Complement Inhibition: Attacking the Overshooting Inflammation @fter Traumatic Brain Injury ( A phase II trial on the safety and efficacy of C1 esterase inhibitor Cinryze for the acute management of severe traumatic brain injury)

Traumatic Brain Injury (TBI) is a major cause of death and disability across
all ages in all countries with the number of elderly people with TBI increasing
due to falls and in the younger population due to road traffic incidents (1).
In Europe, one million TBI patients are admitted to the hospital yearly, of
whom 75.000 people die. This debilitating morbidity leads to enormous societal
costs (2, 3). Therapies and guidelines that have been demonstrated to improve
outcome after TBI are still limited, especially in the management of severe TBI
(s-TBI). Patients with s-TBI have a high mortality rate, estimated at 30-40% in
observational studies on unselected populations (1). Survivors of s-TBI
experience a substantial burden of physical, psychiatric, emotional and
cognitive disabilities, that disrupt their lives and their surroundings. Due to
the huge burden after trauma, adequate treatment of these patients is
important. TBI comprises a dynamic pathophysiology that evolves in time,
consisting of primary injury, followed by systemic disorders which leads to
secondary injury (4). These secondary injuries complicate the period of
admission and recovery (5).

The human immune system comprises the adaptive and innate immune responses. The
complement system forms the first line of defence against microorganisms and is
critical in sensing tissue damage. Complement activation can be mediated by
three distinct pathways: the classical pathway, the alternative pathway and the
lectin pathway (6) Figure 1. Multiple experimental studies have identified a
pathophysiologic role of the complement system in contributing to posttraumatic
neuro-inflammation, disruption of the blood-brain barrier, secondary neuronal
damage and neuronal cell death after traumatic brain injury (7-11). In TBI
patients, elevated complement factors have been found in serum (12) and in
ventricular cerebrospinal fluid (CSF) directly after the initial trauma (13,
14). Activation of the complement system in TBI results in a cascade of events
including increased vascular permeability and activation of microglia and
astrocytes, ultimately resulting in inflammatory reactions in and around
contusion areas (15, 16). The overshooting inflammatory response, formation of
brain edema and elevated intracranial pressures (ICP) cause secondary brain
injury and is subsequently related to late morbidity and mortality in TBI (17).
The neuro-inflammation can persist for years after the initial trauma and
secondary damage in (sub)acute phase (18) even causing predisposition for other
neurological afflictions, like early onset dementia, later in life (19). In TBI
patients, elevated complement factors have been found in serum (12) and in
ventricular cerebrospinal fluid (CSF) directly after the initial trauma (13,
14).

Complement inhibition is therefore considered to be a potentially important
aspect of TBI treatment. Most modalities of complement inhibitions have
focussed on interfering with the cascade at the central level of the C3 or C5
convertases (20-22). The Membrane Attack Complex (MAC), the final product of
the terminal pathway, initiates the NLRP3 inflammasome causing production of
IL-1B, which activates microglia, astrocytes and causing infiltration of
macrophages from the periphery (23). As shown in our previous experiments in
TBI animal models (Baas group), inhibition of MAC formation appears to be
sufficient to prevent secondary neurologic damage and improve neurologic
performance in mice by reducing microglia activation, apoptosis and axonal loss
(24). Studies directed at MAC formation are therefore promising as potential
therapeutic intervention after TBI (25). Although specific MAC inhibition might
seem a good approach to prevent secondary brain injury, there are also studies
showing that C3 activation (upstream of MAC) is negatively influencing the
inflammation overshooting of TBI. Activation of C3 triggered a sustained
degenerative mechanism of microglial and astrocyte activation, reduced
dendritic and synaptic density and inhibited neuroblast migration several weeks
after TBI in animal models (26).

Complement 1 inhibitor (C1-INH) controls activation of multiple plasma mediator
pathways by binding to two of the active sub-units of the first component of
the complement system (C1r and C1s). In addition, it is a known inhibitor of
kinin generation (kallikrein), fibrinolytic (plasmin) and contact activation
(factor XIIa, XIIf, X1a) (27). It has also been shown to be an inhibitor of the
mannan-binding lectin pathway of complement activation, inhibiting
mannan-binding lectin-associated serine proteases (MAPSs) in the pathway (28).
C1 inhibitor also interacts with C2b to inhibit binding of factor B to C3b and
therefore it is also a down-regulator the alternative pathway convertase (29).

Cinryze is a human C1 esterase inhibitor (serine protease inhibitor), isolated
from human plasma and is approved for treatment of hereditary angioedema (30).
Because of its excellent safety profile, it has also been used to treat other
inflammatory diseases, such as sepsis or ischemic reperfusion injury (31, 32).
C1 esterase inhibitors are the only approved drugs that can inhibit most of the
complement pathway activity. An alternative for Cinryze is to use the
recombinant C1 esterase inhibitor Ruconest (33). However, the cost per
treatment is higher for Ruconest and the elimination half-life of the
recombinant C1 esterase inhibitor Ruconest is 2.5 hours, compared to 56-62
hours in the natural human protein Cinryze (34, 35). Due to the longer
circulation time and the lower cost of treatment, Cinryze is the preferred
treatment to use in this clinical trial to reduce ICP and secondary brain
injury.

Currently, there is clinical and preclinical evidence available regarding the
role of the complement system in developing secondary brain injury in TBI
patients. Moreover, high quality pre-clinical evidence exists for years
concluding that interfering in this complement cascade alters the brain damage
dramatically. Complement inhibition is considered to be a potentially important
aspect of TBI treatment. Therefore, C1-INH can be beneficial in treatment of
s-TBI, downsizes the detrimental and overshooting inflammation and preventing
secondary brain injury to ensure a more favorable functional outcome and
quality of life for patients with TBI.

This study has been transitioned to CTIS with ID 2024-514488-24-01 check the CTIS register for the current data.

Primary Objective: To determine the safety and efficacy of 6000 IU Cinryze in
patients with moderate and severe TBI (GCS <13 with a clinical indication for
ICP measurements).

Primary hypothesis: The hypothesis is that random assignment to Cinryze in
patients with moderate and severe TBI will experience a reduction in ICP
directed therapy intensity levels (TIL) compared to random assignment to
placebo (difference of 2.2). Secondary, if efficacy is proven on the TIL scale,
a difference of the GOSE at six months will be evaluated. Furthermore, no
difference should be detected in complication rate after one month between the
two groups.

Secondary Objective: To determine differences between Cinryze and placebo
treatment in the following outcomes for patients with moderate and severe TBI:
- Clinical outcomes: ICP burden, CT midline shift, GOSE, mortality, hospital
and ICU length of stay, ventilator days, hospital disposition, quality of life
(as expressed by QoLiBri), health-related quality of life (as expressed by
SF-36 and EQ-5D-5L)
- Cost-effectiveness
- Neurological damage: BANYAN (GFAP/UCHL-1) blood biomarker
- Complement activation: human serum (WIESLAB assay) and total terminal
complement activity levels (CH50) and protein levels of complement component.
- Level of Cinryze (C1 inhibitor activity) in plasma and RNA expression

Specifically, it is hypothesized that patients randomly assigned to Cinryze
will have a lower ICP burden, measured as minutes of ICP>20 mM Hg as compared
to the control group. Furthermore, we hypothesize that random assignment to
Cinryze in patients with TBI will change the proportion of patients with a
favourable long-term neurologic outcome compared to random assignment to
placebo, based on the GOS-E at six months. The outcome biomarkers (complement
activation and BANYAN) serve to get a better mechanistic understanding of the
pathophysiological neuro-inflammation and of the possible therapeutic effect of
Cinryze.

The CIAO@TBI trial is a prospective, multicenter, randomized, double-blind,
placebo-controlled phase II trial, with one group receiving one dose of Cinryze
intravenously (IV) and one group receiving a placebo injection IV. The study
will be performed in different Dutch level 1 trauma centers. Patients diagnosed
with TBI on admission to the emergency departments between
01/08/2020-01/08/2022 with a Glasgow coma score of < 13 (with intracranial
deviations on scan) and indication for ICU treatment with ICP monitoring will
be eligible for inclusion. If informed consent can be obtained within 12 hours
after the accident, patients are randomly assigned to one of the two study arms
and will receive a single dose of 6000 IU Cinryze or placebo IV. (1 IU = the
average endogenous level C1-esterase inhibitor in 1 ml human plasma). Neither
the participants, nor the experimenters will know who is receiving the Cinryze
or placebo. All patients will receive standard care. Blood and CSF samples (CSF
only when drain is placed) will be taken from both patient groups before
administration of Cinryze or placebo. Additional blood samples will be taken at
6, 12, 24, 48, 72, 96 hours after dosing of the Cinryze or placebo. Additional
CSF samples will be taken at 24, 48, 72, 96 hours after dosing of the C1-INH or
placebo. The timing of these blood samples is based on the estimated activity
and elimination time of the test compound and the timing of the onset of
neuroinflammation. In most TBI patients a crisis can occur due to the onset of
neuro-inflammation about 3 days after the trauma (*troisième jour*) (36). The
brain tends to swell the most on the third day after a traumatic injury. The
patients* blood samples will be used to asses biomarkers for neurological
damage (BANYAN), to measure qualitative levels of functional classical, MBL and
alternative complement pathways in human serum, total terminal complement
activity levels, protein levels of complement component using different assays
and additional inflammatory markers like TNF-alpha and intraleukin. Patients
will receive routine CT scans as part of the standard care.

Clinical scores routinely used in the standard clinical care and follow-up
during recovery of TBI will be registered up to one year after discharge. Most
patients that will match the inclusion criteria are most likely not able to
give informed consent themselves due to the nature of the injury (incapacitated
due to loss of conscience, mental confusion). Informed consent can therefore be
obtained by patient or proxy or deferred consent. Informed consent must be
obtained within 12 hours as the efficacy of the C1-INH is suspected to be
limited 12 hours after trauma. The LUMC will function as a data coordination
and analysis center. All neurosurgeons, intensive care physicians/nurses and
other people concerned will be instructed with regard to the study.

- CINRYZE is a heat-treated, nanofiltered C1 inhibitor product, purified from
human plasma for fractionation. It is presented as lyophilized powder and
solvent for solution for injection for the proposed indication. The product is
reconstituted with Water for Injections (5 ml) resulting in 100 U/ml
C1-inhibitor, with one unit (U) of C1-inhibitor corresponding to the
C1-inhibitor activity in one millilitre of normal human plasma.
The starting material for the manufacturing process of CINRYZE active drug
substance is "plasma for fractionation" according to Ph. Eur. monograph
07/2008:0853. Plasma is provided in the Netherlands by Sanquin, and in Belgium
by CAF-DCF cvba-scrl (CAF-DCF, a subsidiary of Sanquin) and in Finland by the
Finnish Red Cross Blood Service (FRC). Information on the source, collection,
separation and control of the human plasma used as the starting material of
CINRYZE is provided in the relevant Plasma Master Files of Sanquin and CAF-DCF,
respectively.
The primary function of Cinryze is to regulate the activation of the complement
and contact pathways. This regulation is performed through the formation of
pathway-specific complexes that result in inactivation of the target protease
and consumption of C1 INH. C1 inhibitor inhibits the complement system by
binding C1r and C1s, two of the active enzyme subunits of the first component
of the complement system (C1) in the classical pathway. The primary substrate
of the activated C1 enzyme is C4; uninhibited C1 results in diminished C4
levels. An increase in C4 levels is therefore a surrogate measure of the
biological effect of Cinryze.
C1 inhibitor regulates the contact system and the intrinsic coagulation
(kallikrein-kinin) pathway by binding to and inactivation kallikrein and factor
XIIa. Because these pathways are part of enzyme amplification cascades, without
C1 INH, spontaneous or trigger-induced activation of these pathways leads to
production of the vasoactive peptide bradykinin.

- Placebo: physiological saline (0.9%) in equal volume dosed intravenously (60
ml)

The treatment itself consists out of a single intravenous injection (6000 IU)
Cinryze or equal injection volume of placebo (physiological saline. Cinryze is
a C1 esterase inhibitor isolated from human plasma. As such it is a *human
blood product* and cannot be used by people sensitive for human blood products.
When medicinal products prepared from human blood or plasma are administered,
the possibility of transmitting infective agents cannot be totally excluded.
This also applies to unknown or emerging viruses and other pathogens.
Nevertheless, this risk is considered very low. The risks associated with short
term complement inhibition by Cinryze are a theoretical increase in
susceptibility for bacterial meningitis. This is controlled in this study by
the short term of the treatment with complement inhibitors and careful
monitoring of the patient for such infections. This risk is considered low. A
decrease in coagulation time because of Cinryze treatment is possible. This can
result in thrombosis and might pose extra risks in patients with indwelling
catheters. However, most TBI patients experience increased coagulation times
due to the consumption of coagulation factors after trauma. Since the drug is
given only once this risk is considered low and patients with a history of
thrombosis will be excluded. The potential benefit for the patient is that
complement inhibition will delay neuroinflammation. The delay of the
neuroinflammation might improve the clinical outcome and leads to a decrease of
secondary brain injury in the TBI patient. Closely monitoring of the patients
can be beneficial in the treatment of TBI and the information gathered from
this research project will help to introduce better treatment of TBI patients
in the future. Furthermore, C1-inh has been investigated to have an excellent
safety profile in multiple trials.