# Intra and Extra-Chromosomal Probes for Mutagenesis by Carcinogens

> **NIH NIH R01** · MASSACHUSETTS INSTITUTE OF TECHNOLOGY · 2024 · $368,363

## Abstract

Project Summary/Abstract
Human cancer genomes show a complex pattern of mutations that, upon computational deconvolution, resolves
into a systematic series of over 80 “Mutational Signatures” (the pattern of mutations across all possible
trinucleotide sequence contexts). Some signatures are common to many cancers (e.g., CGàTA in CpG sites)
whereas others appear in single tumor types (e.g., signatures involving GCàTA mutations that are presumably
associated with aflatoxin B1 (AFB1) exposure in hepatocellular carcinoma (HCC)). Current sequencing efforts
sometimes provide hints, via examination of the details of mutational signatures, as to cancer mechanistic
etiology. The first two Aims of the present proposal are a bottom up approach to determine which specific
chemical insults to DNA cause mutational patterns that match mutational signatures in tumors. We have
formulated a model that describes three variables that contribute to the complexity of mutational spectra: The
likelihood that a DNA lesion will form in a particular context, the likelihood that it will evade repair, and the
likelihood that it will miscode when traversed by a polymerase. Aim 1 uses a combination of synthetic and
analytical chemistry to generate the adduct-formation spectrum of a host of DNA damaging agents (AFB1,
sterigmatocystin, N-methyl-N-nitrosourea, streptozotocin, temozolomide, chloroacetaldehyde and the oxidant
SIN-1). A strategy involving the use of heavy isotope containing defined-sequence oligonucleotides will be used
along with mass spectrometry to map the binding specificities of electrophiles derived from these agents. Aim 2
takes a two-pronged approach to define the biological effects of DNA damage from the studied agents. Since
the DNA adducts of the compounds evaluated are known, we shall insert those adducts one at a time into
defined-sequence oligonucleotides in all 16 possible 3-base contexts (i.e., 5’-NXN-3’, where X is the adduct and
N is A, G, C or T). The oligonucleotides will be inserted into viral genomes, replicated in cells of defined repair
or replication status, and the areas of the genome that contained the lesion will be characterized to define the
type, amount, genetic requirements for - and sequence context dependency of - mutagenesis. The second part
of Aim 2 is to use a newly developed mouse embryo fibroblast (MEF) line to define, using Duplex Consensus
Sequencing (DCS), the high-resolution mutational spectra (HRMS) of the agents under study. These data are
compared via informatics techniques (e.g., cosine similarity) to human cancer mutational signatures and to the
DNA binding (Aim 1) and mutagenic properties of individual adducts. Lastly, a damaged nucleotide pool is an
often-overlooked source of mutations. Aim 3 uses the MEF line of Aim 2 to probe the opportunity of a pool
damaged with products of oxidative stress (e.g., 8-oxoG and 5-chlorocytosine) to impact the HRMS of the MEF
line. Further, we have custom-designed a pool mutagen (fKP1...

## Key facts

- **NIH application ID:** 10857294
- **Project number:** 5R01CA080024-27
- **Recipient organization:** MASSACHUSETTS INSTITUTE OF TECHNOLOGY
- **Principal Investigator:** JOHN M ESSIGMANN
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $368,363
- **Award type:** 5
- **Project period:** 1998-07-01 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10857294

## Citation

> US National Institutes of Health, RePORTER application 10857294, Intra and Extra-Chromosomal Probes for Mutagenesis by Carcinogens (5R01CA080024-27). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10857294. Licensed CC0.

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