Safety and Feasibility of Nasal Delivery of Human Dental Follicle Mesenchymal Stem Cell-Derived Exosomes for Negative Symptoms in Treatment-Resistant Schizophrenia: A Pilot Study
Phase: EARLY_PHASE1
Conditions: Schizophrenia Negative Type
Sponsor: Zigong Mental Health Center

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ORIGINAL STUDY PROTOCOL
Safety and feasibility of intranasal delivery of human
dental follicle mesenchymal stem cel-derived exosomes
for negative symptoms in treatment-resistant
schizophrenia: A pilot study
Name of the ethic committee: Zigong Mental Health Center Ethics
Committee
Approved No. of ethics committee: 20260101
Principal Investigators: Kezhi Liu (liukezhi@swmu.edu.cn)
Version No.: V1.0
Version Date: Jan 26, 2026

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Contents
Study Summary...........................................................................................3
1.Introduction............................................................................................ 10
2. Study Objectives................................................................................... 14
3. Study Subjects.......................................................................................15
3.1 Inclusion Criteria...........................................................................15
3.2 Exclusion Criteria..........................................................................15
3.3 Withdrawal Criteria.......................................................................16
3.4 Termination Criteria......................................................................16
4. Study Design and Procedures............................................................... 17
4.1 Study Design................................................................................. 17
4.2 Study Procedures...........................................................................17
5. Research Methods................................................................................. 18
5.1 Exosome Preparation and Characterization..................................18
5.2 Intervention Protocol.....................................................................19
5.3 Assessment Content and Schedule............................................... 20
6. Outcome Measures................................................................................21
6.1 Primary Outcome Measures..........................................................21
6.2 Secondary Outcome Measures......................................................22
6.3 Exploratory Outcome Measures................................................... 22
7. Statistic analysis .................................................................................23
8 .References ........................................................................................... 24

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Study Summary
Title
Safety and feasibility of intranasal delivery of human dental
follicle mesenchymal stem cel-derived exosomes for
negative symptoms in treatment-resistant schizophrenia: A
pilot study
Objective
Primary Objective
To assess the safety and feasibility of hDFSCs-Exo in the
treatment of treatment-resistant schizophrenia.
Secondary Objective
To evaluate the preliminary efficacy of hDFSCs-Exo on
negative symptoms of treatment-resistant schizophrenia.
Participant
Treatment-resistant schizophrenia
Study DesignSingle-center, open-label, single-arm, 3+3 dose-escalation
design
hDFSCs-Exo
Drug Name: Human Dental Follicle Mesenchymal Stem
Cell-derived Exosomes (hDFSCs-Exo)
Route of Administration: Intranasal spray
Specification: 30 billion particles / 2 ml (particle
concentration)
Storage Conditions: Store at -80°C
Manufacturer: Chengdu Shilian Kangjian Biotechnology Co.,

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Ltd.
Dosage
regimen
This study plans to include five dose groups:
1×10⁹ particles/dose group
2×10⁹ particles/dose group
4×10⁹ particles/dose group
8×10⁹ particles/dose group
1.6×10¹⁰ particles/dose group
Nasal spray device: Disposable intranasal spray injector
(Tianzhou Medical, model TZ-AN30)
The exosomes will be prepared in 0.2 ml of normal saline and
administered via intranasal spray twice weekly (with a 3-day
interval between doses) for 8 weeks, totaling 16
administrations.
Note: Prior to each treatment, nasal hygiene should be
performed by irrigating both nasal cavities with a normal
saline nasal spray. After waiting for 5 minutes, gently clean
any secretions using a cotton swab.
Inclusion
criteria
1. Diagnosed with schizophrenia according to ICD-10
criteria;
2. Aged between 18 and 60 years;
3. Long-term inpatients with a disease duration of more than
5 years;
4. No acute exacerbation in the past 6 months, and no
change in medication regimen in the past 2 months;
5. Poor response to adequate dose and duration of treatment
with at least two different antipsychotics, including
clozapine;
6. Positive and Negative Syndrome Scale – Factor Score for
Negative Symptoms (PANSS-FSNS) ≥24;
7. At least two of the three core negative symptom items on

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the PANSS (N1, N4, and N6) scored ≥4;
8. Clinical Global Impression – Severity of Illness (CGI-S)
score ≥4;
9. Signed written informed consent.
Exclusion
criteria
1.History of severe allergic reactions;
2.Definite organic brain lesions;
3.Comorbid severe physical diseases (e.g., unstable coronary
heart disease, malignant arrhythmia, hepatic or renal
insufficiency, bronchial asthma, acute exacerbation of
chronic obstructive pulmonary disease, autoimmune
diseases, etc.);
4.Current diagnosis of other mental disorders according to
ICD-10 criteria (e.g., schizoaffective disorder,
schizophreniform disorder, bipolar I disorder, bipolar II
disorder, pervasive developmental disorder, mental
retardation, delirium, dementia, amnestic disorder, or
other cognitive disorders);
5.Unstable condition requiring adjustment of medication
regimen;
6.Non-compliance with treatment;
7.Severe rhinitis or nasal allergies;
8.History of modified electroconvulsive therapy (MECT)
within the past 3 months;
9.Individuals at risk of suicide;
10.Pregnant or lactating women;
11.Other conditions deemed unsuitable for enrollment.
Sample size
15-30

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Measurement
1. Primary Outcome Measures
To assess the safety and feasibility of intranasal delivery of
human dental follicle mesenchymal stem cell-derived
exosomes.
1) Safety Measures:
(1) Adverse Events: Incidence and severity of adverse events
(AEs) and serious adverse events (SAEs), graded according
to CTCAE version 5.0.
(2) Dose-Limiting Toxicity: Incidence of dose-limiting
toxicities (DLTs), used to determine the maximum tolerated
dose (MTD) and recommended Phase II dose (RP2D).
(3) Vital Signs: Incidence of abnormalities in body
temperature, pulse, respiration, and blood pressure.
(4) Physical Examination: Incidence of abnormalities in
general condition, head, skin, mucous membranes, lymph
nodes, neck, chest, abdomen, spine/extremities, and
neurological examination.
(5) Laboratory Tests: Incidence of abnormalities in complete
blood count, urinalysis, blood biochemistry (liver and kidney
function), infectious disease markers, inflammatory factors,
and allergy markers (IgE).
(6) Electrocardiogram (ECG): Incidence of ECG
abnormalities.
2) Feasibility Measures:
(1) Treatment Completion Rate: Proportion of subjects
completing the full 8-week (16 sessions) intranasal

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administration regimen.
(2) Treatment Adherence: Evaluation of subjects' acceptance
of and compliance with the intranasal administration method.
2. Secondary Outcome Measures
Changes from baseline to the end of Week 8 in the following
measures:
1) Change in Positive and Negative Syndrome Scale
(PANSS) total score and factor scores.
2) Change in Clinical Global Impression – Severity of Illness
(CGI-S) score.
3) Change in Scale for the Assessment of Negative
Symptoms (SANS) total score and subscale scores.
4) Change in Calgary Depression Scale for Schizophrenia
(CDSS) score.
5) Change in Montreal Cognitive Assessment (MoCA) total
score.
6) Change in Trail Making Test Part A/B (TMT-A/B)
completion time.
7) Change in Digit Span Test (DST) score.
8) Change in Verbal Learning and Memory Test (VLMT)
score.
3. Exploratory Outcome Measures
To explore changes in brain function from baseline to the
end of Week 8, assessed by:
1) Electroencephalography (EEG):

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Changes in resting-state power spectrum in the prefrontal
theta band (4–8 Hz).
Changes in functional connectivity strength of the default
mode network (DMN) based on source localization.
2) Functional Near-Infrared Spectroscopy (fNIRS):
Changes in oxygenated hemoglobin (Oxy-Hb) concentration
in the bilateral dorsolateral prefrontal cortex (DLPFC) during
a verbal fluency task.
3) Magnetic Resonance Imaging (MRI):
Changes in resting-state functional connectivity strength of
the default mode network (DMN) and salience network
(SN).
Statistic
analysis
This study is a Phase I exploratory clinical trial. Statistical
analyses will be primarily descriptive, with appropriate
statistical inference methods applied for preliminary analyses
of secondary and exploratory outcome measures. All
statistical analyses will be performed using SPSS version
26.0 (IBM Corp., Armonk, NY, USA) and GraphPad Prism
version 9.0 (GraphPad Software, San Diego, CA, USA). All
statistical tests will be two-sided, and statistical significance
will be set at P < 0.05. Given the exploratory nature of this
study, no adjustment for multiple comparisons will be made
for the results of secondary and exploratory outcome
measures; findings should be interpreted as
hypothesis-generating rather than confirmatory.
All primary outcome measures, assessing safety and
feasibility, will be analyzed using descriptive statistics only.

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Secondary outcome measures consist of changes from
baseline to Week 8 in various scale scores. For continuous
variables, normality will first be assessed using the
Shapiro-Wilk test.
If data are normally distributed, changes from baseline will
be analyzed using the paired t-test, and the mean change with
95% confidence interval will be reported. If data are not
normally distributed, changes from baseline will be analyzed
using the Wilcoxon signed-rank test, and the median change
with interquartile range will be reported.
Changes in brain function parameters from baseline to Week
8 will be analyzed using the same statistical methods
described above for continuous variables, with normality
testing guiding the choice between parametric (paired t-test)
and non-parametric (Wilcoxon signed-rank test) approaches.

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1.Introduction
Schizophrenia is a severe chronic mental illness with high disability
rate, affecting approximately 1% of the global population and imposing a
substantial disease burden on patients, families, and society [1].
Schizophrenia is primarily characterized by positive symptoms (e.g.,
hallucinations, delusions, thought disorder, behavioral disturbances),
negative symptoms (e.g., blunted affect, avolition, social withdrawal,
alogia), and cognitive symptoms (e.g., memory impairment, executive
dysfunction) [2]. The Treatment Response and Resistance in Psychosis
(TRRIP) working group defines treatment-resistant schizophrenia (TRS)
as persistent symptoms and functional impairment despite at least two
adequate trials of antipsychotic medications [3], with approximately 25%
of schizophrenia patients meeting TRS criteria [4]. Negative symptoms
are considered a primary contributor to TRS [5] and a core factor leading
to impaired social function and reduced quality of life [6]. Studies report
that approximately 30%–60% of schizophrenia patients exhibit prominent
negative symptoms [7]. Currently, schizophrenia treatment primarily
relies on antipsychotics modulating dopamine and serotonin systems.
However, both conventional and novel antipsychotics have shown limited
efficacy in improving negative symptoms [8]. The pathophysiological
mechanisms underlying negative symptoms are complex, involving
interactions among prefrontal cortex and limbic system dysfunction,
neuroinflammation,
synaptic
dysfunction,
and
neurotrophic
factor
deficiency [9]. This complexity renders medications targeting single
neurotransmitter systems unlikely to achieve optimal efficacy.
Mesenchymal stem cells (MSCs) have garnered significant attention
due
to
their
potent
anti-inflammatory,
immunomodulatory,
and
neurotrophic properties [10]. Gobshtis N et al. demonstrated that a single
intracerebroventricular
injection
of
bone
marrow-derived
MSCs

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successfully
reversed
schizophrenia-like
behaviors
and
promoted
hippocampal neurogenesis in a ketamine-induced schizophrenia mouse
model [11]. Similarly, in an amphetamine-induced schizophrenia mouse
model, a single intravenous injection of human umbilical cord-derived
MSCs
reversed
schizophrenia-like
behaviors
and
suppressed
neuroinflammation
[12].
Furthermore,
studies
have
found
that
MSC-derived exosomes exhibit therapeutic effects comparable to those of
MSCs,
with
advantages
including
low
immunogenicity,
reduced
tumorigenic risk, enhanced ability to cross the blood-brain barrier,
suitability for large-scale production and long-term storage, and improved
safety profiles. These properties position MSC-derived exosomes as
highly promising agents for treating central nervous system (CNS)
disorders [13-15]. Zhong XL et al. reported that intranasal delivery of
exosomes derived from olfactory ecto-mesenchymal stem cells restored
synaptic plasticity and neurogenesis while reducing neuroinflammation in
a methoxy methanol (MAM)-induced schizophrenia mouse model [16].
Similarly, in a phencyclidine (PCP)-induced schizophrenia mouse model,
intranasal administration of MSC-derived exosomes rapidly accumulated
in
the
prefrontal
cortex,
successfully
reversed
schizophrenia-like
behaviors, and attenuated neuronal loss [17].
To our knowledge, although no clinical studies have yet reported the
use of MSCs or their derived exosomes in schizophrenia patients, such
approaches have been investigated in other neuropsychiatric disorders,
including Alzheimer's disease [18, 19], Parkinson's disease [20], and
autism [21]. For example, Professor Duk L. Na's team at Sungkyunkwan
University School of Medicine in South Korea conducted a Phase I
clinical trial involving intracerebroventricular injections (once monthly
for
three
doses)
of
human
umbilical
cord
blood-derived
MSCs
(hUCB-MSCs) in patients with mild-to-moderate Alzheimer's disease,

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demonstrating the safety and feasibility of this approach [19]. Similarly, a
recent Phase I clinical trial led by Professor Gang Wang's team at Ruijin
Hospital,
Shanghai
Jiao
Tong
University
School
of
Medicine,
investigated
intranasal
administration
of
human
allogeneic
adipose-derived MSC exosomes (twice weekly for 12 weeks) in
mild-to-moderate Alzheimer's disease patients, finding good safety and
potential cognitive decline mitigation [18]. Additionally, a Phase II
randomized controlled trial revealed that three intravenous injections of
MSCs (10 × 10⁶/kg) significantly improved motor function in patients
with mild-to-moderate Parkinson's disease [20]. Given the substantial
preclinical evidence supporting the antipsychotic efficacy of MSCs and
their derived exosomes, it is reasonable to hypothesize that these
interventions may also prove effective in schizophrenia patients.
Dental
mesenchymal
stem
cells
(DMSCs),
originating
from
ectodermal neural crest cells, possess enhanced potential for neural
lineage differentiation, promote neuronal axon elongation, exhibit low
immunogenicity, and offer advantages in accessibility and reduced ethical
concerns [22, 23]. Due to their unique origin and superior biological
properties, DMSCs have emerged as "star cells" in regenerative medicine
for
neurological
disorders,
distinguishing
themselves
from
other
mesoderm-derived MSCs (e.g., bone marrow, adipose, umbilical cord)
[24-26]. DMSCs demonstrate capacity for neural tissue regeneration and
cerebral ganglion formation, expressing cellular markers and controlling
growth factors that stimulate angiogenesis, thereby holding significant
therapeutic potential in neurodegenerative diseases [27]. For instance, in
a rat stroke model, Song M et al. compared the efficacy of human dental
pulp MSCs (hDPSCs) and human bone marrow MSCs (hBMSCs),
finding that both stem cell types effectively reduced infarct area
compared to controls, with the hDPSC group showing significantly

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greater improvement [28]. Furthermore, Pagella P et al. reported that
hDPSCs expressed higher levels of neurotrophic factors (including NGF,
BDNF, GDNF, and NT-3) than hBMSCs, and neurons co-cultured with
hDPSCs in microfluidic systems developed longer axons than those
co-cultured with hBMSCs [29]. In Alzheimer's disease animal models,
DPSCs
have
also
demonstrated
anti-neuroinflammatory
effects,
mitochondrial repair, and cognitive improvement [30]. Additionally, Li F
et al. reported that dental pulp stem cell-derived exosomes promoted
axonal repair in spinal cord injury mice by inhibiting neuroinflammation
and apoptosis [31]. Therefore, we hypothesize that dental-derived MSCs
and their exosomes may offer superior therapeutic effects in CNS
disorders compared to other MSC types. Dental follicle tissue, derived
from unerupted tooth germs, offers the distinct advantage of abundant
tissue availability, which is essential for obtaining sufficient cells to meet
cell therapy pharmaceutical requirements. Previous research indicates that
dental follicle stem cells (DFSCs) exhibit higher proliferation rates and
clonogenic capacity compared to other dental-derived stem cells,
suggesting that DFSCs may better satisfy clinical translation requirements
in both quantity and quality [32]. Studies show that DFSCs display
protein expression profiles more similar to cranial neural crest cells,
suggesting potential advantages in treating neurological disorders [33].
Moreover, DFSCs demonstrate stronger inhibitory capacity against
inflammatory cells and enhanced promotion of anti-inflammatory cells,
indicating superior immunomodulatory capabilities [34].
Intranasal administration, as a non-invasive brain-targeted drug
delivery route, is emerging as a novel strategy for treating CNS disorders.
This pathway bypasses the blood-brain barrier, delivering drugs directly
to the brain via olfactory and trigeminal nerve pathways, thereby
enhancing brain bioavailability while reducing systemic side effects [35].

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Preclinical studies have confirmed that intranasal administration of
MSC-derived exosomes effectively improves neurological function in
animal models of Alzheimer's disease [36], Parkinson's disease [37],
stroke [38], and depression [39]. Additionally, intranasal delivery of
olfactory
ecto-mesenchymal
stem
cell-derived
exosomes
restored
synaptic plasticity and neurogenesis while reducing neuroinflammation in
schizophrenia mouse models [16]. Similarly, intranasal MSC-derived
exosomes rapidly reached the prefrontal cortex, successfully reversed
schizophrenia-like behaviors, and attenuated neuronal loss [17]. Tsivion
H et al. suggest that intranasal delivery of MSC-derived exosomes offers
advantages in feasibility and rapid access to brain lesion sites, potentially
becoming
a
valuable
future
approach
for
treating
schizophrenia,
particularly negative symptoms [40]. However, no studies to date have
reported intranasal delivery of dental-derived MSC exosomes for
schizophrenia treatment.
Based on these findings, we hypothesize that intranasal delivery of
human
dental
follicle
mesenchymal
stem
cell-derived
exosomes
(hDFSCs-Exo) may target the brain via olfactory neural pathways,
modulating neuroinflammation, promoting neurogenesis, and restoring
synaptic
plasticity,
thereby
ameliorating
negative
symptoms
in
treatment-resistant schizophrenia patients. Therefore, this study will be
the first to explore the clinical application prospects of intranasal
hDFSCs-Exo delivery for treatment-resistant schizophrenia, potentially
offering a novel therapeutic strategy for this patient population.
2. Study Objectives
1) Primary Objective
To investigate the safety and feasibility of intranasal delivery of human
dental follicle mesenchymal stem cell-derived exosomes (hDFSCs-Exo).

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2) Secondary Objective
To explore the preliminary efficacy of intranasal delivery of hDFSCs-Exo
in treating negative symptoms of treatment-resistant schizophrenia.
3. Study Subjects
3.1 Inclusion Criteria
(1) Diagnosed with schizophrenia according to ICD-10 criteria;
(2) Aged between 18 and 60 years;
(3) Long-term inpatients with a disease duration of more than 5 years;
(4) No acute exacerbation in the past 6 months, and no change in
medication regimen in the past 2 months;
(5) Poor response to adequate dose and duration of treatment with at least
two different antipsychotics, including clozapine;
(6) Positive and Negative Syndrome Scale – Factor Score for Negative
Symptoms (PANSS-FSNS) ≥24;
(7) At least two of the three core negative symptom items on the PANSS
(N1, N4, and N6) scored ≥4;
(8) Clinical Global Impression – Severity of Illness (CGI-S) score ≥4;
(9) Signed written informed consent.
3.2 Exclusion Criteria
(1) History of severe allergic reactions;
(2) Definite organic brain lesions;
(3) Comorbid severe physical diseases (e.g., unstable coronary heart
disease,
malignant
arrhythmia,
hepatic
or
renal
insufficiency,
bronchial
asthma,
acute
exacerbation
of
chronic
obstructive
pulmonary disease, autoimmune diseases, etc.);
(4) Current diagnosis of other mental disorders according to ICD-10
criteria (e.g., schizoaffective disorder, schizophreniform disorder,
bipolar I disorder, bipolar II disorder, pervasive developmental

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disorder, mental retardation, delirium, dementia, amnestic disorder, or
other cognitive disorders);
(5) Unstable condition requiring adjustment of medication regimen;
(6) Non-compliance with treatment;
(7) Severe rhinitis or nasal allergies;
(8) History of modified electroconvulsive therapy (MECT) within the
past 3 months;
(9) Individuals at risk of suicide;
(10)
Pregnant or lactating women;
(11)
Other conditions deemed unsuitable for enrollment.
3.3 Withdrawal Criteria
(1) Intolerance to the investigational treatment or occurrence of serious
adverse reactions requiring discontinuation of treatment, in the
judgment of the investigator;
(2) Development of a serious physical illness requiring discontinuation of
treatment, in the judgment of the investigator;
(3) Request for discontinuation of treatment by the subject and/or their
legal guardian;
(4) Any other circumstances under which the investigator deems it
necessary for the subject to withdraw from the study.
3.4 Termination Criteria
(1) Occurrence of significant cluster serious adverse events during the
trial that, in the investigator's judgment, necessitate study termination;
(2) Identification of major errors in the clinical trial protocol that
compromise the evaluation of drug effects, or occurrence of
significant
deviations
during
implementation
of
an
otherwise
well-designed
protocol
that,
if
continued,
would
impair
the
assessment of drug efficacy and safety;

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(3)Requirement for termination/suspension of the trial by the regulatory
authority.
4. Study Design and Procedures
4.1 Study Design
This is a single-center, open-label, single-arm, 3+3 dose-escalation design
comprising five dose groups. Dose escalation will be determined based
on dose-limiting toxicities (DLTs). It is anticipated that 15–30 subjects
will be enrolled.
4.2 Study Procedures
Five dose groups will be established, with dose escalation decisions
guided by the occurrence of DLTs. DLTs will be assessed according to
NCI-CTCAE version 5.0 criteria.
A DLT is defined as any toxicity possibly related or related to the
investigational drug occurring during the DLT observation period (within
28 days after a single administration) that meets any of the following
criteria:
1) Hematological Toxicity (lasting ≥7 days): Grade 4 neutropenia
(absolute neutrophil count < 0.5 × 10⁹/L); Grade 3 thrombocytopenia
with bleeding, or Grade 4 thrombocytopenia (platelet count < 25 × 10⁹/L);
Grade 3 or Grade 4 anemia (hemoglobin < 8.0 g/dL).
2)
Non-Hematological
Toxicity:
Any
Grade
3
or
Grade
4
non-hematological toxicity. Local Toxicity: Grade 3 or Grade 4 nasal
mucosal inflammation, ulceration, or bleeding; Severe nasal congestion
preventing nasal breathing. Systemic Allergic/Immune Reactions: Grade
3 or Grade 4 allergic reactions, systemic rash, or angioedema.
Neurological Toxicity: New-onset, drug-related Grade 3 or Grade 4
headache,
dizziness,
sensory
or
motor
nerve
disorders.
Systemic
Inflammation: Grade 3 or Grade 4 fever (body temperature > 39.0°C and
drug-related); Grade 3 or higher symptoms related to cytokine release

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syndrome. Serious Adverse Events (SAEs): Any drug-related SAE,
regardless of grade.
3) Laboratory Abnormalities: Grade 3 or Grade 4 hepatic or renal
function abnormalities (e.g., ALT/AST elevation > 5 × upper limit of
normal, creatinine elevation > 3 × upper limit of normal), judged by the
investigator to be related to the investigational drug.
5. Research Methods
5.1 Exosome Preparation and Characterization
Clinical-grade human dental follicle mesenchymal stem cell-derived
exosomes (hDFSCs-Exo) will be provided by Chengdu Shilian Kangjian
Biotechnology Co., Ltd. (specification: 30 billion particles/2 mL). Each
batch has a shelf life of 30 days. The exosomes will be transported via
dry ice refrigeration and stored at -80°C upon arrival at the hospital.
Before use, they will be thawed at room temperature and gently shaken to

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ensure uniformity.
Exosome
morphology will be observed using
transmission electron microscopy (TEM). Particle size distribution and
concentration will be determined using nanoparticle tracking analysis
(NTA). Marker protein expression will be detected via Western blotting,
including positive markers CD9, CD63, CD81, Alix, and TSG101, and
negative markers GM130 and Calnexin.
5.2 Intervention Protocol
1) Dose Determination:
The intranasal dose for exosome treatment in schizophrenia mouse
models was reported as 2.4×10⁷ particles per administration [17]. Based
on body surface area conversion (0.007 m² for a 20 g mouse and 1.53 m²
for a 60 kg adult), the equivalent human dose was calculated as 5.54×10⁹
particles per administration. Applying a 5-fold safety factor, the starting
dose was reduced to 1.1 × 10 ⁹
particles per administration. This was
further considered alongside the safe dose of 8 × 10 ⁸ particles per
administration used in intranasal exosome treatment for Alzheimer's
disease patients [18]. To explore the maximum tolerated dose (MTD) and
recommended Phase II dose (RP2D), five dose groups are planned: 1×10
⁹, 2× 10⁹, 4×10⁹, 8×10⁹, and 1.6×10¹⁰ particles per administration.
2) Administration Method:
Exosomes will be dissolved in 0.2 mL of normal saline and administered
via intranasal spray, alternating between both nostrils. Treatment will be
administered twice weekly (with a 3-day interval between doses) for 8
weeks, totaling 16 administrations. Nasal hygiene will be performed prior
to each treatment session.
3) Exosome Skin Test:
To avoid potential allergic reactions, subjects will undergo a wrist skin
test prior to the first treatment. The skin test will be performed by a

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clinical nurse, with results assessed 20 minutes after administration.
Results will be classified as negative (no reaction at the test site; no rash
with itching, no papules, no surrounding urticaria, redness, or swelling) or
positive (presence of marked rash, papules, or urticaria; surrounding
congestion, swelling, blisters, itching, etc.) [38].
5.3 Assessment Content and Schedule
1) Safety and Feasibility Assessments:
(1) Vital signs monitoring
(2) Electrocardiogram (ECG)
(3) Complete blood count
(4) Liver and kidney function tests
(5) Allergy markers (IgE)
(6) Inflammatory markers
(7) Recording of adverse events (AEs) and serious adverse events (SAEs)
(8) Treatment completion rate
2) Efficacy Assessments:
(1) Positive and Negative Syndrome Scale (PANSS) negative subscale
score
(2) Scale for the Assessment of Negative Symptoms (SANS) score
(3) Calgary Depression Scale for Schizophrenia (CDSS) score
(4) Montreal Cognitive Assessment (MoCA) score
(5) Clinical Global Impression (CGI) scale score
(6) Trail Making Test Parts A & B
(7) Digit Span Test
(8) Verbal Learning and Memory Test
3) Other Assessment Tools:
(1) Electroencephalography (EEG)
(2) Functional near-infrared spectroscopy (fNIRS)
(3) Functional magnetic resonance imaging (fMRI)

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4) Assessment Schedule
Screen
-ing
Base
-line
Intervention
Follow-up
Visiting
V1
V2
V3
V4
V5
V6
V7
V8
V9
Week
-2
0
2
4
8
12
16
20
24
Informed consent
X
Enrollment
X
Demographic data
X
Medical history
X
Physical
examination
X
X
X
X
X
X
X
X
Vital signs
X
X
X
X
X
X
X
X
Lab examination
X
X
X
X
X
X
X
X
PANSS
X
X
X
X
X
SANS
X
X
X
X
X
CDSS
X
X
X
X
X
CGI
X
X
X
X
X
MoCA
X
X
X
X
X
TMT-A
X
X
X
X
X
TMT-B
X
X
X
X
X
DST
X
X
X
X
X
VLMT
X
X
X
X
X
Inflammatory factor
X
X
X
X
X
X
X
ECG
X
X
X
X
X
X
X
X
EEG
X
X
fNIRS
X
X
MRI
X
X
AEs & SAEs
X
X
X
X
X
X
X
X
6. Outcome Measures
6.1 Primary Outcome Measures
1) Safety Measures:
(1) Adverse Events: Incidence and severity of adverse events (AEs) and
serious adverse events (SAEs), graded according to CTCAE version 5.0.
(2) Dose-Limiting Toxicity: Incidence of dose-limiting toxicities (DLTs),
used to determine the maximum tolerated dose (MTD) and recommended
Phase II dose (RP2D).

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(3) Vital Signs: Incidence of abnormalities in body temperature, pulse,
respiration, and blood pressure.
(4) Physical Examination: Incidence of abnormalities in general condition,
head, skin, mucous membranes, lymph nodes, neck, chest, abdomen,
spine/extremities, and neurological examination.
(5) Laboratory Tests: Incidence of abnormalities in complete blood count,
urinalysis, blood biochemistry (liver and kidney function), infectious
disease markers, inflammatory factors, and allergy markers (IgE).
(6) Electrocardiogram (ECG): Incidence of ECG abnormalities.
2) Feasibility Measures:
(1) Treatment Completion Rate: Proportion of subjects completing the
full 8-week (16 sessions) intranasal administration regimen.
(2) Treatment Adherence: Evaluation of subjects' acceptance of and
compliance with the intranasal administration method.
6.2 Secondary Outcome Measures
(1) Change in Positive and Negative Syndrome Scale (PANSS) total
score and factor scores.
(2) Change in Clinical Global Impression --Severity of Illness (CGI-S)
score.
(3) Change in Scale for the Assessment of Negative Symptoms (SANS)
total score and subscale scores.
(4) Change in Calgary Depression Scale for Schizophrenia (CDSS) score.
(5) Change in Montreal Cognitive Assessment (MoCA) total score.
(6) Change in Trail Making Test Part A/B (TMT-A/B) completion time.
(7) Change in Digit Span Test (DST) score.
(8) Change in Verbal Learning and Memory Test (VLMT) score.
6.3 Exploratory Outcome Measures
(1) Electroencephalography (EEG): Changes in resting-state power
spectrum in the prefrontal theta band (4 – 8 Hz), and changes in

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functional connectivity strength of the default mode network based on
source localization.
(2)
Functional
Near-Infrared
Spectroscopy
(fNIRS):
Changes
in
oxygenated
hemoglobin
(Oxy-Hb)
concentration
in
the
bilateral
dorsolateral prefrontal cortex (DLPFC) during a verbal fluency task.
(3) Magnetic Resonance Imaging (MRI): Changes in resting-state
functional connectivity strength of the default mode network (DMN) and
salience network (SN).
7. Statistic analysis
This study is a Phase I exploratory clinical trial. Statistical analyses
will be primarily descriptive, with appropriate statistical inference
methods applied for preliminary analyses of secondary and exploratory
outcome measures. All statistical analyses will be performed using SPSS
version 26.0 (IBM Corp., Armonk, NY, USA) and GraphPad Prism
version 9.0 (GraphPad Software, San Diego, CA, USA). All statistical
tests will be two-sided, with statistical significance set at P < 0.05. Given
the exploratory nature of this study, no adjustment for multiple
comparisons will be made for the results of secondary and exploratory
outcome
measures;
findings
should
be
interpreted
as
hypothesis-generating rather than confirmatory.
Primary outcome measures (safety and feasibility) will be analyzed
using descriptive statistics exclusively.For secondary outcome measures,
which assess changes from baseline to Week 8 in various scale scores, the
following approach will be applied to continuous variables: normality
will first be assessed using the Shapiro-Wilk test. If data are normally
distributed, changes from baseline will be analyzed using the paired t-test,
with the mean change and 95% confidence interval reported. If data are
not normally distributed, changes from baseline will be analyzed using
the Wilcoxon signed-rank test, with the median change and interquartile

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range reported. Exploratory outcome measures (changes in brain function
parameters assessed by EEG, fNIRS, and MRI) will be analyzed using
the same statistical methods described above for continuous variables,
with normality testing guiding the selection between parametric (paired
t-test) and non-parametric (Wilcoxon signed-rank test) approaches.
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