Chikungunya virus (CHIKV) is the causative agent of an outbreak that began in La Réunion in 2005 and remains a major public health concern in India, Southeast Asia, and southern Europe. CHIKV is transmitted to humans by mosquitoes and the associated disease is characterized by fever, myalgia, arthralgia, and rash. As viral load in infected patients declines before the appearance of neutralizing antibodies, we studied the role of type I interferon (IFN) in CHIKV pathogenesis. Based on human studies and mouse experimentation, we show that CHIKV does not directly stimulate type I IFN production in immune cells. Instead, infected nonhematopoietic cells sense viral RNA in a Cardif-dependent manner and participate in the control of infection through their production of type I IFNs. Although the Cardif signaling pathway contributes to the immune response, we also find evidence for a MyD88-dependent sensor that is critical for preventing viral dissemination. Moreover, we demonstrate that IFN-α/β receptor (IFNAR) expression is required in the periphery but not on immune cells, as IFNAR−/−→WT bone marrow chimeras are capable of clearing the infection, whereas WT→IFNAR−/− chimeras succumb. This study defines an essential role for type I IFN, produced via cooperation between multiple host sensors and acting directly on nonhematopoietic cells, in the control of CHIKV.

5510 Background: Simlukafusp alfa (SIM; FAP-IL2v) comprises an interleukin-2 variant (IL-2v) moiety and an antibody against fibroblast activation protein-α (FAP). The binding of SIM to FAP, expressed on cancer-associated fibroblasts, accounts for retention and accumulation in malignant lesions. The engineered IL-2v moiety has an abolished binding to IL-2Rα while the affinity to IL-2Rβγ is retained, resulting in activation of immune effector CD8 T and NK cells, but not of regulatory T cells, and therefore may augment activity of PD-(L)1 inhibitors. Methods: The clinical activity and safety of the SIM and atezolizumab (ATZ) combination in patients with recurrent or metastatic (R/M) cervical squamous cell carcinoma (SCC) were evaluated in a phase 2 basket study (NCT03386721). Patients (pts) were treated with SIM 10 mg IV and ATZ 1200 mg IV once every 3 weeks. The primary endpoint was objective response rate (ORR) by RECIST v1.1 assessed by investigators. Secondary endpoints were: disease control rate (DCR), duration of response (DoR), progression free survival (PFS). Results: 47 Pts with ECOG ≥1 and median age of 53 years (range: 25-69) were enrolled. All pts were checkpoint inhibitor naïve and 40 (85%) had ≥ 1 previous lines of therapy in the metastatic setting. The median number of cycles was 6 (range: 1-29). The ORR was 27% (90% CI: 18, 39) and DCR was 71% (90% CI: 58, 80) in 44 response-evaluable patients: 2 (5%) had complete response, 10 (23%) partial response, and 19 (43%) stable disease. Responses were observed across PD-L1 subgroups (SP142 assay, IC/TC cut-off ≥ 1%) with 8/22 and 4/18 responders in PD-L1+ and PD-L1- patients, respectively. Responses were durable, the median DoR was 13.3 months (95% CI: 7.6, 14.7). PFS probability at 6 months was 0.4 (95% CI: 0.27, 0.59). The most common adverse events (AE) (reported in > 30% patients), irrespective of relatedness to treatment and severity, were pyrexia (74.5%), anemia (48.9%), asthenia (48.9%), AST increased (44.7%), nausea (42.6%), ALT increased (42.6%), vomiting (36.2%) and diarrhea (31.9%). Grade 3 and 4 AEs related to SIM were observed in 63.8 % and 29.8 % of pts, respectively, while serious AEs (SAE) related to SIM were reported in 40.4%. The most common SAEs (reported in > 5% pts) irrespective of relatedness to treatment were infusion related reaction (14.9%), pyrexia (6.4%) and hydronephrosis (6.4%). One Grade 5 event occurred, which was unrelated to treatment. Conclusions: SIM in combination with ATZ demonstrated an acceptable safety profile in pts with R/M cervical SCC. The anti-tumor activity compares favorably to the approved PD-1 inhibitors in this setting and supports further exploration of IL-2v and checkpoint inhibition in this patient population of high unmet medical need. Clinical trial information: NCT03386721.

4556 Background: Simlukafusp α ([SIM], FAP-IL2v) is a novel IL-2v immunocytokine engineered to preferentially activate effector CD8 T and NK cells, but not regulatory T cells (Tregs), due to abolished binding to Interleukin-2 receptor α (IL-2Rα) and retained affinity to IL-2Rβγ. High affinity binding of SIM to fibroblast activation protein (FAP), expressed on cancer-associated fibroblasts, mediates its accumulation in malignant lesions. Methods: The Dose-Escalation (DE) consisted of: Arm A: SIM 5-25 mg weekly for 4 weeks, and every 2 weeks (Q2W) thereafter in combination with atezolizumab [ATZ] 840mg Q2W; and Arm B: same as Arm A + bevacizumab [BEV] 10 mg/kg Q2W. Patients (pts) not previously treated were evaluated in the Extension Part: Arm C (n=3): SIM + ATZ every 3 weeks (Q3W); or Arm D (n=25): SIM + ATZ + BEV (“triplet”) Q3W. Primary objectives were: finding the recommended dose of SIM and assessment of objective response rate (ORR) by RECIST v1.1. Results: We enrolled 69 pts with unresectable advanced/ metastatic clear-cell and/or sarcomatoid RCC. Median age of patients was 57 years (range: 35-78). The recommended dose for extension of SIM was 10 mg. Median treatment duration in days in each arm were: A: 106 (range: 1-877); B: 324 (8-940); C: 659 (71-768); D: 437 (1-682). Twenty-five pts are evaluable for therapeutic activity in Arm A [ORR: 24% (6 PR; 90% CI 12.95, 40.12)]; 15 in Arm B [46.7% (1 CR, 6PR; 90% CI 27.67, 66.68)]; 3 in Arm C [33.3% (1PR; 90% CI 7.83, 74.65)]; and 23 in Arm D [47.8% (2 CR, 9 PR; 90% CI 35.74, 68.15)]. Twelve patients are ongoing on study treatment. Treatment related grade 3 and 4 adverse events (AE) occurred respectively in 69.7% and 9.1% patients. The most common serious AEs were pyrexia (10.6 %) and infusion-related reactions (9.1%). 65.2% Of the patients reported at least one AE of elevations in liver transaminases/GGT/ alkaline phosphatase/bilirubin. Drug-related AEs led to dose modification/interruption in 37.9 % of the pts, and treatment discontinuation in 3% of the patients. SIM led to preferential expansion and activation of NK and CD8 T cells (but not Tregs) in peripheral blood and augmented tumor infiltration and tumor inflammation. Intriguingly responses were observed not only in pts with PD-L1 positive or inflamed tumors, but also in pts with PD-L1 negative tumors (n=13) or poorly infiltrated tumors classified as immune deserts (n=2). Conclusions: The combination of SIM with ATZ ± BEV was feasible with an acceptable safety profile. Clinical activity was more favorable for the triplet among the study Arms, but comparable to the ATZ + BEV combination in the IMmotion151 (Rini B, et al 2019). Observed pharmacodynamic findings were consistent with the expected effects. Clinical trial information: NCT03063762.

BackgroundFibroblast activation protein alpha (FAP) is frequently over-expressed in the tumor microenvironment (TME) while exhibiting limited expression in normal tissues. FAP expression was reported to be immunosuppressive in tumor mouse models and generally associated with worse prognosis in clinical studies. Therefore, it is important to understand the context in which FAP both exhibits immunosuppressive characteristics and be a useful target for immunotherapy.MethodsComprehensive immunohistochemistry (IHC) analyses on formalin-fixed paraffin-embedded tissue specimens with emphasis on lymph nodes and primary and metastatic tumor lesions spanning a wide range of indications were undertaken in this study. FAP staining of tumor tissues was performed with an optimized IHC robust-prototype-assay (RPA) and manually scored. The area (normal stroma & neoplastic) staining positively relative to the total tumor area at each intensity level was recorded and an H-score calculated (FAP-intensity score).These were supplemented by gene expression analysis using public as well as Roche phase 1, 2 and 3 cancer immunotherapy (CIT) clinical trial data sets.ResultsAnalysing FAP expression on normal tissue confirmed the general absence of FAP apart from a subset of pancreatic islet cells. Unlike the more homogenous expression of typical protein targets on tumor cells, FAP expression in the TME is heterogeneous in both pattern and intensity, requiring the analysis of a large sample set. Therefore, we evaluated 1216 samples from 23 tumor indications and 70 sub-indications. FAP expression exhibited a significant spread ranging from indications with highly abundant expression to those with low coverage.Using data from matching IHC and gene expression samples we confirmed FAP mRNA expression to significantly correlate with RPA H-scores (Spearman correlation: 0.62) (N=289, P=1.2E-31). Gene expression data from 12 atezolizumab clinical studies, including standard of care (SOC) randomized studies, with more than 6000 samples from 4 major indications were interrogated for the association between FAP expression and clinical response as evaluated by overall and progression free survival. This analysis suggests that FAP expression is generally associated with higher hazard ratios across all atezolizumab-treated samples (OS: 95% CI 1.04–1.09; PFS: 1.04–1.08), with the highest effect observed in Renal Cell Carcinoma (OS: 95% CI 1.08–1.31; PFS: 1.05–1.21), indicating a potential role of FAP in limiting CIT.ConclusionsData from these analyses can tailor indication and patient enrichment strategies for achieving optimal FAP-targeting. We propose to select indications with FAP-levels that are high enough to enable drug accumulation, yet low enough to reduce immunosuppressive effects that can hamper successful immunotherapy.