet expression of CD40 ligand which promotes thrombosis
• Inhibits NADPH oxidase subunit p47phox expression in microvascular endothelium
• Influences dendritic cell function.
• Effects of vitamin C on intra cytoplasmic cytokine production in human whole blood monocytes and lymphocytes. exposed to LPS or immune complexes
• Inhibition of the induction of inducible nitric oxide sythetase (iNOS) and TNF- , IL-1 and IL-6 in astrocytes, microglia and macrophages stimulated with LPS or cytokines
• Vitamin C inhibits NO-induced stabilization of HIF-1alpha in HUVECs
• Cobalt-induced oxidant stress in cultured endothelial cells: prevention by ascorbate in relation to HIF1alpha.

2. Intact animal studies

• Attenuates LPS induced acute lung injury.
• Attenuates lung injury in a cecal ligation and puncture model of peritonitis.
• Corrects capillary blood flow in septic skeletal musculature.
• Inhibits iNOS expression in septic vasculature.
• Attenuates iNOS expression in IFN gamma-stimulated rat skeletal muscle endothelial cells
• Attenuates hepatic fibrosis by upregulating peroxisome proliferators-activated receptor-gamma
• Inhibits both flow- and agonist-induced EDHF in the rat mesentery
• Multiple molecular transporters responsible for movement of ascorbate intracellular
• Attenuates peroxidative damage and tissue edema in ischemia/reperfused gut.
• Ascorbate supplementation significantly decreases plasma IL-6 levels in an animal model of hemorrhagic shock.
• Vitamin C deficiency causes the collagen-disassembly disease scurvy
• Ascorbic acid prevents testosterone-induced hyperplasia of rat prostate by down-regulating HIF1alpha
• Plasma AA maintains the stability of "acellular Hb " susceptible to oxidation
• Endogenous ascorbate on oxidation, oxygenation, and toxicokinetics of cell-free modified hemoglobin after exchange transfusion in rat and guinea

Human studies

• Normalizes monocyte adhesion to endothelium in vitamin C deficient subjects.
• Supplementation in chronic hemodialysis patients reduce lymphocyte 8-OHdG levels and intracellular ROS production
• Increase in muscle blood flow during dynamic exercise with acute AA administration in older adult humans
• Intravenous ascorbate improves outcomes during percutaneous myocardial intervention
• Plasma vitamin C level positively associated with serum pre-albumin levels and negatively associated with high sensitivity C-Reactive Protein levels in patients with chronic renal
  failure.
• Low plasma ascorbate levels are associated with enhanced proinflammatory responses and impaired vascular function in lean and obese men.
• Reduced inflammatory tissue damage in patients subjected to cardiac surgery with extracorporeal circulation
• Ascorbate promotes iron utilization for erythropoiesis in patients with chronic renal failure
• High dose vitamin C infusion into surgically critically ill daily for 28 days attenuated the incidence of acute lung injuy/ARDS.
  

  

C.1. Ventilator Management

1. Any mode of ventilation capable of delivering the prescribed tidal volume (VT, predicted body weight, ) may be used, provided the VT target is monitored and adjusted appropriately. If airway pressure release ventilation (APRV) is used, tidal volume is defined as the sum of the volume that results from the ventilator pressure- release and an estimation of the average spontaneous VT.
2. VT Goal: predicted body weight.
3. Predicted body weight (PBW) is calculated from age, gender, and height (heel to crown)
   a. according to the following equations:
   b. Males: PBW (kg) [height (inches) -60 ]
   c. Females: PBW ( kg ) [height (inches) -60 ]
4. Measure and record inspiratory plateau pressure (Pplat) according to ICU routine (at least every four hours and after changes in VT and PEEP recommended)
5. If Pplat , reduce VT to and then to if necessary to decrease Pplat to .
6. If and Pplat , raise by to a maximum of .
7. If "severe dyspnea" (more than 3 double breaths per minute or airway pressure remains at or below PEEP level during inspiration), then raise VT to 7 or if Pplat remains below . If Pplat exceeds with VT of 7 or , then revert to lower VT and consider more sedation.
8. If , VT may be raised and Pplat limit suspended (not required).
9. Oxygenation target: [ ] or [88% < SpO2 ]. When both and are available simultaneously, the criterion will take precedence.
10. Minimum
11. Adjust FiO 2 or PEEP upward within 5 minutes if there are consistent measurements below the oxygenation target range
12. Adjust or PEEP downward within 30 minutes if there are consistent measurements above the oxygenation target range.
13. There are no requirements for maintaining a specific PEEP to ratio. The lower higher table represents a consensus approach developed by ARDS Network investigators in 1995. The higher PEEP/lower table (ALVEOLI) yielded equivalent results in a randomized trial and would be acceptable and perhaps preferable in patients who appear to respond with a substantial increase in arterial oxygenation in the transition from lower to higher PEEP.

Lower PEEP/Higher FiO2 Treatment Group

C.2. Weaning

1. At least 12 hours since enrollment in the trial
2. and or and
3. Values of both PEEP and values from previous day
4. Not receiving neuromuscular blocking agents and without neuromuscular blockade
5. Patient exhibiting inspiratory efforts. If no efforts are evident at baseline, ventilator set rate will be decreased to of baseline level for up to 5 minutes to detect inspiratory efforts.
6. Systolic arterial pressure without vasopressor support ( dopamine will not be considered a vasopressor)

Spontaneous Breathing Trial Procedure and Assessment for Unassisted Breathing

1. Pressure support (PS) , PEEP
3. T-piece
4. Tracheostomy collar (mask)

tracheostomy mask, or T-piece) at any time during the spontaneous breathing trial.

1. and or
2. Mean spontaneous tidal volume PBW (if measured)
3. Respiratory Rate / min
4. (if measured)
5. No respiratory distress (defined as 2 or more of the following):
   a. Heart rate of the 0600 rate ( at may be tolerated)
   b. Marked use of accessory muscles
   c. Abdominal paradox
   d. Diaphoresis
   e. Marked subjective dyspnea

Decision to remove ventilatory support:

C.3. Definition of Unassisted Breathing

1. Spontaneously breathing with face mask, nasal prong oxygen, or room air, OR
2. T-tube breathing, OR
3. Tracheostomy collar (mask) breathing, OR
4. CPAP without PS or IMV assistance
5. Use of CPAP or BIPAP solely for sleep apnea management

C.4. Definition of Extubation

1. Removal of an oral or nasotracheal tube
2. If a patient receives a tracheostomy, the time of extubation is defined as the time when the patient achieves unassisted breathing as defined in section C. 3

C.5. Completion of Ventilator Procedures

1. Death
2. Hospital discharge
3. Alive 28 days after enrollment

C.6. Removal from the Ventilator Management Protocol

APPENDIX D: FACTT LITE Conservative Fluid Management Approach

• Discontinue maintenance fluids.
• Continue medications and nutrition.
• Manage electrolytes and blood products per usual practice.
• For shock, use any combination of fluid boluses and vasopressor(s) to achieve MAP as fast as possible. Wean vasopressors as quickly as tolerated beginning four hours after blood pressure has stabilized.
• Withhold diuretic therapy in renal failure and until 12 hours after last fluid bolus or vasopressor given.

CVP (recommend)	PAOP (optional)	MAP AND off vasopressors for hours
		Average urine output < 0.5 ml/kg/hr	Average urine output ml/kg/hr
>8	> 12		Furosemide* Reassess in 4 hours
4-8	8-12	Give fluid bolus as fast as possible Reassess in 1 hour
< 4	< 8		No intervention Reassess in 4 hours

APPENDIX E: Time Events Schedule

Assessments	Hour	024 hrs 7 days or in ICU	Hour	Hour 96	Hour 168	Day 28	Day 60
VS: BP, HR, MAP, RR, Temp., O2 sats, CVP, Glasgow Coma Scale	X		X	X	X
Body Weight	A		A	A
Suspected or known site of sepsis	A		A	A	A	A	A
I/Os Total and Urine only
Assessment of Acute/Chronic Renal Failure and Use of Dialysis			X	X
Calculate SOFA Score (post study by biostatistician)	X		X	X	X
Calculate VE40 Score (post study by biostatistician)	A		A	A	A
Calculate Oxygenation Index (post study by biostatistician)	A			A	A
Ventilator Data: Tv, FiO2, PEEP, Plateau Pressure, Peak Inspiratory Pressure, Mean Airway Pressure, Minute Ventilation			A	A
Labs: Arterial Blood Gasses, Na+, K+, BUN, Cr, WBC, Hgb, Hct, Platelets, PT/INR
Bilirubin Total
Vasopressors or Inotropes: Epi, Nor-epi, Phenylephrine, Vasopressin, Dopamine			A	A
Concomitant Medications: Methylprednisone, Hydrocortisone	X		X	X	X
AE/SAE Assessments	X
Blood for Biomarkers	X			X	X
Ventilator Free Days						X
All Cause Mortality
ICU Free Days						X
Hospital Free Days
Glucose Monitoring

APPENDIX F: Adverse Events

Procedures for Reporting Adverse Events

• Any clinically significant abnormal laboratory findings or other abnormal safety assessments that are associated with the underlying disease, unless judged by the investigator to be more severe than expected for the subject's condition.
• The disease/disorder being studied, or expected progression, signs, or symptoms of the disease/disorder being studied, unless more severe than expected for the subject's condition.
• Medical or surgical procedure (e.g., endoscopy, appendectomy); the condition that leads to the procedure is an AE .
• Situations where an untoward medical occurrence did not occur (social and/or convenience admission to a hospital).
• Anticipated day-to-day fluctuations of pre-existing disease(s) or condition(s) present or detected at the start of the study that do not worsen;
• "Lack of efficacy" or "failure of expected pharmacological action" would not be reported as an AE or SAE. However, the signs and symptoms and/or clinical sequelae resulting from lack of efficacy will be reported if they meet the definition of an AE or SAE.
  If an event does not meet the definition of an AE per Section 11, then it cannot be an SAE even if serious conditions are met (e.g., hospitalization for signs/symptoms of the disease under study, death due to progression of disease, etc).

SAE Follow-up language:

1. Serious, Expected, AND Study-Related Adverse Events: adverse events occurring from the time of informed consent through study hour 168 or until discharged from the hospital, withdrawal from the study, or death will not be considered 'reportable' Adverse Events (AE)/Serious Adverse Events (SAE) unless the Investigator has a reasonable doubt regarding the relatedness of the event to the investigational product (refer to Table: Anticipated AE/SAEs for the SEPSIS ALI Study Population).
2. Serious, Unexpected, AND Study-Related Adverse Events: Investigators will report all serious, unexpected, AND study-related adverse events from the time of informed consent through study hour 168, to the Clinical Coordinating Center within 24 hours by email. The local Institutional Review Board must also be notified in a timely manner. The investigator will then submit a detailed written report to the Clinical Coordinating Center and the local Institutional Review Board no later than 5 calendar days after the investigator discovers the event.
3. Definitions of Adverse Events
   a. A serious adverse event is any event that is fatal or immediately life threatening, is permanently disabling, or severely incapacitating, or requires or prolongs inpatient hospitalization. Important medical events that may not result in death, be life threatening, or require hospitalization may be considered serious adverse events when, based upon appropriate medical judgment, they may jeopardize the patient or subject and may require medical or surgical intervention to prevent one of the outcomes listed above. Life-threatening means that the patient was, in the view of the investigator, at immediate risk of death from the reaction as it occurred. This definition does not include a reaction that, had it occurred in a more serious form, might have caused death. Assessment of the cause of the event has no bearing on the assessment of the event's severity.
   b. An unexpected event is any experience not identified by the type, severity, or frequency in the current study protocol or an event that is unexpected in the course of treatment for ALI or ARDS.
   c. Adverse events will be considered to be study-related if the event follows a reasonable temporal sequence from a study procedure and could readily have been produced by the study procedure.
   d. Organ failures or death related to ALI or ARDS or the patient's underlying condition that are systematically captured by the protocol should not be reported as adverse events unless they are considered to be study related.

4. Assigning Causality

- Unrelated:

• Event occurred before dosing;
• Event or concomitant illness due to factors other than drug or study procedure;

- Possibly:

• Reasonable sequential relationship with study procedure or drug treatment;
• Event could be explained by patient's clinical state or other factors
• Probably:
• Reasonable sequential relationship with study procedure or drug treatment;
• Likely to be a known reaction to study agent or chemical group, or predicted by known pharmacology;
• Event cannot easily be explained by patient's clinical state or other factors.

- Definitely:

• Distinct sequential relationship with study procedure or drug treatment;
• Known reaction to study agent or chemical group, or predicted by known pharmacology;
• Event cannot be explained by patient's clinical state or other factors.

ANTICIPATED AE/SAEs for the SEPSIS ALI Study Population