First-in-Human Phase I Trial of a Tumor-Targeted Cytokine (NHS-IL12) in Subjects with Metastatic Solid Tumors

Abstract

Purpose: The NHS-IL12 immunocytokine is composed of two IL12 heterodimers fused to the NHS76 antibody. Preclinical studies have shown that this antibody targets IL12 to regions of tumor necrosis by binding histones on free DNA fragments in these areas, resulting in enhanced antitumor activity. The objectives of this phase I study were to determine the maximum tolerated dose (MTD) and pharmacokinetics of NHS-IL12 in subjects with advanced solid tumors.

Patients and methods: Subjects (n = 59) were treated subcutaneously with NHS-IL12 in a single ascending-dose cohort followed by a multiple ascending-dose cohort (n = 37 with every 4-week dosing).

Results: The most frequently observed treatment-related adverse events (TRAE) included decreased circulating lymphocytes, increased liver transaminases, and flu-like symptoms. Of the grade ≥3 TRAEs, all were transient and only one was symptomatic (hyperhidrosis). The MTD is 16.8 μg/kg. A time-dependent rise in IFNγ and an associated rise in IL10 were observed following NHS-IL12. Of peripheral immune cell subsets evaluated, most noticeable were increases in frequencies of activated and mature natural killer (NK) cells and NKT cells. Based on T-cell receptor sequencing analysis, increases in T-cell receptor diversity and tumor-infiltrating lymphocyte density were observed after treatment where both biopsies and peripheral blood mononuclear cells were available. Although no objective tumor responses were observed, 5 subjects had durable stable disease (range, 6-30+ months).

Conclusions: NHS-IL12 was well tolerated up to a dose of 16.8 μg/kg, which is the recommended phase II dose. Early clinical immune-related activity warrants further studies, including combination with immune checkpoint inhibitors.See related commentary by Lyerly et al., p. 9.

Figure 1.

A) Pharmacokinetic and pharmacodynamic parameters for NHS-IL12 dosed at the MTD of 16.8 µg/kg every 4 weeks. A time-dependent rise in IFN-γ was noted after administration and a subsequent rise of IL-10. These corresponded to a rise in IL-12 similar to what was seen with the PK values, peaking around 36 hours, and then falling to near baseline levels around day 8. When subjects were re-dosed at 4 weeks, in most cases, despite a similar rise in the serum IL-12, a diminished rise was noted in IFN-γ and IL-10 with the second dose. B) Analysis of serum levels of IP-10 following treatment with NHS-IL12 at different dose levels. Serum levels of IP-10 were measured by ELISA pre‒ and at multiple time points post‒NHS-IL12.

Figure 1.

A) Pharmacokinetic and pharmacodynamic parameters for NHS-IL12 dosed at the MTD of 16.8 µg/kg every 4 weeks. A time-dependent rise in IFN-γ was noted after administration and a subsequent rise of IL-10. These corresponded to a rise in IL-12 similar to what was seen with the PK values, peaking around 36 hours, and then falling to near baseline levels around day 8. When subjects were re-dosed at 4 weeks, in most cases, despite a similar rise in the serum IL-12, a diminished rise was noted in IFN-γ and IL-10 with the second dose. B) Analysis of serum levels of IP-10 following treatment with NHS-IL12 at different dose levels. Serum levels of IP-10 were measured by ELISA pre‒ and at multiple time points post‒NHS-IL12.

Figure 2.

The frequencies of five refined immune cell subsets changed significantly 1 week post‒cycle 1 vs. pre‒NHS-IL-12 treatment for 10 subjects at dose level 8 (16.8 µg/kg). Flow cytometry was performed to evaluate 123 discrete immune cell subsets, including nine standard subsets and 114 refined subsets A). All values shown are the frequency of the subset out of all PBMCs shown as median (IQR). B-D) Graphs depicting three of these subsets.

Figure 3.

Effect of NHS-IL12 on remodeling of T-cell repertoire in four subjects from dose level 8 expansion with varying IFN-γ responses. A) Subject characteristics, peak IFN-γ levels, and tissues assayed by TCRseq are indicated. B and C) TCRβ clonotype frequency plots are depicted from the tumor biopsy (B) and PBMCs (C) of subject 58, before and day 35 after treatment with NHS-IL12. Clones remaining stable in abundance are found on the X=Y diagonal, while those that increase or decrease following treatment are found above or below the diagonal, respectively. D) TCR diversity, measured by the metric of repertoire size, in the biopsy of subjects before and after NHS-IL12. Values indicate the number of individual clonotypes comprising the top 25^th percentile by ranked molecule count after sorting by abundance. E) Overall TIL density calculated from TCRseq data using the formula: TIL density = (# Productive Templates) / (DNA input (pg) / 6.6). This formula is based on the human genome weighing 6.6 pg/cell.