First clinical trials of a new heteropolymer technology agent in normal healthy volunteers and patients with systemic lupus erythematosus: safety and proof of principle of the antigen-heteropolymer ETI-104

Abstract

Background: The heteropolymer technology was developed to remove pathogens from the circulation.

Objectives: To evaluate the safety and tolerability of a single administration and to establish proof of principle for ETI-104 in normal healthy volunteers (NHV) and patients with systemic lupus erythematosus (SLE) METHODS: The drug was given intravenously to 11 NHV and six patients with SLE. Over 28 days, vital signs were noted, a haematological and chemical analysis of blood and urine was carried out, and adverse events were recorded. CR1 receptor numbers, the ability of antigen based heteropolymers to bind to red blood cells (RBCs), and the clearance of high avidity and total anti-dsDNA antibodies were measured by Farr assays and FACS analysis.

Results: No safety measure differed significantly from normal in both groups; no drug related serious adverse events occurred. ETI-104 rapidly bound to RBCs in NHV and patients with SLE. Binding of the drug to RBCs of patients with SLE also caused a rapid reduction of circulating anti-dsDNA antibodies in the plasma 15 minutes after administration, with a maximum reduction of 55% (range 43-62). At 28 days statistically significant decreases were maintained in three patients, while in the other three the values had returned to baseline levels.

Conclusion: These clinical trials established the safety and the proof of principle of the new immunoconjugate ETI-104. This provides the basis for further development of this technology for numerous indications-for example, therapeutic options for autoimmune diseases or viral and bacterial infections.

Figure 1

Scheme of ETI-104 bound to an RBC.

Figure 2

FACS analysis establishes binding of ETI-104 to RBCs of NHV. (A) Binding of ETI-104 to RBCs of each individual was confirmed by FACS analysis in vitro (goat antimouse for detection of the murine CR1 antibody and the PicoGreen DNA component of ETI-104). (B) RBCs from NHV 103 were bound in vitro to a saturating amount of Alexa-488 labelled anti-CR1 monoclonal antibody (mAb; 7G9) in the presence of varying concentrations of AHP. Increasing concentrations of AHP blocked 7G9 binding to the cells, demonstrating that AHP binds specifically to CR1. Thus, 250 ng AHP was not sufficient to block all available CR1. (C) RBCs from NHV 103 were stained after drug infusion to detect binding of the murine CR1 mAb (antimouse) and DNA (PicoGreen) components of ETI-104. The average of the geometric mean fluorescence intensity after subtracting non-specific fluorescence is given. The pattern of staining was similar in the other 10 NHV. All values were measured in duplicate.

Figure 3

FACS analysis establishes binding of ETI-104 and target anti-dsDNA antibodies to RBCs in patients with SLE in vivo. RBCs from patients were stained after drug infusion to detect binding of the murine mAb (antimouse) and DNA (PicoGreen) components of ETI-104 and binding of the target anti-dsDNA antibody (antihuman). Average of the geometric mean fluorescence intensity of stained samples, after subtracting non-specific fluorescence, at each time was normalised to the 15 minute post-infusion value, which was set at 100%. The patients shown here represent the three patterns of association of anti-dsDNA antibodies with the RBCs; anti-dsDNA antibody tracked with the murine mAb component of ETI-104 (patient 1), with the DNA component of ETI-104 (patient 5), or non-parallel to either ETI-104 component (patient 6). Patients 2 and 3 showed essentially the same pattern as patient 1 and patient 4 as patient 5 (not shown). All values were measured in duplicate.

Figure 4

Administration of ETI-104 reduces the high avidity anti-dsDNA antibodies (Farr activity) and total dsDNA antibodies (FACS analysis) from the circulation of patients with SLE. (A) High avidity anti-dsDNA antibodies (Farr titre) reductions from 15 minutes to 4 hours after ETI-104 administration. The Farr titre was reduced from the initial values by 43–62% at 15 minutes and by 15–62% at 4 hours after ETI-104 administration; it was maximally reduced in patient No 1 at this time. The reduction was ∼200 IU/ml in patients with starting titres of 306 and 484 IU/ml (starting titres of each patient are given in brackets). All Farr values were measured in quadruplicate. (B) Total anti-dsDNA antibody (FACS analysis) reductions from 15 minutes to 4 hours after ETI-104 administration. Patient plasma samples were incubated in vitro with naïve AHP opsonised RBCs (blood group O), washed, and stained with Alexa antihuman IgG and IgM antibody. Total dsDNA antibody levels were reduced from their initial value by 40–79% at 15 minutes and by 11–71% at 4 hours after ETI-104 administration. All FACS values were measured in duplicate.