XmAb™ Fc Engineered Anti-CD19 Monoclonal Antibody with Enhanced In Vitro Efficacy Against Lymphoma and Leukemia.
Authors: Zhukovsky Eugene , Chu Seung , Bernett Matthew , Karki Sher , Richards John , Pong Erik , Peipp Matthias , Repp Roland , Desjarlais John 
Published: April 2020

Abstract
CD19 is a pan-B cell surface receptor expressed from early stages of pre-B cell development through terminal differentiation into plasma cells. It is an attractive target for cancers of lymphoid origin since it is expressed on the vast majority of Non-Hodgkin Lymphoma (NHL) as well as different types of leukemia, including those (e.g. pediatric ALL) lacking CD20 expresion. The majority of NHL patients will relapse after the current treatment regimen of chemotherapy combined with rituximab (CHOP-R) despite major improvements in response rates and progression free survival. Thus salvage regimens with novel non-cross resistant antibody therapies are warranted. Here we present the characterization of a novel Fc-engineered and humanized anti-CD19 antibody (XmAb™ CD19) that was generated using our XmAb™ antibody engineering technologies. This novel antibody is highly cytotoxic against a panel of lymphoma and leukemia cell lines as well as primary cancer cells. The main features of this antibody are:
increased affinity for Fc gamma receptors (FcgR), improved effector function, and significantly increased antitumor potency.
Humanization and affinity maturation technologies were applied to this antibody in order to:
decrease immunogenicity, increase affinity, and increase stability of the engineered antibody.
Since internalization is expected to impact a naked antibody’s effector functions, we assayed its internalization rate using Eu-labeled XmAbCD19 and observed an unexpectedly low rate of internalization. Therefore, we proceeded to investigate several direct and indirect (Fc-mediated) mechanisms of antibody-mediated cytotoxicity. The potency of XmAbCD19 in antibody-dependent cell-mediated cytotoxicity (ADCC) increased 10- to 100-fold relative to a native/non Fc-engineered version (CD19-IgG1) of the antibody in a screen of 16 NHL and leukemia cell lines (chronic lymphocytic leukemia [CLL], B-cell acute lymphoblastic leukemia [B-ALL], hairy cell leukemia [HCL], follicular lymphoma [FL], mantle cell lymphoma [MCL], chronic myelogenous leukemia [CML], and Burkitt’s lymphoma [BL]). ADCC potency (EC50) and efficacy (% Lysis) of the Fc-engineered anti-CD19 antibody were superior to that of rituximab: in CLL - 10- and 1.5-fold higher, in B-ALL - 10- and 100-fold higher, and in HCL - 6- and 1.2-fold higher, respectively; unlike XmAbCD19 native CD19-IgG1 mediated little ADCC. Moreover, XmAbCD19 mediated potent ADCC of primary patient-derived ALL cells that was also significantly increased in potency and efficacy relative to rituximab and CD19 IgG1. Furthermore, XmAbCD19 demonstrated 50-fold increased antibody-dependent cellular phagocytosis (ADCP) relative to CD19-IgG1. Finally, XmAbCD19 exhibited robust anti-proliferative activity that was 10-fold more potent than that of rituximab. In order to facilitate preclinical toxicology studies with XmAbCD19, we assayed its cross-reactivity with non-human primate (NHP) species CD19 (cynomolgus and rhesus monkeys). In contrast to the parental murine antibody, the affinity-optimized humanized XmAbCD19 reacted with B cells from both NHP species. In summary, our data suggest that our anti-CD19 Fc variant antibody engineered for increased effector function is a promising next-generation immunotherapeutic for a variety of leukemias and lymphomas.