Interleukin-1 Antagonist Anakinra in Amyotrophic Lateral Sclerosis—A Pilot Study

Preclinical studies show that blocking Interleukin–1 (IL–1) retards the progression of Amyotrophic Lateral Sclerosis (ALS). We assessed the safety of Anakinra (ANA), an IL–1 receptor antagonist, in ALS patients. In a single arm pilot study we treated 17 ALS patients with ANA (100 mg) daily for one year. We selected patients with dominant or exclusive lower motor neuron degeneration (LMND) presentation, as peripheral nerves may be more accessible to the drug. Our primary endpoint was safety and tolerability. Secondary endpoints included measuring disease progression with the revised ALS functional rating scale (ALSFRSr). We also quantified serum inflammatory markers. For comparison, we generated a historical cohort of 47 patients that fit the criteria for enrolment, disease characteristics and rate of progression of the study group. Only mild adverse events occurred in ALS patients treated with ANA. Notably, we observed lower levels of cytokines and the inflammatory marker fibrinogen during the first 24 weeks of treatment. Despite of this, we could not detect a significant reduction in disease progression during the same period in patients treated with ANA compared to controls as measured by the ALSFRSr. In the second part of the treatment period we observed an increase in serum inflammatory markers. Sixteen out of the 17 patients (94%) developed antibodies against ANA. This study showed that blocking IL–1 is safe in patients with ALS. Further trials should test whether targeting IL–1 more efficiently can help treating this devastating disease. Trial Registration ClinicalTrials.gov NCT01277315

Scientific background for the study: ALS is a neurodegenerative disease of adult onset caused by selective degeneration of motor neurons in the cortex and the spinal cord. The degeneration of motor neurons results into progressive paralysis of the limbs, speech, swallowing and respiratory muscles. ALS is always fatal and patients die of respiratory insufficiency between 3 and 5 years after diagnosis. So far, Riluzol (Sanofi-Synthelabo-Aventis) is the only approved medication that results into a minor prolongation of life (1.5 to 2.5 months). Riluzol is an inhibitor of the presynaptical release of glutamate. Another effect of Riluzol is the inhibition of glutamate release in activated microglia.
Neuroinflammation is an essential pathogenicity factor of motor neuron degeneration. Inflammatory changes can be found in sporadic ALS, the autosomal dominant form of ALS as well as in the SOD1-mouse model of the ALS. This can be demonstrated morphologically through the accumulation of activated microglia, astrocytes, and T-cells. At the biochemical level, these changes are accompanied by the activation and release of the proinflammatory cytokine Interleukin (IL)-1. The inflammatory mediator IL-1 mediates the recruitment and activation of phagocytic cells and induces an immune response. The bioactivity of IL-1 is physiologically controlled by the naturally occurring, highly selective Interleukin-1-Receptor-Antagonist (IL-1RN).
On the basis of the neuroinflammatory changes in ALS there is a scientific rationale for a clinical trial with ANA, which has anti-inflammatory pharmacological effects. ANA is a recombinant IL-1RN (r-metHuIL-1RN) which directly and selectively inhibits IL-1.

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The primary study target implies the safety and tolerance of ANA in combination with Riluzol in ALS patients. An open tolerability study aims at testing whether a subcutaneous injection of 100 mg ANA is tolerated. This tolerability study will be the basis of a phase IIb/III efficacy study.
Tested medication: Trial medication: ANA (Kineret®) 100 mg of injectable solution in a readyto-use pre-filled syringe in combination with an oral dosis of 100 mg Riluzol. Open treatment with 100 mg ANA + 100 mg Riluzol.
Comparative medication: none Study design: One arm study to assesst the safety and tolerance of 100 mg ANA for the duration of 52 weeks in combination with Riluzol. • clinical diagnosis of ALS (revised El Escorial criteria) with dominant affection of the 2nd motor neurons or the clinical ALS variant of a progressive muscle atrophy (PMA).
• clinical signs of a dysfunction of the 2nd motorneuron in at least one anatomic region outside the bulbar region. • sporadic and familiar ALS. • onset of symptoms with pareses from six months up to four years before inclusion in the study.
• treatment with Riluzol 100 mg/day at least for 3 months before termination of study. • written informed consent (according to AMG §40 (1) 3b). • ability to communicate, either in person or by an authorized person. • ability to communicate, either verbally, manually or through an electronic communication system. • internet access, either personally or by an authorized person. • refrigerator facilities for ready-to-use pre-filled ANA-syringes at home. • patienten with known hypersensitivity to ANA, Riluzol or one of the adjuvants.
• clinical severe hypoventilation syndrome with a vital capacity < 50 %. • pregnant or breast-feeding women.
• women of childbearing age who are not using or cannot use any highly effective contraceptives (Pearl-index <1).
• clincal significant modifications of the ECG including symptomatic bradycardia. • continuous non-invasive ventilation with a ventilation free period < 2 hours. • Tracheotomy and mechanical ventilation. • Laboratory parameters outside the standard range and equivalent to a clinical significant. cardiovascular, pulmonological, hematological, hepatological, metabolic and kidney disease. • malignant tumors. • severe renal impairment (creatinine-clearance < 30 ml/min). • previous history of recurrent infection or an underlying disease that predisposes to infections.
• patients with severe clinical co-morbidities including psychiatric diseases. • dementia and lack of capacity for consent. • clinicial diagnosis of epilepsy or an epileptic seizure.
• treatment with any other study medication < 3 months prior to study inclusion. • diseases or malfunctions which exclude participation at the study at the treating investigator's discretion.
• non-cooperation. • all contraindications concerning the study medication (inclusive adjuvants of the dosage form).
• non-cooperation regarding storage and disclosure of pseudonomized data in the context of the clinical trial.

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Targets: The primary target of the study is the examination of safety and tolerability of a subcutaneous injection of ANA together with an oral dosis of Riluzol in ALS patients. The choice of sample size is not appropriate for securing the effectiveness of ANA in combination with Riluzol in ALS. However, by means of secondary endpoints the influence of the ANA treatment on the ALS progression rate and life quality will be studied.

Safety Information, consent, and documentation:
• the patient will be given written information on the application of the medication, its risks, the importance of a contraception, and appropriate methods for safe contraception.
• written consent of the patient to participate in the study. • written confirmation from the patient that the information on the study has been read and understood.
• documentation of information and written consent.
Women in childbearing age who did not have a hysterectomy, and whose menopause was less than 24 months before the initiation of the study will have to have a pregnancy test prior to treatment with the study. Women in childbearing age also have to ise a highly effective contraception method (defined as Pearl Index < 1) (pill, contraceptive coil, hormone implant, transdermal patch, a combination of two barrier methods (condom and diaphragm)), a sterilization or sexual abstinence at least one month prior to the first application of the study medication and for the whole duration of the treatment.
Measures to be taken prior to the treatment of women in childbearing age • pregnancy test in the urine • discontinuation of therapy in case of proof of pregnancy Page 12 in the German text Termination criteria Termination or premature discontinuation of the study will be documented for each patient. The reason for the termination will be examined and documented. The following criteria can be sufficient for the discontinuation of the study: • laboratory parameters outside the normal range, e. g. o neutropenia (ANZ < 1,5 x 109/l). o severe renal impairment (CLCr < 30 ml/minute). o ALT/AST values are more than 5 fold higher than the upper limit. • pregnancy. • significant violation of study plan . • lack of compliance. • severe infection.
• clinical signs for an allergic reaction including maculopapular or urticarial rashes or unclear increase in body temperature. • severe skin reactions, especially bullous skin reactions including Stevens-Johnsonsyndrome and toxic-epidermal necrolysis • pathological examination findings, including electrocardiography. • personal request of the patient. • withdrawal of informed consen.t • loss of contact.
•if the patient moves to another place and can no longer participate in the study.
Page 13 in the German text • death of the patient Termination of the study: A premature termination of the study is possible under the following conditions: • a patients suffers of a severe infection which is possibly, likely, or certainly caused by the study medication and cannot be explained in the context of the underlying clinical condition.
• new and reproducibly scientific findings on ANA during the course of the clinical study show the inferiority of the treatment with ANA • evidence of increasing number of serious adverse events The PI of the clinical trial will decide on the termination of the study. After termination the following events will be documented and communicated to the sponsor of the study: • all serious adverse events which have incurred within 30 days after the last treatment with ANA • Any pregnancy which has been confirmed within 84 days (12 weeks) after the last dosis of ANA .

Statistical evaluation:
The primary objective of this pilot study is tolerability and the secondary endpoints should allow, concurrently, to establish conditions to plan a statistically justifiable phase II efficacy study, is a proof of tolerability and several secondary endpoints. Primary and several secondary endpoints will first be examined exploratively and evaluated descriptively. The conceptual study will be carried out with n=20 patients. Similar scientific questions have been described in the literature with a comparable number of patients. Secondary endpoints will be examined by parametric or non-parametric tests depending on scaling and distribution type. The biometric evaluation will be done by SAS®.
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Aims of the clinical trial
The primary target of the study is the examination of safety and tolerability of a subcutaneous injection of ANA combined with Riluzol in ALS patients. The chosen sample size is not appropriate for proving the effectiveness of ANA in combination with Riluzol in ALS. However, by means of secondary study endpoints the influence of ANA on the ALS progression rate can be investigated.

Main target criterion
Primary objective of the study: Examination of number and severity of adverse events (UE), severe adverse events (SUE), adverse drug reactions (ADR), unexpected adverse drug reactions (uADR), severe adverse drug reactions (SADR), Suspected Unexpected Serious Adverse Reaction (SUSAR), and pathological laboratory parameters.

Other criteria
Secondary objectives of the study: • Evaluation of long term tolerability and drug safety of ANA in combination with Riluzol. • Number of patients in need of continuous non-invasive ventilation or tracheostoma supported invasive ventiliation. • number of patients in need of a percutaneous endoscopic gastrostomy (PEG). • number of patients who have concluded a treatment with ANA in combination with Riluzol. • evaluation of clinical effectiveness of ANA on the progression rate of pareses . and functional impairments in ALS by comparative analysis of the intraindividual progression rate of the revised ALS Functional Rating Scale (ALS-FRS-R) in a study period of 52 weeks compared to the progression rate before participation in the study (Δ ALS-FRS-R).
• examination of effectiveness of ANA on the respiratory function in ALS which is assessed by a forced vital capacity in the course of 52 weeks. • examination of effectiveness of ANA on the respiratory function in ALS patients in need of a non-invasive mask ventilation (determination of ventilation time per day). • determination of survival time or timing for a tracheotomy by respiratory insufficiency (exclusive elective tracheotomy or tracheotomie for control of secretion).

Study design
The study design is to evaluate of safety and tolerability of 100 mg ANA in combination with Riluzol for the duration of 52 weeks in.