# Defining Pre-treatment Correlates of Patient GD2 CAR T Cell Exhaustion and Memory Using Multi-Dimensional Immune Profiling

> **NIH NIH K08** · STANFORD UNIVERSITY · 2024 · $212,080

## Abstract

PROJECT SUMMARY
 While chimeric antigen receptor T cells (CAR-Ts) have provided impressive responses in hematologic
malignancies, children and young adults with metastatic or relapsed solid tumors have not yet benefited from
CAR-Ts and continue to suffer dismal outcomes. A major barrier to CAR-T efficacy in solid tumors is inadequate
CAR-T expansion, driven in part by CAR-T exhaustion and poor memory potential. Preclinically, CAR-T
exhaustion results from excessive CAR signaling, and can be rescued by eliminating the CD28 T cell
costimulatory domain from CAR design. Clinically, in samples from general pediatric oncology patients, poor
memory potential in T cells is associated with chemotherapy exposure. Preclinical models further suggest that
CAR-T exhaustion can be reduced by overexpressing a transcription factor, cJun, and that memory potential can
be enhanced by exposing T cells to a drug, Ibrutinib. Based on these observations, Dr. Ramakrishna will apply
novel single-cell and multi-dimensional technologies to CAR-T patient samples to assess the impact of CAR
costimulatory domain or pre-apheresis chemotherapy exposure on CAR-T exhaustion and memory potential and
ultimately on patient CAR-T fitness, defined as CAR-T molecular signature paired with functionality. To
accomplish her aims, Dr. Ramakrishna will innovatively compare CAR-T samples across three GD2 CAR-T
clinical trials. In Aim 1, with training in multi-dimensional data analysis, Dr. Ramakrishna will integrate phenotypic
(CyTOF), epigenetic (ATACseq), and transcriptomic (RNAseq) molecular signature with CAR-T functional
assessments to determine whether GD2.Ox40.CD28.z CAR-Ts (NCT02107963) confer exhaustion, thereby
affecting CAR-T fitness, as compared to GD2.41BB.z CAR-Ts (NCT04539366) in osteosarcoma patients. In Aim
2, with training on cancer immune biology, Dr. Ramakrishna will use CyTOF, ATACseq, and in vitro cytokine
production, to test the hypothesis that apheresis and CAR-T products from chemotherapy-naïve patients (diffuse
midline glioma; NCT04196413) have improved CAR-T fitness as compared to those from chemotherapy-treated
patients (osteosarcoma; NCT04539366), followed by single-cell RNAseq to track persistent CAR-T
transcriptional profiles in patients. In Aim 3, with training in immune regulation, Dr. Ramakrishna will evaluate
whether CAR-T fitness in chemotherapy-treated patient aphereses can be enhanced through modulating
molecular pathways by cJun or Ibrutinib. To build upon her substantial prior CAR-T research and clinical
experience, Dr. Ramakrishna has developed a strong training plan and mentorship team, including her primary
mentor, Dr. Crystal Mackall, a pioneer in translational immunotherapy research; co-mentor, Dr. Sean Bendall,
an innovator in multi-dimensional immune assays; advisory committee, Dr. David Miklos, Dr. Holden Maecker,
and Dr. Michelle Monje; and collaborators, Dr. Rosie Kaplan and Dr. Steven Feldman. In completing her
proposed plan with this team, ...

## Key facts

- **NIH application ID:** 10828387
- **Project number:** 5K08CA267057-03
- **Recipient organization:** STANFORD UNIVERSITY
- **Principal Investigator:** Sneha Ramakrishna
- **Activity code:** K08 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $212,080
- **Award type:** 5
- **Project period:** 2022-05-01 → 2027-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10828387

## Citation

> US National Institutes of Health, RePORTER application 10828387, Defining Pre-treatment Correlates of Patient GD2 CAR T Cell Exhaustion and Memory Using Multi-Dimensional Immune Profiling (5K08CA267057-03). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10828387. Licensed CC0.

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