# In situ assay for topoisomerase 2-mediated DNA damage

> **NIH NIH R43** · VIVID TECHNOLOGIES · 2021 · $236,684

## Abstract

In situ assay for topoisomerase 2-mediated DNA damage
Abstract
 Topoisomerase 2 (Top2) is essential for DNA replication and transcription, but this enzyme
generates double-strand DNA breaks during its normal catalytic cycle. This activity of Top2 is a key
contributor to cancer initiation and progression in several tumor types. Paradoxically, amplification of
Top2 DNA damage became the basis of the most successful strategy for cancer treatment. It
produced some of the best therapeutic agents used to treat human malignancies. Virtually every form
of cancer can be treated with regimens targeting Top2. The development of novel non-toxic and
selective Top2 inhibitors became one of the prioritized targets in precision anticancer medicine.
 Therefore, a technology which detects and quantifies Top2-produced DNA breaks will be broadly
important in both molecular and clinical research fields. The most advantageous method should detect
its target directly in sections of normal and cancer tissues. Such an in situ assay would enable efficient
deciphering of cancer initiation mechanisms and would enhance discovery of new Top2 anticancer
drugs. However, currently there are no in situ assays specific for Top2-mediated DNA breakage.
 The goal of this project is to introduce the first in situ assay for Top2 DNA damage. The new
technology will quantitatively visualize Top2-generated DNA breaks directly in tissue sections and in
individual cells. This new ability is essential for molecular pathology studies and for the assessment of
anticancer therapies. The assay will have high commercial potential because it will offer advantages of
the new detection format, speed, specificity and cost over the current biochemical approaches. It will
use a novel molecular labeling method specific for signature Top2 breaks, which carry homodimeric
adducts linked to extruding 4-nucleotide overhangs on the 5-end of DNA.
 The Specific Aims of this project are:
1. To develop and validate in the context of commercial use the first tissue section technology
specifically labeling Top2-produced DNA breaks with 5’Top2 adducts and 4-base overhangs. The
technology will use the new and original in situ 5’ tyrosine enabled labeling method. To employ model
systems with controlled production of Top2 DNA breaks to validate labeling specificity of the new
approach, and to ensure its adequate sensitivity and reliability of detection.
2. To apply the new imaging technology to fixed tissue sections. To validate the assay in the context of
commercial use in models related to cancer. To assess and verify the specific, robust, and sensitive
detection of Top2 DNA cleavage by using tissue sections with Top2 DNA breaks. To test and optimize
sensitivity, specificity and commercial applicability of the new assay.

## Key facts

- **NIH application ID:** 10323156
- **Project number:** 1R43GM143957-01
- **Recipient organization:** VIVID TECHNOLOGIES
- **Principal Investigator:** KM Wahidur Rahman
- **Activity code:** R43 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $236,684
- **Award type:** 1
- **Project period:** 2021-09-20 → 2023-09-19

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10323156

## Citation

> US National Institutes of Health, RePORTER application 10323156, In situ assay for topoisomerase 2-mediated DNA damage (1R43GM143957-01). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10323156. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*