Abstract
Background/Aim: The emergence of novel DNA damage repair (DDR) pathways in molecular-target therapy drugs (MTTD) has shown promising outcomes in treating patients with metastatic castration-resistant prostate cancer (mCRPC). About 25% of mCRPC patients have actionable deleterious aberrations in DDR genes, primarily in the homologous recombination (HR) pathway. However, the response rate in patients with BRCA1/2 or mutations in HRR-related genes is only 45%-55%, when exposed to poly ADP ribose polymerase (PARP) inhibitor-based therapy (PARPi). A frequent characteristic feature of prostate cancer (PC) is the occurrence of genomic rearrangement that affects the transmembrane protease serine 2 (TMPRSS2) and E26 transformation-specific (ETS)- transcription factor-related gene (ERG). Materials and Methods: In this study, a total of 114 patients with mCRPC had their RNA and DNA sequenced using next-generation sequencing. Results: Based on their genetic profile of deleterious gene alterations of BRCA1/2 or ATM, six patients were selected for PARPi. Patients with TMPRSS2:ERG gene fusion and homozygous alteration in ATM or BRCA2 (n=2) or heterozygous alterations (BRCA1 or BRCA2) and lack of TMPRSS2:ERG gene fusion (n=2) did not show clinical benefit from PARPi (treatment duration <16 weeks). In contrast, patients (n=2) without TMPRSS2:ERG gene fusion and homozygous deleterious alterations in ATM or BRCA2 all had clinical benefit from PARPi (treatment duration ≥16 weeks). Conclusion: The TMPRSS2:ERG transcript product might be used as a PARPi resistance biomarker.
- Prostate cancer
- targeted therapeutics
- next-generation sequencing
- gene fusion
- DNA repair gene
- PARPi
- ATRi
- resistance
Prostate cancer (PC) accounts for a high number of cancer-related deaths in men worldwide. In the USA, the PC incidence rate was 110.5 per 100,000 men during 2014-2018, while the death rate was 18.9 per 100,000 men during 2015-2019 (1). These figures are comparable to those observed in the Danish population.
A hallmark of PC is the high dependency on the androgen receptor (AR) for growth and survival (2). Androgen deprivation therapy (ADT) remains the backbone of systemic therapy in patients with localized and advanced disease. In addition, novel AR-targeting agents such as abiraterone acetate and enzalutamide are now administered in patients with metastatic castration-resistant PC (mCRPC) (3). Abiraterone acetate inhibits CYP17A1 (both 17a–hydroxylase and 17,20-lyase), which is responsible for androgen biosynthesis, whereas enzalutamide binds to the androgen receptor, reduces the efficiency of its nuclear translocation, and impairs both DNA binding to androgen response elements and recruitment of coactivators (4, 5). Treatment options in mCRPC also include the use of taxanes (docetaxel and cabazitaxel), which inhibit mitosis by stabilizing microtubules, but also AR nuclear translocation, thereby reducing AR signaling (2). Although the introduction of radiopharmaceuticals such as Radium-223 and Lutetium-177-PSMA-617 has further expanded available treatment options, overall survival in patients with mCRPC remains poor (6).
The emergence of novel DNA damage repair (DDR) pathways in molecular-target therapy drugs has shown promising outcomes in mCRPC patients (7-9). Approximately 25% of mCRPC patients have actionable deleterious aberrations in the DDR genes where BRCA1/BRCA2 somatic or germline variants account for 6-14% of the cases (10). The DDR mechanisms include single-strand break (SSB) repair via base excision repair (BER), intra-strand crosslinks bulky adducts repair by the nucleotide excision repair (NER), inter-strand crosslinks repair by the Fanconi Anemia (FA) pathway, and replication error repair via mismatch repair (MMR). The repair of double-strand breaks (DSB) in DNA relies on two distinct mechanisms, classic-non homologous end joining (c-NHEJ) and homologous recombination (HR). HR is activated during the S/G2 phase of the cell cycle and requires a sister chromatid as a template. On the other hand, NHEJ is a non-allelic homologous recombination between non-allelic, or non-identical, DNA sequences and is typically limited to the G1 phase of the cell cycle (11). This process may lead to the formation of PC-specific gene fusions, where segments of two different genes are joined together (12).
Defects in the FA, BER, and NER pathways are infrequently observed in mCRPC, though PARP1 has a distinct role as a DNA damage sensor in SSB repair and protects against the generation of excessive SSB during BER (13). Exome and genome sequencing of patients with mCRPC has revealed a low tumor mutational burden (TMB) even in heavily pre-treated patients. However, these tumors have a higher burden of chromosomal structure variants compared to many other cancers, including insertions, deletions, inversions, translocations, gene-fusions, and tandem duplications (14).
Three main kinase signal transducers, the ataxia-telangiectasia mutated kinase (ATM), Ataxia telangiectasia and Rad3-related kinase (ATR), and DNA-dependent protein kinase (DNA-PKcs) transmit the DNA damage through different repair pathways (15). The ATM kinase acts on DNA with DSB that have single-stranded overhangs and can initiate the HR repair pathway. On the other hand, the DNA-PKcs kinase acts on DNA with DSB that has blunt ends and can initiate the c-NHEJ repair pathway. Finally, the ATR kinase acts on DNA with SSB and initiates repair mechanisms for this kind of DNA damage. Two molecular-target therapy drugs (MTTDs) that have shown promising benefits in mCRPC act on different repair pathways. The TRESR trail showed that the ATR inhibitors (ATRi) are synthetic lethal with the loss of function of ATM kinase, with the highest clinical benefit in tumors with biallelic loss of function alterations (9). The phase III PROFOUND trial demonstrated that mCRPC patients with BRCA1, BRCA2, or ATM mutations may benefit from PARPi (10). However, patients with ATM and BRCA2 mutations may respond to PARPi in a more divergent fashion (16) and responses to PARPi have been observed in patients with other loss-of-functions mutations in DDR pathway genes (17). Furthermore, the inactivation of both alleles in the CRPC cells may increase the likelihood of clinical benefit from PARPi (18).
A common genetic rearrangement found in PC involves the transmembrane protease serine 2 (TMPRSS2) and a member of the erythroblastosis virus E26 transformation-specific (ETS) transcription factor family. In about 50% of PC cases, the 5′ untranslated end of TMPRSS2 is observed to be translocated to the ETS-related gene (ERG) (19, 20). TMPRSS2 has also been found to fuse with other ETS members (ETV1, ETV4, and ETV5) but at a much lower frequency in PC cases (20). Several studies have shown that the TMPRSS2:ERG fusion protein is present not only in mCRPC but also in high-grade prostate intra-epithelial neoplasia and benign prostatic hyperplasia, linking this gene fusion to an early and common event in PC development and progression (21). This ERG gene fusion with TMPRSS2 is regulated by androgens (22). Almost all tumors that over-express ERG are found to have TMPRSS2:ERG fusion (20).
Studies on the TMPRSS2:ERG fusion gene have mostly focused on its role in PC initiation, and progression, and its use as a diagnostic tool. However, its direct impact on DNA repair mechanisms has been less studied. Mass spectrometry has revealed interactions between PARP1, ERG, Ku80, Ku70, and the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) in ERG gene fusion-positive cases, suggesting an important role of ETS members in DNA repair (23). This interaction is facilitated by the Ets-1 (ETS domain) on the ERG gene and the domain BRCT and SAP on the PARP1 and Ku70, respectively. This interaction has been found to play a role in cancer (24). The BRCT domain of PARP1 is involved in PARylation, which is important in the recruitment of DNA repair enzymes at the SSB and DSB sites. In contrast, the recruitment of DNA repair to c-NHEJ relies on the phosphorylation of DNA-PK for activity, and PARylation of the DNA-PK by PARP1 can inhibit the activity of DNA-PK (25-27). This study aimed to examine possible explanations of differences in clinical response to PARPi in patients with mCRPC presenting a mutation in DDR genes.
Materials and Methods
Patients. From 1 October 2015 to 31 June 2021 a total of 114 patients with mCRPC were referred to the Experimental Cancer Therapy Unit at Herlev and Gentofte Hospital. Out of the 114 patients, a total of 8 patients were selected for treatment with MTTD. One of the patients treated with olaparib was excluded due to inclusion in an on-going sponsored trial, thus seven patients were included in the analysis. All patients had experienced disease progression following available standard treatments including AR-targeting agents (Abiraterone and/or Enzalutamide), chemotherapy (docetaxel and cabazitaxel), and Radium-223. The MTTD consisted of either PARPi alone (olaparib) off-label therapy, an ATR inhibitor (ATRi) alone (camonsertib) in the TRESR trial (NCT04497116) or immunotherapy (avelumab) in combination with a PARPi (talazoparib) in the Javelin PARP trial (NCT03330405). Treatment duration (PARPi) of 16 weeks or more was defined as indicative of clinical benefit.
The research was carried out following the guidelines of the International Conference of Harmonization of Good Clinical Practice and the Declaration of Helsinki. This retrospective review was approved by the Danish Patient Safety Authority (3-3013-2793/1).
DNA and RNA extraction. An area of cancerous tissue that consists of a minimum of 20% tumor cells was identified by a specialized pathologist and subsequently extracted by 1-mm disposable punchers to ensure a high content of tumor cells. Genomic DNA and RNA were extracted using Maxwell® RSC DNA FFPE Kit (Promega, Madison, WI, USA) or Maxwell® RSC RNA FFPE Kit (Promega) following the manufacturer’s instructions. DNA and RNA concentration was quantified using Qubit™ dsDNA High-Sensitive Assay kit (Thermo Fisher Scientific, Waltham, MA, USA) or Qubit™ RNA High Sensitivity (HS) Assay kit (Thermo Fisher Scientific) on a Qubit fluorometer (Thermo Fisher Scientific).
NGS library preparation and sequencing. NGS sequencing libraries were prepared for therapy treatment decisions using one of three assays; Oncomine Focus Assay targeting 53 unique genes (pt: 1) (Thermo Fisher Scientific), Oncomine Comprehensive Assay v2 targeting 143 unique genes (pt. 2) (Thermo Fisher Scientific) or Oncomine Comprehensive Assay v3 targeting 161 unique genes (pt. 3, 4, 5, 6, and 7) (Thermo Fisher Scientific). All library preparations were performed manually, and PCR amplification was carried out using a DNA concentration of 20 ng as input or 20 ng RNA treated with SuperScript VILO cDNA synthesis kit (Thermo Fisher Scientific), according to manufacturer’s instructions MAN0015819 for the Oncomine Focus Assay and MAN0015885 for the Oncomine Comprehensive Assays. The prepared libraries were loaded onto Ion 540™ Chips before 2017 and Ion 550™ Chips (Thermo Fisher Scientific) thereafter, using the Ion Chef™ System according to the manufacturer’s instructions (MAN0010851 and MAM0017275, respectively). Sequencing was performed using the Ion S5™ XL Sequencer (Thermo Fisher Scientific) MAN0010811.
Data analysis. All data analyses were carried out using Ion Reporter Software (Thermo Fisher Scientific). The analyses were conducted using the human hg19 as the standard reference genome. Two workflows – Oncomine Focus – DNA and Fusion or Oncomine Comprehensive – DNA and Fusion were utilized to identify somatic variants [single nucleotide variation (SNV), multi nucleotide variation (MNV), insertion-deletion (indel), copy number variation (CNV)] and RNA gene fusions. The Ion reporter’s coverage analysis reports were also used to evaluate the quality of the sequencing reactions by measuring mapping reads, mean depth, uniformity, and alignment over a target region. The genetic variants observed were clinically annotated using the implemented ClinVar database or the database available at VarSome following the ACMG Guideline (28).
Results
AR gene amplification status and MTTD treatment. None of the seven patients who received MTTD had any pathogenic or likely pathogenic variants in the AR gene (data not shown). Six patients (pt. 2, 3, 4, 5, 6, and 7) with an AR copy number greater than 5 (CNV >5) showed no association with clinical benefit from MTTD. Additionally, a single patient (pt. 1) with CNV below 5 showed no clinical benefit from MTTD (Table I). Furthermore, there was no association between TMPRSS2:ERG gene fusion and the presence of AR gene amplification, regardless of the clinical benefit from MTTD treatment (Table I).
Patient treatment and genetic data.
Mutated BRCA1/2 and ATM gene status effect on MTTD treatment. In this analysis, it was observed that three patients (pt. 3, 5, and 7) had a homozygous genetic alteration in the ATM gene, where gene deletion results in the loss of the unaffected ATM allele. Of these patients, two (pt. 3 and 5) received a PARPi, but only one showed clinical benefit. Another patient (pt. 7) with a homozygous genetic alteration in the ATM gene was treated with an ATRi and showed clinical benefit. Two other patients (pt. 1 and 2) treated with PARPi had a germline heterozygous BRCA1 gene variant that caused premature termination of the BRCA1 gene but still had a functional unaffected second BRCA1 allele (pt. 1). The PARPi did not provide any clinical benefit in this patient. The second patient (pt. 2) had a deletion of the entire BRCA2 gene with a functional, unaffected second BRCA2 allele remaining. The PARPi did not show any clinical benefit for pt. 2. Three patients (pt. 4, 5, and 6) had genetic alternations in the BRCA2 gene. Among these, one patient (pt. 5) had a somatic deletion of the entire BRCA2 gene but still had an unaffected second BRCA2 allele. This patient also had a homozygous ATM mutation, which might be the reason for the clinical benefit from PARPi. The other two patients (pt. 4 and 6) with BRCA2 mutations had a deletion of the unaffected BRCA2 allele. One patient (pt. 6) showed clinical benefit from PARPi, whereas pt. 4 had no clinical benefit from PARPi (Table I and Figure 1).
Schematic presentation of the PARP inhibitor-based therapy (PARPi) and ATR inhibitors-based therapy (ATRi) flow and genetic alteration. Black persons: metastatic castration-resistant prostate cancer (mCRPC) patients without BRCA1, BCRA2, or ATM genetic aberrations, which allow molecular-target therapy drug (MTTD) treatment. Red persons: mCRPC patients treated with MTTD with a lack of clinical benefit. Green persons: mCRPC patients treated with MTTD showing clinical benefit. Black person with question mark: excluded due to inclusion in an on-going clinical trial. Light blue and dark blue arrows: patients that might have benefit from ATRi.
TMPRSS2:ERG gene fusion status and MTTD treatment. Of the seven patients (pt. 1, 2, 3, 4, 5, and 6) who received PARPi, only two had TMPRSS2:ERG gene fusions (pt. 3 and 4). Two patients (pt. 1 and 2) with heterozygous BRCA1 mutation or heterozygous BRCA2 deletion, were both negative for TMPRSS2:ERG gene fusion and lacked a response to PARPi. Two patients (pt. 4 and 6) had homozygous alternations of the BRCA2 gene and loss of the unaffected BRCA2 allele. The main genetic difference between the two patients was the presence of the TMPRSS2:ERG gene fusion in the patient who had no clinical benefit from PARPi (pt. 4), whereas pt. 6 had clinical benefit. Two other patients (pts. 3 and 5) in the study had homozygous alterations of the ATM gene and loss of the unaffected ATM allele. One of them (pt. 3) showed no clinical benefit from PARPi, whereas the other (pt. 5) had clinical benefit. The main genetic difference between the two patients was the absence of the TMPRSS2:ERG gene fusion in the patient who had clinical benefit (pt. 5). A patient (pt. 7) had ATM aberration and TMPRSS2:ERG gene fusion, which is a profile identical to pt. 3 who had no clinical benefit from PARPi. Pt.7 received an ATRi and was treated for more than 16 weeks, indicating a clinical benefit from this treatment (Table I and Figure 1).
Discussion
Precision medicine holds great potential for cancer patients but identifying DNA repair mechanisms through biomarkers like homologous recombination deficiency (29), microsatellite instability (30), and tumor mutational burden (31) is challenging. To better understand how recurrent TMPRSS2:ERG gene fusion affects treatment with MTTD, we manually datamined the clinical outcomes of seven mCRPC patients who underwent experimental treatment with PARPi or ATRi. Most PC patients treated with ADT initially responded but unfortunately progressed to mCRPC over time. During this transition, the PC cells adapt to the low levels of testosterone, which can lead to the restoration of AR signaling through various mechanisms, such as AR amplification, AR mutation, or generation of AR splice variants (32). Indeed, AR amplification is a common genetic alteration, particularly observed in mCRPC patients as compared to patients with untreated PC (33). Our study showed that 6 out of 7 patients (86%) who underwent treatment with MTTD had AR amplification (CNV>5), which is consistent with the results from other studies (25, 34).
Research has shown that the clinical benefit of PARPi is predominantly observed in patients with tumors harboring a biallelic loss of function alterations (35). Therefore, the two patients (pt. 1 and 2) with only mono-allelic loss or loss of function may have low clinical benefit.
Several studies have shown that the TMPRSS2:ERG fusion-gene is present in PC patients who have not yet received treatment. This indicates that the gene fusion is not an AR resistance-related event. Most research on this gene fusion has focused on its role in PC initiation, progression, and potential as a diagnostic tool. However, less attention has been given to its impact on DNA repair. It has been shown that the TMPRSS2:ERG protein interacts with PARP1 and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) in a DNA-independent manner (23).
DNA repair mechanisms are crucial for maintaining genomic stability and preventing the accumulation of alterations that could lead to cancer. DDR signaling recognizes DNA breaks and arrests cell cycle progression to promote DNA repair; the three main kinase signal transducers ATM, ATR, and DNA-PKcs mediate this DRR signal. The ATM is recruited to DSBs in response to repair through HR, the ATR is recruited to ssDNA sites to activate single-stranded repair, and the DNA-PKcs are responsible for repairing through c-NHEJ (36). Some genetic changes, such as gene fusions, can impact DNA repair, but it is not yet clear how TMPRSS2:ERG affects this process. Our results may indicate one possible explanation for how TMPRSS2:ERG may affect DNA repair mechanisms.
The over-expression of the TMPRSS2:ERG protein can significantly increase the level of PARylation through its interaction with PARP1, independent of DNA. However, this effect can be reversed by introducing PARPi (37). When repairing DNA damage caused by platinum chemotherapies, the NER repair pathway is typically used for bulky lesions. Unlike PARP1, which acts as a sensor for DNA damage, the DDB1 and DDB2 complex is responsible for bulky adduct detection in NER. In cases where TMPRSS2:ERG fusion leads to hyper-activated PARylated PARP1, it can interact with the DDB1/DDB2 complex, enhancing the activity of other DNA repair proteins to complete NER (13). This could in part explain why many patients with mCRPC do not respond well to platinum-based chemotherapy (38).
It has been observed that the increased activity of PARP1 due to TMPRSS2:ERG expression leads to a significant increase in the activity of BER, which repairs non-bulky lesions. This happens through the recruitment of XRCC1 and LIG3 by PARylated PARP1 (37, 39). Additionally, the abortive TOP1cc can also cause SSB repair that is fixed by BER (13). The classical c-NHEJ operates in the G1 phase of the cell cycle, where the Ku70/Ku80 heterodimer recognizes DSBs, followed by DNA-PKcs recruitment. The DSB is then shielded by 53BP1, which inhibits extensive end resection (40). In the presence of TMPRSS2:ERG, the c-NHEJ repair is blocked by the interaction with PARP1 through DNA-PKcs in a DNA-independent event though DNA-PKcs is activated by phosphorylation (26), which prevents the recruitment of XRCC4 (23, 41). This interaction only occurs in TMPRSS2:ERG gene fusion-positive cases (23). As a result, the alt-NHEJ pathway is activated, which uses MRE11-RAD50-NBS1 as a sensor and DNA end processing and is recruited by PARylated PARP1 (13, 40, 42). However, this often leads to small deletions (43). If the resection generates short single-strand DNA, the overhang is not protected by RPA or RAD51, which makes the alt-NHEJ independent of ATRIP/ATR and BRCA2/PALB2 (44). On the other hand, if the extensive resection generates a large single-strand DNA overhang, it is recognized by the RPA proteins, and the removal of the non-homologous ssDNA overhang is mediated by RAD52 and ERCC1/XPF. Any gaps left over are filled with polymerase to complete the single-strand annealing, which often results in a large deletion (43).
During the S and G2 phases of the cell cycle, BRCA1 works against 53BP1 to promote HR through its interaction with the PARylated PARP1 and the BRCA1 partner protein BRAD1 (45). This interaction limits the resection size of the DSB (46). The BRCA1/BARD1 complex binds to the DSB and recruits BRCA2/PALB2 to replace RPA with RAD51, thus making the DSB ready for strand invasion (13).
Of the seven patients who received PARPi treatment, two were selected due to their heterozygous germline BRCA1 mutation or heterozygous deletion of BRCA2, respectively. As expected, they did not respond to the treatment because an effect on tumor suppressor genes typically requires a secondary event on the unaffected allele (47).
One of the two patients who were homozygous BRCA2 mutated and treated with PARPi had clinical benefit. Only the patients without TMPRSS2:ERG gene fusion demonstrated clinical benefit. In patients with TMPRSS2:ERG gene fusion the c-NHEJ is inactivated by the ERG/DNA-PKcs interaction with PARylated PARP1 and the HR is blocked due to the lack of BRCA2, which makes the cell incapable of exchanging RPA with RAD51. However, the RPA-protected single-strand DNA might instead promote the use of single strand alignment (SSA) and/or alt-NHEJ due to the activation by PARylated PARP1. Additionally, the BER pathway is highly activated due to the PARylated PARP1 interaction with DDB1/DDB2, which lowers the possibility of DSB due to replication fork stress. Treatment with PARPi will impair the PARylation of PARP1, and thereby allow the PARP1/ERG/DNA-PKcs to form a c-NHEJ active complex. The RPA-bound single-stranded DNA will still promote the use of SSA, which can exchange the RPA with RAD52 and initiate repair independent of PARP1. This suggests that TMPRSS2:ERG helps the cell to escape the synthetic lethality in BRCA2 mutated mCRPC by either reactivating the c-NHEJ pathway in the presence of PARPi or inducing an alternative DNA repair pathway other than c-NHEJ and HR by changing the PARylated stage of PARP1 to a normal level of uPARylated PARP1 (37). This is in line with the observation that PARPi only inhibits ETS-positive, but not ETS-negative, prostate cancer xenograft growth (23).
The response to PARPi treatment differed between two homozygous ATM mutated patients. Only the patient without TMPRSS2:ERG gene fusion had a response. It appears that the TMPRSS2:ERG gene fusion in combination with an PARPi might reactivate the c-NHEJ pathway or trigger an alternative DNA repair pathway, which is different from c-NHEJ and HR. These pathways are blocked by the ERG/DNA-PKcs interaction with PARylated PARP1 or by the lack of ATM to phosphorylate the H2AX and activate the HR pathway (36). This might promote the BER pathway to be activated due to the PARylated PARP1 interaction with DDB1/DDB2 complex leading to SSB at a low level and reducing the possibility of DSB due to replication fork stress. In the absence of the HR pathway, RAD51 is incapable of protecting the single-stranded DNA, and instead, RPA promotes the use of alt-NHEJ and/or SSA. Administering PARPi treatment to the patient with homozygous ATM mutation and TMPRSS2:ERG fusion might normalize the PARylated stage of PARP1 and reactivate the c-NHEJ, thus enabling the cell to evade synthetic lethality (37) (Figure 1). We also observed that patients with homozygous ATM mutations and potentially BRCA2 mutations in combination with TMPRSS2:ERG gene fusion might benefit better from an alternative approach such as ATRi (Table I and Figure 1), due to the expected higher level of RPA single-stranded DNA.
While these results are interesting, it is important to acknowledge several limitations of this study, including the very small sample size and lack of confirmatory data on de- or up-regulation of the DDR pathway in the cases studied. The important use of TMPRSS2:ERG gene-fusion in patient selection has been demonstrated in an exploratory TALAPRO-2 study, which also highlighted the need of a control arm when analyzing response data (48). We have not studied the significance of the various types of TMPRSS2-ERG gene-fusions. These fusions can occur in different ways, such as through a 3.0 Mbp interstitial deletion, translocation, or through microdeletion and simultaneous translocation. The majority of cases are generated by deletions, and these cases appear to have a poorer prognosis compared to cases with a fusion resulting from translocation (49). The significantly shorter time to recurrence observed in patients with high expression of the TMPRSS2-ERG fusion transcript was not taken into consideration, as reported (50). To gain more insight, validation in larger cohorts will be needed with supportive markers for DDR pathway activity.
In conclusion, we observed a limited clinical benefit when administering PARPi to patients with homozygous ATM or BRCA2 mutations in combination with TMPRSS2:ERG gene fusion. It is important to confirm the findings in larger studies, as they may have a potential impact on the selection of patients likely to benefit from PARPi.
Footnotes
Authors’ Contributions
T.S.P., A.N.L., P.K., R.L.E., M.H., and E.V.H. all contributed to the conceptualization, design, review, and editing of the drafted manuscript of the study. T.S.P. performed analysis, visualizations, and writing of the original draft. All Authors approved the final version.
Conflicts of Interest
The Authors have declared no conflicts of interest in relation to this study. MH has served as a principal investigator on the TRESR trial for Repare Therapeutics. MH has received research funding (institutional) from Repare Therapeutics.
- Received July 24, 2024.
- Revision received August 20, 2024.
- Accepted August 21, 2024.
- Copyright © 2024 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).
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