Elevated Axicabtagene Ciloleucel (CAR-19) Expansion By Immunophenotyping Is Associated with Toxicity in Diffuse Large B-Cell Lymphoma
Authors: Spiegel Jay Y. , Sahaf Bita , Hossain Nasheed , Frank Matthew J. , Claire Gursharan , Abramian Matthew , Latchford Theresa , Villa Bertha , Cancilla Juancarlos , Oak Jean , Natkunam Yasodha , Long Steven R. , Arai Sally , Johnston Laura J , Lowsky Robert , Meyer Everett H , Muffly Lori S , Negrin Robert S. , Rezvani Andrew R , Shizuru Judith A , Weng Wen-Kai , Kong Katherine A. , Mackall Crystal L. , Miklos David B. 
Published: October 2019

Background:

Axicabtagene ciloleucel (axi-cel), an autologous anti-CD19 chimeric antigen receptor (CAR-T), showed significant clinical responses in patients with relapsed-refractory large-B cell lymphomas in the Zuma-1 trial (Neelapu et al, NEJM 2017). Zuma-1 analysis showed blood CAR-T cell expansion was associated with clinical response and toxicity. Herein, we report on 25 patients treated with commercial axi-cel and describe CAR-T expansion by immunophenotyping and its correlation with clinical outcomes.

Methods:

Twenty-five patients with aggressive lymphoma consecutively apheresed at Stanford University prior to June 30, 2018 were studied on an IRB approved biorepository-clinical outcome protocol. Cytokine release syndrome (CRS) was graded by Lee criteria (Blood 2014) and neurotoxicity according to Neelapu et. al (Nat. Rev. Clin. Onc. 2017). CAR-T cell immunophenotyping was assessed by peripheral blood flow cytometry on days 7, 14, 21 and 28 and then monthly. CAR-T cells were identified by gating on singlet+, live+, CD45+, CD14-, CD3+, anti-CD19-specific CAR mAb (clone 136.20.1; Jena et. al Plos 2013) and characterized as either CD4+ or CD8+.

Results:

Of 25 apheresed patients, 3 patients died prior to axi-cel infusion due to progressive lymphoma. Of 22 infused patients, 14 (64%) would have been eligible for the Zuma-1 trial. Reasons for ineligibility included symptomatic DVT (n=2), renal insufficiency (n=1), transaminitis (n=1), thrombocytopenia (n=1), MDS (n=1), pleural effusion (n=1) and 1 was ineligible by multiple criteria.

Median time from initial clinic visit to infusion was 47 days (range 34-117); median time from apheresis to infusion was 22 days (range 19-38). Nine patients received bridging therapy prior to lymphodepletion chemotherapy (chemo = 4, radiation = 2, high dose dexamethasone = 3). Axi-cel infusion occurred in hospital and patients were followed expectantly for a minimum of 7 days or until adverse events resolved to <Grade 2; median hospitalization was 13.5 days (range 7-44).

Ninety-five percent of patients developed CRS (Grade 2 = 73%, none ≥Grade 3). Median number of tocilizumab doses was 1 (range 0-4). Neurotoxicity occurred in 64%, Grade 3 or 4 in 27%. Corticosteroid therapy was required in 82% (77% received both tocilizumab and steroids). Median duration of steroids was 8.5 days (range 1-30); 12 patients required at least 1 week and 4 patients ≥2 weeks. Of patients infused, complete response (CR) at day 28 was 45% (ORR 86%). Of 15 patients evaluable at 3 months, ORR was 53% (CR = 7, PR = 1) and 47% progressed, similar to Zuma-1. Ineligibility for Zuma-1 was not associated with inferior outcomes.

Overall, median day 7 peak in vivo axi-cel expansion using anti-CAR19 flow cytometry was 38 CAR-T/ul (Fig. 1A), matching RT-PCR measured levels reported in Zuma-1. As shown in Fig. 1A, the majority of CAR-T cells were CD8+. Patients with Grade 2 CRS had significantly higher peak expansion of CAR-T cells (both CD4+ and CD8+) as compared to those with either Grade 0 or 1 CRS (Fig. 1B). Grades 2-4 neurotoxicity were significantly associated with peak total and CD8+ CAR-T but not CD4+ (Fig. 1B). Illustratively, 2 patients with the most robust CAR-T expansion (▪, ▼ Fig. 1A) experienced Grade 4 neurotoxicity including status epilepticus requiring multiple anti-epileptics and intubation. Peak CAR-T expansion in blood did not correlate with CR or ORR at day 28; expansion did not differ between patients who did or did not require steroids.

Fine needle aspirates (FNA) on a subset of patients with FDG-avid lymph nodes 2-3 days post axi-cel showed significant CAR-T expansion within the node despite low detectable circulating CAR-T. Figure 1C depicts a 76-year-old male with double expressor DLBCL who attained a CR at day 28; day 14 blood CAR-T expansion was below average (6 CAR-T/ul), while his day 2 FNA showed >35% of CD3+ T-cells expressed CAR19. As of submission, 34 patients were apheresed and updated blood and FNA results will be presented.

Conclusion:

Our analysis of 22 infused axi-cel patients showed an ORR of 86% and CR of 45%, despite 36% Zuma-1 ineligibilities and steroid use in 82%. Blood CAR-T expansion was associated with both CRS and neurotoxicity but not clinical response. Detection of high concentration of CAR-T cells in affected lymph nodes 2 days post infusion suggests quantification of CAR-T cells at disease sites could be predictive of clinical responses.

J.Y.S and B.S are co-first authors

Disclosures

Latchford: Kite a Gilead Company: Speakers Bureau. Muffly:Adaptive Biotechnologies: Research Funding; Shire Pharmaceuticals: Research Funding. Miklos:Kite - Gilead: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; Genentech: Research Funding; Novartis: Consultancy, Research Funding; Pharmacyclics - Abbot: Consultancy, Research Funding; Adaptive Biotechnologies: Consultancy, Research Funding.