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Clinical Trial
. 2021 Oct 27:12:778459.
doi: 10.3389/fimmu.2021.778459. eCollection 2021.

Radiotherapy and High-Dose Interleukin-2: Clinical and Immunological Results of a Proof of Principle Study in Metastatic Melanoma and Renal Cell Carcinoma

Jenny Bulgarelli  1 , Claudia Piccinini  1 , Elisabetta Petracci  2 , Elena Pancisi  1 , Anna Maria Granato  1 , Francesco de Rosa  1 , Massimo Guidoboni  1 , Massimiliano Petrini  1 , Valentina Ancarani  1 , Giovanni Foschi  1 , Antonino Romeo  3 , Luca Tontini  3 , Ugo De Giorgi  4 , Cristian Lolli  4 , Giorgia Gentili  2 , Linda Valmorri  2 , Alice Rossi  5 , Fabio Ferroni  5 , Carla Casadei  6 , Pietro Cortesi  7 , Laura Crudi  8 , Laura Ridolfi  1
Affiliations
Clinical Trial

Radiotherapy and High-Dose Interleukin-2: Clinical and Immunological Results of a Proof of Principle Study in Metastatic Melanoma and Renal Cell Carcinoma

Jenny Bulgarelli et al. Front Immunol. .

Abstract

High-dose interleukin-2 (HD IL-2) has curative potential in metastatic melanoma (MM) and renal cell carcinoma (RCC). Radiotherapy (RT) kills cancer cells and induces immunomodulatory effects. Prospective trials exploring clinical and immunological properties of combined RT/HD IL-2 are still needed. We designed a phase II, single-arm clinical trial for patients with MM and RCC. The treatment schedule consisted of 3 daily doses of 6-12 Gy of RT to 1-5 non-index metastatic fields, before IL-2 at the first and third treatment cycle. HD IL-2 was administered by continuous infusion for 72 hours and repeated every 3 weeks for up to 4 cycles, thereafter every 4 weeks for a maximum of 2 cycles. The primary endpoint was the immunological efficacy of the combined RT/HD IL-2 treatment (assessed by IFN-γ ELISPOT). Nineteen out of 22 patients were evaluable for immunological and clinical response. Partial response occurred in 3 (15.7%) patients and stable disease was observed in 7 (36.8%). The disease control rate was 52.6% after a median follow up of 39.2 months. According to Common Terminology Criteria for Adverse Events 4.0 (CTCAE 4.0), the majority of toxicities were grade 1-2. Immunological responses were frequent and detected in 16 (84.2%) patients. Increased levels of IL-8 and IL-10 in melanoma, circulating effector memory CD4+ and intratumoral CD8+ T cells in both tumor types were detected after therapy. Overall the treatment was well tolerated and immunologically active. Immunomonitoring and correlative data on tumor and peripheral blood cell subsets suggest that this combination treatment could be a promising strategy for patients progressing after standard treatments.

Keywords: IFN-γ ELISPOT assay; clinical immunomonitoring; high dose IL-2; metastatic melanoma; radiotherapy; renal cell carcinoma.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Schematic representation of the therapeutic schedule and biological sample collection. The therapeutic strategy was based on a sequence of therapeutic cycles performed every 3 weeks up to cycle 4 (C4) and every 4 weeks thereafter for a further 2 cycles, up to a maximum of 6 months of treatment. At the first (C1) and third (C3) cycle, the administration of high-dose interleukin-2 (HD IL-2) was preceded by radiotherapy. The second (C2), the fourth (C4), the fifth (C5) and the sixth (C6) cycles consisted in the administration of HD IL-2 only. Biological samples were collected at established times during the treatment.
Figure 2
Figure 2
Trend of immunological response to treatment. (A) Heatmaps highlighting the gradient of the immunological response to treatment represented as % of variation in the number of INF-γ SFCs for each TAA compared to baseline values. Patients are divided by tumor type (MM upper panel and RCC lower panel). For each tested TAA, immune responses ≥10% compared to baseline values are highlighted in red and the therapy cycle for which the best response was observed is reported. Immune responses <10% are shown in blue. (^ after the first RT cycle, ^^ after the second RT cycle). (B) The overall trend of the immunological response for each patient is shown in a spider plot (orange dots/line for MM and black dots/lines for RCC). Each dot represents the median variation in the number of INF-γ SFCs for each TAA, considering the baseline value as 0. Treatment cycles are indicated in the x axis.
Figure 3
Figure 3
Cytokines and proangiogenic factor modulation after treatment. (A) Dot plot with bar represents IL-8 concentration (pg/mL) in the entire patient cohort (left panel), in MM patients (middle panel) and in RCC patients (right panel). (B) Dot plot with bar represents IL-10 concentration (pg/mL) in the entire patient cohort (left panel), in MM patients (middle panel) and in RCC patients (right panel). Dot plots with bar represent pre- and post-therapy concentration levels (pg/mL) of (C) IL-1β, (D) IL-6, (E) TNF-α, (F) IL-12 p70, (G) VEGF and (H) fibronectin in ng/mL. Data represent individual values, mean (central bar) ± SEM (upper and lower bar). The black circular dots represent pre- therapy samples, while the red squares represent post-therapy samples. Statistical analysis was performed with the nonparametric Wilcoxon signed rank test; *p value < 0.05, **p value < 0.01.
Figure 4
Figure 4
Peripheral blood cell biomarkers of immune modulation. (A) Scatter plots with bars represent leukocyte, neutrophil, platelet, lymphocyte and monocyte counts (109/L) in peripheral blood collected before and after therapy. Data represent individual values, mean (central bar) ± SEM (upper and lower bar). The black circular dots represent pre- therapy samples, while the red squares represent post-therapy samples. (B) Boxes with floating bars represent neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR) and platelet-to-lymphocyte ratio (PLR). The black and red boxes represent pre- and post-therapy values, respectively. (C) Trend of the absolute count of lymphocytes and neutrophils (upper panel) and the NLR (lower panel) during treatment cycles for patient #002 (RCC, partial responder). The black line and dots refer to the lymphocyte count, the blue line and dots to the neutrophil count and the red line and dots to the NLR. Statistical analysis was performed with the nonparametric Wilcoxon signed rank test; *p value < 0.05.
Figure 5
Figure 5
CD8+ and CD4+ T cell subsets. A manual gating strategy was used to define T cell subsets within CD8+ and CD4+ lymphocytes. Briefly, naïve T cells were identified as CCR7+CD45RA+ (TN), central memory as CCR7+CD45RA- (TCM), effector memory as CCR7-CD45RA- (TEM) and effector as CCR7-CD45RA+ (TE). (A) Percentages of different CD8+ T cell subpopulations in pre- and post-treatment patient samples (n=19). (B) Percentages of different CD4+ T cell subpopulations in pre- and post-therapy patient samples (n=19). The black circular dots represent pre- therapy samples and the red squares represent post-therapy samples. (C) Scatter plots representing the percentage of Treg cells (CD3+CD4+CD25highFOXP3+) in the entire patient cohort (left panel), in MM patients (middle panel) and in RCC patients (right panel). The black circular dots represent pre-therapy samples, the orange triangles represent on-treatment samples (C3 or C4), the red squares represent post-therapy samples. Data represent individual values, mean (central bar) ± SEM (upper and lower bar). Statistical analysis was performed with the nonparametric Wilcoxon signed rank test; *p value < 0.05.
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