Kenderian, Saad S.

Abstract The receptor tyrosine kinase AXL is a member of the TYRO3, AXL, and proto-oncogene tyrosine-protein kinase MER family and plays pleiotropic roles in cancer progression. AXL is expressed in immunosuppressive cells, which contributes to decreased efficacy of immunotherapy. Therefore, we hypothesized that AXL inhibition could serve as a strategy to overcome resistance to chimeric antigen receptor T (CAR T)–cell therapy. To test this, we determined the impact of AXL inhibition on CD19-targeted CAR T (CART19)–cell functions. Our results demonstrate that T cells and CAR T cells express high levels of AXL. Specifically, higher levels of AXL on activated Th2 CAR T cells and M2-polarized macrophages were observed. AXL inhibition with small molecules or via genetic disruption in T cells demonstrated selective inhibition of Th2 CAR T cells, reduction of Th2 cytokines, reversal of CAR T-cell inhibition, and promotion of CAR T-cell effector functions. AXL inhibition is a novel strategy to enhance CAR T-cell functions through two independent, but complementary, mechanisms: targeting Th2 cells and reversing myeloid-induced CAR T-cell inhibition through selective targeting of M2-polarized macrophages.

Abstract Glioblastoma multiforme (GBM) is the most aggressive and common form of brain tumor, and it is characterized by an immunosuppressive tumor microenvironment (TME) that supports tumor growth and pathology. An integral part of the GBM stroma is the mesenchymal stem cells (MSC), which promote GBM growth. Receptor tyrosine kinase EphA3, a member of the Eph family, is overexpressed in GBM tumors and in MSCs within TME, making it an attractive target for Chimeric antigen receptor T-cell (CART) therapy. CART has proven incredible promise in certain hematological malignancies therapy but remained short in solid tumors, particularly GBM. We hypothesize that concurrent targeting of cancers and TME by EphA3-CART can be a potential therapeutic option in GBM. We first showed the expression of the EphA3 receptor by Immunohistochemistry and western blot on GBM patient-derived xenograft (PDX), as well as on the SNB-19 and U87 human GBM cell lines and adipose-derived MSCs using flow cytometry. We then generated EphA3-CAR construct derived from a clinically validated anti-EphA3 scFv cloned in a lentiviral vector containing CD28 and CD3ζ, and generated EphA3 CART cells with over 90% efficiency. EphA3 CART cells demonstrated potent antigen specific effector functions. To demonstrate the antitumor activity in vivo, we subcutaneously grafted 1 × 106 GBM cells per NOD-SCID-γ-/- mice and at ~150-300 mm3 tumor volume, mice were then treated with 5 × 106 EphA3-CART or un-transduced (UTD) T cells control. Tumors were monitored by bioluminescence imaging (BLI) and caliper measurements, and mice were followed for survival. Mice treated with EphA3 CART cells exhibited a significant reduction in tumor volume (p<0.005), and prolonged overall survival (p=0.0015). Finally, we tested the ability of EphA3-CART cells to overcome MSC induced CART cell suppression by co-culturing EphA3-CART cells with MSC and EphA3+ GBM target cells while using the CD19+ mantle cell lymphoma cell line JeKo-1 and CART19 as control. MSC conditioned coculture of GBM and CART showed significant suppression of CAR19 in the CART19 - JeKo-1 system (p=000.3), but no significant suppression of EphA3-CART cells. In summary, our results indicate that EphA3 CART cells exhibited potent and specific antitumor activity in vitro and in vivo and ameliorated MSC induced inhibition of CART cell functions, representing a potentially promising therapeutic option in GBM. Citation Format: Ekene J. Ogbodo, Michael W. Ruff, Reona L. Sakemura, Claudia ManriquezRoman, Truc Huynh, James H. Girsch, Olivia L. Sirpilla, Kun Yun, Carli M. Stewart, Ismail Can, Lionel K. Fonkoua, Mehrdad Hefazi, Elizabeth L. Siegler, Saad S. Kenderian. Simultaneous targeting of EphA3 on glioblastoma and tumor microenvironment to overcome resistance to CART cell therapy in brain cancer. [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 5017.

12057 Background: AlthoughCLL primarily affects older adults (median age 71 years), limited data exists about the outcomes of adults who are ≥80 years old because they are under-enrolled on clinical trials. Methods: The Mayo Clinic CLL Database was used to conduct a retrospective cohort study of adults ≥80 years at the time of frontline CLL treatment. Kaplan-Meier analysis was used to plot OS, and cumulative incidence was used to plot TTNT, accounting for competing risk of death. The Mayo Clinic IRB approved the study. Results: Our study included 216 patients with CLL who were age ≥80 years at the time of frontline CLL therapy between 1/1995 and 11/2022 (Table). The median time from CLL diagnosis to initiation of frontline therapy was 3.1 years. The median OS after start of frontline therapy was 3.9 years. The median OS was 3.3 years (95% CI 2.6–4.2) for patients who received alkylating agents (e.g., chlorambucil, n=96), 3.8 years (95% CI 2.8–not reached [NR]) for purine analogues (e.g., fludarabine, n=11), 3.9 years (95% CI 3.2–4.9) for anti-CD20 monoclonal antibody monotherapy (e.g., obinutuzumab, n=64), and not reached (95% CI 3.3-NR) for novel agents (n=43) (P=0.02). The types of novel agents included ibrutinib (n=19), acalabrutinib (n=16), venetoclax (n=7), and orelabrutinib (n=1). There was no difference in OS between 36 patients who received a BTKi compared to the 7 patients who received venetoclax-based therapy. At a median follow-up time of 6.7 years from the first visit, 143 patients died from the following: progressive CLL (n=63), infections (n=12), other cancer (n=11), non-CLL reasons (n=21). Cause of death was unavailable for 36 patients. On multivariable analyses, older age (i.e., any 5-year increase in age) was associated with a 50% increased risk of death (HR 1.5, 95% CI 1.2-1.9, p=0.001), and treatment with non-novel agents was associated with 3.2 times increased risk of death (HR 3.2, 95% CI 1.2-8.9, p=0.02). Of the 216 patients, 76 patients required second-line therapy; the median TTNT was 4.2 years (95% CI 2.9-NE). The most used second-line agents were monoclonal antibodies alone (20/76, 26%) and single-agent alkylators (18/76, 24%). Conclusions: When compared to all other CLL treatments, novel agents such as BTKi and venetoclax-based treatments are associated with significantly improved OS in patients with CLL ≥80 years old when used in the frontline setting. [Table: see text]

e19507 Background: Patients (pts) with CLL are at elevated risk of developing non-hematologic malignancies. Radiation therapy (RT) is a critical component of management in localized solid organ malignancies. Novel agents (NA) are widely used in CLL treatment; however, safety data are lacking to guide treatment of pts requiring concurrent RT and NA. Methods: We identified pts treated concurrently with NA for CLL and RT for second primary malignancy (SPM) at Mayo Clinic from 2014-2022. Adverse events within 3 months of RT were evaluated using the International Workshop on Chronic Lymphocytic Leukemia (iwCLL) criteria for hematological toxicity and the Common Terminology Criteria for Adverse Events (CTCAE) version 5 for non-hematological toxicity. Patients without seamless, concurrent treatment with NA and RT were excluded. Results: Twenty-six pts were included. NA therapy consisted of ibrutinib (n = 23), acalabrutinib (n = 2), and venetoclax (n = 1). Median follow up time since RT completion was 1.5 years. The median age at NA initiation was 67 years (range:46-85) and at RT start was 68 years (range: 51-85); 92% of pts were male. NA was first treatment in 23% of pts; the median number of prior lines of therapy was 1 (range: 0-4). IGHV was unmutated in 82% and TP53 aberration was present in 24% of evaluable patients. SPM sites included prostate (n = 9), head/neck (n = 8), lung (n = 3), and brain, breast, bone, skin, kidney, and thymus (n = 1 each). The most common conventionally fractionated prescriptions (cGy/fx) were 6000/30 (n = 6) and > 6000-7020/20-33 (n = 7). SBRT ranged from 3300/5 to 5400/3 (n = 8). Five pts received other, varied RT prescriptions. Overall, 3 (12%) pts experienced at least one grade 3/4 toxicity. Grade 3/4 thrombocytopenia was observed in 2 (8%) pts and grade 3/4 neutropenia was observed in 2 pts per iwCLL criteria. Grade ≥3 non-hematological toxicities occurred in two pts that required hospitalization for critical illness, including 1 pt with grade 5 respiratory failure within 6 weeks after RT. Dose de-escalation of ibrutinib was reported in one case for pancytopenia attributed to use of a radiosensitizing agent. There were no cases of RT dose modification. Conclusions: Continuing NA CLL therapy during RT did not result in unexpected toxicities. This approach may maintain CLL disease control without compromising management of the SPM. [Table: see text]

7501 Background: In patients (pts) with R/R CLL/SLL that progressed on BTKi and failed venetoclax (ven)-based regimens, achieving CR with current treatment is uncommon. New therapies that achieve deep and durable responses are needed. We report the primary analysis of the phase 1/2, single-arm, multicenter TRANSCEND CLL 004 (NCT03331198) study evaluating liso-cel in pts with R/R CLL/SLL. Methods: Pts must have received at least 2 prior lines of therapy, including a BTKi. Eligible pts received liso-cel at a target dose of either 50 (DL1) or 100 (DL2) × 106 CAR+ T cells. The primary endpoint was rate of CR and CR with incomplete marrow recovery (CRi) by IRC per 2018 iwCLL criteria in the prespecified subset of efficacy-evaluable pts with disease progression on BTKi and ven failure (primary efficacy analysis set [PEAS]) at DL2 (null hypothesis [H0]: ≤ 5%). Key secondary endpoints were ORR (H0: ≤ 40%) and rate of undetectable minimal residual disease (uMRD; 10−4) in blood (H0: ≤ 5%). Results: Of 137 leukapheresed pts, 117 received liso-cel (safety set), 96 (DL1 = 9; DL2 = 87) were efficacy evaluable, and 53 (DL1 = 4; DL2 = 49) were in the PEAS. In the safety set, median (range) age was 65 y (49–82), 83% had high-risk features, median (range) lines of prior therapy was 5 (2–12), and all pts had prior BTKi. Median (range) on-study follow-up was 21.1 mo (0.4–55.6) for the safety set. In the PEAS at DL2, the primary endpoint of CR/CRi rate was met at 18.4% (95% CI, 8.8–32.0; 1-sided P = 0.0006; Table). ORR was 42.9% and was not statistically significant (95% CI, 28.8–57.8; 1-sided P = 0.3931). The uMRD rate was 63.3% in blood and 59.2% in marrow. Median (95% CI) DOR was 35.3 mo (11.01–not reached [NR]) with a median follow-up of 19.7 mo. Median duration of CR/CRi was NR. In the safety set, rate of any-grade CRS was 84.6% (gr 3, 8.5%; no gr 4/5) and neurological events (NE) was 45.3% (gr 3, 17.9%; gr 4, 0.9%; no gr 5); 69.2% received tocilizumab and/or corticosteroids for CRS/NEs. Rate of gr ≥ 3 infections, hypogammaglobulinemia, and prolonged cytopenia was 17.1%, 15.4%, and 53.8%, respectively. One death related to liso-cel was due to hemophagocytic lymphohistiocytosis. Liso-cel exhibited rapid in vivo expansion and was detected by qPCR in blood up to 36 mo after infusion. Conclusions: Liso-cel demonstrated durable CR/CRi, high uMRD rates, and a manageable safety profile in pts with heavily pretreated, high-risk R/R CLL/SLL and high unmet need. Clinical trial information: NCT03331198 . [Table: see text]