# Regulation of Zbtb44-Eomes complex in CD8+T cells and anti-tumor immunity

> **NIH NIH R01** · UT SOUTHWESTERN MEDICAL CENTER · 2021 · $421,793

## Abstract

ABSTRACT
CAR-T cell therapy is an emerging option for cancer treatment, but its efficacy is limited, especially in solid tumors
because the effector CD8+T cells become dysfunctional and exhausted in the tumor microenvironment (TME).
However, the key pathways that define the delicate balance between the effector vs exhausted state of CD8+T
cells remain unclear.
Our preliminary studies demonstrate that sumoylation of the T-box transcription factor, Eomesodermin (Eomes),
facilitates its association with Zbtb44, a member of the ThPOK family of transcription factors. The Zbtb44-Eomes
complex promotes the effector function and anti-tumor activity of CD8+ tumor infiltrated lymphocytes (TILs). In
exhausted CD8+ TILs, the ubiquitin ligase Trim47 targets Zbtb44 for degradation and disrupts the Zbtb44-Eomes
complex. Furthermore, CRISPR-Cas9-mediated inhibition of Trim47 rescues exhausted CD8+ TILs and restores
their effector function. These preliminary findings led us to hypothesize that ubiquitination and sumoylation of
the Zbtb44/Eomes complex are critical molecular events that dictate the effector vs exhaustion of CD8+ TILs
which can be therapeutically targeted.
In Aim1, we will determine the mechanism by which the Zbtb44-Eomes complex promotes effector CD8+T cell
function and anti-tumor immunity. We will use newly generated Zbtb44-/- mice to investigate how sumoylation of
Eomes at Lys(K)-446 facilitates the formation of the Zbtb44-Eomes complex via the SUMO interacting motif
(SIM) within Zbtb44. Further, we will delineate the mechanism by which the Zbtb44-Eomes complex
cooperatively binds to and transactivates the IFN- promoter. In Aim 2, we will determine the mechanism by
which Trim47-mediated ubiquitination of Zbtb44 leads to dysfunction of CD8+T cells. We will investigate how
Trim47, which is upregulated in exhausted (PD1+Tim3+) CD8+ TILs, targets Zbtb44 for ubiquitination at K139 and
promotes its degradation. Using newly generated Trim47-/- mice, we will determine how disruption of the Zbtb44-
Eomes complex leads to the inhibitory transcriptional profile of exhausted CD8+T cells. In Aim 3, we will target
the Zbtb44-Trim47 pathway to promote anti-tumor immunity. We will test the therapeutic potential of blocking
Zbtb44 ubiquitination in CAR-T cells against carcinoembryonic antigen (CEA) in the MC38 and in a patient-
derived xenograft (PDX) colon cancer model.
Completion of these studies will lead to: 1) dissection of the novel Zbtb44-Eomes complex that is critical for
effector CD8+ T cell function, 2) determination of how Trim47-mediated ubiquitination disrupts this complex
leading to alternate transcription profile in exhausted CD8+ TILs, and 3) evaluate the means to target the Zbtb44-
Trim47 pathway to overcome the current limitations of CAR-T cell therapy for solid tumors.

## Key facts

- **NIH application ID:** 10099504
- **Project number:** 1R01AI155786-01
- **Recipient organization:** UT SOUTHWESTERN MEDICAL CENTER
- **Principal Investigator:** Venuprasad K Poojary
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $421,793
- **Award type:** 1
- **Project period:** 2021-03-24 → 2026-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10099504

## Citation

> US National Institutes of Health, RePORTER application 10099504, Regulation of Zbtb44-Eomes complex in CD8+T cells and anti-tumor immunity (1R01AI155786-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10099504. Licensed CC0.

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