Safety Evaluation of 3K3A-APC in Ischemic Stroke (RHAPSODY)

• Maximum Observed Plasma Concentration (Cmax) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Area Under the Plasma Concentration-time Curve (AUC) From Time 0 to Infinity [AUC(0-inf)] for 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Elimination Rate Constant (λz) for 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Half-life (t1/2) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Total Clearance (CL) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Volume of Distribution (V) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Relationship between the change from baseline in activated partial thromboplastin time (aPTT) and the concurrently measured 3K3A-APC plasma concentration [ Time Frame: Following a single dose, on Day 1, 2 or 3: 60 minutes following EOI ]
• Presence of measurable bleeds in the brain (hemorrhage and microbleeds) as determined by 1.5T MRI [ Time Frame: Day 30 ]
  MRI examination to include—at minimum—T1 and T2 weighted images, as well as diffusion weighted imaging (DWI) and susceptibility weighted imaging (SWI) sequences. Post-tPA microbleeds—defined as hypointensities less than 5mm in diameter seen on SWI—will be counted as tPA-related only if found within the ischemic territory. All other areas of hypointensity on SWI larger than 5mm diameter will be counted as tPA-related regardless of the territory in which they are found. All treated subjects (regardless of dose) will be compared to all placebo subjects using a Pearson chi-square test.
• Number of subjects who confirm positive for anti-drug antibody formation [ Time Frame: Day 30 ]

• Maximum Observed Plasma Concentration (Cmax) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Area Under the Plasma Concentration-time Curve (AUC) From Time 0 to Infinity [AUC(0-inf)] for 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Elimination Rate Constant (λz) for 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Half-life (t1/2) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Total Clearance (CL) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Volume of Distribution (V) of 3K3A-APC by Compartmental Analysis [ Time Frame: Following a single dose, on Day 1, 2 or 3: End of infusion (EOI) and 20, 40, 60 and 80 minutes following EOI ]
  Measured following a single dose; primary parameters (CL, V) were used to fit the model. Secondary parameters (Cmax, AUC(inf), λz, t1/2) were estimated from the primary parameters.
• Relationship between the change from baseline in activated partial thromboplastin time (aPTT) and the concurrently measured 3K3A-APC plasma concentration [ Time Frame: Following a single dose, on Day 1, 2 or 3: 60 minutes following EOI ]
• Number of measurable bleeds in the brain (symptomatic & asymptomatic) as determined by 1.5T MRI [ Time Frame: Day 7 (or discharge) ]
  MRI examination to include—at minimum—T1 and T2 weighted images, as well as diffusion weighted imaging (DWI) and susceptibility weighted imaging (SWI) sequences. Post-tPA microbleeds—defined as hypointensities less than 5mm in diameter seen on SWI—will be counted as tPA-related only if found within the ischemic territory. All other areas of hypointensity on SWI larger than 5mm diameter will be counted as tPA-related regardless of the territory in which they are found. All treated subjects (regardless of dose) will be compared to all placebo subjects using a Pearson chi-square test.
• Number of measurable bleeds in the brain (symptomatic & asymptomatic) as determined by 1.5T MRI [ Time Frame: Day 30 ]
  MRI examination to include—at minimum—T1 and T2 weighted images, as well as diffusion weighted imaging (DWI) and susceptibility weighted imaging (SWI) sequences. Post-tPA microbleeds—defined as hypointensities less than 5mm in diameter seen on SWI—will be counted as tPA-related only if found within the ischemic territory. All other areas of hypointensity on SWI larger than 5mm diameter will be counted as tPA-related regardless of the territory in which they are found. All treated subjects (regardless of dose) will be compared to all placebo subjects using a Pearson chi-square test.
• Number of subjects who confirm positive for anti-drug antibody formation [ Time Frame: Day 30 ]

• National Institutes of Health Stroke Scale (NIHSS) score [ Time Frame: Day 7 ]
  The 7-day NIHSS mean scores will be compared using a Kruskal-Wallis test. The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• Modified Rankin Scale (mRS) score [ Time Frame: Day 90 ]
  The 90-day mRS will be compared using a Pearson chi-square test. The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• Barthel Index (BI) score [ Time Frame: Day 90 ]
  The 90-day BI will be compared using a Kruskal-Wallis test. The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• Infarct volume [ Time Frame: Day 90 ]
  The mean infarct volume at 90 days (MRI, or CT if unable to obtain MRI) will be compared using a linear regression model. Median and categorical infarct volumes (based upon quartiles) will be compared. The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• National Institutes of Health Stroke Scale (NIHSS) score [ Time Frame: Day 7 ]
  The change from Baseline NIHSS to 7-day NIHSS will be compared using a linear regression model, with baseline NIHSS score as a covariate. The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• The Relationship between the 7-day NIHSS score and the 90-day mRS [ Time Frame: Days 7 and 90 ]
  The relationship between the 7-day NIHSS score and the 90-day mRS will be examined using a linear regression model. The degree of fit and the R-square will be determined, and the model will be used to assess the potential for using the 7-day outcome in subsequent studies.The comparisons will evaluate all treated subjects (regardless of dose) versus all placebo subjects.
• National Institutes of Health Stroke Scale (NIHSS) score [ Time Frame: Day 7 ]
  The 7-day NIHSS mean scores will be compared using a Kruskal-Wallis test. The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.
• Modified Rankin Scale (mRS) score [ Time Frame: Day 90 ]
  The 90-day mRS will be compared using a Pearson chi-square test. The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.
• Barthel Index (BI) score [ Time Frame: Day 90 ]
  The 90-day BI will be compared using a Kruskal-Wallis test. The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.
• Infarct volume [ Time Frame: Day 90 ]
  The mean infarct volume at 90 days (MRI, or CT if unable to obtain MRI) will be compared using a linear regression model. Median and categorical infarct volumes (based upon quartiles) will be compared.The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.
• National Institutes of Health Stroke Scale (NIHSS) score [ Time Frame: Day 7 ]
  The change from Baseline NIHSS to 7-day NIHSS will be compared using a linear regression model, with baseline NIHSS score as a covariate. The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.
• The Relationship between the 7-day NIHSS score and the 90-day mRS [ Time Frame: Days 7 and 90 ]
  The relationship between the 7-day NIHSS score and the 90-day mRS will be examined using a linear regression model. The degree of fit and the R-square will be determined, and the model will be used to assess the potential for using the 7-day outcome in subsequent studies.The comparisons will evaluate all subjects receiving the dose selected as the MTD versus all placebo subjects.

This is a multicenter, prospective, randomized, controlled, double-blinded Phase 2 study intended to evaluate the safety, PK and preliminary efficacy of 3K3A-APC following treatment with tPA, mechanical thrombectomy or both in subjects with moderate to severe acute ischemic stroke.

Approximately 115 subjects will be randomized, which includes the planned 88 subjects in groups of four to either 3K3A-APC or placebo (in a 3:1 ratio) and the additional placebo subjects who will be enrolled during safety review pauses. This study will utilize a modified version of the continual reassessment method (CRM) in order to establish a maximum tolerated dose (MTD).

Eligible subjects will receive 3K3A-APC or placebo every 12 hours for up to 5 doses (approximately 3 days), or until discharge from the hospital, whichever occurs first. Subjects will be monitored for safety evaluations through Day 7 (or discharge, if earlier) and are expected to be seen on Day 7, 14, 30 and 90 for safety and outcome evaluations.

• Biological: 3K3A-APC
  3K3A-APC, diluted in 0.9% sodium chloride in water, given as 100 mL IV infusion
• Drug: Placebo
  Matching placebo, 0.9% sodium chloride in water, given as 100 mL IV infusion

• Active Comparator: 120 µg/kg of 3K3A-APC
  3K3A-APC, q12h for up to 5 doses
  Intervention: Biological: 3K3A-APC
• Active Comparator: 240 µg/kg of 3K3A-APC
  3K3A-APC, q12h for up to 5 doses
  Intervention: Biological: 3K3A-APC
• Active Comparator: 360 µg/kg of 3K3A-APC
  3K3A-APC, q12h for up to 5 doses
  Intervention: Biological: 3K3A-APC
• Active Comparator: 540 µg/kg of 3K3A-APC
  3K3A-APC, q12h for up to 5 doses
  Intervention: Biological: 3K3A-APC
• Placebo Comparator: Placebo
  Matching placebo, q12h for up to 5 doses
  Intervention: Drug: Placebo

• Lyden P, Levy H, Weymer S, Pryor K, Kramer W, Griffin JH, Davis TP, Zlokovic B. Phase 1 safety, tolerability and pharmacokinetics of 3K3A-APC in healthy adult volunteers. Curr Pharm Des. 2013;19(42):7479-85.
• Lyden P, Weymer S, Coffey C, Cudkowicz M, Berg S, O'Brien S, Fisher M, Haley EC, Khatri P, Saver J, Levine S, Levy H, Rymer M, Wechsler L, Jadhav A, McNeil E, Waddy S, Pryor K. Selecting Patients for Intra-Arterial Therapy in the Context of a Clinical Trial for Neuroprotection. Stroke. 2016 Dec;47(12):2979-2985. Epub 2016 Nov 1.

Inclusion Criteria:

• Acute ischemic stroke
• Able to receive IV tPA, mechanical thrombectomy or both
• National Institutes of Health Stroke Scale (NIHSS) score of ≥ 5
• Signed informed consent
• Mechanical thrombectomy subjects only: onset time to arterial puncture time < 6 hours

Exclusion Criteria:

• History of stroke or penetrating head injury within 90 days prior to enrollment
• History of previous or current diagnosis of intracranial hemorrhage that represents a potential for re-hemorrhage if subjected to thrombolytic therapy or mechanical thrombectomy
• Moyamoya disease, cerebral arterio-venous malformation (AVM), known unsecured aneurysm requiring intervention during the acute study period
• Presence of other neurological or non-neurological co-morbidities that may lead, independently of the current stroke, to further deterioration in the subject's neurological status during the trial period
• Presence of premorbid neurological deficits and functional limitations assessed by a retrospective Modified Rankin Scale (mRS) score of ≥ 2
• Mechanical thrombectomy subjects only: baseline non-contrast CT scan revealing a large core occlusion as defined by local protocol
• Prolonged prothrombin time (PT) or aPTT
• Severe hypertension or hypotension
• Glomerular filtration rate (GFR) <35 mL/min
• Blood glucose concentration < 50 mg/dL
• Prior exposure to any exogenous form of APC

• National Institute of Neurological Disorders and Stroke (NINDS)
• Cedars-Sinai Medical Center
• Massachusetts General Hospital
• University of Iowa