Pilot Study of Non-Viral, RNA-Redirected Autologous Anti-CD19 Chimeric Antigen Receptor Modified T-Cells in Patients with Refractory/Relapsed Hodgkin Lymphoma (HL)
Authors: Svoboda Jakub , Rheingold Susan R. , Gill Saar I , Grupp Stephan A , Lacey Simon F , Melenhorst J. Joseph , Kulikovskaya Irina , Loudon Brandon , Kerr Naseem , Marcucci Katherine T , Shea Kim-Marie , Schmidt Aliza , Lledo Lester , Marshall Amy , Mato Anthony R. , Dwivedy Nasta Sunita , Landsburg Daniel J. , Youngman Matthew R. , Levine Bruce L , Porter David L , June Carl H , Schuster Stephen J. 
Published: April 2021

Abstract
Background: Cellular therapy using anti-CD19 autologous chimeric antigen receptor modified T (CART19) cells demonstrates promising outcomes in several hematologic malignancies of B-cell origin, but this therapy has not been studied in HL patients (pts). While neoplastic HL Reed-Sternberg (HRS) cells are considered CD19 negative, circulating CD19 positive clonal HRS cell precursors and CD19 positive reactive cells within the HRS tumor microenvironment represent potential therapeutic targets for CART19 in HL.
Methods: We designed companion pediatric (NCT02624258) and adult (NCT02277522) open-label pilot studies to estimate the feasibility, safety, and efficacy of CART19 cell infusions in pts with relapsed/refractory HL lacking curative treatment options. To allow transient CD19 targeting and limit the window for acute toxicity and B cell aplasia, we used autologous T-cells electroporated with mRNA encoding chimeric anti-CD19 immunoreceptor scFv (RNA CART19) cells in lieu of permanently modified cells engineered by viral transduction. The scFv is derived from a murine monoclonal antibody. Following pheresis and manufacturing of RNA CART19 cells, pts undergo up to 6 infusions of 8x105-1.5x106 RNA CART19 cells/kg/dose for pts &lt;80kg and 1x108 RNA CART19 cells/dose (±20%) for pts ≥80kg. Intravenous cyclophosphamide (30mg/kg) is administered prior to the first and fourth infusion to enhance engraftment. Safety, response assessments (Cheson 2007 criteria), and ancillary studies are measured at defined time points. The primary objective is to describe manufacturing feasibility, safety, and persistence of RNA CART19 cells in HL. Secondary objectives are to estimate efficacy by overall response rates (ORR) and the effect of RNA CART19 on immune factors.
Results: To date, 5 pts have been enrolled and had RNA CART19 manufactured. Four pts were infused with RNA CART19 and are evaluable for toxicity/response. Characteristics of the 5 pts include: median age 24 years (range 21-42), 4 (80%) with stage IV, median number of previous therapies 5 (range 4-9), 4 (80%) had stem cell transplant (SCT): 3 had auto SCT, 1 had both auto and allo SCT. Three pts previously progressed on PD-1 inhibitor. All 5 pts underwent successful manufacturing of RNA CART19. One pt developed MDS prior to RNA CART19 infusions and was taken off study. Four patients who underwent RNA CART19 infusions were treated with cyclophosphamide as per protocol. The median number of infused CART19 cells/kg/dose was 1.5x106 (range 7.3x105 -1.5x106). Each patient (pt) received 6 infusions over 2 weeks. Using qRT-PCR, RNA CART19 was detected in 80% of peripheral blood samples drawn within 2 hours after each infusion (Figure 1). RNA CART19 was also detected in 20% of samples drawn immediately prior each infusion reflecting persistence of RNA CART19 from previous infusion at 48 or more hours ago. No pt had RNA CART19 detected by Day (D) 21. There were no study related deaths or grade (G) 3/4 non-hematologic toxicities. Most common G1/2 toxicities at least possibly related to the RNA CART19 therapy occurring in &gt; 1 pt included transient headache in 3 and insomnia in 2. There was no evidence of cytokine release syndrome or significant elevation in cytokines at any point. The ORR at D28 was 50%: 1 complete (CR) and 1 partial response (PR). One pt had stable disease (SD) and one pt had progressive disease (PD). The CR pt (#3) was noted to have persistent RNA CART19 cells in 3 samples drawn at about 48 hours after the prior infusion compared with the PD pt (#1) who had no RNA CART19 detected by this time point on any measurement. In 2 pts who responded (#3 and #4), number of CD19-positive B-cells by flow cytometry as % of total leukocytes declined by 50% on D28 when compared to baseline. There was no evidence of B cell aplasia. The pt in CR progressed at 3 months and the pt in PR was taken off study. As part of routine clinical care, the 2 responding pts (#3 and #4) are currently in CR on PD-1 inhibitor (one underwent auto SCT). The pt with SD is in PR on lenalidomide and the pt with PD died of disease progression.
Conclusion: Targeting CD19 positive cells with non-viral, RNA-electroporated, transiently expressed CART19 cells is a feasible and safe strategy in pts with relapsed/refractory HL. We saw encouraging responses, but these were short-lived. We are planning a study for HL pts utilizing virally transduced CART19 cells that are capable of in vivo expansion in combination with PD-1 inhibitors.

Disclosures
Svoboda: Merck: Research Funding; BMS: Consultancy, Research Funding; Seattle Genetics: Consultancy, Research Funding; Celgene: Research Funding; Kite: Consultancy; Pharmacyclics: Research Funding. Gill: Novartis Pharmaceuticals: Patents & Royalties, Research Funding. Grupp: Adaptimmune: Consultancy; University of Pennsylvania: Patents & Royalties; Jazz Pharmaceuticals: Consultancy; Novartis Pharmaceuticals Corporation: Consultancy, Other: grant. Lacey: Novartis: Research Funding; Genentech: Honoraria. Melenhorst: Novartis: Research Funding. Mato: DTRM: Research Funding; AstraZeneca: Consultancy; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Research Funding; Regeneron: Research Funding; Acerta: Research Funding; Portola: Research Funding; Kite: Consultancy; AbbVie: Consultancy, Research Funding; Gilead Sciences, Inc.: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy. Dwivedy Nasta: Takeda: Research Funding; Incyte: Research Funding; Immunogen: Research Funding. Landsburg: Takeda: Research Funding; Curis: Consultancy, Research Funding. Levine: GE Healthcare: Consultancy; Tmunity Therapeutics: Equity Ownership, Research Funding; Brammer Bio: Consultancy; Novartis Pharmaceuticals Corporation: Patents & Royalties, Research Funding. Porter: Immunovative Therapies: Other: Member DSMB; Novartis: Honoraria, Patents & Royalties, Research Funding; Genentech/Roche: Employment, Other: Family member employment, stock ownship - family member; Servier: Honoraria, Other: Travel reimbursement; Incyte: Honoraria. June: Novartis: Patents & Royalties, Research Funding; Tmunity Therapeutics: Equity Ownership, Research Funding; WIRB/Copernicus Group: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celldex: Honoraria, Membership on an entity's Board of Directors or advisory committees; Immune Design: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Schuster: Genentech: Consultancy, Research Funding; Merck: Research Funding; Celgene: Consultancy, Research Funding; Nordic Nanovector: Consultancy; Novartis: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Seattle Genetics: Consultancy; Gilead: Consultancy, Research Funding; Janssen: Consultancy, Honoraria.