Phase I Trial of CD8-Depleted Human Leukocyte Antigen (HLA) Mismatched Unrelated Donor Lymphocyte Infusion (DLI) to Achieve Remissions in Myelodysplastic Syndrome (MDS) and Secondary Acute Myeloid Leukemia (sAML)

Background Hypomethylating agents (HMAs) are the standard of care in higher risk MDS, yielding a median survival of 12-18 months. After HMA failure or progression to sAML. median survival is only 4-6 months. We developed a novel cellular therapy using HLA mismatched, unrelated, CD8-depleted, CD4+ DLI to induce remission in patients with MDS and sAML who have not previously undergone allogeneic hematopoietic cell transplant (HCT). The DLI is intentionally class II HLA mismatched to activate donor CD4+ T cells which then provide immunologic "help" to reverse recipient immune exhaustion and, thus, promote a potent antitumor response (Symons. BBMT. 2008). Further, HLA mismatched donors are easily identified and allow the potential for an "off the shelf" product. By depleting CD8+ T cells, it is intended that the DLI does not engraft, thus negating the risk of graft-versus-host disease (GVHD). Instead, this therapy is intended to induce remission as a bridge to curative allogeneic HCT.

Methods This completed phase I trial enrolled patients with MDS refractory to at least 4 cycles of HMA or sAML at the Moffitt Cancer Center from 2020-2022. Patients who received prior allogeneic HCT were excluded. DLI donors were selected from a pre-existing pool of unrelated donors at the New York Blood Center (NYBC) on the basis of class II HLA mismatch, CMV compatibility, and ABO compatibility. Patients were treated with induction chemotherapy as chosen by their primary physician. In parallel, donors underwent leukapheresis at the NYBC and the cell product was shipped to the Moffitt Cancer Center. The DLI was depleted of CD8⁺ T cells using the CliniMACS® system with CliniMACS® CD8 reagent in the Moffitt Cell Therapy Laboratory. HLA-mismatched CD4+ DLI was infused fresh within 24-36 hours after completing induction chemotherapy. The trial followed a 3+3 design to determine the recommended phase II dose (RP2D) of DLI where dose level (DL) 1 was 1x10⁶ CD4 cells/kg, DL2 was 1x10⁷ CD4 cells/kg, and DL3 was 5x10⁷ CD4 cells/kg. Dose limiting toxicities (DLTs) were defined as grade 2-4 acute GVHD, grade 3-4 cytokine release syndrome (CRS), grade 2-4 infusion reactions, aplasia beyond 56 days in the presence of detectable donor chimerism, or grade 4 organ toxicity by CTCAE 5.0. Mechanistic studies were pursued to evaluate markers of immune tolerance, cytokine profiles, and confirm the DLI did not engraft.

Results Nine patients (8 sAML, 1 MDS) were treated. All patients had progressed through at least one prior line of therapy (Table 1). The median age was 66. All AML patients had adverse risk disease by ELN risk stratification including 4 patients with TP53 mutation and complex cytogenetics. The MDS patient had very high-risk disease by R-IPSS. A suitable donor was identified for all patients despite a donor pool of only 30 donors. Cell collections, shipments, and processing for all patients were successful with all products achieving <3% CD8+ T cells and >70% cell viability. No patients experienced DLTs at any dose level. One patient at DL3 experienced grade 2 CRS that resolved with tocilizumab. There were no cases of grade 3-4 CRS nor GVHD (Table 2). Complete remission (CR) was achieved in 2/3 (66%) patients at DL3, 1/3 (33%) patients at DL2, and 0/3 (0%) patients at DL1. All 3 patients who reached CR proceeded to allogeneic HCT. At the time of marrow recovery post-treatment, donor DNA in the blood and marrow was no longer detectable in any of the patients. Patients who achieved CR (n=3) had significantly higher levels of IL6 on day +1 (p=0.01) and a trend towards higher levels of IL2, IL15, IFN-gamma, and TNF-alpha in the first 7 days after DLI compared to patients who did not achieve CR.

Conclusions Induction chemotherapy followed by CD8-depleted HLA mismatched, unrelated donor DLI was feasible and well tolerated with no DLTs, confirming the RP2D of 5x10⁷ CD4 cells/kg. At DL3, 2 of 3 patients achieved CR, including one with TP53 mutation and complex cytogenetics, which is generally resistant to induction chemotherapy and compares favorably to 20-30% historical CR rates with chemotherapy alone in those pts. Mechanistic studies correlated increased post-treatment cytokine levels with depth of remission. Assessment of markers of immune tolerance are pending and will be presented at the conference. Based on these promising results, this novel cellular therapy is being investigated now in a phase II clinical trial at the Moffitt Cancer Center.

Elmariah:Bristol Myers Squibb: Research Funding. Kuykendall:Pharmaessentia: Consultancy, Honoraria, Speakers Bureau; Imago Biosciences: Consultancy, Honoraria, Speakers Bureau; Incyte: Consultancy, Honoraria, Speakers Bureau; Blueprint: Consultancy, Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Abbvie: Consultancy, Honoraria, Speakers Bureau; GSK - Sierra Oncology: Consultancy, Honoraria, Other: Research Support, Speakers Bureau; Prelude Pharmaceuticals: Other: Research Support; BMS: Consultancy, Honoraria, Other: Research Support, Speakers Bureau; Morphosys: Other: Research Support; Protagonist: Other: Research Support; CTI Biopharma: Consultancy, Honoraria, Speakers Bureau. Lancet:Agios/Servio: Consultancy; Boxer Capital: Consultancy; Jasper Therapeutics: Consultancy; Astellas: Consultancy; Syntrix Pharmaceuticals: Research Funding; Dava Oncology: Consultancy; Dedham Group: Consultancy; Novartis: Consultancy; Servier: Consultancy; Jazz: Consultancy; BerGenBio: Consultancy; Millenium Pharma/Takeda: Consultancy; ElevateBio Management: Consultancy; Daiichi Sankyo: Consultancy; Celgene/BMS: Research Funding; AbbVie: Consultancy. Komrokji:Taiho Oncology: Consultancy, Membership on an entity's Board of Directors or advisory committees; Servier: Consultancy, Honoraria, Speakers Bureau; PharmaEssentia, Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Geron: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Acceleron Pharma: Consultancy; AbbVie: Consultancy, Honoraria, Speakers Bureau; CTI BioPharma, Innovent: Honoraria, Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Consultancy, Honoraria, Speakers Bureau; Bristol Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Sallman:Takeda: Consultancy; Shattuck Labs: Membership on an entity's Board of Directors or advisory committees; Kite: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Speakers Bureau; Syndax: Membership on an entity's Board of Directors or advisory committees; Magenta: Consultancy; Intellia: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Aprea: Membership on an entity's Board of Directors or advisory committees, Research Funding; Agios: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees; Syntrix Pharmaceuticals: Research Funding; Lixte: Patents & Royalties: LB-100; Nemucore: Membership on an entity's Board of Directors or advisory committees. Chan:Syntrix Pharmaceuticals: Research Funding. Sweet:Gilead Sciences, Inc.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; berGenBio: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; AROG: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Syntrix Pharmaceuticals: Research Funding; Mablytics: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Curis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Faramand:Novartis: Research Funding; Kite/Gilead: Research Funding. Pidala:Takeda: Research Funding; Abbvie: Research Funding; CTI Biopharma: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Regeneron: Consultancy, Membership on an entity's Board of Directors or advisory committees; Pharmacyclcis: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Johnson and Johnson: Research Funding; Novartis: Research Funding; Syndax: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Research Funding. Fuchs:Iyuda Therapeutics, LLC: Consultancy, Current Employment, Other: Co-Founder. Jain:MyeloidTx: Consultancy; BMS: Consultancy; Novartis: Consultancy; Incyte: Research Funding; Kite Pharma: Consultancy, Research Funding. Locke:), National Cancer Institute: Research Funding; Daiichi Sankyo: Consultancy; Sana: Consultancy; CERo Therapeutics: Research Funding; Leukemia and Lymphoma Society: Research Funding; Takeda: Consultancy; Aptitude Health: Other: Education or editorial activity; ASH: Other: Education or editorial activity; BioPharm Communications: Other: Education or editorial activity; CAREducation: Other: Education or editorial activity; Clinical Care Options Oncology: Other: Education or editorial activity; Imedex: Other: Education or editorial activity; Society for Immunotherapy of Cancer: Other: Education or editorial activity; BMS: Research Funding; A2: Consultancy; Celgene: Consultancy; Other: Patents & Royalties: patents, royalties, other intellectual property from several patents held by the institution in my name (unlicensed) in the field of cellular immunotherapy.; Wugen: Consultancy; Umoja: Consultancy; Novartis: Consultancy, Research Funding; Legend Biotech: Consultancy; Kite, a Gilead Company: Consultancy, Research Funding; Janssen: Consultancy; Iovance: Consultancy; GammaDelta Therapeutics: Consultancy; Emerging Therapy Solutions Gerson Lehrman Group: Consultancy; EcoR1: Consultancy; Cowen: Consultancy; Calibr: Consultancy; Cellular Biomedicine Group: Consultancy; Bristol Myers Squibb/Celgene: Consultancy; Bluebird Bio: Consultancy, Research Funding; Allogene: Consultancy, Research Funding; Amgen: Consultancy. Bejanyan:Medexus Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Magenta Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; CareDX Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees; CTI BioPharma: Consultancy, Membership on an entity's Board of Directors or advisory committees. DeZern:Novartis: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Bristol Myers Squibb: Consultancy, Honoraria; CTI BioPharma: Consultancy, Honoraria; Syntrix Pharmaceuticals: Research Funding; GERON: Other: DSMB.