Single Agent CD45-Targeted Antibody Drug Conjugate Enables Full Mismatch Allogeneic Hematopoietic Stem Cell Transplantation in a Murine HSCT Model
Authors: Hyzy Sharon L. , Proctor Jennifer L , Gillard Geoff O , Hammond Katelyn J. , Sarma Ganapathy N. , Saha Asim , Clark Nicholas , Bhat Anjali , Lamothe Tahirih L , Palchaudhuri Rahul , Pearse Bradley R , McDonagh Charlotte F , Kiem Hans-Peter , Wagner John E , Blazar Bruce R. , Boitano Anthony E , Cooke Michael P , Davis John C. 
Published: November 2020

Introduction
Allogeneic hematopoietic stem cell transplant (Allo-HSCT) is a potentially curative treatment for malignant and non-malignant blood disorders. However, current conditioning regimens limit the use of this curative procedure in many eligible patients due to regimen-related mortality and morbidities, including organ toxicity, infertility, and secondary malignancies. We are developing novel antibody drug conjugates (ADC) as conditioning agents that can achieve full myeloablation as a single agent that may reduce toxicity associated with current conditioning regimens. We have generated an anti-murine ADC targeting CD45 and assessed its effectiveness as single agent conditioning regimen in a fully allogeneic murine HSCT model.
Methods
Our tool CD45 ADC is engineered for rapid clearance (t1/2=1.7hr) to enable HSCT after conditioning. A single dose of 3 mg/kg is fully myeloablative in C57BL/6 mice. To determine if the tool CD45-ADC could successfully condition recipients for fully mismatched allo-HSCT, we evaluated the ability of a single dose of 5 mg/kg of the tool CD45-ADC to condition C57BL/6 hosts (H-2b, CD45.2+) for transplant with cells from CByJ.SJL(B6) donors (H-2d, CD45.1+). A matched dose of an isotype ADC (Iso-ADC) was used as a negative control, while 9 Gy TBI was used as a conventional conditioning positive control. Conditioned mice were transplanted with 4x107 whole BM cells, and peripheral blood chimerism was assessed over 22 weeks. At 22 weeks, donor hematopoietic cell chimerism was evaluated in the spleen, bone marrow, and thymus of recipients.
Results
In the fully mismatched Balb/c → C57Bl/6 allo-HSCT model, conditioning with a single dose of 5 mg/kg of CD45-ADC as a single agent was well tolerated and enabled full allogeneic donor chimerism (n=2 separate experiments). Peripheral blood chimerism was observed in mice conditioned with CD45-ADC at week 4 and maintained through week 22 (Figure 1). Multilineage reconstitution was observed in the T-, B-, and myeloid cell compartments with &gt;90% donor chimerism seen in each compartment, indicative of HSC engraftment. These results were comparable to chimerism seen in the 9 Gy TBI positive control. Treatment with a non-targeting isotype ADC at a matched dose was not effective (Figure 1). For all groups, stem cell chimerism in the bone marrow matched that in the periphery. Splenic and thymic donor immune cell reconstitution was similar between CD45-ADC and TBI conditioning at week 22 (Figure 1), demonstrating that CD45-ADC efficiently depletes host lymphocytes in secondary lymphoid organs while preserving the capacity of the host thymus to support de novo generation of donor-derived T cells after transplantation.
Conclusion
Conditioning with CD45-ADC was well-tolerated, fully myeloablative, and enabled complete chimerism in a full mismatch allo-HSCT model as a single agent. This targeted, readily translatable approach for safer conditioning could improve the risk-benefit profile for allogenic and haploidentical HSCT and may extend the curative potential of HSCT to more patients suffering from blood cancers and other diseases that may benefit from HSCT.

Disclosures
Hyzy: Magenta Therapeutics: Current Employment, Current equity holder in publicly-traded company. Proctor:Magenta Therapeutics: Current Employment. Gillard:Magenta Therapeutics: Current Employment. Hammond:Magenta Therapeutics: Current Employment, Current equity holder in publicly-traded company. Sarma:Magenta Therapeutics: Ended employment in the past 24 months. Clark:Magenta Therapeutics: Current Employment. Bhat:Magenta Therapeutics: Current Employment. Lamothe:Magenta Therapeutics: Current Employment. Palchaudhuri:Magenta Therapeutics: Current Employment. Pearse:Magenta Therapeutics: Ended employment in the past 24 months. McDonagh:Magenta Therapeutics: Ended employment in the past 24 months. Kiem:Magenta Therapeutics: Consultancy; CSL: Consultancy; Homology Medicines: Membership on an entity's Board of Directors or advisory committees; Rocket Pharma: Membership on an entity's Board of Directors or advisory committees; Umoja: Membership on an entity's Board of Directors or advisory committees; Enochian: Membership on an entity's Board of Directors or advisory committees; Vor Biopharma: Membership on an entity's Board of Directors or advisory committees. Wagner:Rocket Pharmaceuticals, Inc.: Consultancy, Current equity holder in publicly-traded company; Novartis: Research Funding; Magenta Therapeutics: Consultancy, Research Funding; BlueRock: Research Funding; Gadeta: Membership on an entity's Board of Directors or advisory committees. Blazar:Magenta Therapeutics: Consultancy; Fate Therapeutics Inc.: Research Funding; BlueRock Therapeutics: Research Funding; Childrens' Cancer Research Fund: Research Funding; KidsFirst Fund: Research Funding; Tmunity: Other: Co-founder; BlueRock Therapeuetic: Consultancy. Boitano:Magenta Therapeutics: Ended employment in the past 24 months, Patents & Royalties. Cooke:Magenta Therapeutics: Ended employment in the past 24 months, Patents & Royalties. Davis:Magenta Therapeutics: Current Employment, Current equity holder in publicly-traded company.