Background/Purpose: Excessive formation and impaired clearance of Neutrophil Extracellular Traps (NETs) have been linked to autoimmune and inflammatory diseases, notably systemic lupus erythematosus (SLE). DNASE1-like 3 (DNASE1L3) is a circulating endonuclease that degrades NETs, and loss-of-function mutations are associated with severe autoimmune phenotypes in patients, including childhood-onset SLE, vasculitis, and systemic sclerosis. Based on this biology, Neutrolis developed a platform of engineered recombinant DNASE1L3 analogues, including NTR-441, a DNASE1L3 analogue fused to human serum albumin. In this first-in-human study, we aimed to validate the novel approach of NET inactivation by DNASE1L3 as a therapeutic strategy with three main objectives:

(1) assess safety, tolerability, and pharmacokinetics (PK) in healthy subject lacking the pharmacological target (NETs)

(2)   evaluate safety, tolerability, PK, and target engagement in patients with a high NET burden

(3)   establish pharmacodynamic and clinical proof-of-concept in a patient with SLE and congenital DNASE1L3 deficiency

Methods: A first-in-human Phase 1a/b, placebo-controlled, double-blind trial was conducted, including a single ascending dose study in 35 healthy adults and 12 hospitalized patients with severe COVID-19, representing a high NET burden disease (NCT04941183) across dose levels of 0.01–10 mg/kg. Proof-of-concept was investigated in a 16.5-year-old male with a homozygous DNASE1L3 loss-of-function mutation (c.643del) and severe, longstanding, treatment-refractory monogenic SLE, who received weekly doses of NTR-441 (3 mg/kg) as add-on therapy to corticosteroids, antimalarials, and tofacitinib through a compassionate use protocol (Ethics Committee approval no. 0120-553/2024-2711-8).

Results: NTR-441 was well tolerated across all studies and produced robust increases in DNASE1L3 levels that remained supraphysiologic for >48 hours (Fig. 1). Infusion reactions occurred in one healthy subject at the highest dose and at the fifth dose in the patient with SLE, the latter linked to anti-drug antibodies but managed with desensitization and adjusted dosing. Target engagement was demonstrated in patients with COVID-19 and SLE by rises in NET-degradation products (MPO–DNA complexes, cell-free DNA) peaking 4–8 hours post-infusion, while absent in healthy volunteers (Fig. 2). In the SLE patient, the first dose of NTR-441 resulted in a rapid (≤6 hours) and sustained clinical improvement across multiple organ systems, including urticarial rash, episcleritis, and arthritis except for a transient rash recurrence that subsequently subsided (Fig. 3).

Conclusion: These combined studies validate DNASE1L3 as a therapeutic strategy to inactivate NETs. In a patient with treatment-refractory SLE with DNASE1L3-deficiency, pharmacologic degradation of NETs with NTR-441 induced rapid, sustained remission, providing the first clinical proof-of-concept for this approach. The temporal link between NET clearance and clinical benefit supports NETs as a central pathogenic driver in autoimmunity. These findings warrant broader trials in SLE and other autoimmune disorders.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/early-evidence-of-proof-of-concept-of-an-albumin-dnase1l3-fusion-protein-ntr-441-for-the-rapid-enzymatic-inactivation-of-nets-in-sle-with-dnase1l3-deficiency/