Transcranial Electrical Stimulation in Stroke EaRly After Onset Clinical Trial_ Bridging and Adjunctive Neuroprotection
Phase: NA
Conditions: Acute Ischemic Stroke
Sponsor: University of California, Los Angeles

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Study Protocol 
 
Transcranial electrical Stimulation in Stroke EaRly After onset Clinical Trial_ Bridging and Adjunctive 
neuroprotection (TESSERACT-BA) 
 
 
 
 
 
 
 
 
Version 1.4 
1/12/2021 
 
 

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Investigational Plan:  
Protocol: Transcranial electrical Stimulation in Stroke EaRly After onset Clinical Trial – Bridging and 
Adjunctive neuroprotection (TESSERACT-BA)  
 
 
Overview of Protocol of TESSERACT-BA 
 
Title  
Transcranial electrical Stimulation in Stroke EaRly After onset 
Clinical Trial - Bridging and Adjunctive neuroprotection 
(TESSERACT-BA) 
Objective 
To evaluate the safety, tolerability, and feasibility, and 
preliminarily explore the potential efficacy, of 
cathodal 
transcranial direct current stimulation (C-tDCS) as an innovative 
non- invasive neuroprotection technique in acute ischemic stroke 
patients before and after endovascular thrombectomy  
Study Design  
A single site, sham- controlled, dose escalation study 
Population Studied  
24-48 acute ischemic stroke patients from Ronal Regan Medical 
Center (RRMC) Emergency Department or RRMC Inpatient 
hospital with:  
1) New focal neurologic deficit consistent with acute ischemic 
stroke; 2) NIHSS ≥4; 3) Age ≥18; 4) Last known well time within 
24 hours of randomization stimulation initiation; 5) ICA or M1 or 
M2 MCA occlusion on MRA/CTA; 6) Pre-stroke modified Rankin 
Scale≤ 3; 7) Ineligible for intravenous thrombolysis; 8) Patient 
have undergone thrombectomy with less than a complete 
reperfusion (TICI < 2c,3); 9) Undergoing endovascular 
thrombectomy for patients enrolled in 5 and 6 dose tiers; 10) 
Signed informed consent obtained from the patient or patient’s 
legally authorized representative.  
Intervention 
3:1 randomization to active treatment with C-tDCS vs sham 
control  
Device  
Transcranial Direct Current Stimulation (tDCS) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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A. Specific Aims 
Stroke is a devastating condition and continues to be a leading cause of adult death and disability.1 
Endovascular thrombectomy (ET) has become standard of care for acute ischemic stroke due to large vessel 
occlusion. 2However, ET has limitations that would be aided by neuroprotective interventions. ET fails to achieve 
substantial reperfusion in 20-30% of patient. Even among reperfusing patients, rates of excellent outcome are 
low - only 20-25% achieve freedom from disability (modified Rankin Scale 0-1), due to infarct growth prior to 
reperfusion and delayed and reperfusion injury after reperfusion. 3-5 Therefore, bridging neuroprotective 
therapies prior to ET, can preserve more viable penumbra to rescue intervention by reperfusion and, therefore, 
enhancing its clinical benefit. While adjunctive neuroprotective strategies after ET can protect the brain against 
reperfusion injury and infarct growth due to incomplete reperfusion.6   
Multiple candidate neuroprotective agents have failed in translation from animal to human studies.7,8 
Amongst the missing intervention properties in the failed neuroprotection trials, two offer the most significant 
prospect of success: 1) having a pleiotropic effect, interdicting multiple pathways in the ischemic cascade, and 
2) delivery by a direct transcranial, rather than intravascular, route, with fast and direct delivery to neural tissues 
regardless of cerebral perfusion. Transcranial direct current stimulation (tDCS) offers promise as just such an 
approach. In pre-clinical acute ischemic stroke models, cathodal tDCS (C-tDCS) applied to the ischemic 
hemisphere, has been shown to have a substantial neuroprotective effect with widely pleiotropic neuroprotective 
molecular mechanisms of action and can be delivered to the salvageable ischemic tissue beyond the occlusive 
thrombus despite reduced blood flow.9,10  
This study proposes to conduct the first-in-human, sham-controlled, dose-escalation, safety, feasibility, 
and potential efficacy trial of C-tDCS as a bridging and adjunctive neuroprotection before and after thrombectomy 
in acute ischemic stroke. To fully assess the potential safety aspects of adjunct C-tDCS to ET, we will exclude 
patients who receive intravenous thrombolysis.  
 
Specific Aim I: To establish the safety and tolerability of C-tDCS in AIS as an adjunct and bridge to 
endovascular thrombectomy (EVT).  
   Hypothesis I-a: C-tDCS will be safe in AIS patients receiving EVT with no increased risk of symptomatic 
intracranial hemorrhage (SICH) within the 24-hour period after EVT. Hypothesis I-b: C-tDCS will not increase 
the rates of asymptomatic intracranial hemorrhage, early neurological deterioration (NIHSS during the 24-hour 
period after stimulation), 90-day mortality, and all serious adverse events.  Hypothesis I-c: Majority of AIS patients 
(≥ 90%) will tolerate the C-tDCS by completing at least 75% of the stimulation period with no major stimulation 
related pain, discomfort and skin changes.  
 
Specific Aim II: To determine the feasibility of delivering C-tDCS as an adjunct and bridge to EVT.  
   Hypothesis II-a: Adjunctive C-tDCS after endovascular thrombectomy will be started more rapidly after the end 
of ET procedure over the course of the study and in the last 10 patients, median time from end of ET procedure 
to C-tDCS initiation will be ≤ 10 minutes. Hypothesis II-b: Bridging C-tDCS prior to endovascular thrombectomy 
will be started more rapidly over the course of the study and in the last 4 patients, median time from randomization 
to C-tDCS initiation will be ≤ 10 minutes. 
 
Specific Aim III: To explore the potential imaging and clinical efficacy of tDCS in AIS an adjunct and 
bridge to EVT.  
1) Early and late penumbral salvage, defined as proportion of penumbral tissue on baseline multimodal MRI that 
does not proceed to infarct on 2-hour and 24-hour MRI (PWI); 2) Early and late collateral enhancement, 
evidenced by reduced perfusion lesion volume on 2-hour and 24-hour perfusion-weighted MRI (PWI); 3) Early 
and late reduction in infarct growth from baseline to 2-hour and 24-hour MRI; 4) Early improvement in neurologic 
deficit (NIHSS at 24h); 5) Improvement in final 3-month disability and quality of life (modified Rankin scale, AMC 
Linear Disability Scale, EQ-5D, and Barthel Index) 
Impact  
This in-hospital safety trial, will identify C-tDCS dose regimens that are feasible, well-tolerated, and preliminarily 
safe in patients with acute ischemic stroke who undergo ET. Study finding will identify the most promising dose 
regimens to advance to phase 2b and pivotal phase 3 efficacy trials for three patient populations with pressing 
need for improved therapies: 1) thrombectomy-eligible patients arriving directly to thrombectomy-capable 
hospitals who would benefit from post-arrival brain protection; 2) thrombectomy-eligible patients arriving initially 

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to non-thrombectomy-capable hospitals who would benefit from brain protection during interfacility transfer; and 
3) thrombectomy-eligible patients initially care for in Mobile Stroke Units who would benefit from brain protection 
during transport to, and early post-arrival course at, thrombectomy-capable hospitals.  
 
B. Background and Significance  
    Transcranial direct current stimulation (tDCS) is a non-invasive neuromodulation method that delivers a weak 
electrical current to the brain via scalp electrodes.11 Rather than directly eliciting a neuronal response, tDCS 
modulates neuronal excitability in regions of the brain depending on the polarity of stimulation. By altering the 
resting membrane potential cathodal tDCS (C-tDCS) reduces neuronal excitability [in contract to anodal tDCS 
(A-tDCS) which increases excitability]. 12,13 tDCS has been extensively investigated in humans for decades, as 
a neuromodulatory intervention to treat depression and diverse other neuropsychiatric disorders, as a 
neuroplasticity-enhancing intervention for chronic stroke patients, and as a tool to change cognition and behavior 
in healthy individuals. 14-17Thus far, tDCS has been found safe and tolerable with no reported serious adverse 
events across multiple clinical and preclinical studies. 18,19 In addition to these established applications of tDCS,  
tDCS studies in patients with acute ischemic stroke. C-tDCS is of substantial promise for acute cerebral ischemia 
due to large vessel occlusion, based on preclinical studies.   
   Stroke is a leading cause of mortality and morbidity across the world. Large vessel occlusion (LVO) is 
accounted for 30-40% of acute ischemic strokes and foretells a poorer prognosis compared to non-LVO 
strokes.20 While endovascular therapy (EVT) has fundamentally improved outcomes of AIS patients with LVO, 
patient outcomes even with EVT remain suboptimal; with only 20-25% achieving a disability-free outcome.3,5 
Therefore, even in the modern thrombectomy era, the need to develop complementary neuroprotective 
therapies exists. These therapies interrupt the cellular, biochemical and metabolic process that mediate hypoxic 
and reperfusion cellular injury.  
   There are two stages of care in endovascular thrombectomy patients in which effective neuroprotection 
therapies could synergistically further improve tissue salvage and functional outcome: 1) bridging 
neuroprotection prior to EVT start, and 2) adjunctive neuroprotection immediately after EVT completion. Bridging 
neuroprotective treatments applied prior to start of the endovascular thrombectomy procedure could preserve 
neural parenchyma and the neurovascular unit until reperfusion and resolution of ischemia is achieved by EVT. 
Bridging neuroprotective therapies would be beneficial in at least 3 settings: i) started soon after patient arrival 
at the thrombectomy-capable hospital during the 60-120 minute window from door to recanalization; ii) started 
in Mobile Stroke Units prior to arrival at thrombectomy-capable hospitals; and iii) started at outside, non-
thrombectomy hospitals prior to patient interfacility transfer to a thrombectomy center. If a neuroprotective 
treatment could completely “freeze” stroke progression until reperfusion was achieved, with every 15 minutes in 
which therapy was delivered, among 1000 treated patients, an estimated 39 patients would have less disabled 
outcome at 3 months, including 25 more who will be functionally independent (mRS 0-2).3 Similarly, adjunctive 
neuroprotective therapies, begun soon after EVT procedure completion, could be helpful in three additional ways 
:i) among patients with incomplete reperfusion (less than TICI 2c-3), adjunctive neuroprotective therapies could 
permit tissue tolerance of ongoing ischemia while spontaneous further recanalization proceeds; ii) among 
patients with substantial (TICI 2b) or excellent/complete (TICI 2c-3) reperfusion, adjunctive neuroprotective 
therapies could prevent delayed post-ischemia elaboration of injury through apoptotic and secondary injury 
cascades; and iii) adjunctive neuroprotective therapies could prevent reperfusion injury arising from reperfusion 
oxidative injury, inflammation, and other mechanisms.6  
   While many past attempts at developing neuroprotective therapies in human have been disappointing, they 
had the following shortcomings:  
1) The great preponderance tested pharmacologic agents that affected only one or two molecular ischemic 
pathways. 2) The majority of the neuroprotection therapies were administered intravenously, reducing delivery 
to brain areas with low blood flow, and, even there, requiring passage through the blood-brain barrier.7,21 3) Few 
trials of neuroprotection in human have used penumbral imaging for patient selection. 22 Therefore, the use of 
penumbral imaging to limit the patient selection to those for whom the intervention is most likely to be of benefit 
is crucial in studies of neuroprotection, and lastly 4) The neuroprotection therapies have been tested as stand-
alone treatments without definitive reperfusion. While neuroprotection is intrinsically a temporizing therapy, 
enabling brain cells to tolerate an ischemic episode longer, reperfusion is the ultimate therapy for AIS that finally 
resolves the ischemic episode. Hence, combining neuroprotection with reperfusion therapies in the post-
thrombectomy era is not only feasible but also increases the chance of proving clinical benefits.  

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Our study addresses all these challenges by: a) Testing C-tDCS as a multi-mechanistic neuroprotective method, 
b) Delivering the neuroprotection transcranially to ischemic tissue beyond occlusive thrombus, c) Using 
penumbral imaging prior to enrollment, and d) Using C-tDCS as a bridge and adjunct to thrombectomy.  
In pre-clinical studies, multiple investigators in several independent labs worldwide have found evidence of a 
neuroprotective effect of C-tDCS in different animal models of acute cerebral ischemia:  
 
B.1. Proposed neuroprotective mechanisms of cathodal direct current electrical stimulation based on 
experimental pre-clinical models of acute cerebral ischemia  
B.1.1. Inhibition of peri-infarct excitatory depolarizations 
   During the acute stages of cerebral ischemia, an excitotoxic cascade will be triggered by the excess glutamate 
and other excitotoxic amino acids that are released as the result of cellular necrosis.23 Experimental models of 
acute middle cerebral artery occlusion (MCAO) have shown that the excitotoxicity generates recurrent 
spontaneous waves of depolarization also known as peri-infarct depolarizations (PIDs).24,25 The PIDs occur soon 
after the MCA occlusion and spread across the penumbra to the normally perfused tissue. The infarct growth 
correlates with the number and duration of PIDs and the basis for this relationship has been related to: 1) An 
abnormal vasoconstriction in response to depolarization 2) An imbalance between increased metabolic overload, 
induced by the depolarization, and blood supply in acute ischemic stroke.26 Notturno et al. studied the effect of 
cathodal stimulation on PIDs in 3-vessel permanent occlusion rat model, with cumulative stimulation durations 
of 120 and 180 mins (15' on-15' off cycles).9 They found that cathodal stimulation applied to the ischemic MCA 
territory significantly reduced PIDs, and reduced infarct volume by 20-30%. They found no effect on brain edema 
between the stimulated and control groups and no stimulation induced macroscopic or microscopic lesion or 
hemorrhagic transformation. 
B.1.2. Anti-inflammatory, Anti-apoptotic, and Angiogenic effect 
  Beyond the activation of the excitotoxic cascade, an inflammatory response and programmed cellular apoptosis 
will result in a secondary damage and expansion of the irreversibly damaged core (Figure1). 23 Therefore, 
suppression of the innate pro-inflammatory cells and the cellular apoptotic cascade results in the reduction of 
the infarct size and cerebral edema. Peruzzotti et al. studied cathodal direct current stimulation in acute stroke  
mice models with temporary MCAO (reperfusion models) applied for 40 minutes (20'on-20off-20’on). 10 Cathodal 
stimulation of the ischemic hemisphere reduced final infarct size, with lowering of cortical glutamate synthesis, 
downregulation of N-methyl-D- aspartate (NMDA) receptor (NR2B) expression, and reduction in peri-ischemic 
inflammatory response and apoptotic markers.   
 
 
Figure 1. This schematic shows 
the events leading to ischemic 
brain injury. 

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Furthermore, a significant functional amelioration and improvement of cerebral edema were observed even when 
stimulation was applied hours after the MCAO. No macroscopic or microscopic lesion or hemorrhagic 
transformation induced by cathodal stimulation was found.  
   Baba et al. showed in their study that a low-frequency (2 and 10 Hz) electrical cortical stimulation may exert 
neuroprotective effects reflecting by attenuation in both necrotic and apoptotic cell deaths, blockade of 
microglial/astrocytic activation and expression of trophic factors. They also demonstrated an increase in cerebral 
blood flow of the electrically stimulated animals. 27 No neuroprotective effect was observed at higher frequency 
(50Hz). 
B.1.4. Preservation of neuronal axons 
   Kim et al. studied the neuroprotective effect of cathodal and anodal direct current stimulation on axons and 
myelin integrity. They delivered stimulation for 30 minutes two days after MCAO. They found significant 
amelioration of axonal damage and preservation of white matter axonal integrity in rat models of cerebral 
ischemia mainly after anodal stimulation. 28 
B.1.3. Direct vasodilatory effect 
   In addition to the direct neuroprotective effects, direct current may confer benefit in AIS via vasodilation and 
collateral blood flow enhancement. In fact, Fox et al. found a direct vasodilatory effect of electrical stimulation 
when applied over the basilar artery. 29This effect was more pronounced with cathodal compared to anodal 
stimulation. 
C. Preliminary studies  
C.1. Experience with transcranial neuromodulation in human subjects 
    
Our group at UCLA has extensive 
experience with transcranial neuromodulation 
using direct electrical and magnetically 
induced currents in human subjects. Over the 
past 10 years, we have performed transcranial 
neuromodulation in over 500 subjects.30-35 
This substantial experience provides a firm 
foundation for undertaking 
C.2. A meta-analysis of preclinical studies 
using 
electrical 
stimulation 
as 
neuroprotection in acute cerebral ischemia 
To assess the neuroprotective effect of 
electrical stimulation in AIS, we performed a 
systematic review of all preclinical acute 
cerebral ischemia studies using electrical 
stimulation as a neuroprotective method. Our 
systematic search identified 28 experiments in 
21 studies, including a total of 350 animals, of 
electrical stimulation in preclinical acute 
cerebral ischemia. Studies were categorized 
among 6 strategies. Three strategies applied 
different stimulation subtypes to tissues within 
the ischemic zone [cathodal hemispheric 
stimulation 
(CHS), 
anodal 
hemispheric 
stimulation (AHS), and pulsed hemispheric 
stimulation (PHS)] and three strategies 
applied deep brain stimulation to different 
neuronal targets remote from the ischemic 
zone [fastigial nucleus stimulation (FNS), 
subthalamic vasodilator area stimulation 
(SVAS), and dorsal periaqueductal gray 
stimulation (DPAGS)]. Random effects 
meta-analysis 
assessed 
electrical 
Figure 2. Forest plot shows the neuroprotective effect of electrical stimulation across 28 
pre-clinical experiments. Within the highlighted square are the studies using direct current 
stimulation as neuroprotection.  

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stimulation modification of final infarct volume.  Study-level risk-of-bias and intervention-level readiness-for-
translation were assessed using formal rating scales. Overall, in animals undergoing electrical stimulation, 
compared with control, final infarct volumes were reduced by 37% (CI 95%; 34-40% P< 0.00001). There was 
evidence of heterogeneity of efficacy among stimulation mode subgroups (I2 = 93.1%, pheterogeneity < 0.0001). 
Amongst the within-ischemic zone stimulation techniques, only CHS significantly reduced the infarct volume (27 
%, CI 95%; 22-33%, P< 0.00001); amongst the remote-from ischemic zone techniques, all (FNS, SVAS, and 
DPAGS) reduced infarct volumes by approximately half.  On formal rating scales, CHS studies had the lowest 
risk-of-bias and CHS had the highest overall quality of intervention-level evidence supporting readiness to 
proceed to clinical testing. We concluded that electrical stimulation reduces final infarct volume across preclinical 
studies. Cathodal hemispheric direct current stimulation shows the most robust evidence and is potentially 
appropriate for progression to early stage human clinical trial testing.   
C.3. Vascular response associated with Direct Current 
   
 A preliminary study in human volunteers by Marom Bikson (Co-mentor of the current proposal) 
demonstrated that direct current produces vasodilation of cerebral vessels (Figure 3). Furthermore, the 
vasodilatory response to tDCS is evident by its known mild dose-dependent effect in causing skin erythema. The 
cerebral vasodilatation is likely partially due to non-specific polarization of vascular system. Bikson and 
colleagues also demonstrated that 10 minutes of tDCS resulted in up-regulation of endothelial nitric oxide 
synthase (eNOS) gene expression and increase production of nitric oxide (NO), a known vasodilator. 36 Samdani 
et al. have demonstrated in their study that the endothelial NO upregulation favorably affects outcomes by the 
accentuation of the cerebral blood flow and attenuation of platelet aggregation, platelet adhesion, and NMDA 
current. 37 These findings support that tDCS could augment blood flow through the stimulated vasculature. 
Leptomeningeal collateral networks, peripherally located, are particularly accessible to the electric field. The 
potential vasoactive effects of tDCS also raise the possibility that stimulation will affect the blood-brain barrier. 
While no hemorrhagic transformation with C-tDCS was noted in preclinical models, this potential effect supports 
the approach of undertaking dose escalation safety trial as the first study in AIS patients, even though all studied 
dose tiers are within ranges safe in chronic stroke and other brain disease patients.11 19  
 
Figure 3. Shows models of intra-cranial current flow during tDCS is 
concentrated by the vasculature 
 
 
 
 
C.4. Feasibility of neuroprotection therapies in acute ischemic stroke 
    
Our group has extensive experience with the conduct of clinical trials of drug and device neuroprotective 
therapies for acute ischemic stroke, both as standalone interventions and as a complement to reperfusion 
treatment. We recently completed the NIH Field Administration of Stroke Therapy-Magnesium phase 3 trial, 
enrolling 1700 patients. 8 Of particular relevance to the current proposal, we have conducted trials of transcranial 
delivery of acute neuroprotective intervention using laser 38, and trials of collateral enhancement interventions 39 
and trials and studies using MRI penumbral imaging and permeability imaging as technical efficacy and safety 
end points.  
 
D. Trial Design and Methods 
D.1. Study overview 
This proposal is a prospective, single-center, dose-escalation safety, tolerability and feasibility study of 
C-tDCS as adjunctive and bridging neuroprotection in acute stroke patients undergoing endovascular 
thrombectomy. Patients with less than a complete EVT reperfusion (the thrombolysis in cerebral infarction (TICI) 
grading of <2c and 3) will be receiving at least one cycle of adjunct C-tDCS after EVT and at higher dose tiers, 
one cycle of bridging C-tDCS will be administered before EVT and if EVT results in TICI < 2c/3, then two cycles 
of adjunct EVT will be applied after EVT. The primary safety endpoints will be symptomatic intracranial 
hemorrhage during the 24-hour period after thrombectomy. Secondary measures of safety will include 
asymptomatic intracranial hemorrhage, early neurological deterioration, 3-month mortality and all serious 
adverse events. Tolerability will be judged based on the percentage of the patients completing the protocol-
assigned stimulation treatment and secondarily, the rate and severity of cutaneous, neurologic, nociceptive or 

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other adverse effects will be assessed. Feasibility endpoints will analyze the speed with which tDCS will be 
implemented. Finally, we will explore signals of potential efficacy by examining the imaging evidence of infarct 
growth, and clinical outcomes of early neurologic deficit evolution, and 3-month global disability and health-
related quality of life.  
D2. Technology 
D.2.1. tDCS Device and Stimulation Parameters  
The study will employ a Soterix™ high-definition DC-Stimulator, owned by the principal investigator. This tDCS 
unit consists of a stimulator, 4x1 HD interface and an adjustable cap with pre-maid openings that quickly and 
easily fits different head sizes. (Figure 5). The cap will be loaded with plastic electrode holders. These will be 
filled with conductance gel (Signa® gel), 1 cm2 electrodes will be placed in the holders, and the holder then 
locked. 
D.2.2. Montage 
The electrode positioning montage will be a 4 to 1-ring configuration, with the center or active electrode 
connected to cathode and the 4 reference or return electrodes connected to anode. The 4 to 1-ring configuration 
is chosen to focus the stimulation to the penumbral region. (Fig 6)  
For electrodes positioning location, we use one of the 3 predefined positionings according to the part of the 
middle cerebral artery (MCA) territory at risk. These electrode positionings are based on computational modeling 
of 3 electrical fields concentrated over different parts of the MCA territory: electrical field covering territory of 
MCA-M1 branch (Fig 5A, a); electrical field covering territory of MCA-M2 superior branch (Fig 5B,b); and 
electrical field covering territory of MCA-M2 inferior branch (Fig 5C,c).  
 
 
 
Figure 5.  Shows an example of electrode positioning in a 4 to 1-ring configuration in a patient with left M1 occlusion. A) The 
reference electrodes (anode-blue) are positioned on F3, T7, Cz and P3 and the center electrode (cathode-red) is positioned 
over the C3 (central sulcus). B) Shows the computational modeling of the electrical field, concentrated over the MCA territory. 
C) Shows the electrode positioning on the tDCS HD cap, the schematic of the tDCS stimulator connected to the HD interface, 
and the Soterix HD tDCS unit (stimulator + interface) that will be used in this study. This stimulator connects to the adjustable 
cap. D) Shows the penumbral region on the perfusion MRI of the patient with L MCA-M1 occlusion.   

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D.2.3. Sterility and Quality Assurance 
   Prior to stimulation, all the electrodes will be monitored for any sign of damage such as chipping. The 
electrodes will be discarded after being exposed to 5 cycles of stimulation. We will avoid any metal to contact 
the electrodes. The stimulation cap and the electrodes will be sanitized prior to each subject use.  
D.2.4. Device training  
  Device use will be performed by physician investigators, technologists, and research staff who have completed 
training and certification in safe tDCS use. Training will include instructions on different components of the device, 
installing and assembling the components, charging the device prior to the first use, recharging the device after 
each procedure, verifying the device is charged prior to each procedure, recognizing and addressing the different 
Warning/Error indications (e.g., battery light flashing blue, indicating that the battery is low but can still perform 
at least one procedure, battery light steady red, indicating battery low error – not enough battery power to run a 
complete procedure, etc.) and specific instructions for returning the device to the sponsor in the event of an error 
notification that cannot be addressed by the site personnel, or a failure of the device to charge after three hours. 
All device training will be documented in a training log that will be maintained in the site regulatory binder. 
D3. Subjects 
   Twenty four to 48 acute ischemic stroke patients who are undergoing endovascular therapy and meet study 
inclusion/exclusion criteria will be enrolled from Ronald Reagan Medical Center (RRMC) Emergency Department 
or RRMC inpatients. At last two dose tiers, patients arriving in Mobile Stroke Unit (MSU) with large vessel 
occlusion who are candidates for EVT will also be included for bridging stimulation en route to RRMC.  
    Based upon acute stroke patient referral rates to UCLA over the past 3 years, we anticipate 24 enrollments 
per year. Therefore, we estimate that the study enrollment will take 2 years to complete. Enrolled subjects will 
be randomized to active versus sham stimulation in 3:1 ratio.  
D.3.1 Entry Criteria 
Inclusion criteria  
1) New focal neurologic deficit consistent with AIS 
2) Age≥18 
3) NIHSS ≥ 4 
4) ICA or M1 or M2 MCA occlusion on pre-thrombectomy MRA or CTA  
5) Onset (last-seen-well) time to randomization time within 24 hours 
6) Pre-stroke modified Rankin Scale≤ 3 
8) Patient ineligible for IV tPA, per national AHA/ASA Guidelines   
9) Having undergone endovascular thrombectomy with less than a complete reperfusion (<TICI 2c, 3) for 
receiving post-thrombectomy adjunct C-tDCS 
10) Undergoing endovascular thrombectomy, per national AHA/ASA Guidelines for patients who are assigned 
to pre-thrombectomy bridging session at Tiers 5, 6 
11) A signed informed consent is obtained from the patient or patient’s legally authorized representative  
Exclusion criteria  
1) Acute intracranial hemorrhage 
2) Evidence of a large Ischemic core volume (ADC < 620 µm2/s or rCBF< 30%) ≥ 100 ml  
3) Presence of tDCS contraindications - electrically or magnetically activated intracranial metal and non-metal 
implants 
4) Pregnancy 
5) Severe contrast allergy or absolute contraindication to iodinated contrast preventing endovascular 
intervention 
6) History of seizure disorder or new seizures with presentation of current stroke 
7) Evidence of any other major life-threatening or serious medical condition that would prevent completion of the 
study protocol including attendance at the 3-month follow-up visit 
8) Concomitant experimental therapy 
9) Preexisting scalp lesion at the site of the stimulation or presence of skull defects (may alter current flow 
pattern) 
10) Preexisting coagulopathy, consist of platelet count of ≤ 100, INR ≥ 3, PTT ≥ 90  
11) Patients suspected of having infective endocarditis and ischemic stroke related to septic emboli 
12) Patients suspected or known to be infected with coronavirus 2019 (CoVID-19) 

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13) Patient with radiographic evidence or suspicion of chronic conditions that may predispose them to 
intracranial 
hemorrhage 
including 
brain 
arteriovenous 
malformations, 
cerebral 
cavernous 
malformations, cerebral telangiectasia, multiple previous intracerebral hemorrhages (amyloid 
angiopathy) 
14) Suspected cerebral vasculitis based on medical history and CTA/Magnetic Resonance Angiogram 
(MRA) 
15) Suspected cysticercosis 
16) Suspected cranial dural arteriovenous fistula 
17) Cerebral venous thrombosis 
18) Head trauma causing loss of consciousness, concussion, confusion, or a headache within the past 
30 days 
19) Patient has suffered a hemorrhagic or ischemic stroke within the last three (3) months 
20) History of a cancer known to cause hemorrhagic metastases, e.g., melanoma, renal cell carcinoma, 
choriocarcinoma, thyroid carcinoma, lung carcinoma, breast carcinoma, and hepatocellular carcinoma 
21) History of left atrial myxoma 
22) Evidence of dissection in the intracranial cerebral arteries 
23) Suspicion of aortic dissection  
24) Significant mass effect with midline shift 
25) The patient is in a coma 
 
 
Additional Inclusion Criterion for Penumbral Salvage Substudy 
1) Presence of salvageable penumbra with Tmax> 6 sec/ ischemic core volume (ADC < 620 µm2/s or rCBF< 
30%) ≥ 1.2  
2) Dose tier to which patient will be assigned includes a pre-thrombectomy bridging active or sham 
stimulation session 
D.3.2.Enrollment and Consent 
   All acute ischemic stroke patients within 24 hours of their symptoms onset who present to Roland Reagan 
Medical Center (Emergency Department or inpatient hospital if they are already admitted for a different 
indication) and meet the study inclusion/exclusion criteria will be identified by a study physician-investigator and 
subsequently offered enrollment in the trial. Prospective subjects will be provided with written and verbal 
information regarding the nature of the study, the procedures and evaluations involved, and the potential risks 
and benefits. All participating patients or their legally authorized representatives will provide explicit written 
informed consent.  Not all adult subjects will have the capacity to give informed consent. The likely range of 
impairment includes stupor and aphasia. By interviewing the patient, the investigators will assess whether the 
affected individual understands the central elements of the study procedures and has the capacity for informed 
consent, using the recommended approach of the institution’s Institutional Review Board, such as the UCLA 
Office of Human Research Protection Decision-Making Capacity Assessment Tool.  
    Patients with capacity to consent will be invited to participate by the PI. In patients without the capacity to 
consent, the patient’s legally authorized representative will be asked to provide consent for participation. The 
investigator will inform the patient or legally authorized representative of the availability of the study as follows: 
“You (your relative) is having a stroke. We are doing a research study of a new treatment for stroke. Here is an 
informed consent form that describes the study. Please read it. After you are finished, I will answer any questions 
you may have.” Once subjects or their legally authorized representatives have read and understood the IRB-
approved consent form, and had all their questions answered, written informed consent will be elicited. 
D.4. Dose Tiers and Randomization 
   We will implement a traditional 3+3 (rule-based, modified Fibonacci) dose escalation design, with 3:1 
randomization to active treatment vs sham control. There will be 6 dose tiers, reflecting increasing intensity and 
duration of stimulation:  

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Tier 1 – 1 mA, single 20 - min cycle, will start at the conclusion of the EVT procedure in patients with less than 
a complete EVT recanalization (the thrombolysis in cerebral infarction (TICI) grading of <2c and 3) and negative 
immediate post-EVT CT scan for a definitive evidence of intracranial hemorrhage (ICH).   
 
Tier 2– 2 mA, single 20 min cycle, will start at the conclusion of the EVT procedure in patients with TICI<2c and 
3 and negative immediate post-EVT CT scan for a definitive evidence of ICH.    
 
Tier 3 – 1 mA, in 2 treatment cycles. First cycle will be a 20 min cycle, administered after the conclusion of the 
EVT procedure in patients with TICI<2c and 3 and negative immediate post-EVT CT scan for a definitive 
evidence of ICH. The second cycle will start 20 minutes after the end of the first cycle, and will last 20 minutes. 
 
Tier 4– 2 mA, in 2 treatment cycles. First cycle will be a 20 min cycle, administered after the conclusion of the 
EVT procedure in patients with TICI<2c and 3 and negative immediate post-EVT CT scan for a definitive 
evidence of ICH. The second cycle will start 20 minutes after the end of the first cycle, and will last 20 minutes. 
 
Tier 5 - 1 mA, in 3 treatment cycles. First cycle will be a 20 min cycle, administered after initial imaging either in 
the Roland Reagan Medical Center or mobile stroke unit and prior to arterial puncture (cycle will be shorter than 
20 minutes if EVT puncture is ready to proceed, in order not to delay EVT). The second cycle will start at the 
conclusion of the EVT procedure in patients with TICI<2c and 3 and negative immediate post-EVT CT scan for 
a definitive evidence of ICH (and not sooner than 20 minutes after the end of the first cycle), and will last 20 
minutes. The third cycle will start 20 minutes after the end of the second cycle, and will last 20 minutes. 
 
Tier 6 - 2 mA, in 3 treatment cycles. First cycle will be a 20 min cycle, administered after initial imaging either in 
the Roland Reagan Medical Center or mobile stroke unit and prior to arterial puncture (cycle will be shorter than 
20 minutes if EVT puncture is ready to proceed, in order not to delay EVT). The second cycle will start at the 
conclusion of the EVT procedure in patients with TICI<2c and 3 and negative immediate post-EVT CT scan for 
a definitive evidence of ICH (and not sooner than 20 minutes after the end of the first cycle), and will last 20 
minutes. The third cycle will start 20 minutes after the end of the second cycle, and will last 20 minutes. 
 
   Patients in the sham stimulation arm at all the tiers will have the cap and electrodes in place, but without any 
delivered electrical stimulation. While the highest dose tier in this study is expected to be fully safe, based on 
preclinical and clinical studies 11,18, since this is a first-in-human study in acute stroke, a formal escalation to 
higher dose tiers is prudent.  

12 
 
 
 
 
 
 
D.4.Management pre, during, and post adjunctive stimulation period  
As soon as the endovascular thrombectomy (EVT) ends, a CT head will be obtained in patients with less than a 
complete EVT reperfusion (TICI< 2c,3) and if negative for definitive evidence of intracranial hemorrhage, then 
the patient will be randomized, and the adjunct stimulation will be initiated in neurosurgical intensive care unit 
according to the tier that patient is assigned to.  
After randomization and during the stimulation, all patients will be monitored closely by a physician-investigator. 
NIHSS will be obtained at the end of each 20-minute stimulation cycle and a visual inspection of the skin and 
rate degree of any potential erythema under the electrode will be performed. Then, a tolerability form will be 
completed based on validated cutaneous, neurological, and pain items of the PRO-CTAE (Patient-Reported 
Outcomes version of the Common Terminology Criteria for Adverse Events) (Figure 8).40,41  
After the stimulation, nursing neurologic assessments will be performed by Neurointensive Care Unit with 
extensive experience in monitoring acute stroke patients. Specifically, patients will be monitored for any signs of 
neurological worsening and will be queried on development of new or worsening neurological symptoms, 
including headache, dizziness, numbness, or weakness, etc. If any sign or symptom of neurological worsening 
is suspected, the patient will be immediately assessed by the on-call neurologist and an immediate brain imaging 
(CT or MRI) will be obtained. Neuro-checks will be performed at 1h and 2h after stimulation and then every 2 
hours.  Subsequent care will be continued in these settings, including medical management per national 
guidelines for acute ischemic stroke management issued by the American Stroke Association.42  Patients will 
not receive NSAIDs as concomitant therapy for the first 7 days after enrollment. All patients will undergo a 
multimodal MRI or CT including standard parenchymal images, non-invasive angiography, and perfusion studies 
and formal NIHSS assessment at 2-hour and 24-hour following the end of the stimulation tier as part of the study 
protocol. 
D.5. Management pre, during, and post bridging stimulation 
At tiers 5 and 6, hospital arriving patients who are candidates for EVT with an LVO detected on CT angiography 
and are being transferred to agio suite for EVT procedure will receive up-to-20 minutes of stimulation. See section 
D.4. for care during and post-stimulation.  
Figure 7. This figure shows the 6 escalating tiers. Blue represents 1 mA, purple shows 2 mA of C-tDCS. The black/white 
neuroangio suite icon represents the endovascular thrombectomy (EVT) procedure, occurring before the adjunctive C-tDCS 
at all tiers if EVT results in a less than a complete recanalization (TICI <2c, 3). At tiers 5 and 6, bridging C-tDCS occurs for up 
to 20’ before EVT. Symptomatic ICH (SICH) in none of the three C-tDCS patients in a tier will result in escalation. However, if 
SICH occurs in one C-tDCS patient, 3 more C-tDCS patients and 1 more sham will be enrolled at the same tier. 

13 
 
 
 
 
D.6. Study Visits and Data Acquisition  
D.6.1 Schedule of Events  
* Skin Inspection will be performed before each stimulation period  
D.6.2. Baseline Evaluation 
• 
Demographics (age, sex, race) 
• 
Last known well time 
• 
Past medical/surgical history including vascular diagnoses and risk factors (stroke, TIA, carotid stenosis, 
myocardial infarction, atrial fibrillation, peripheral arterial disease, hypertension, diabetes, dyslipidemia) 
• 
Medications, including antithrombotics, antihypertensives, statins, anti-arrhythmics      Family history of 
vascular disease; 
• 
Tobacco (including timing, duration, amount), alcohol, and illicit drug use;  
• 
Vital signs (systolic blood pressure, diastolic blood pressure, pulse, respiratory rate, oxygen saturation); 
• 
Premorbid global disability (modified Rankin Scale);   
• 
Neurological deficits severity (NIHSS);  
• 
Laboratory results (CBC, platelet count, glucose, lytes, INR, PTT, LFTs.  
D.6.3. C-tDCS Treatment  
• 
Skin inspection before each cycle of stimulation  
• 
Tolerability/AE Form – after each cycle of stimulation 
• 
NIH Stroke Scale (NIHSS) after each cycle of stimulation 
Baseline End of each 20-min stimulation cycle* 
Before/ 
After ET 
2-hr 24-hr Day 4 Day 30 Day 90 
1( Bridging)  2 (adjunctive) 3 (adjunctive) 
Consent 
 
History / Exam 
 
Routine Labs 
 
Vital Signs 
 
 
 
 
 
Tolerability form/Skin Inspection*  
 
 
 
Multimodal MRI/CT 
 
 
 
NIHSS 
 
 
 
 
 
 
 
 
Modified Rankin Scale (mRS) 
 
 
 
 
Barthel Index (BI) 
 
EuroQol (EQ-5D) 
 
AMC Linear Disability Scale 
 
Interval serious adverse events 
 
 
 
 
 
 
 
 
 
Concomitant/Interval 
medications/procedures  
 
 
 
 
 
 
 
 
 
 
Figure 
8. 
Shows 
the 
technician-filled 
tolerability 
form. The selected items are 
based on the most common 
reported 
adverse 
effects 
associated with tDCS. 
 

14 
 
D.6.4. Endovascular thrombectomy (EVT)  
• 
Records of all the time metrics related to EVT: time of decision to take to catheter angiography suite for 
EVT, time of arrival at the catheter angiography suite, time of Groin Puncture, the time that C-tDCS was 
aborted prior to EVT, door to puncture time, on clot time, time first device deployed, time of first device 
passed (the times for each pass and deployment will be recorded), Total duration of EVT.  
• 
Type of device uses ( stent-retriever vs aspiration catheter)  
• 
Total number of device passes  
• 
NIHSS before and after EVT  
• 
Interval serious adverse events during EVT  
• 
Interval medications/procedures during EVT  
D.6.4. Early (2 Hours) After EVT 
• 
NIHSS,  
• 
Multimodal MRI/CT 
• 
Interval serious adverse events  
• 
Interval medications/procedures  
D.6.5. Late (24 Hours) After EVT 
• 
NIHSS 
• 
Multimodal MRI/CT 
• 
Interval serious adverse events 
• 
Interval medications/procedures  
D.6.6. Day 4  
• 
Modified Rankin Scale (mRS) 
• 
Interval serious adverse events 
• 
Interval medications/procedures  
D.6.7. Day 30  
• 
mRS (phone),  
• 
Interval serious adverse events 
• 
Interval medications/procedures  
D.6.8. Day 90  
• 
NIHSS,  
• 
Modified Rankin Scale (mRS) 
• 
AMC Linear Disability Scale (granular disability)  
• 
Barthel Index (BI) (activities of daily living) 
• 
EuroQol (EQ-5D) (health-related quality of life) 
• 
Interval serious adverse events 
• 
Interval medications/procedures  
D.6.9. Imaging Assessments  
• 
Baseline 
Emergent multimodal MR and CT is currently acquired as the routine initial imaging study in all acute stroke 
patients at UCLA. In patients receiving MRI, a 1.5 T or 3 T scanner equipped with echo-planar imaging capability 
is used for rapid acquisition of diffusion and perfusion scans. The standard clinical MRI protocol includes Gradient 
Recall- Echo (GRE), DWI, FLAIR (Fluid-Attenuated Recovery Image), PWI and MR angiography. The ADC 
(apparent -diffusion coefficient) values derived from DWI acquisition (b=0, 1000 s/mm2 applied in each of three 
principal gradient directions) will be used to delineate the volume of ischemic core. The tissue with ADC values 
of ≤ 620 µm2/s will be considered ischemic core, indicative of tissue with advanced, irreversible bioenergetic 
compromise. FLAIR delineates early parenchymal signal abnormality associated with ischemia and slow 
retrograde flow in leptomeningeal collaterals appearing as FLAIR vascular hyperintensity (FVH). The GRE 
sequence is used to evaluate the presence of intracranial hemorrhage and deoxygenated leptomeningeal 
collaterals appearing as GRE vascular hypointensity (GVH). PWI is acquired with sequential T2*-weighted 
(gradient echo) EPI time sequence scanning. Early in the time series, a bolus (0.1 mmol/kg) of MRI contrast 
material is rapidly infused (5 ml/sec through an 18 or larger gauge angiocatheter) using a power injector. The 
perfusion lesion volume (tissue at risk volume) will be the region with Tmax  6sec (Tmax = the time delay from 

15 
 
the arrival of contrast to its peak concentration at the tissue vasculature). 43,44 Intracranial CEMRA (Contrast-
Enhanced MRA) will identify large vessel occlusion.  
In patients receiving CT, the standard clinical CT protocol includes non-contrast CT (NCCT), multiphase CT 
angiography, CT perfusion (CTP) processed through RAPID software. NCCT is used to rule out acute intracranial 
hemorrhage appearing as hyperdensity. The tissue with rCBF < 30% on CTP will be considered ischemic core 
and the perfusion lesion volume (tissue at risk volume) will be the region with Tmax  6sec. CT angiography 
(CTA) will identify large vessel occlusion.  
• 
Endovascular Thrombectomy 
Patients will be undergoing cerebral angiography as part of the endovascular thrombectomy for treatment of 
acute ischemic stroke due to large vessel occlusion. The following data from pre-reperfusion cerebral 
angiography images will be collected:  Target vessel occlusion for mechanical thrombectomy procedure, 
collateral grade prior to EVT (measured with the ASITN collateral grading scale). The following data will be 
collected from the post-reperfusion cerebral angiography images: arterial occlusion lesion: 1) Reperfusion, as 
measured with the expanded thrombolysis In cerebral infarction scale (eTICI) after final device pass (TICI 0, 
TICI 1, TICI 2a, TICI 2b,TICI 2c, TICI3); and 2) Recanalization, as measured with the arterial occlusive lesion 
(AOL) scale (0-No reduction of target thrombus, 1 – Reduced thrombus size but no recanalization, 2 – Partial 
recanalization, 3 – Complete recanalization).  
TICI scale will be defined as the following: 0 – No perfusion, 1 – Antegrade reperfusion past the initial 
occlusion, but limited distal branch filling with little or slow distal reperfusion, 2a – Antegrade reperfusion of less 
than half of the occluded target artery previously ischemic territory (eg, in 1 major division of the MCA and its 
territory), 2b – Antegrade reperfusion of 51-90% of the previously occluded target artery ischemic territory, 2c – 
Anterograde perfusion of >90% but less than TICI 3 (i.e. near complete reperfusion), 3 – Complete antegrade 
reperfusion of the previously occluded target artery ischemic territory, with absence of visualized occlusion in 
all distal branches.45  
• 
2h and 24h Post- Endovascular Thrombectomy 
Follow-up MR/CT multimodal imaging will be obtained routinely at UCLA at 2h and 24h post-thrombectomy 
(supported 
by 
Radiology 
Dept. 
research 
funds). 
MRI 
Imaging 
sequences 
will 
include 
DWI/FLAIR/GRE/PWI/CEMRA and CT imaging sequences will consist of NCCT/CTA/CTP. RAPID image 
processing software will be applied to the above images to quantify, at the baseline, 2h, and 24h time-points: 1) 
Early penumbra preservation: Volume of baseline penumbra tissue not progressing to ischemic core at 2h; 2) 
Penumbral salvage: Volume of baseline penumbra tissue not progressing to ischemic core at 24h; 3) Early 
collateral flow enhancement: Perfusion lesion volume at baseline – Perfusion lesion volume at 2h; 4) Sustained 
collateral flow enhancement: Perfusion lesion volume at baseline – Perfusion lesion volume at 24h; 5) Early 
infarct growth: Ischemic core lesion volume at 2h – Ischemic core lesion volume at baseline; and 6) Final infarct 
growth: Ischemic core lesion volume at 24h – Ischemic core lesion volume at baseline. Additional location and 
extent of ischemic injury at baseline, 2h, and 24h will be rated using the ASPECTS scoring system.  
D.6.10. Adverse Events 
D.6.10.1 Serious Adverse Events (SAEs) 
   All serious adverse events occurring during the 90 days of study participation will be recorded. A serious 
adverse event is any adverse event that is fatal, is life-threatening, is permanently or substantially disabling, 
requires or prolongs hospitalization, or requires medical or surgical intervention to prevent one of the above 
outcomes.18  
D.6.10.2 Symptomatic intracranial hemorrhage 
   The lead safety endpoint adverse event is symptomatic intracranial hemorrhage (SICH), defined using the 
SWIFT PRIME trial criteria 46: an increase of 4 or more points on the NIHSS within 24 hours of stimulation 
associated with parenchymal hematoma type 1 (PH1), parenchymal hematoma type 2 (PH2), remote 
intraparenchymal hemorrhage (RIH), subarachnoid hemorrhage (SAH), or intraventricular hemorrhage (IVH). In 
addition, all hemorrhages, both symptomatic and asymptomatic, will also be separately classified and analyzed 
by radiologic subtype, as hemorrhagic infarction type 1 (HI1), hemorrhagic infarction type 2 (HI2), PH1, PH2, 
RIH, SAH, or IVH.46 A central neuroimaging core lab, blinded to treatment assignment, will review all brain scans 
obtained at 24h and rate presence and type of radiologic hemorrhagic transformation. In addition, in patients 
who experienced worsening by 4 or more NIHSS points in the first 24 hours, they will review any and all additional 
brain MRI or CT scans obtained during the 24h time period,   

16 
 
   SICH is the primary safety endpoint of the current trial, but trial results will be assessed on a variety of additional 
safety, feasibility, and tolerability results as well. It is anticipated that, after completion of the current trial, the 
judgement of whether to proceed directly to a pivotal trial, to proceed to a larger safety and preliminary efficacy 
trial, or to not proceed with further development, will rest on a considered and informed assessment of all 
outcome measures. It is important, therefore, to collect data on a wide range of safety endpoints and a wide 
range of measures of functional outcome, as is planned in this proposal. In making the selection of a primary 
safety endpoint, an emphasis was placed on ensuring patient safety throughout the course of the trial by 
choosing an endpoint with uncontestable clinical relevance (Symptomatic Intracranial Hemorrhage).  
D.6.10.3 Additional Adverse Events with Specific Interrogation 
    In addition to general screening for all serious adverse events and focused elicitation of symptomatic 
intracranial hemorrhage events, the following events will be specifically interrogated for and recorded in the case 
report forms: skin redness, scalp rash, hair loss, seizure, headache, sensitivity to light, new ischemic stroke, 
deep venous thrombosis, pulmonary embolism, pneumonia, acute MI.  
E. Safety Monitoring 
E.1. Data and Safety Monitoring Plan 
   The trial will be monitored by an Independent Data and Safety Monitoring Board (DSMB). Dr. Bruce Dobkins 
from department of neurorehabilitation will lead the DSMB. DSMB will assess for the causal relationship of the 
serious adverse event to the study treatment as definite, probable, possible, unlikely, or unrelated.  
DSMB will meet at the completion of each dose tier, review all safety data, and determine whether the study will 
proceed to the next dose tier. DSMB deliberations will be guided by: 1) a formal stopping/escalation rule, based 
on the occurrence of the lead safety endpoint, symptomatic intracranial hemorrhage (SICH), and 2) The DSMB 
members’ clinical judgement upon review of all other safety outcomes. In addition, the Food and Drug 
Administration (FDA) will be notified if there is one occurrence of SICH prior to repeating the tier or escalating to 
a higher tier. The DSMB, along with the investigators, will also continuously monitor door-to-puncture and 
imaging-to-puncture times in enrolled patients, comparing them with national guidelines from American 
Heart/American Stroke Association (AHA/ASA), the Society of Neurointerventional Surgery (SNIS), the Society 
of Vascular and Interventional Neurology (SVIN) and academic medical center comparator hospitals to ensure 
that start of thrombectomy is not being delayed by study procedures. 
E. 2. Statistical Design and Analyses Plan 
E.2.1. Sample size 
   The study sample size derives from the use of the 3+3, rule-based, modified Fibonacci, dose escalation design, 
with 3:1 randomization to active treatment vs sham control. There will be 6 dose tiers, reflecting increasing 
intensity and duration of stimulation (Figure 7). The 3+3 study design (3 patients and 1 sham) with 6 dose tiers 
yields a sample size of at least 24 and potentially up to 48. The 3+3 design is the classical approach to dose 
escalation in first-in-human studies. 47,48 While newer, adaptive designs for dose escalation trials have been 
developed, they are more complex and have limited advantages when major toxicities are not expected. 49 
Therefore, the rule-based, 3+3 design remains the dominantly employed approach in current dose-escalation 
studies.  We will take every step to avoid missing data by scheduling of the follow-up visits early in visit time 
windows and readiness to travel to the patient’s location to perform needed assessments. Should any missing 
data occur, primary analyses will be performed using multiple imputation, with sensitivity analyses including 
complete case and worst possible outcome analyses. 50  
E.2.2. Baseline characteristics 
   The demographic and baseline clinical characteristics of the study population will be delineated with standard 
descriptive statistics. Categorical variables describing the clinical history, examination findings, and initial 
treatment will be summarized by frequencies. Continuous variables such as vital signs, laboratory results, and 
time variables will be characterized by means, standard deviations, and 95% confidence intervals (CI). Ordinal 
and non-normally distributed variables (such as the NIHSS) will be characterized by medians and interquartile 
ranges. Baseline characteristics will be compared between the tDCS stimulation group with sham group to 
assess covariate balance. Wilcoxon Rank-Sum tests will be used for continuous or ordinal variables; Fisher’s 
exact tests and chi-square tests will be used for grouped or nominal categorical variables.   
E.2.3. Safety and Tolerability (Specific Aim 1) 
E.2.3.1 Dose Escalation 
   The study Data and Safety Monitoring Board (DSMB) will meet at the completion of each dose tier, review all 
safety data, and determine whether the study will proceed to the next dose tier. DSMB deliberations will be 

17 
 
guided by: 1) a formal stopping/escalation rule, based on the occurrence of the lead safety endpoint, symptomatic 
intracranial hemorrhage (SICH), and 2) The DSMB members’ clinical judgement upon review of all other safety 
outcomes. The formal dose escalation rule uses SICH frequency to gate the occurrence and pace of escalation 
through the 6 dose tiers. If no SICH occurs in the 3 active patients at a dose tier, enrollment may escalate to the 
next dose tier. If one SICH occurs, 3 more active (and 1 more control) patient will be enrolled at that dose tier 
before escalation. If 2 SICHs occur at a dose tier, further study enrollment is held until detailed review by the 
DSMB (Fig 6). In addition, whenever the formal SICH criteria for dose escalation has been met, the DSMB will 
formally meet, review the SICH data and all other safety data, and advice regarding proceeding to the next tier, 
continuing at the current tier, or placing the study on hold. 
E.2.3.2. Primary Safety Endpoint Analysis 
   For the final statistical analysis of the primary SICH safety endpoint, a chi-square test will be used to detect 
differences in the rate of SICH between the active treatment and sham patients and higher and lower dose tiers. 
The treatment will be considered to have exhibited adequate safety in the current trial to proceed to future, larger, 
pivotal efficacy trials if tDCS results in lower or equivalent rates of SICH compared to sham.  
E.2.3.3. Secondary Safety Endpoints Analysis 
   In secondary safety endpoint analyses, the following will be compared between the active treatment and sham 
patients, and between higher and lower dose tiers, using chi-squared tests: 1) Asymptomatic ICH by 24h 
(intracranial hemorrhage not associated with NIHSS worsening ≥ 4); 2) Early neurologic deterioration (worsening 
≥ 4 on NIHSS during the 24-hour period after stimulation, with or without intracranial hemorrhage); 3) All-cause 
mortality at day 90 (mRS); and 4) All serious adverse events.  
E.3.3. Tolerability Endpoint Analysis 
    The lead tolerability endpoint is completion of the protocol-assigned stimulation treatment without early 
cessation due to cutaneous, neurologic, nociceptive or other adverse effects. Experience with tDCS in post-
stroke patients indicates only infrequent cutaneous (itching, tingling) adverse effects are likely to occur. 
Accordingly, for the current study, a patient will be considered to have tolerated the procedure if at least 75% of 
the stimulus period was completed. The treatment will be considered generally tolerable if, among all enrolled 
patients, tolerated procedures are achieved in ≥90% of patients, assessed with a one-sided p value of 0.025. 
Secondary tolerability endpoints will be the rate and severity of cutaneous, neurological, and pain items of the 
technician-filled (Figure 7), descriptively compared between active treatment and sham patients, and between 
higher and lower dose tiers.   
E.3.4. Feasibility Endpoint Analysis (Specific Aim 2) 
   A time-motion analysis and mock run-throughs will be conducted prior to first enrollment. Nonetheless, it is 
anticipated that processes to optimize rapid placement of the cap and electrodes will continue to improve with 
experience gained from initially enrolled patients. The predefined success threshold for feasibility will be median 
times from randomization to bridging C-tDCS initiation and the time form end of EVT procedure to adjunctive C-
tDCS initiation ≤ 10 minutes in the last 4 and 10 enrolled patients, respectively.  
E.3.5. Exploratory Efficacy Endpoints Analysis (Specific Aim 3) 
E.3.5.1. Exploratory Clinical Efficacy Endpoints 
   This study is underpowered to definitively determine efficacy, so all clinical efficacy analyses will be purely 
exploratory and descriptive. Four clinical outcome measures were selected based on their reliability, familiarity 
to the neurologic community, and adaptability for use in patients who have had a stroke. These endpoints are: 
the modified Rankin Scale (mRS), a rating of global disability; the Barthel Index (BI), a measure of instrumental 
activities of daily living; the National Institutes of Health Stroke Scale (NIHSS), a measure of neurologic deficit 
severity; and the EuroQol (EQ-5D), an assessment of health-related quality of life; and AMC Linear Disability 
Scale, a granular degree of disability.  
Clinical efficacy endpoints will be characterized in the active and sham patients, and in higher and lower dose 
tiers. Early course clinical efficacy endpoints of greatest interest that will be explored are: 1) Normalized change 
in neurologic deficit from baseline to 24h (normalized delta NIHSS – linear variable, analyzed with means and 
95% CIs; and 2) Degree of neurologic deficit at 24h (NIHSS – quasi-linear variable, analyzed with means and 
95% CIs). Final outcome clinical efficacy endpoints of greatest interest that will be explored are: 1) Degree of 
disability at 90 days, assessed across all 7 levels of the modified Rankin Scale – ordinal variable, analyzed with 
medians (IQRs) and means (95% CIs); 2) Functional independence (mRS 0-2) at 90 days – binary variable, 
analyzed with rates and 95% CIs; 3) Granular degree of disability at 90 days (AMC Linear Disability Scale) – 

18 
 
linear variable, analyzed with means and 95% CIs; and 4) Health-related quality of life (EQ-5D) – linear variable, 
analyzed with means and 95% Cis.  
E.3.5.2. Exploratory Imaging Biomarker Efficacy Endpoints 
   This study is underpowered to definitively determine efficacy, so all imaging efficacy analyses will be purely 
exploratory and descriptive. The imaging efficacy analysis will be done on the subgroup of patients (including 
tier 5 and 6 patients who have presence of salvageable penumbra in their entry criteria) with baseline pre-EVT 
penumbral imaging demonstrating presence of salvageable penumbra with Tmax> 6 sec/ ischemic core volume 
of ≥ 1.2.  
Imaging biomarker efficacy endpoints will be characterized in the active and sham patients, and in higher and 
lower dose tiers, using means and 95% confidence intervals. The six imaging efficacy endpoints of greatest 
interest that will be explored are: early and late penumbral salvage, early and late collateral flow enhancement, 
and early and late infarct growth.  
 
 
 
 

19 
 
 
 
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