Immunological effects and activity of multiple doses of zolbetuximab in combination with zoledronic acid and interleukin-2 in a phase 1 study in patients with advanced gastric and gastroesophageal junction cancer

Abstract

Purpose: Zolbetuximab (IMAB362) is engineered to induce antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity. We evaluated ADCC activity and the impact of the immune-modulating drugs zoledronic acid (ZA) and interleukin-2 (IL-2) as co-treatment with zolbetuximab on relevant immune cell populations and ADCC lysis activity.

Methods: This phase 1, multicenter, open-label study investigated the immunological effects and activity, safety, tolerability, and antitumor activity of multiple doses of zolbetuximab alone (n = 5) or in combination with ZA (n = 7) or with ZA plus two different dose levels of IL-2 (low dose: 1 million international units [mIU] [n = 9]; intermediate dose: 3 mIU [n = 7]) in pretreated patients with advanced gastric and gastroesophageal junction (G/GEJ) adenocarcinoma.

Results: Twenty-eight patients with previously treated advanced G/GEJ adenocarcinoma that was CLDN18.2-expressing were enrolled into four treatment arms. Treatment with zolbetuximab + ZA + IL-2 induced short-lived expansion and activation of ADCC-mediating cell populations, namely γ9δ2 T cells and natural killer cells, within 2 days after administration; this effect was more pronounced with intermediate-dose IL-2. Expansion and activation of regulatory T cells treated with either IL2 dose was moderate and short-lived. Strong ADCC activity was observed with zolbetuximab alone. Short-lived ADCC activity was observed in several patients treated with ZA + intermediate-dose IL-2, but not lower-dose IL-2. In the clinical efficacy population, the best confirmed response was stable disease (n = 11/19; 58%).

Conclusions: Zolbetuximab mediates proficient ADCC in patients with pretreated advanced G/GEJ cancers. Co-treatment with ZA + IL-2 did not further improve this effect.

Trial registration: NCT01671774.

Keywords: Antibody-dependent cell-mediated cytotoxicity; Claudin 18.2; Esophagogastric junction cancer; Gastric cancer; Interleukin-2; Zolbetuximab.

Fig. 1

Definition of time frames for the evaluation of immunomodulatory effect of ZA/IL-2 in combination with zolbetuximab. D Day

Fig. 2

Kinetic profile of γ9δ2 T-cell frequency in lymphocytes. Kinetic profile of γ9δ2 T-cell frequency in lymphocytes. Peripheral blood mononuclear cells from patients were analyzed by flow cytometry at nine different time points (D1, D3, D8, D22, D43, D45, D50, D64, and D85). a Fold changes of median γ9δ2 T-cell frequencies (Vd2⁺CD3⁺ in lymphocytes) relative to pretreatment (D1). Arms 2 and 3 are shown in red; the lower error bar represents Q1 and the upper error bar Q3. b Median ratios in (b) were normalized to the median ratios of the control arm on the respective days. Red, bold font indicates the ZA and ZA + IL-2 treatment days. Table lists medians of Arms 1 to 3 vs. Arm 4 (control). Error bars represent mean ± SD. Arm 1: zolbetuximab + ZA; Arm 2: zolbetuximab + ZA + IL-2 (1 mIU); Arm 3: zolbetuximab + ZA + IL-2 (3 mIU); Arm 4: zolbetuximab. D Day, IL-2 interleukin 2, mIU million international units, SD standard deviation, ZA zoledronic acid

Fig. 3

Kinetic profile of activated CD69⁺ γ9δ2 T cells in lymphocytes. Peripheral blood mononuclear cells from patients were analyzed by flow cytometry at nine different time points (D1, D3, D8, D22, D43, D45, D50, D64, and D85). a Fold changes of the median frequencies of activated CD69⁺Vd2⁺CD3⁺. b Median values normalized to the median values of the control arm on the respective days. Medians of the same treatment arm at different time points are connected and the red, bold font indicates the ZA and ZA + IL-2 treatment days. Error bars represent mean ± SD. Arm 1: zolbetuximab + ZA; Arm 2: zolbetuximab + ZA + IL-2 (1 mIU); Arm 3: zolbetuximab + ZA + IL-2 (3 mIU); Arm 4: zolbetuximab. D Day, IL-2 interleukin 2, mIU million international units, SD standard deviation, ZA zoledronic acid

Fig. 4

Kinetic profile of activated CD69⁺ NK cells. Peripheral blood mononuclear cells from patients were analyzed by flow cytometry at nine different time points (D1, D3, D8, D22, D43, D45, D50, D64, and D85). a Median frequencies relative to pretreatment (D1) of activated NK cells (CD69⁺ in CD16⁺CD56⁺). Arms 2 and 3 are shown in red; the lower error bar represents Q1 and the upper error bar Q3. b Median ratios shown in (a) were normalized to the median ratios of control Arm 4 on respective days. Red, bold font indicates the ZA and ZA + IL-2 treatment days. Error bars represent mean ± SD. Arm 1: zolbetuximab + ZA; Arm 2: zolbetuximab + ZA + IL-2 (1 mIU); Arm 3: zolbetuximab + ZA + IL-2 (3 mIU); Arm 4: zolbetuximab. D Day, IL-2 interleukin 2, mIU million international units, NK natural killer, SD standard deviation, ZA zoledronic acid

Fig. 5

Kinetic profile of T_reg cell frequencies in T lymphocytes. Peripheral blood mononuclear cells from patients were analyzed by flow cytometry at nine different time points (D1, D3, D8, D22, D43, D45, D50, D64, and D85). Fold changes of the median frequencies of (CD3⁺CD4⁺CD25⁺FoxP3⁺CD127^–) T_reg cells (a), activated (FoxP3^highCD45RA⁻) T_reg cells (b), and non-suppressive (FoxP3^lowCD45RA⁻) T_reg cells (c) relative to respective medians of D1 are presented. Arms 2 and 3 are combined (red); the lower error bar represents Q1 and the upper error bar Q3. Red, bold font indicates ZA and ZA + IL-2 treatment days. Error bars represent mean ± SD. Arm 1: zolbetuximab + ZA; Arm 2: zolbetuximab + ZA + IL-2 (1 mIU); Arm 3: zolbetuximab + ZA + IL-2 (3 mIU); Arm 4: zolbetuximab. D Day, IL-2 interleukin 2, mIU million international units, SD standard deviation, ZA zoledronic acid

Fig. 6

ADCC lysis activity during the course of treatment. Peripheral blood mononuclear cells were purified from patient blood samples and zolbetuximab-specific ADCC was determined using NUGC-4 target cells. a Proportion of patients with measurable ADCC-specific lysis (lysis > 19%) for all treatment arms on all study days until Day 85. Bracketed numbers indicate maximum to minimum patient numbers during the study for each treatment arm. b Corresponding patient values (symbols) grouped by treatment arm. Each line represents a single patient. The shaded area < 19% indicates the minimum threshold for specific lysis. Red, bold font indicates the ZA and ZA + IL-2 treatment days. Arm 1: zolbetuximab + ZA; Arm 2: zolbetuximab + ZA + IL-2 (1 mIU); Arm 3: zolbetuximab + ZA + IL-2 (3 mIU); Arm 4: zolbetuximab. ADCC antibody-dependent cell-mediated cytotoxicity, IL-2 interleukin 2, mIU million international units, ZA zoledronic acid