Intermediate-affinity Interleukin-2 Receptor Research Articles

Abstract A099: Preliminary pharmacodynamic characterization in patients with platinum-resistant ovarian cancer treated with nemvaleukin in combination with pembrolizumab

Abstract A099: Preliminary pharmacodynamic characterization in patients with platinum-resistant ovarian cancer treated with nemvaleukin in combination with pembrolizumab

ARTISTRY-6: Nemvaleukin alfa monotherapy in patients with advanced mucosal and cutaneous melanoma.

TPS9592 Background: Despite improved outcomes in melanoma with the introduction of checkpoint inhibitors (CPIs), ≈50% of patients do not respond. A subset of responders ultimately progress and have limited treatment options, underscoring a high unmet need for novel treatments with durable benefit. Patients with mucosal melanoma exhibit response rates and progression-free survival ≈2 times lower than those with cutaneous melanoma. Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel, engineered cytokine that selectively binds to the intermediate-affinity interleukin-2 receptor complex to preferentially activate CD8+ T and natural killer cells, with minimal expansion of regulatory T cells. Nemvaleukin has been granted Orphan Drug designation for the treatment of mucosal melanoma by the US FDA. In the ARTISTRY-1 study, the intravenous (IV) recommended phase 2 dose (RP2D) for nemvaleukin monotherapy (6 µg/kg on days 1-5 of a 21-day cycle) demonstrated durable antitumor activity in patients with advanced melanoma, including mucosal melanoma, previously treated with a CPI (N = 46, overall response rate [ORR] 13.0% [95% CI 4.9-26.3], median duration of response 8.1 weeks [range 6.1-79.0]). In the ARTISTRY-2 study, the subcutaneous (SC) RP2D was identified as 3 mg every 7 days, which demonstrated pharmacodynamic effects consistent with those of IV delivery. Data from these studies support further evaluation of nemvaleukin monotherapy among patients with advanced mucosal or cutaneous melanoma. Additionally, a less frequent IV nemvaleukin dosing schedule is being evaluated in patients with advanced solid tumors in the ARTISTRY-3 study. Methods: ARTISTRY-6 (NCT04830124) is an ongoing phase 2, global, multicenter, open-label study that is currently enrolling. Eligible patients will have had prior anti–PD-(L)1 therapy with or without anti–CTLA-4 therapy, an Eastern Cooperative Oncology Group performance status of 0 or 1, and adequate hematologic reserve and hepatic and renal function. In Cohort 1, patients with advanced cutaneous melanoma will receive nemvaleukin at the SC RP2D of 3 mg every 7 days. In Cohort 2, patients with advanced mucosal melanoma will receive nemvaleukin at the IV RP2D of 6 µg/kg on days 1-5 of a 21-day cycle. In Cohort 3 (added as a protocol amendment), patients with advanced cutaneous melanoma will receive IV nemvaleukin in 1 of 2 less frequent dosing schedules of 1 or 2 doses per cycle. The dosing schedules for Cohort 3 will be determined when a less frequent IV RP2D has been established in ARTISTRY-3. Patients will receive nemvaleukin until progression or intolerable toxicity. The primary objective is to evaluate the antitumor activity of nemvaleukin monotherapy defined by ORR. Additional objectives include evaluation of safety, health-related quality of life, predictive biomarkers, pharmacokinetics, immunogenicity, and pharmacodynamic effects. Clinical trial information: NCT04830124 .

Abstract CT133: Trial in progress: a multicenter phase 1/1b dose escalation study of WTX-124 as a monotherapy and in combination with pembrolizumab in patients with selected advanced or metastatic solid tumors

Abstract Background: High-dose recombinant human IL-2 (HD IL-2) is an FDA-approved therapy that produces durable complete remissions in a subset of patients with advanced or metastatic cutaneous melanoma and renal cell carcinoma. However, the clinical benefit of HD IL-2 is counterbalanced by life-threatening toxicities such as capillary leak syndrome. WTX-124 is an engineered cytokine prodrug composed of a wild-type IL-2 fused via proprietary cleavable linkers to an inactivation domain and to a half-life extension domain. In the circulation and in normal tissues, WTX-124 is designed to remain inactive and not bind to high- (αβγ) and intermediate-affinity (βγ) IL-2 receptors. In contrast, proteolytic activation of WTX-124 in the tumor microenvironment (TME) is expected to liberate the IL-2 cytokine to stimulate antitumor immune responses. Preclinical data show that WTX-124 is activated by a range of dissociated human tumors in vitro but not by normal human cells or by patient serum. Moreover, WTX-124 demonstrates potent antitumor activity in multiple murine syngeneic tumor models with substantial expansion and activation of tumor-infiltrating lymphocytes. In the MC38 model, WTX-124 has a large therapeutic window of >19-fold compared to ~3-fold for recombinant human IL-2. Methods: This first-in-human, open-label phase 1/1b trial is investigating the safety, tolerability, pharmacokinetics, pharmacodynamics, immunogenicity, and antitumor activity of WTX-124 administered as a monotherapy or in combination with the anti-PD-1 antibody pembrolizumab to patients with advanced or metastatic solid tumors (NCT05479812). All patients must have ≥ 1 measurable lesion per RECIST 1.1 and a tumor type for which immune checkpoint inhibitor (ICI) therapy is indicated. Dose escalation uses a modified toxicity probability interval-2 study design with WTX-124 administered intravenously (IV) every two weeks in 28-day cycles. If the safety of WTX-124 as a monotherapy is established in the first three cohorts, the study will then initiate dose escalation of WTX-124 in combination with pembrolizumab 400mg IV every six weeks. Dose expansion is anticipated to include four arms that will enroll patients with either advanced or metastatic cutaneous melanoma or renal cell carcinoma to receive WTX-124 monotherapy or combination therapy (N=20 patients per arm). Patients with melanoma assigned to the WTX-124/pembrolizumab combination arm may be naïve to all prior systemic therapy for advanced disease, but all other patients in expansion must have previously received a standard of care ICI regimen. Pre- and on-treatment tumor biopsies, archival tumor tissue, and blood samples will be used to investigate multiple exploratory biomarkers. As of January 12, 2023, WTX-124 monotherapy dose escalation is proceeding. Citation Format: Ildefonso Ismael Rodriguez-Rivera, Justin C. Moser, Mateusz Opyrchal, Brendan Curti, Mehmet A. Bilen, Saero Park, Marissa Bruno, Paul Windt, Kulandayan K. Subramanian, Sameer S. Chopra, Randi Isaacs. Trial in progress: a multicenter phase 1/1b dose escalation study of WTX-124 as a monotherapy and in combination with pembrolizumab in patients with selected advanced or metastatic solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT133.

Abstract 1801: STK-012, an a/b-selective IL-2 activates tumor antigen specific CD25+ CD8 T cells to reject tumors without acute vascular toxicity

Abstract High-dose Interleukin-2 (IL-2) monotherapy induces complete responses in cancer patients but its use is limited by acute vascular toxicities including capillary leak syndrome and severe hypotension 1,2. IL-2 activates lymphocytes and NK-cells through the intermediate-affinity dimeric IL-2 receptor, IL-2Rβ/γ (CD122/CD132), while antigen activated T-cells and regulatory T-cells (Tregs) have increased sensitivity to IL-2 by expressing the high-affinity trimeric IL-2 receptor, IL-2Rα/β/γ (CD25/CD122/CD132)3. CD25-independent IL-2s (“non-α-IL-2s”) aim to increase the therapeutic efficacy of IL2 in cancer patients by avoiding Treg activation through selective binding to IL-2Rβ/γ 4. However, those molecules still are reported to induce fever and hypotension and have limited efficacy as a monotherapy or in combination with anti-PD-1 5,6. Here we show that a novel α/β-IL-2 agonist that was designed to preferentially bind to the IL-2Rα/β/γ receptor highly upregulated on antigen activated T-cells can greatly improve on the efficacy of IL-2 while avoiding the vascular toxicity commonly associated with IL-2 treatment. In syngeneic tumor models, this α/β-IL-2 agonist significantly reduced exhaustion of tumor infiltrating T cells compared to WT-IL-2 or a non-α-IL-2 leading to improved expansion of tumor antigen specific CD25+PD-1+CD8+ T cells systemically and in the tumor microenvironment. This resulted in complete responses and tumor immune memory with α/β-IL-2 monotherapy as well as improved outcomes in combination with anti-PD-1 therapy in PD-1 refractory syngeneic tumors. In contrast, WT-IL-2 reduced T cell exhaustion and drove antigen specific T cell responses to a lesser degree, resulting in reduced combinatorial efficacy with anti-PD-1, while the non-α-IL-2 failed to do either. Furthermore, the α/β-IL-2 agonist reduced intratumoral Tregs compared to treatment with WT-IL-2 or PBS improving the intratumoral CD8 to Treg ratio. In non-human primates and mice, WT-IL-2 and a non-α-IL-2 led to broad extravasation of lymphocytes and NK cells and activation of intra-pulmonal T cells resulting in systemic tissue inflammation and NK cell-mediated lethal capillary leak syndrome whereas the α/β-IL-2 agonist, which avoids binding the dimeric IL-2Rβ/γ expressed on NK cells, avoided systemic lymphocyte activation which facilitated continuous treatment without acute vascular toxicities. Overall, through selective engagement of CD25+ T cells, this α/β-IL-2 agonist demonstrated improved efficacy and tolerability of IL-2 in preclinical tumor models. Clinical trials with STK-012, a human α/β-IL-2 agonist, are in progress.1 Atkins, et al.; JCO 1999, 2 Dutcher, et al.; JITC 2014, 3 Liao, et al.; Immunity 2013, 4 Levin, et al.; Nature 2012, 5 Janku, et al.; Cancer Research 2021; 6 Diab, et al.; Cancer Disc. 2020 Citation Format: Martin Oft, Navneet Ratti, Sandro Vivona, Jan Emmerich, Romina Riener, Ievgen O. Koliesnik, Scott McCauley, Michele Bauer, Marie Semana, Deepti Rokkam, Bhargavi Jayaraman, Rene de Waal Malefyt, Paul-Joseph Aspuria, Michael Totagrande, Anita Mehta-Damani, Patrick J. Lupardus, Rob A. Kastelein. STK-012, an a/b-selective IL-2 activates tumor antigen specific CD25+ CD8 T cells to reject tumors without acute vascular toxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1801.

Selective Agonism of Intermediate-Affinity IL-2 Receptor Promotes Systemic Antitumor Responses in Combination with Radiotherapy in Metastatic Lung Cancer

<h3>Purpose/Objective(s)</h3> For immune checkpoint inhibitor-resistant patients, adding interleukin-2 (IL-2) may aid antitumor responses by stimulating effector T cells and NK cells. Unfortunately, high-dose IL-2 therapy has limited clinical utility due to adverse events and stimulation of immunosuppressive regulatory T cells (T_regs). Nemvaleukin alfa (nemvaleukin) is a novel engineered cytokine that selectively binds to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8⁺ T and NK cells, with minimal expansion of T_regs. Thus, Nemvaleukin may be beneficial in combination with radiation therapy (RT) and immunotherapy. This study aimed to evaluate the antitumor activity and mechanism of action of the combination of RDB 1462 (the murine ortholog of Nemvaleukin, a.k.a. mNemvaleukin), RT, and immunotherapy in multiple mouse tumor models. <h3>Materials/Methods</h3> In a bilateral 344SQ lung adenocarcinoma murine model, primary tumors were treated with high-dose RT (HDRT; 12 Gy per fraction for 3 fractions), and secondary tumors were observed for evidence of abscopal effects. In another model for low absolute lymphocyte count (ALC), we used 2 Gy total body irradiation (TBI) three days before HDRT and RDB 1462. In a third MC38-gp100 mouse model, tumors were irradiated with low-dose RT (LDRT; 3 fractions of 1 Gy each) followed by pmel-1 T cell infusion after 24 hours, and RDB 1462 administration 12 hours thereafter. We performed immune phenotyping by flow cytometry; analyzed 770 immune-related genes using a multiplex analysis platform; also performed T cell receptor (TCR) repertoire analysis. Serum pro-inflammatory cytokine markers were analyzed by 23-plex kit. <h3>Results</h3> In 344SQ-parental lung adenocarcinoma, we combined HDRT with escalating doses of RDB 1462 at various timepoints, resulting in significant tumor reduction in both irradiated and unirradiated (abscopal) tumors compared with HDRT or RDB 1462 therapy alone. HDRT+RDB 1462 increased the numbers and activity of tumor-specific CD8⁺ T cells, NK cells, and memory T cells without drastically increasing T_regs, as evidenced by peripheral and tumor-infiltrating immune phenotypic and functional analyses. T cells showed increased receptor clonality and diversity in groups receiving HDRT+RDB 1462. The addition of anti-PD-1 to HDRT+RDB 1462 further increased antitumor control of both irradiated and abscopal tumors, reduced lung metastases, and prolonged survival. The antitumor effects of HDRT and RDB 1462 persisted even in mice with low ALC. The addition of LDRT contributed to increased tumor-specific T cells that were expanded by RDB 1462 with increased infiltration of Thy1.1⁺ CD8⁺ T cells. <h3>Conclusion</h3> The addition of Nemvaleukin therapy may enhance responses to RT alone and in combination with anti-PD-1 or adoptive T cell therapy.

IL-2K35C-moFA, a Long-Acting Engineered Cytokine with Decreased Interleukin 2 Receptor α Binding, Improved the Cellular Selectivity Profile and Antitumor Efficacy in a Mouse Tumor Model.

Simple SummaryThe application of IL-2 for treating cancer is limited owing to its toxicity and short half-life. Its high binding ability to IL-2 receptor α expands immunosuppressive Treg cells, which represents an undesirable toxicity in cancer immunotherapy. Moreover, its small molecular size is the reason for its short half-life. We solved these problems by using a covalent modification strategy of IL-2 variant IL-2K35C with fatty acid by maleimide chemistry, namely, IL-2K35C-moFA. The experiments performed in vitro and in vivo proved that IL-2K35C-moFA is a novel immunotherapeutic agent with the potential to selectively stimulate CD8+ T cells and NK cells. Compared to IL-2WT, IL-2K35C-moFA showed a specifically reduced potency for the stimulation of Treg cells. Our results also showed that fatty acid conjugation appears to be effective in half-life extension. The combination of selective lymphocyte expansion and its long half-life means IL-2K35C-moFA should be evaluated as a potential human immunotherapeutic in the future.Human interleukin 2 (IL-2) has shown impressive results as a therapeutic agent for cancer. However, IL-2-based cancer therapy is limited by strong Treg amplification owing to its high binding affinity to IL-2 receptor α (IL-2Rα) and its short half-life owing to its small molecular size. In this study, we solved these problems using a covalent modification strategy of the IL-2 variant, i.e., substituting cysteine (C) for lysine (K) at position 35, using octadecanedicarboxylic acid through maleimide chemistry, creating IL-2K35C-moFA. IL-2K35C-moFA was equipotent to human IL-2 wild type (IL-2WT) in activating tumor-killing CD8+ memory effector T cells (CD8+ T) and NK cells bearing the intermediate affinity IL-2 receptors, and less potent than IL-2WT on CTLL-2 cells bearing the high-affinity IL-2 receptors. Moreover, it was shown to support the preferential activation of IL-2 receptor β (IL-2Rβ) over IL-2Rα because of the mutation and fatty acid conjugation. In a B16F10 murine tumor model, IL-2K35C-moFA showed efficacy as a single dose and provided durable immunity for 1 week. Our results support the further evaluation of IL-2K35C-moFA as a novel cancer immunotherapy.

ARTISTRY-7: A phase 3, multicenter study of nemvaleukin alfa, a novel engineered cytokine, in combination with pembrolizumab versus chemotherapy in patients with platinum-resistant epithelial ovarian, fallopian tube, or primary peritoneal cancer (326)

Objectives: ARTISTRY-7 will evaluate the novel engineered cytokine nemvaleukin alfa (nemvaleukin, ALKS 4230) in patients (pts) with gynecologic cancers. Nemvaleukin was designed to selectively bind to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8+ T and NK cells with minimal expansion of Tregs. This selectivity may enhance tumor killing and improve safety/tolerability compared with high-dose IL-2. In clinical studies, nemvaleukin monotherapy and combination with pembrolizumab have shown evidence of clinical benefit in multiple tumor types. In ARTISTRY-1, four responses are observed in pts with ovarian cancer (OC), including one complete response in a pt with platinum-resistant OC and five prior lines of therapy who remains on treatment. Epithelial OC is the 7th most common cause of cancer mortality in women. The standard frontline treatment is platinum-based chemotherapy, to which many pts become resistant or are refractory. There is a high unmet medical need for platinum-resistant OC. Methods: ARTISTRY-7 is a phase III, multicenter, open-label randomized study of nemvaleukin and/or pembrolizumab versus chemotherapy (ClinicalTrials.gov registration forthcoming). Eligible pts included females (>18 y) with histologically confirmed epithelial OC (high-grade serous, endometrioid, clear cell), fallopian tube cancer, or primary peritoneal cancer. Pts must have received >1 prior line of systemic anticancer therapy in the platinum-sensitive setting, ≤5 prior lines in the platinum-resistant setting, and prior bevacizumab, with evidence of radiographic progression on most recent therapy. Primary platinum-refractory disease (progression during first-line platinum-based therapy) or primary platinum resistance (progression <3 months after completion of first-line platinum-based therapy) is exclusionary. Pts must have an ECOG performance status of 0 or 1, an estimated life expectancy of >3 months, and adequate hematologic reserve and hepatic and renal function. Approximately 376 pts will be randomized (3:1:1:3) to receive intravenous nemvaleukin (6 μg/kg) on days 1-5 and pembrolizumab (200 mg) on day 1 of each 21-day cycle, pembrolizumab monotherapy, nemvaleukin monotherapy, or investigator’s choice chemotherapy (pegylated liposomal doxorubicin, paclitaxel, topotecan, or gemcitabine) and stratified according to PD-L1 status, histologic subtype (high-grade vs non-high-grade serous), and chemotherapy (paclitaxel vs other). Pts will continue treatment in the absence of disease progression or intolerable toxicity (maximum 35 cycles for pembrolizumab; nemvaleukin can be continued). Investigator-assessed progression-free survival (RECIST v1.1) in pts treated with nemvaleukin/pembroli- zumab versus chemotherapy (primary endpoint), characterization of antitumor activity of nemvaleukin and pembrolizumab in combination and as monotherapy, safety, HRQOL, and pharmacokinetic/ pharmacodynamic effects will be collected. Objectives: ARTISTRY-7 will evaluate the novel engineered cytokine nemvaleukin alfa (nemvaleukin, ALKS 4230) in patients (pts) with gynecologic cancers. Nemvaleukin was designed to selectively bind to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8+ T and NK cells with minimal expansion of Tregs. This selectivity may enhance tumor killing and improve safety/tolerability compared with high-dose IL-2. In clinical studies, nemvaleukin monotherapy and combination with pembrolizumab have shown evidence of clinical benefit in multiple tumor types. In ARTISTRY-1, four responses are observed in pts with ovarian cancer (OC), including one complete response in a pt with platinum-resistant OC and five prior lines of therapy who remains on treatment. Epithelial OC is the 7th most common cause of cancer mortality in women. The standard frontline treatment is platinum-based chemotherapy, to which many pts become resistant or are refractory. There is a high unmet medical need for platinum-resistant OC. Methods: ARTISTRY-7 is a phase III, multicenter, open-label randomized study of nemvaleukin and/or pembrolizumab versus chemotherapy (ClinicalTrials.gov registration forthcoming). Eligible pts included females (>18 y) with histologically confirmed epithelial OC (high-grade serous, endometrioid, clear cell), fallopian tube cancer, or primary peritoneal cancer. Pts must have received >1 prior line of systemic anticancer therapy in the platinum-sensitive setting, ≤5 prior lines in the platinum-resistant setting, and prior bevacizumab, with evidence of radiographic progression on most recent therapy. Primary platinum-refractory disease (progression during first-line platinum-based therapy) or primary platinum resistance (progression <3 months after completion of first-line platinum-based therapy) is exclusionary. Pts must have an ECOG performance status of 0 or 1, an estimated life expectancy of >3 months, and adequate hematologic reserve and hepatic and renal function. Approximately 376 pts will be randomized (3:1:1:3) to receive intravenous nemvaleukin (6 μg/kg) on days 1-5 and pembrolizumab (200 mg) on day 1 of each 21-day cycle, pembrolizumab monotherapy, nemvaleukin monotherapy, or investigator’s choice chemotherapy (pegylated liposomal doxorubicin, paclitaxel, topotecan, or gemcitabine) and stratified according to PD-L1 status, histologic subtype (high-grade vs non-high-grade serous), and chemotherapy (paclitaxel vs other). Pts will continue treatment in the absence of disease progression or intolerable toxicity (maximum 35 cycles for pembrolizumab; nemvaleukin can be continued). Investigator-assessed progression-free survival (RECIST v1.1) in pts treated with nemvaleukin/pembroli- zumab versus chemotherapy (primary endpoint), characterization of antitumor activity of nemvaleukin and pembrolizumab in combination and as monotherapy, safety, HRQOL, and pharmacokinetic/ pharmacodynamic effects will be collected.

Clinical outcomes of ovarian cancer patients treated with the novel engineered cytokine nemvaleukin alfa in combination with the PD-1 inhibitor pembrolizumab: recent data from ARTISTRY-1 (077)

<b>Objectives:</b> ARTISTRY-1 (NCT02799095) is an ongoing multi-cohort study evaluating nemvaleukin alfa (nemvaleukin, ALKS 4230) alone and with pembrolizumab in patients (pts) with advanced solid tumors, including those with platinum-resistant ovarian cancer (PROC). The standard frontline treatment for OC is platinum-based chemotherapy, to which many tumors become resistant or are refractory. Conventional therapies in the platinum-resistant setting offer a low likelihood of durable response, with a median progression-free survival of ~3.5 months. Nemvaleukin is a novel engineered cytokine that selectively binds to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8⁺ T and NK cells, with minimal expansion of regulatory T cells. Recent data support the potential for nemvaleukin to work with other agents to enhance antitumor activity in tumors with high unmet medical needs such as PROC. <b>Methods:</b> Pts with OC who had progressed on prior therapy were enrolled into a cohort of mixed tumor types that were not otherwise approved for anti-PD-L1 treatment (Cohort 1). Pts received intravenous nemvaleukin (3 μg/kg) on days 1-5 and pembrolizumab (200 mg) on day 1 of each 21-day cycle. Outcomes include antitumor activity via RECIST v1.1, safety, and description of CD8⁺ T and NK cell populations as of August 2021. <b>Results:</b> Fifteen pts with OC were enrolled in Cohort 1 of ARTISTRY-1. Pts were aged 48-83 years and were mostly White (<i>n</i>=11), had an ECOG performance status of 0 (<i>n</i>=4) or 1 (<i>n</i>=11), and had a median of four prior lines of therapy (range: 2-7), including PARP (<i>n</i>=6) and checkpoint (<i>n</i>=4) inhibitors and bevacizumab (<i>n</i>=9). All pts had previously received platinum-based therapy. Five pts with PROC—of whom four were on treatment >1 y—experienced clinically meaningful benefit: one confirmed complete response, three partial responses (two confirmed, one unconfirmed), and one pt with stable disease on treatment for ~1.5 y. In all evaluable pts with OC, the objective response rate was 28.6%, and the clinical benefit rate was 71.4%. Frequent (≥40%) treatment-related adverse events (AEs) in the full cohort (<i>n</i>=42) were chills (78.6%), pyrexia (71.4%), and fatigue (45.2%). Among pts with OC (<i>n</i>=15), grade ≥3 treatment-related AEs occurring in ≥10% were anemia (<i>n</i>=6; 40.0%) and fatigue (<i>n</i>=3; 20.0%); one pt had grade 3 CRS requiring dose reduction. Two serious AEs (grade 3 fatigue possibly related to treatment and grade 3 lethargy not related to treatment) led to treatment discontinuation in pts with OC; no AEs led to death. Expansions of CD8⁺ T cells (~2-fold) and NK cells (near 5-fold) were observed. <b>Conclusions:</b> Nemvaleukin/pembrolizumab was generally well-tolerated and provided evidence of tumor response and disease control in pts with heavily pretreated PROC. A phase III study (ARTISTRY-7) is initiated in pts with PROC to further evaluate this combination, representing a new therapeutic option for these pts. Exploration of other treatment combinations is warranted.

Abstract 4225: IL-2Rβ/IL-2R γsynthetic cytokines induce activation of human T and NK cells

Abstract Introduction: Heterodimerization of the intermediate affinity Interleukin-2 Receptors (IL-2R), IL-2Rβ and IL-2Rγ, initiates a signaling cascade in T and NK cells that ultimately results in proliferation and production of Interferon-gamma (IFN-γ). Binding of IL-2 in a dimeric complex with intermediate affinity or in the high affinity trimeric complex, which also includes IL-2Rα and is predominantly expressed on antigen activated T cells and Tregs leads to trans-phosphorylation of signaling motifs on the IL-2Rβ and IL-2Rγ intracellular domains by the associated JAK kinases. The function of the wild type IL-2 is to bring the receptor chains in close proximity to produce an optimal level of phosphorylation and activation of associated STAT transcription factors. Here we established a way to bring IL-2Rβ and IL-2Rγ chains together in a cytokine-independent manner that results in functional activation. Experimental Procedures: Human IL-2Rβ and IL-2Rγ specific heavy chain single domain antibodies (VHH) were generated by camel immunization and screening of VHH libraries prepared from peripheral blood cells for binding. Ten IL-2Rβ VHH segments and six IL-2Rγ VHH segments, all with low nanomolar affinity, were identified and coupled as IL-2Rβ/IL-2Rγ VHH dimers in all possible combinations in both amino-carboxy and carboxy-amino orientations yielding 120 different proteins that were tested in human T and NK cell functional assays. Results: IL-2Rβ/IL-2Rγ VHH dimers were biologically active and induced pSTAT-5 phosphorylation in the IL-2 dependent NK cell line NKL at various levels. The biological activity of these IL-2Rβ/IL-2Rγ VHH dimers was further confirmed on primary cells. IL-2Rβ/IL-2Rγ VHH dimers induced pSTAT5 phosphorylation on NK cells isolated from human peripheral blood and culture with IL-2Rβ/IL-2Rγ VHH dimers resulted in proliferation and production of high but varied levels of IFN-γ. The activity of IL-2Rβ/IL-2Rγ VHH dimers on primary T cells was also examined. Similar to NK cells, IL-2Rβ/IL-2Rγ VHH dimers induced pSTAT5 phosphorylation, proliferation and IFN-γ production by CD4 positive and CD8 positive T cell blasts generated after CD3/CD28 activation of human PBMC. Conclusions: In conclusion, we have generated a series of functional IL-2Rβ/IL-2Rγ VHH dimers representing surrogate cytokine agonists each with unique signaling strengths that will now be further analyzed for potential therapeutic application in tumor immunology and auto-immunity Citation Format: Rene de Waal Malefyt, PJ Aspuria, Sandro Vivona, Priyanka Balasubrahmanyam, Cindy Buffone, Mahalakshmi Ramadass, Romina Riener, Martin Oft, Patrick Lupardus, Robert Kastelein. IL-2Rβ/IL-2R γsynthetic cytokines induce activation of human T and NK cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4225.

Nemvaleukin alfa monotherapy and in combination with pembrolizumab in patients (pts) with advanced solid tumors: ARTISTRY-1.

2500 Background: Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel, engineered cytokine that selectively binds to the intermediate-affinity interleukin-2 (IL-2) receptor (IL-2R) to preferentially activate antitumor CD8+ T cells and natural killer (NK) cells with minimal expansion of immunosuppressive regulatory T cells. It is sterically occluded from binding to the high-affinity IL-2R, leveraging antitumor effects of the IL-2 pathway but mitigating toxicity associated with preferential binding of IL-2 to the high-affinity IL-2R. Methods: ARTISTRY-1 (NCT02799095) is a 3-part, first-in-human, phase 1/2 study of IV nemvaleukin alone and in combination with pembrolizumab in pts with advanced solid tumors. Parts A (dose escalation to 10 µg/kg/day), B (monotherapy in pts with melanoma or renal cell carcinoma [RCC]), and C (combination) included nemvaleukin 3 or 6 µg/kg/day ´ 5 and pembrolizumab every 21 days. Investigator-assessed antitumor activity (confirmed responses as per RECIST v1.1) and safety are reported as of 29 October 2021. Results: In Part A (N = 46), nemvaleukin recommended phase 2 dose was 6 µg/kg/day IV; maximum tolerated dose not reached. One pt had dose-limiting toxicity (grade 4 acute kidney injury) at 10 µg/kg. Pts in Parts B and C were heavily pretreated (1–9 prior lines of therapy, including prior checkpoint inhibitor therapy). Durable antitumor activity was observed for nemvaleukin monotherapy, including in RCC (objective response rate [ORR], 18.2% [4/22]) and in melanoma (ORR, 8.7% [4/46]), with 2 partial responses (1 unconfirmed) in 30 pts with cutaneous melanoma (ORR, 6.7%) and 2 PRs (1 unconfirmed) in 6 pts with mucosal melanoma (ORR, 33.3%). Durable antitumor activity was also observed for combination therapy (ORR, 16.1% [22/137]; disease control rate [DCR], 59.9%), including in platinum-resistant ovarian cancer (PROC; ORR, 28.6% [4/14]; DCR, 71.4%), with 2 complete responses and 2 PRs (1 unconfirmed) in 14 pts. Forty-three pts remain on therapy. The most frequent grade 3/4 treatment-related adverse events in Parts B and C, respectively, were anemia (9%, 10%), neutropenia (34%, 9%), and decreased neutrophil count (12%, 9%). Safety was consistent with previous reports. In pharmacodynamic studies, nemvaleukin monotherapy induced robust expansion of CD8+ T and NK cells, with minimal effect on regulatory T cells. Conclusions: ARTISTRY-1 showed proof of principle for preferential expansion of CD8+ T cells and NK cells by nemvaleukin. Nemvaleukin was generally well tolerated and demonstrated promising efficacy. Durable responses were observed with monotherapy and combination therapy in heavily pretreated pts across a range of tumors, warranting further investigation. The US FDA granted nemvaleukin Fast Track designation for treatment of mucosal melanoma and PROC, and Orphan Drug designation for mucosal melanoma. Clinical trial information: NCT02799095.

ARTISTRY-6: Nemvaleukin alfa monotherapy in patients with advanced mucosal and cutaneous melanoma.

TPS9609 Background: Despite improved outcomes for melanoma patients with the introduction of checkpoint inhibitors (CPIs), ̃50% of patients do not respond. A subset of responders ultimately progress and have limited treatment options, underscoring a high unmet need for novel treatments with durable benefit. Patients with mucosal melanoma exhibit response rates and progression-free survival times ̃2 times lower than those with cutaneous melanoma. Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel, engineered cytokine that selectively binds the intermediate-affinity interleukin-2 receptor complex to preferentially activate CD8+ T and NK cells with minimal expansion of regulatory T cells. Nemvaleukin has been granted Orphan Drug designation for the treatment of mucosal melanoma by the FDA. In ARTISTRY-1, the intravenous (IV) recommended phase 2 dose (RP2D) of 6 µg/kg nemvaleukin monotherapy demonstrated durable antitumor activity in patients with advanced melanoma, including mucosal melanoma, previously treated with a CPI. In ARTISTRY-2, the subcutaneous (SC) RP2D of 3 mg q7d was identified demonstrating pharmacodynamic effects consistent with IV delivery. Data support further evaluation of nemvaleukin monotherapy among patients with advanced mucosal and cutaneous melanoma. Methods: ARTISTRY-6 is a phase 2, global, multicenter, open-label study. Eligible patients have had prior treatment with an anti–PD-(L)1 therapy with or without anti–CTLA-4 therapy and have an ECOG performance status of 0 or 1 and adequate hematologic reserve and hepatic and renal function. Patients with advanced cutaneous (Cohort 1) and mucosal (Cohort 2) melanoma will receive nemvaleukin at the SC and IV RP2D, respectively. Patients will receive nemvaleukin until progression or intolerable toxicity. The primary objective is to evaluate the antitumor activity of nemvaleukin monotherapy defined by overall response rate. Additional objectives include the evaluation of safety, health-related quality of life, predictive biomarkers, pharmacokinetics, immunogenicity, and pharmacodynamic effects. Clinical trial information: NCT04830124.

A first-in-human, multicenter, phase 1/2, open-label study of XTX202, a masked and tumor-selective recombinant human interleukin-2 (IL-2) protein, in patients with advanced solid tumors.

TPS2697 Background: High-dose interleukin-2 (IL-2) has been approved in metastatic renal cell carcinoma (RCC) and metastatic melanoma, and can result in durable complete responses, including cures. Its use has been limited by life-threatening treatment-related multisystem toxicities, consisting mainly of vascular leak syndrome. XTX202 is a masked, tumor-selective IL-2 that is designed to be pharmacologically inactive in non-tumor tissue when circulating systemically and unmasked by matrix metalloproteases (MMPs) found preferentially in the tumor microenvironment (TME). XTX202 is expected, therefore, to achieve a wider therapeutic index resulting in increased efficacy and lower toxicity compared to existing non-tumor selective IL-2 based-therapies. The IL-2 domain of the XTX202 molecule is modified to reduce binding to the high-affinity IL-2 receptor while maintaining binding to the intermediate-affinity IL-2 receptor, thereby decreasing activation of regulatory T-cells relative to wild-type IL-2, while still activating effector T cells. A masking domain is designed to pharmacologically inactivate IL-2 until it is activated by MMPs that are enriched in the TME. Cleavage at a protease cleavage site in the XTX202 linker by MMPs results in an active IL-2 moiety when the masking domain is released. XTX202 is designed with the goal of producing a localized anti-tumor immune response and limiting exposure of the active form of XTX202 in non-tumor tissue. Preclinically, XTX202 exhibited tumor-selective activity in mice without peripheral toxicities in non-human primates [O’Neil et al. ASCO 2021]. Methods: XTX202-01 trial (NCT05052268) is a first-in-human Phase 1/2 study to determine a recommended Phase 2 dose of XTX202 (Phase 1) and to evaluate the efficacy of XTX202 monotherapy in patients with metastatic RCC and unresectable or metastatic melanoma (Phase 2). Patients with advanced solid tumors are eligible for Phase 1 and will receive XTX202 monotherapy administered every 21 days in an accelerated and standard 3+3 dose escalation design. Phase 2 will consist of 2 parts: Part 2a will enroll patients with metastatic RCC who have received a prior tyrosine kinase inhibitor therapy and have been treated and progressed on an anti-PD-1 therapy. Part 2b will enroll patients with unresectable or metastatic melanoma who have received immune-checkpoint therapy with an anti-PD-1 therapy and an anti-CTLA-4 therapy to determine the efficacy of XTX202 monotherapy in this population. Enrollment to the study began in January 2022. Clinical trial information: NCT05052268.

Nemvaleukin alfa in patients with advanced renal cell carcinoma: ARTISTRY-1.

330 Background: Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel engineered cytokine that selectively binds to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8+ T and NK cells, with minimal expansion of Tregs. Nemvaleukin is under investigation for the treatment of advanced solid tumors, including renal cell carcinoma (RCC), in the ARTISTRY-1 trial (NCT02799095). Demonstration of single-agent activity is essential to validate the potential therapeutic benefit of nemvaleukin, both as monotherapy and in combination with a checkpoint inhibitor (CPI), particularly in patients (pts) with limited treatment options, such as CPI-experienced pts with RCC. Methods: Pts with advanced RCC who had previously responded (either objective response or stable disease [SD]) to a CPI alone or as part of a combination were enrolled into an RCC-specific cohort of ARTISTRY-1 Part B. Nemvaleukin 6 µg/kg IV monotherapy was administered daily for 5 days every 14 days in cycle 1 and every 21 days in cycles 2+. Pts with disease progression (PD; after ≥2 cycles) or SD (after ≥4 cycles) could be enrolled into Part C to receive nemvaleukin and pembrolizumab combination therapy. Outcomes presented include antitumor activity (RECIST v1.1), pharmacodynamics, and safety as of August 2021. Results: Twenty-seven pts with RCC received nemvaleukin monotherapy in Part B. Median age was 69 y (range, 39-77); median prior lines of therapy was 2 (range, 1-7). Four of 21 evaluable pts had a best overall response of partial response (1 confirmed, 1 unconfirmed, 2 awaiting confirmation); 11 had SD. Nemvaleukin induced robust expansion of CD8+ T and NK cells with minimal effect on Tregs. The most frequently reported adverse events (AEs), regardless of causality (N = 27), were pyrexia (59.3%), chills (55.6%), nausea (29.6%), and anemia (29.6%). The most frequently reported grade ≥3 nemvaleukin-related AEs are shown in the table. One pt had an AE (nemvaleukin-related bronchospasm) resulting in treatment discontinuation. There were no deaths due to treatment-related AEs. Four pts are continuing monotherapy in Part B (up to 1 y) and 10 have rolled over to Part C (combination therapy). Of the 8 evaluable pts with PD on Part B who subsequently received combination therapy in Part C, 1 had a confirmed partial response and has now been on treatment for almost 1 y and 5 had SD. No additional safety signals were observed. Conclusions: Nemvaleukin was generally well tolerated as monotherapy and in combination with pembrolizumab, and provided evidence of single-agent tumor response and disease control in CPI-experienced pts with advanced RCC. Clinical evaluation of nemvaleukin among pts with advanced RCC is ongoing. Clinical trial information: NCT02799095. [Table: see text]

Nemvaleukin alfa combination therapy for gastrointestinal (GI) cancers: Preclinical evidence and clinical data from the ARTISTRY-1 trial.

659 Background: Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel engineered cytokine that selectively binds to the intermediate-affinity IL-2 receptor, preferentially activating and expanding antitumor CD8+ T and NK cells, with minimal expansion of regulatory T cells. Inherently active, nemvaleukin neither requires metabolic/proteolytic conversion nor degrades into native IL-2. Preclinical characterization confirmed receptor selectivity and antitumor activity of nemvaleukin alone and with checkpoint inhibitors (CPIs) and informed dose selection for the first-in-human study ARTISTRY-1; other combination regimens are ongoing evaluation. In ARTISTRY-1, responses with nemvaleukin in combination with pembrolizumab were observed in a variety of tumors, including GI. In addition to the clinical outcomes of these patients (pts), we describe preclinical evaluation of nemvaleukin with tyrosine kinase inhibitors (TKIs)—a drug class indicated for some GI cancers. Methods: Pts with GI cancers who had progressed on prior therapy were enrolled into cohorts of mixed tumor types in ARTISTRY-1 (NCT02799095) and received IV nemvaleukin (3 µg/kg) and pembrolizumab (200 mg). Outcomes presented include antitumor activity and safety as of August 2021. Combinations of nemvaleukin with TKIs were evaluated in mouse tumor models, including colorectal adenocarcinoma (MC38). Results: Clinically, 26 pts with GI cancer (colon/colorectal, n = 14; esophageal, n = 5; hepatocellular, n = 3; pancreatic, n = 3; gastric, n = 1) received nemvaleukin + pembrolizumab (median 3 cycles [range: 1-24]). Median age was 55 y (range: 26-82), ECOG performance status was 0 (n = 6) or 1 (n = 20), and median prior lines of therapy was 3 (range: 1-6). Four pts had a partial response, 2 with esophageal, 1 with MSI-H colorectal, and 1 with pancreatic cancer, with target lesion decreases of 37% to 63%. Six pts had stable disease. Two responders remain on treatment (&gt; 36 and &gt; 80 wks for colorectal and esophageal cancer, respectively). Frequent (&gt; 25%) nemvaleukin-related adverse events (AEs) among all pts receiving nemvaleukin combination (n = 156) were chills (58%), pyrexia (53%), nausea (29%), and fatigue (29%). Grade ≥3 nemvaleukin-related AEs (≥8%) were reported in 48%, including anemia (12%), neutrophil count decreased (10%), and neutropenia (8%). In mice, the combination of the mouse ortholog of nemvaleukin with VEGF TKIs showed improved antitumor activity and survival, along with increased immune activation and angiogenesis blockade in the tumor microenvironment compared with any compound alone. Conclusions: Emerging clinical data show responses in GI tumors may be achieved with nemvaleukin/pembrolizumab with manageable safety. Preclinical evidence of antitumor activity of nemvaleukin with CPIs or TKIs warrants further exploration of nemvaleukin in new combinations for pts with GI cancers. Clinical trial information: NCT02799095.

Preclinical and clinical researches of Denileukin Diftitox (Genetical Recombination) (Remitoro®), a novel agent for T-cell lymphoma

Denileukin Diftitox (DD) is a recombinant fusion protein of diphtheria toxin (DT) fragments and human interleukin-2 (IL-2). DD binds to IL-2 receptor (IL-2R) expressed on tumor cells and is taken up into the cells. Subsequently, DT fragments with adenosine diphosphate ribosylation enzyme inhibit protein synthesis, then ultimately trigger cell death. DD binds to both high- and intermediate-affinity IL-2Rs via IL-2 domain and inhibits growth of human T-cell lymphomas cell lines. E7777, which contains DD as an active component, has improved purity and an increased percentage of active monomer compared with the approved drug E7272 (ONTAK in the US, not approved in Japan). In the phase I clinical study in Japanese patients with relapsed or refractory peripheral T-cell lymphoma (PTCL) and cutaneous T-cell lymphoma (CTCL), the maximum tolerated dose and recommended dose of E7777 were 9 μg/kg/day (administered on Days 1-5 of each cycle) based on the evaluation of dose-limiting toxicity. In the phase II clinical study, the objective response rate was 36.1%, showing comparable efficacy to existing therapies. E7777 showed anti-tumor activity observed across the range of CD25 expression. Grade 3 or higher adverse events (AE) occurred in 94.6%, and serious AE such as capillary leak syndrome and rhabdomyolysis were reported. Therefore, safety monitoring activities have been continued along with alerting related events. Based on these results, E7777 obtained a new drug approval in Japan in March 2021 for the indication of relapsed or refractory PTCL/CTCL.

FLT3L Release by Natural Killer Cells Enhances Response to Radioimmunotherapy in Preclinical Models of HNSCC.

Natural killer (NK) cells are type I innate lymphoid cells that are known for their role in killing virally infected cells or cancer cells through direct cytotoxicity. In addition to direct tumor cell killing, NK cells are known to play fundamental roles in the tumor microenvironment through secretion of key cytokines, such as FMS-like tyrosine kinase 3 ligand (FLT3L). Although radiotherapy is the mainstay treatment in most cancers, the role of radiotherapy on NK cells is not well characterized. This study combines radiation, immunotherapies, genetic mouse models, and antibody depletion experiments to identify the role of NK cells in overcoming resistance to radiotherapy in orthotopic models of head and neck squamous cell carcinoma. We have found that NK cells are a crucial component in the development of an antitumor response, as depleting them removes efficacy of the previously successful combination treatment of radiotherapy, anti-CD25, and anti-CD137. However, in the absence of NK cells, the effect can be rescued through treatment with FLT3L. But neither radiotherapy with FLT3L therapy alone nor radiotherapy with anti-NKG2A yields any meaningful tumor growth delay. We also identify a role for IL2 in activating NK cells to secrete FLT3L. This activity, we show, is mediated through CD122, the intermediate affinity IL2 receptor, and can be targeted with anti-CD25 therapy. These findings highlight the complexity of using radio-immunotherapies to activate NK cells within the tumor microenvironment, and the importance of NK cells in activating dendritic cells for increased tumor surveillance.

1034TiP ARTISTRY-6: Nemvaleukin alfa monotherapy in patients with advanced mucosal and cutaneous melanoma

Despite improved outcomes for melanoma patients with the introduction of checkpoint inhibitors (CPIs), ∼50% of patients do not respond. A subset of responders ultimately progress and have limited treatment options, underscoring a high unmet need for novel treatments with durable benefit. Patients with mucosal melanoma exhibit response rates and progression-free survival times ∼2 times lower than those with cutaneous melanoma. Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel, engineered cytokine that selectively binds the intermediate-affinity interleukin-2 receptor complex to preferentially activate CD8+ T and NK cells with minimal expansion of regulatory T-cells.

Pharmacokinetics and Pharmacodynamic Effects of Nemvaleukin Alfa, a Selective Agonist of the Intermediate-Affinity IL-2 Receptor, in Cynomolgus Monkeys.

Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel cytokine created by the fusion of circularly permuted interleukin-2 (IL-2) to the IL-2Rα subunit of the IL-2 receptor (IL-2R) complex that confers selectivity for the intermediate-affinity IL-2R expressed on CD8+ T cells and natural killer (NK) cells. The pharmacokinetics and selective pharmacodynamic properties of nemvaleukin have been demonstrated using in vitro and in vivo mouse models. The pharmacokinetic/pharmacodynamic effects of nemvaleukin on immune cell subtypes were evaluated in cynomolgus monkeys after intravenous and subcutaneous administration to inform dose selection and predict pharmacodynamic effects in humans. Male drug-naïve cynomolgus monkeys (N = 15) were administered either single-dose (0.3 mg/kg i.v.; 0.3 mg/kg or 1.0 mg/kg s.c.) or repeated doses (0.1 mg/kg i.v. on days 1-5 or 0.5 mg/kg s.c. on days 1 and 4) of nemvaleukin. Serial blood samples were collected for pharmacokinetic assessment, immunophenotyping by flow cytometry, and profiling of serum cytokines. Repeat-dose subcutaneous administration of nemvaleukin with less frequent dosing resulted in total systemic exposure and trough serum concentrations comparable to those seen with intravenous administration, with lower peak serum concentrations. Transient elevation of interferon-γ and IL-6 peaked at 2 and 8 hours after intravenous and subcutaneous administration, respectively. Selective expansion of immunoprotective central memory, effector memory, terminal effector CD8+ T cells, and CD56+ NK cells, and minimal expansion of immunosuppressive CD4+CD25+FoxP3+ regulatory T cells was observed after both intravenous and subcutaneous administration. These data support the ongoing clinical evaluation of intravenous and subcutaneous nemvaleukin. SIGNIFICANCE STATEMENT: Administration of the novel interleukin-2 receptor agonist nemvaleukin alfa to cynomolgus monkeys resulted in selective expansion of immune effector cells, including CD8+ T and natural killer cells with minimal effects on immunosuppressive CD4+ regulatory T cells, confirming the design of nemvaleukin and highlighting its potential as a cancer immunotherapy. Subcutaneous administration of nemvaleukin achieved systemic exposure and immunostimulatory effects similar to those observed after more frequent intravenous dosing and may represent a practical alternative in a clinical setting.

Selection of the recommended phase 2 dose (RP2D) for subcutaneous nemvaleukin alfa: ARTISTRY-2.

2552 Background: Nemvaleukin alfa (nemvaleukin, ALKS 4230) is a novel engineered cytokine that selectively binds the intermediate-affinity interleukin-2 receptor to preferentially activate CD8+ T and natural killer (NK) cells with minimal expansion of regulatory T cells (Tregs),designed for use as a cancer immunotherapy. ARTISTRY-2 (NCT03861793) is an ongoing phase 1/2 study evaluating the safety, efficacy, and pharmacokinetic and pharmacodynamic (PD) responses of subcutaneous (SC) nemvaleukin in combination with pembrolizumab in patients (pts) with advanced solid tumors. Methods: In phase 1, cohort-specific doses of SC nemvaleukin are administered on an every-7-day (q7d) or every-21-day (q21d) schedule during a 6-week monotherapy lead-in period, followed by combination with pembrolizumab 200 mg q21d. We present safety, PD effects, and preliminary clinical activity outcomes as of 12/02/2020. Results: 57 pts received nemvaleukin doses ranging from 0.3 mg to 6 mg q7d or 1 mg to 10 mg q21d. The most frequent tumor types (&gt; 5 pts) were colorectal, pancreatic, ovarian, and lung; median number of prior therapies was 4. Treatment-related adverse events (TRAEs) in &gt; 30% pts overall were pyrexia (43.9%), chills (38.6%), injection site erythema (33.3%), injection site reaction (33.3%), and fatigue (31.6%). 3 mg q7d (n = 7) had no drug-related dose reductions, discontinuations, or deaths during the monotherapy or combination periods. 6 mg was declared the maximum tolerated dose (MTD) for q7d dosing as 2 of 8 pts experienced dose-limiting toxicities (DLTs). For 6 mg q21d (n = 7), no drug-related dose reductions, discontinuations, or deaths have occurred during the monotherapy period; combination period data are not mature. 10 mg was declared the MTD for q21d dosing as 1 of 9 pts experienced DLTs and 3 had TRAEs leading to dose reductions. Systemic exposure to nemvaleukin increased with increasing dose. Increases in NK cells and CD8+ T cells of approximately 16-fold and 3-fold, respectively, at 3 mg q7d, and approximately 8-fold and 3-fold, respectively, at 6 mg q21d were observed, with minimal change in Tregs. 46 pts had at least 1 on-treatment scan as of the data cutoff date, and 30 (65%) had stable disease (SD) on the first scan. Of the 30 pts with ≥2 scans, 13 (43%) had 2+ consecutive scans of SD. 16 of 57 pts remain on therapy. Antitumor activity data for more recent cohorts are still maturing. Based on the totality of the safety, PD effects, and antitumor activity data, 3 mg q7d was selected as the RP2D for SC nemvaleukin. Conclusions: SC nemvaleukin 3 mg q7d was generally well tolerated as monotherapy and in combination with pembrolizumab, and demonstrated robust PD effects on NK cells and CD8+ T cells with minimal expansion of Tregs. These PD effects are similar to or greater than those observed with intravenous nemvaleukin. Thus, 3 mg q7d was selected as RP2D; phase 2 expansion cohorts for combination with pembrolizumab are enrolling. Clinical trial information: NCT03861793.

MYELOID DERIVED SUPPRESSOR CELLS (MDSC) IN KIDNEY RECIPIENTS: IDENTIFICATION OF CD25+EMDSC, AND AUGMENTATION OF MONOCYTIC-MDSC EARLY AFTER TRANSPLANTATION IN PATIENTS LATER DEVELOPING CANCER

Background: MDSC increase in cancer and associate with poor prognosis, however their role in transplantation (Tx) is unknown. We aimed to study the MDSC effects on the evolution of kidney transplant recipients (KTR). Methods: A cohort of229 KTR was prospectively studied at pre- and day 7 and 14, month 1, 3, 6, and 12 post-Tx.Myeloid cells CD11b+CD33+CD14+CD15-HLA-DR-/low(M-MDSC), CD33+CD11b+CD14-HLA-DRlow/-(early stage MDSC, eMDSC) and CD11b+CD33+CD14+CD15-HLA-DR+(monocytes) were enumerated, and their suppressive capacity was tested in cocultures with autologous lymphocytes. Anti-HLA antibodies, total antioxidant capacity (TAC), arginase-1 (Arg-1), matrix metalloproteinase 9 (MMP9) and suppressive cytokines were measured in serum and supernatants. In some experiments, IL2 receptor (IL2R) expression of chains α (CD25), β (CD122) and γ (CD132) and STAT5 phosphorylation (pSTAT5) after incubation with IL2 were tested. The induction of myeloid cell death by basiliximab (Bx, anti-CD25) after adding complement (C´) was also analyzed. Results: Before Tx, HLA-DR-/low and HLA-DR+ myeloid subsets were similar among KTR and healthy volunteers. After Tx, CD11b+CD33+CD14+CD15-HLA-DR-/lowcells expanded and remained high for the first year. In vitro, these cells inhibited CD4 and CD8 T lymphocytes proliferation and IFNγ, TNFα and GM-CSF synthesis, thus confirming them as M-MDSC. The expansion of M-MDSC was not related with post-Tx events excepting for the development of cancer. Patients with M-MDSC>45.2% and >179.2 cell/µl at post-Tx day 14 had a significantly lower cancer-free survival in the 2 years after Tx (p<0.05), and showed a higher risk for cancer than patients with low M-MDSC (p=0.029 and 0.031, respectively). Multivariate Cox regression showed that M-MDSC>179.2 cell/µl was a risk factor for cancer in KTR independent of age, sex and immunosuppression (HR=6.98, p=0.02). Circulating M-MDSCs high counts correlated with high Arg-1 (p<0.01) and IL-10 (p=0.01), and low TAC (p=0.03) in serum. M-MDSC were lower in patients with enhanced alloimmune response as represented by anti-HLA sensitization. eMDSC levels also increased after Tx, excepting in patients who received Bx. CD25+eMDSC counts and the CD25per cell expression (MFI CD25) increased after Tx, but were not detected in patients who received Bx. eMDSC were found to express all three IL2R chains (high affinity IL2R), while M-MDSC and monocytes only expressed β and γ (intermediate affinity IL2R). After incubation of eMDSC with IL2, pSTAT5 was detected. Preincubation with Bx blocked both detection of CD25 and pSTAT5, but did not result in cell death after adding C´. Conclusions: Early post-transplant mobilisation of M-MDSC predicts cancer and adds risk as an independent factor in KTR. M-MDSC may favor an immunosuppressive environment that promotes the tumoral development. A CD25+eMDSC subset has been identified which is under study. ISCIII project 45/2013. CAM project B2017/BMD3731. AECC project IDEAS206PAZ.