# Investigating how chemotherapeutic thiopurines inhibit telomerase elongation of telomeres

> **NIH NIH F32** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2024 · $76,756

## Abstract

Project Summary/ Abstract
Telomere maintenance at chromosome ends is essential for the growth of cancer cells. In humans, telomeres
end in a single strand overhang consisting of GGTTAG repeats that form the substrate for telomerase.
Telomerase is a reverse transcriptase composed of two subunits: a protein component (TERT) and an RNA
component (TR) which contains an integral template used to reverse transcribe multiple telomeric repeats
during a single telomere binding event. Telomerase is an attractive target for cancer therapies because it is
expressed in over 85% of cancer cells, while most adult somatic cells lack telomerase. Thiopurines are a class
of nucleoside analogs used to treat leukemia and some pediatric cancers but are highly toxic. The prodrug 6-
thio-2'-deoxyguanosine (6-thio-dG) was developed to reduce non-specific thiopurine toxicity. This drug was
shown to successfully reduce the growth of mouse tumor xenografts for multiple cancer types. 6-thio-dG
treatment can also cause telomere shortening and dysfunction, however, the mechanism was unknown until
recently. In biochemistry experiments, I discovered that human telomerase can readily add the 6-thio-dG
metabolite, 6-thio-dGTP, to a growing telomere chain, but this insertion then strongly inhibits telomerase’s
ability to add additional telomeric repeats. I found that POT1-TPP1, which normally enhances telomerase
binding and the addition of multiple repeats, cannot restore telomere elongation in the presence of 6-thio-
dGTP. In addition, 6-thio-dGTP has a low micromolar IC50 for human telomerase due to telomerase’s inability
to discriminate between dGTP and 6-thio-dGTP but does not inhibit DNA polymerase progression, reducing its
off-target impact. My preliminary data for this project demonstrate that 6-thio-dG can disrupt telomerase
binding when the modified base is located at the 3’ end of the telomeric overhang. I hypothesize that 6-thio-dG
addition impairs telomerase’s interaction with the telomere, thereby disrupting cycling for repeat addition
synthesis and inhibiting telomerase elongation of telomeres in cells. I will use complementary biochemical,
single-molecule, and cellular approaches to test my hypothesis. Aim 1 will elucidate the mechanism by which
6-thio-dG disrupts the telomerase catalytic cycle. Aim 2 will determine whether telomerase can synthesize
DNA and extend telomeres in cancer cells treated with 6-thio-dG. Aim 3 will determine if 6-thio-dG efficacy in
cancer cells is enhanced by inhibiting the thiopurine sanitase, NUDT15.
Completing this project will improve our understanding of how 6-thio-dG impacts telomerase activity and
telomere maintenance in cancer cells. It will also enhance our understanding of how 6-thio-dGTP nucleotide
impacts telomerase catalysis and its potential utility as a therapeutic for halting cancer cell proliferation. This
fellowship will allow me to acquire new skills related to the project’s aims and outlines specific mentors for
each. Ca...

## Key facts

- **NIH application ID:** 10833022
- **Project number:** 5F32CA275287-02
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Samantha Sanford
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $76,756
- **Award type:** 5
- **Project period:** 2023-05-01 → 2026-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10833022

## Citation

> US National Institutes of Health, RePORTER application 10833022, Investigating how chemotherapeutic thiopurines inhibit telomerase elongation of telomeres (5F32CA275287-02). Retrieved via AI Analytics 2026-06-01 from https://api.ai-analytics.org/grant/nih/10833022. Licensed CC0.

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