CD19 CAR-T Cells Are Highly Effective in Ibrutinib-Refractory Chronic Lymphocytic Leukemia
Authors: Turtle Cameron J , Hanafi Laila-Aicha , Li Daniel , Chaney Colette , Heimfeld Shelly , Riddell Stanley R , Maloney David G 
Published: October 2019

Abstract
BACKGROUND:
Ibrutinib, a Bruton Tyrosine Kinase (BTK) inhibitor causes partial responses (PR) in a majority of patients (pts) with chronic lymphocytic leukemia (CLL). However, complete responses (CR) are rare and high-risk pts who progress on ibrutinib have short survival. Lymphodepletion chemotherapy followed by infusion of CD19-specific chimeric antigen receptor (CAR)-modified T cells has produced encouraging responses in CLL in phase 1 clinical trials, but the majority of pts in those studies had not previously received or failed ibrutinib.
METHODS:
We treated 18 adults with CLL who had previously received ibrutinib with anti-CD19 CAR-T cells that were manufactured from defined CD4+ and CD8+ T cell subsets obtained by immunomagnetic selection of leukapheresis products, formulated in a final 1:1 ratio of CD8+:CD4+ CAR+ T cells, and infused at 1 of 3 dose levels (2x105, 2x106 or 2x107 CAR-T cells/kg) after lymphodepletion chemotherapy.
RESULTS:
Eighteen pts (median age 60; range 40-73) with a median of 5 previous therapies (range 3-9), including 3 pts that failed prior allogeneic stem cell transplant were enrolled and treated on the study. All pts were refractory to or had relapsed after receiving a regimen containing fludarabine and rituximab, and all pts had previously received ibrutinib; 11 were ibrutinib-refractory, 3 were ibrutinib-intolerant, and 4 were refractory to venetoclax. Twelve pts had a complex karyotype and 11 pts had 17p deletion. The median percentage of abnormal B cells in marrow was 77% (range 0.4 Ð 96). All pts had extramedullary disease and 2 had CNS disease. Lymphodepletion chemotherapy consisted of cyclophosphamide (Cy) 30-60 mg/kg x 1 and fludarabine (Flu) 25 mg/m2/day x 3 days (n=15); Flu 25 mg/m2/day x 3 days alone (n=2); and Cy 60 mg/kg alone (n=1). CAR-T cells were manufactured for all pts and 16/18 received CD4+ and CD8+ CAR-T cells in the defined 1:1 ratio. Four pts received 2x105, 13 received 2x106 and 1 received 2x107 CAR-T cells/kg.
Seventeen pts have completed response and toxicity assessment. Analysis of all pts with B cell malignancies treated with Cy/Flu and CAR-T cells on our trial showed that the highest dose level (2x107 CAR-T cells/kg) was too toxic for an initial CAR-T cell infusion, and identified a maximum tolerated first dose of 2x106 CAR-T cells/kg. Cytokine release syndrome (CRS) was graded according to Lee et al (Blood, 2014). After a single cycle of lymphodepletion chemotherapy and CAR-T cell infusion, 8 pts developed grade (gr) 0-1, 5 had gr 2, 3 had gr 3, and 1 had gr 4 CRS. Four pts developed gr ³3 neurotoxicity (NT). No gr 5 events were observed, no pts were intubated, and only 1 pt required pressors. Three pts received tocilizumab and dexamethasone to treat CRS and/or NT. Four pts received a second cycle of lymphodepletion chemotherapy and CAR-T cells at a 10-fold higher dose than the 1st infusion for persistent disease. CRS and NT (gr3) was observed in 3 of 4 pts after the second cycle of therapy.
Restaging was performed 4 weeks after the last CAR-T cell infusion. The ORR was 76% (8 PR and 5 CR). Two of the pts with PR by lymph node size criteria (IWCLL 2008) had negative PET scans after therapy. Among ibrutinib-refractory (n=10) or intolerant pts (n=3), the ORR was 77% (7 PR and 3 CR). In venetoclax refractory pts, 2 of 4 responded (PR). Only 1 of the 3 pts who did not receive Cy/Flu lymphodepletion responded. At day 28, 11 of 13 (85%) pts who received Cy/Flu lymphodepletion and a CAR-T cell infusion at ²2x106 CAR-T cells/kg had complete elimination of marrow disease by flow cytometry; 10/13 (77%) with nodal disease had a PR or CR at restaging, 1 had a mixed response, and 2 had progressive disease (PD).
No malignant IGH sequences were detected in marrow of 4/4 pts in CR who underwent IGH deep sequencing. Pts with CR had a higher peak percentage of CD8+ (p=0.006), but not CD4+ CAR-T cells in blood. Robust CAR-T cell expansion was seen in some non-responders, which in conjunction with the lower response rate in nodal sites compared to blood, suggests that factors in the malignant lymph node environment may inhibit CAR-T cell activity. No pt in CR has relapsed or died with a median follow-up of 8.4 months. For the 13 pts that received Cy/Flu lymphodepletion and ≤ 2 x 106 CAR-T cells/kg, OS is 100% and PFS is shown in Fig. 1.
CONCLUSION:
CD19 CAR-T cells of defined CD4:CD8 ratio are highly active in CLL and can induce high response rates and durable CRs in poor prognosis pts who have previously failed ibrutinib.

Disclosures
Turtle: Juno Therapeutics: Consultancy, Honoraria, Research Funding; Seattle Genetics: Consultancy, Honoraria. Li:Juno Therapeutics: Employment, Equity Ownership. Riddell:Adaptive Biotechnologies: Consultancy, Honoraria; Cell Medica: Consultancy, Honoraria; Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding. Maloney:Juno Therapeutics: Research Funding; Genentech/Roche: Honoraria.