Single low-dose rHuIL-12 safely triggers multilineage hematopoietic and immune-mediated effects

Abstract

Background: Recombinant human interleukin 12 (rHuIL-12) regulates hematopoiesis and cell-mediated immunity. Based on these hematopoietic and immunomodulatory activities, a recombinant human IL-12 (rHuIL-12) is now under development to address the unmet need for a medical countermeasure against the hematopoietic syndrome of the acute radiation syndrome (HSARS) that occurs in individuals exposed to lethal radiation, and also to serve as adjuvant therapy that could provide dual hematopoietic and immunotherapeutic benefits in patients with cancer receiving chemotherapy. We sought to demonstrate in healthy subjects the safety of rHuIL-12 at single, low doses that are appropriate for use as a medical countermeasure for humans exposed to lethal radiation and as an immunomodulatory anti-cancer agent.

Methods: Two placebo-controlled, double-blinded studies assessed the safety, tolerability, pharmacokinetics and pharmacodynamics of rHuIL-12. The first-in-human (FIH) dose-escalation study randomized subjects to single subcutaneous injections of placebo or rHuIL-12 at 2, 5, 10, and 20 μg doses. Due to toxicity, dose was reduced to 15 μg and then to 12 μg. The phase 1b expansion study randomized subjects to the highest safe and well tolerated dose of 12 μg.

Results: Thirty-two subjects were enrolled in the FIH study: 4 active and 2 placebo at rHuIL-12 doses of 2, 5, 10, 12, and 15 μg; 1 active and 1 placebo at 20 μg. Sixty subjects were enrolled in the expansion study: 48 active and 12 placebo at 12 μg dose of rHuIL-12. In both studies, the most common adverse events (AEs) related to rHuIL-12 were headache, dizziness, and chills. No immunogenicity was observed. Elimination of rHuIL-12 was biphasic, suggesting significant distribution into extravascular spaces. rHuIL-12 triggered transient changes in neutrophils, platelets, reticulocytes, lymphocytes, natural killer cells, and CD34+ hematopoietic progenitor cells, and induced increases in interferon-γ and C-X-C motif chemokine 10.

Conclusion: A single low dose of rHuIl-12 administered subcutaneously can elicit hematological and immune-mediated effects without undue toxicity. The safety and the potent multilineage hematopoietic/immunologic effects triggered by low-dose rHuIL-12 support the development of rHuIL-12 both as a radiation medical countermeasure and as adjuvant immunotherapy for cancer.

Trial registration: ClinicalTrials.gov: NCT01742221.

Figures

Figure 1

Plasma concentration profiles of rHuIL-12 over time. A) Plasma concentration vs. time for rHuIL-12 at 2, 5, 10, 12, 15 and 20 μg doses in the FIH study. B) Plasma concentration vs. time for rHuIL-12 (12 μg) and for IFN-γ and CXCL10 in subjects treated with rHuIL-12 at 12 μg in the phase 1b expansion study.

Figure 2

Transient hematological changes with different rHuIL-12 doses and placebo in the FIH study. The percentage changes from baseline count after treatment with rHuIL-12 (2, 5, 10, 12, 15, or 20 μg) or placebo are shown as follows: A) lymphocytes/rHuIL-12; B) lymphocytes/placebo; C) neutrophils/rHuIL-12; D) neutrophils/placebo; E) platelets/rHuIL-12; F) platelets/placebo; G) reticulocytes/rHuIL-12; H) reticulocytes/placebo.

Figure 3

Transient hematological changes with 12 μg dose of rHuIL-12 or placebo in phase 1b expansion study. Standard hematologic methods were used to determine cell counts at the indicated time points. The mean percentage of baseline count after treatment with rHuIL-12 (12 μg) or placebo are shown as follows: A) lymphocytes, neutrophils, platelets, and reticulocytes after treatment with rHuIL-12; B) lymphocytes, neutrophils, platelets, and reticulocytes after treatment with placebo; C) CD45+, CD3+, CD4+ and CD8+ cells after rHuIL-12; D) CD45+, CD3+, CD4+ and CD8+ cells after rHuIL-12; D) NK and CD34+ after placebo; E) NK and CD34+ cells after rHuIL-12; F) NK and CD34+ cells after placebo. NK cells were defined as CD45 + CD16 + CD56+ triple positive lymphocytes.

Figure 4

Effect of rHuIL-12 or Placebo on rHuIL-12Rβ2 Positivity and CD56 Mean Fluorescence Intensity in the Phase 1b Expansion Study. Flow cytometry was used to determine A) changes in the percentage of rHuIL-12Rβ2 positive NK and CD34+ cells in response to rHuIL-12 (12 μg) or placebo; and B) changes in CD56 mean fluorescence intensity on NK cells in response to dose of rHuIL-12 (12 μg) or placebo. NK cells were defined as CD45 + CD16 + CD56+ triple positive lymphocytes.

Figure 5

Effect of rHuIL-12 or Placebo on EPO, IL-18 and CXCL10 levels in the Phase 1b expansion study. EPO, IL-18, and CXCL10 were measured at the indicated time points using validated assays. The mean changes from baseline after treatment with rHuIL-12 (12 μg) or placebo are shown as follows: A) EPO; B) IL-18; C) CXCL10.

Figure 6

Gating strategy for flow cytometry in Phase 1 b expansion study.