Characterization of PARP15 inhibitors
(2025) KEMP30 20251Department of Chemistry
- Abstract
- Introduction: PARP15 belongs to the PARP family, which carries out ADP-ribosylation, a post- translational modification involved in DNA repair. Finding selective inhibitors can help discover more of the role PARP15 plays in our body.
Background: Inhibitors of other PARP family members, such as PARP1, have been studied because of their role in cancer processes. However, PARP15 remains poorly characterized, and no specific inhibitors have been identified so far.
Aim(s): Which molecules are the better PARP15 inhibitors, how do their structures play a role, and how selective are they?
Methods: Human PARP15 ART domain was expressed in E. coli cells and purified with IMAC and SEC. The enzymatic activity of PARP15 was tested with a Western... (More) - Introduction: PARP15 belongs to the PARP family, which carries out ADP-ribosylation, a post- translational modification involved in DNA repair. Finding selective inhibitors can help discover more of the role PARP15 plays in our body.
Background: Inhibitors of other PARP family members, such as PARP1, have been studied because of their role in cancer processes. However, PARP15 remains poorly characterized, and no specific inhibitors have been identified so far.
Aim(s): Which molecules are the better PARP15 inhibitors, how do their structures play a role, and how selective are they?
Methods: Human PARP15 ART domain was expressed in E. coli cells and purified with IMAC and SEC. The enzymatic activity of PARP15 was tested with a Western blot, detecting auto-ADP- ribosylation of the ART domain. A biotinylated NAD+ plate assay was used to test the efficiency of a set of compounds to inhibit PARP15 activity. With the same method, IC50 curves for the best inhibitors were created, and the selectivity of these inhibitors was tested by using them in PARP10 and PARP14.
Results: PARP15 enzymatic activity was confirmed, and compounds TIQ-A, PND, 751, 1093, and 2104 showed the highest inhibition. IC50 curves were created and IC50 values determined for these most potent inhibitors. There was a problem with reproducibility, most likely because of solubility issues, so IC50 values could not be determined for all inhibitors. TIQ-A was found to have the best IC50 value (0,118+/-0,04 μM), followed by PND (1,596+/-0,45 μM) and compound 2104 (2,28+/- 2,8 μM). None of these inhibitors showed selectivity for PARP15.
Conclusion: TIQ-A, PND, and 2104 are the most potent inhibitors for PARP15, although none of them are specific for this enzyme. 751 and 1093 showed the highest inhibition, but the data could be misleading, as the inhibition shown may be due to aggregation. This issue could be solved by replacing the methoxy with a hydroxyl group and analyzing if the loss of activity was due to aggregation. The TIQ-A scaffold with a methoxy group added to the benzene ring would be the most potent structure identified so far, if aggregation were dismissed as the cause of inhibition. (Less) - Popular Abstract
- Testing potential PARP15 inhibitors as a first step towards understanding its biological role
PARP15 is a human enzyme, a type of protein that helps make chemical reactions faster inside our cells. Without enzymes, essential processes in our body would be so slow that they would not be compatible with life. PARP 15 belongs to the PARP family of proteins, known for carrying out ADP-ribosylation, a type of reaction involved in processes like DNA repair. By using alternative repair pathways, some PARPs can help cancer cells avoid the natural process of cell death, known as apoptosis.
Finding effective inhibitors for PARP proteins is necessary, both for understanding the function of PARPs and for developing new treatments. Because of their... (More) - Testing potential PARP15 inhibitors as a first step towards understanding its biological role
PARP15 is a human enzyme, a type of protein that helps make chemical reactions faster inside our cells. Without enzymes, essential processes in our body would be so slow that they would not be compatible with life. PARP 15 belongs to the PARP family of proteins, known for carrying out ADP-ribosylation, a type of reaction involved in processes like DNA repair. By using alternative repair pathways, some PARPs can help cancer cells avoid the natural process of cell death, known as apoptosis.
Finding effective inhibitors for PARP proteins is necessary, both for understanding the function of PARPs and for developing new treatments. Because of their obvious roles in DNA repair and cancer, other PARP members have been well studied, and some of their inhibitors are being tested as potential drugs in cancer therapy. But PARP15's specific role is still unclear, and identifying a selective PARP15 inhibitor would be crucial, as blocking its activity could help uncover its precise function. Given its structural and functional similarities with other well- known PARPs, studying PARP15 could also reveal new insights into cellular mechanisms and potential therapeutic targets.
In this project, we tested a group of compounds with a similar structure to TIQ-A, a known PARP inhibitor previously used to block PARP activity. Our goal was to see if similar molecules could inhibit PARP15. First, we purified PARP15 and confirmed it was active by checking the presence of ADP-ribosylation using a Western blot. Before testing the compounds, we first had to identify a reliable method and set the right conditions to quantify PARP15 activity. This was essential to later see how much the activity was reduced in the presence of inhibitors. We found that several compounds showed low IC50 values, meaning they were effective at inhibiting the enzyme. However, to determine whether these inhibitors were specific to PARP15, we also tested them against PARP10 and PARP14. Unfortunately, none of the most effective compounds were selective for PARP15 alone.
Structural comparisons between inhibitors also showed which type of structures made it easier to inhibit PARP15, and this information can be used for the design of future inhibitors truly selective for PARP15, which could help finally discover its biological role and evaluate its potential as a therapeutic target in the future. (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/9193435
- author
- Milan Fernandez, Ana LU
- supervisor
- organization
- course
- KEMP30 20251
- year
- 2025
- type
- M2 - Bachelor Degree
- subject
- keywords
- Biochemistry, ADP-ribosylation, Biotinylated-NAD+, Inhibitors, PARP15, Selectivity
- language
- English
- id
- 9193435
- date added to LUP
- 2025-06-10 09:22:05
- date last changed
- 2025-06-10 09:22:05
@misc{9193435,
abstract = {{Introduction: PARP15 belongs to the PARP family, which carries out ADP-ribosylation, a post- translational modification involved in DNA repair. Finding selective inhibitors can help discover more of the role PARP15 plays in our body.
Background: Inhibitors of other PARP family members, such as PARP1, have been studied because of their role in cancer processes. However, PARP15 remains poorly characterized, and no specific inhibitors have been identified so far.
Aim(s): Which molecules are the better PARP15 inhibitors, how do their structures play a role, and how selective are they?
Methods: Human PARP15 ART domain was expressed in E. coli cells and purified with IMAC and SEC. The enzymatic activity of PARP15 was tested with a Western blot, detecting auto-ADP- ribosylation of the ART domain. A biotinylated NAD+ plate assay was used to test the efficiency of a set of compounds to inhibit PARP15 activity. With the same method, IC50 curves for the best inhibitors were created, and the selectivity of these inhibitors was tested by using them in PARP10 and PARP14.
Results: PARP15 enzymatic activity was confirmed, and compounds TIQ-A, PND, 751, 1093, and 2104 showed the highest inhibition. IC50 curves were created and IC50 values determined for these most potent inhibitors. There was a problem with reproducibility, most likely because of solubility issues, so IC50 values could not be determined for all inhibitors. TIQ-A was found to have the best IC50 value (0,118+/-0,04 μM), followed by PND (1,596+/-0,45 μM) and compound 2104 (2,28+/- 2,8 μM). None of these inhibitors showed selectivity for PARP15.
Conclusion: TIQ-A, PND, and 2104 are the most potent inhibitors for PARP15, although none of them are specific for this enzyme. 751 and 1093 showed the highest inhibition, but the data could be misleading, as the inhibition shown may be due to aggregation. This issue could be solved by replacing the methoxy with a hydroxyl group and analyzing if the loss of activity was due to aggregation. The TIQ-A scaffold with a methoxy group added to the benzene ring would be the most potent structure identified so far, if aggregation were dismissed as the cause of inhibition.}},
author = {{Milan Fernandez, Ana}},
language = {{eng}},
note = {{Student Paper}},
title = {{Characterization of PARP15 inhibitors}},
year = {{2025}},
}