# Overcoming tumor heterogeneity in glioblastoma with multi-targeted CAR T cells secreting bispecific antibodies

> **NIH NIH R01** · MASSACHUSETTS GENERAL HOSPITAL · 2022 · $556,372

## Abstract

PROJECT SUMMARY
 Glioblastoma (GBM) is uniformly lethal and is the most common malignant primary brain tumor.
Immunotherapy promises a precise approach, and the hope of durability. One way to deliver precision
immunotherapy is with genetically-engineered T cells designed to express a target-specific chimeric antigen
receptor, or CAR. In 2017, CAR T cells targeting CD19 were approved by the FDA for B cell malignancies, and
several CARs targeting GBM have been described recently. We have developed CARs that target the EGFRvIII
tumor mutation, and demonstrated their activity in preclinical studies and in a first-in-human clinical trial. We
found that peripheral infusion of CART-EGFRvIII cells was safe and led to elimination of EGFRvIII-expressing
glioma cells in patients. However, despite CART-EGFRvIII trafficking to intracranial tumors and targeting of
EGFRvIII, the patients ultimately had outgrowth of EGFRvIII-negative disease and tumor progression. Thus,
while the CAR T cell platform certainly holds great promise, a critical barrier to clinical impact for brain tumors is
targeting a single antigen in an inherently heterogeneous disease. In addition, our study also demonstrated an
adaptive increase in immunosuppression within the tumor microenvironment; specifically, the endogenous T cell
infiltrate increased in the tumor, but consisted largely of immune-suppressive regulatory T cells (TRegs), rather
than effector T cells reflective of antitumor epitope spreading. To simultaneously address antigenic heterogeneity
and promote local antitumor activity in GBM, we have now modified CART-EGFRvIII to secrete bispecific
antibodies known as bispecific T cell engagers (BiTEs) against wild-type EGFR, which is not expressed in the
normal brain but is nearly always expressed in GBM. Delivering BiTES to the brain using T cells as carriers is
also attractive because antibodies do not effectively cross the blood-brain barrier. The overall goal of this work
is to develop a safe and effective immunotherapy for patients with GBM. We test the hypothesis that anti-
EGFRvIII CAR T cells designed to secrete anti-EGFR BiTEs (CAR-BiTE) will lead to potent and durable
responses in models of heterogeneous GBM. We will test their mechanism of action by quantifying secretion of
BiTEs and systematically testing the role of bystander T cells (Aim 1). Next, we will determine the optimal route
of administration of CAR-BiTE products, and the pharmacokinetics and biostribution of these two-in-one active
"drugs” (Aim 2). These data are expected to lead to an Investigational New Drug (IND) application and Phase I
clinical trial of CART-vIII/BiTE-EGFR in patients with recurrent glioblastoma. Finally, CAR-BiTEs represent a
platform that can target multiple combinations of antigens. In Aim 3 we will identify targetable antigens in primary
glioma samples and test CAR-BiTEs targeting three or more antigens.

## Key facts

- **NIH application ID:** 10469337
- **Project number:** 5R01CA238268-04
- **Recipient organization:** MASSACHUSETTS GENERAL HOSPITAL
- **Principal Investigator:** Marcela Valderrama Maus
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $556,372
- **Award type:** 5
- **Project period:** 2019-09-20 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10469337

## Citation

> US National Institutes of Health, RePORTER application 10469337, Overcoming tumor heterogeneity in glioblastoma with multi-targeted CAR T cells secreting bispecific antibodies (5R01CA238268-04). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10469337. Licensed CC0.

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