# Optimizing NK Cell-Engaging Bispecific Antibody Therapy Targeting CD33

> **NIH NIH R21** · FRED HUTCHINSON CANCER CENTER · 2021 · $83,621

## Abstract

ABSTRACT
CD33-targeted therapies have long been pursued in acute myeloid leukemia (AML). Longer survival of some
patients treated with the antibody-drug conjugate gemtuzumab ozogamicin (GO) validates this approach, but
many patients with CD33+ AML do not benefit from GO. This has prompted efforts to develop better CD33-
directed therapeutics, including T cell engaging bispecific antibodies (BsAbs). Several agents have recently
entered early phase clinical testing, with initial results indicating some efficacy but also substantial toxicities from
cytokine-release syndrome associated with T cell activation. There is therefore increasing focus on exploring the
utility of other immune cells such as natural killer (NK) cells (e.g. via engagement of CD16), which might
circumvent these limitations. How CD33/CD16-directed therapy can be optimized is unknown. As one limitation
of CD33-targeted therapy, CD33 antibodies (including GO) typically recognize immune-dominant epitope(s)
within the membrane-distal V-set domain. In our preliminary studies with CD33V-set/CD3 BsAbs and artificial
CD33 proteins, however, we have observed enhanced T cell-mediated cytotoxicity with membrane-proximal
binding of CD33. We have therefore generated a series of antibodies against the membrane-proximal C2-set
domain of CD33 that recognize all naturally occurring variants of CD33 (i.e. are “CD33PAN antibodies”) as basis
for novel therapeutics. Our previous studies have also shown that the anti-tumor efficacy of T cell-directed BsAbs
is abrogated by inhibitory T cell co-receptor signaling, e.g. via the PD-L1/PD-1 axis. We further demonstrated
that the use of a paired BsAb which binds and blocks such an inhibitory signal while, in turn, providing co-
stimulation to T cells (e.g. via cross-linking CD28) can convert cellular inhibition into activation and dramatically
increase the efficacy of T cell engaging BsAbs. So far, the role of activating and inhibitory NK receptors and their
corresponding ligands as modulators of BsAb-based NK cell therapy has not been evaluated. Based on our
studies with T cell-engaging BsAbs, we hypothesize CD33/CD16-directed NK cell engaging therapy can be
optimized by membrane-proximal targeting of CD33. We also predict that activating and inhibitory NK cell ligands
modulate the anti-tumor efficacy of CD33/CD16 BsAbs and that, consequently, the resistance to CD33/CD16
BsAbs can be reversed by pairing with a BsAb that binds/blocks an inhibitory NK cell ligand and stimulates an
activating NK cell receptor. We will test these hypotheses in well-controlled preclinical studies in vitro and in vivo.
Upon completion of the proposed research, it is our expectation that we have gained critical insight into how the
anti-AML efficacy of CD33/CD16 NK cell-engaging therapeutics can be maximized. Our work is anticipated to
have an important positive impact because it may form the basis for optimized NK cell engaging therapeutics for
patients with AML and other CD33+ ...

## Key facts

- **NIH application ID:** 10601351
- **Project number:** 6R21CA259779-02
- **Recipient organization:** FRED HUTCHINSON CANCER CENTER
- **Principal Investigator:** Roland Bruno Walter
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $83,621
- **Award type:** 6
- **Project period:** 2021-07-01 → 2023-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10601351

## Citation

> US National Institutes of Health, RePORTER application 10601351, Optimizing NK Cell-Engaging Bispecific Antibody Therapy Targeting CD33 (6R21CA259779-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10601351. Licensed CC0.

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