# Characterization unit

> **NIH NIH U2C** · CHILDREN'S HOSP OF PHILADELPHIA · 2020 · $1,633,223

## Abstract

Abstract
The overarching goal of the Characterization Unit is to capture the molecular and cellular
heterogeneity and clonal evolution of three major pediatric cancers, high-grade glioma
(pHGG), neuroblastoma (NB), and very high-risk acute lymphoblastic leukemia (VHR-
ALL). Although many studies have characterized the interplay between DNA
methylation, chromatin accessibility and transcriptome using bulk tumor samples, how
the epigenome and transcriptome relate at the single-cell level and how such
relationships contribute to cancer heterogeneity is not well understood. In addition to
factors intrinsic to malignant cells, extrinsic factors in the tumor microenvironment also
need to be better characterized at single-cell resolution. With these considerations in
mind, we will apply a two-pronged, spatial multi-omics approach in this project. Our
approach is enabled by the diverse array of cutting-edge single-cell and in situ
technologies developed by our team of investigators, including: massive parallel single
nucleus RNA-Seq, single-cell transposome hypersensitivity site sequencing, multiplexed
single molecule RNA fluorescence in situ hybridization and imaging mass cytometry.
The multidimensional and longitudinal data set will be analyzed and integrated by the
Data Analysis Unit to construct atlases of the tumor ecosystems before and after
therapy. We propose the following four specific aims: 1) To identify somatic and germline
mutations using whole genome sequencing of bulk tumor samples and blood samples.
For each cancer type, we will collect tumor samples from 20 patients before and after
medical therapy. The mutation data will be integrated with single-cell omics data to
construct phylogenetic tree of cancer evolution during therapy; 2) To conduct
transcriptome profiling of both bulk and single-cell tumor samples. For VHR-ALL
samples, we will profile malignant cells only. For NB and pHGG samples, we will profile
malignant cells, immune cells, and stromal cells in the tumor microenvironment; 3) To
conduct epigenomic profiling of both bulk and single-cell tumor samples. We will profile
5-methylcytosine and chromatin accessibility using whole-genome bisulfite sequencing
and transposome hypersensitive site sequencing, respectively; 4) To conduct targeted in
situ RNA and protein assays. A panel of marker genes for each tumor will be nominated
based on computational analysis of the omics data collected in Aims 1-3.

## Key facts

- **NIH application ID:** 10016228
- **Project number:** 5U2CCA233285-03
- **Recipient organization:** CHILDREN'S HOSP OF PHILADELPHIA
- **Principal Investigator:** Kun Zhang
- **Activity code:** U2C (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $1,633,223
- **Award type:** 5
- **Project period:** 2018-09-30 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10016228

## Citation

> US National Institutes of Health, RePORTER application 10016228, Characterization unit (5U2CCA233285-03). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10016228. Licensed CC0.

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