Efficacy and safety of PARP inhibitors in the treatment of BRCA-mutated breast cancer: an updated systematic review and meta-analysis of randomized controlled trials

ABSTRACT Introduction Poly-ADP-ribose polymerase inhibitors (PARPis) have emerged as a new class of therapeutic agents for breast cancer patients with breast cancer susceptibility gene (BRCA) mutations. However, the efficacy and toxicity of PARPis have not been clearly established. Methods This study comprehensively evaluated the efficacy and safety of PARPis in patients with BRCA-mutated breast cancer. Online databases were systematically searched, and six clinical trials were included. The primary endpoint of efficacy was progression-free survival (PFS), whereas the secondary endpoints were overall survival (OS) and objective response rate (ORR). Additionally, we assessed the safety of PARPis. Results The results of the meta-analysis showed that PARPis can effectively improve the PFS and OS in patients compared with the control group. The pooled HR (PARPi vs control groups) was 0.63 (95% CI, 0.55 − 0.73) and 0.83 (95% CI, 0.73 to −0.95) for PFS and OS, respectively. In safety, PARPis demonstrated controllable adverse reactions. There were no significant differences in overall AEs or grade ≥3 AEs between the PARP inhibitor and control arms. Conclusions Our results confirm the efficacy and safety of PARPis in patients with BRCA-mutated breast cancer, and more specifically clarify the efficacy of PARPis alone or in combination with other chemotherapy drugs.


Introduction
Breast cancer is one of the most fatal malignancies affecting the female population and has surpassed lung cancer in being the most commonly diagnosed cancer worldwide [1]. As per the latest statistics, 287,850 new cases of breast cancer and 43,250 related deaths were estimated in the United States in 2022 [2]. In China, breast cancer in women was estimated to account for 16.72% (306,000) of all new cancers in 2016 [3].
Recently, in addition to early screening, the main treatment measures for breast cancer include breast-conserving surgery, radiotherapy, and mastectomy [4][5][6]. According to previous studies, BRCA is the most common oncogene, and patients with BRCA mutations carry a higher risk of breast cancer [7].
BRCA1/2 is a gene associated with breast and ovarian cancers and plays an important role in homologous recombinant DNA repair, and it is known as a tumor suppressor gene [8]. BRCA1/2 plays a role in homologous repair (HR) by participating in the synthesis of multiprotein complexes that recognize and repair certain damaged broken DNA duplexes and is also involved the protection of stalled replication forks [9,10]. When BRCA1/2 is mutated, it causes homologous recombination repair defects (HRDs), impairs DNA repair, and leads to irregular DNA synthesis, which may increase genomic instability, lead to cell cycle arrest and apoptosis, and increase the risk of malignant tumor development [10][11][12][13].
Poly-ADP -ribose polymerase (PARP) is a class of multifunctional enzymes that plays an important role in the DNA repair pathway by participating in DNA base excision repair and DNA single-strand break repair [14][15][16], whereas PARP inhibitors (PARPis) are highly tumor-specific and are only very sensitive to breast cancers with BRCA mutations [17,18]. Based on the synthetic lethality, PARPis treat breast cancer by inhibiting DNA single-strand break repair in tumor cells defective in homologous recombination, producing double-strand breaks that lead to selective death of BRCA mutant cells [18,19].
PARPis play an important role in the treatment of BRCA1/ 2-mutated breast cancer in studies investigating the molecular targets of breast cancer. Several PARPis have been approved by the Food and Drug Administration (FDA) as clinical treatments for BRCA1/2 mutated breast cancer [20]. In recent years, several clinical trials have shown that PARPis, including, olaparib, veliparib, and talazoparib exert better efficacy and safety in the treatment of BRCA-mutated breast cancer and have great prospects for development. This review will provide an up-to-date and comprehensive evaluation of the efficacy and safety of PARPis in BRCA-mutated breast cancer treatment to provide objective basis for the clinical treatment of breast cancer. The meta-analysis will be conducted on the relevant literatures published so far.

Search strategy
RCTs of PARPis in BRCA-mutated breast cancer were searched from PubMed, Embase, Cochrane Library, Web of Science, and CNKI. In addition, we also searched the minutes of the meeting, including: The American Society of Clinical Oncology (ASCO) and the ESMO and the Clinical Trials-Registration website (http://www. ClinicalTrials.gov) to ensure that our search is comprehensive and comprehensive. The following combination of MeSH-terms and keywords strategy was used: 'Breast Neoplasms,' "Poly(ADP-ribose) Polymerases inhibitors, 'PARPis,' 'talazoparib,' 'olaparib,' 'niraparib,' 'rucaparib,' 'veliparib,' 'BRCA mutation,' 'BRCA-mutated.' Qualified clinical studies were screened according to the inclusion criteria. Two researchers, Sun and Xu independently screened the titles and abstracts of all the citations through literature search. Any disagreements between the reviewers were resolved by consensus through discussion. For duplicated clinical trials, only the most complete or up-to-date publications were included. This systematic review and meta-analysis were carried out following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement recommendations (http://www. prisma-statement.org/).

Inclusion and exclusion criteria
Included trials were required to meet the following criteria: (1) Phase II or III RCTs in which PARPis alone or in combination with other drugs were used as intervention and conventional chemotherapy or placebo were used as controls. (2) Women 18 years or older with BRCA mutations in breast cancer. (3) sufficient data to assess efficacy outcomes (PFS, OS, and ORR) and safety outcomes.
Exclusion criteria were mainly as follows: (1) reviews, metaanalysis, commentaries, or conference abstracts. (2) Phase I clinical trials or single-arm trials. (3) trials with incomplete data. (4) the study evaluating the efficacy of PARPis during adjuvant therapy.

Data extraction
Data extraction and recording were performed independently by two investigators according to inclusion criteria. The following information was acquired from each included study: the trial name, first author, publication time, phase, number in each arm, type of PARPis, type of control groups, HR or relative risk (RR) with 95% CI for OS and PFS analysis, ORR and occurrence of AEs. If the PFS is represented only by the Kaplan-Meier curve, the data is digitized and extracted using Engauge digitizer 4.1 software. In case of trials that did not include all survival analysis, we also reviewed each clinical trial's supplement.

Risk of bias assessment
Two reviewers used the Cochrane Risk of Bias tool to conduct a quality assessment of the risk of bias. Seven items were evaluated according to 'yes' (low bias), 'no' (high bias) and 'unclear' (unclear bias), including sequence generation (selection bias); allocation concealment (selection bias); blinding of participants and personnel (performance bias); blinding of outcome assessment (detection bias); incomplete outcome data (attrition bias); selective outcome reporting (reporting bias), and other potential sources of bias. The total result was presented as percentages in a figure.

Statistical analyses
For dichotomous variables (ORR), the RR and 95% CI were calculated for each study. Analysis of event occurrence time variables using HR and 95% CI (PFS and OS). All data were expressed as the combination of HR or RR and 95%CI, and p <0.05 was statistically significant. We assessed the betweenstudy heterogeneity by using the inconsistency index (I [2] statistic), which estimates the percentage of total variability across all studies [21]. I 2 regarded an estimated value applied three fixed knots at 25%, 50% and 75% as an indicator of mild, moderate, and high heterogeneity. If the test showed I 2 >50% or p <0.10, the data were calculated through a random-effects (RE) model [22]. Otherwise, a fixed-effects (FE) model was used to pool effect size [22]. To deeply explore the heterogeneity and its potential influence, subgroup analysis was performed. Meta-regression analysis was employed to examine which characteristics might be the possible source of heterogeneity. In addition to, publication bias was also estimated by Egger's test and Begg's test [23,24]. Sensitivity analysis, which examined the robustness of included trials to different aspects from methodological bias. All p-values were two-sided, and all statistical analyses were performed using Review Manager 5.3 and Stata 12.0 software.

Literature selection and study characteristics
We used our search formula ( Figure 1) to identify 1153 studies, of which 6 were conference abstracts. According to inclusion and exclusion criteria, 6 studies from randomized controlled trials (RCTs) were eventually included in this meta-analysis. The specific search and study selection process is shown in Figure 1. Among them, there are two EMBRACA tests, and one of them has updated the OS [25,26]. It is noteworthy that, iniparib was not included as a PARPi in the search formula because previous evidence suggests that iniparib is not a real PARP inhibitor [27,28].
These included RCTs published in 2016-2021. A total of 1477 patients with BRCA mutations from phase II and III clinical trials were included in this meta-analysis. The characteristics of the studies and enrolled patients included in this meta-analysis are listed in Table 1. Three of these studies investigated the PARPi in combination with chemotherapy, and three evaluated the efficacy of PARPis alone. BROCADE, which is a randomized, partially blind phase II clinical trial (NCT01506609) assessed the safety and efficacy of intermittent veliparib with carboplatin/paclitaxel (VCP) or temozolomide (VT) [29]. BROCADE3 is a randomized, double-blind, placebocontrolled phase III clinical trial. Therefore, veliparib or placebo in combination with carboplatin and paclitaxel to evaluate the efficacy of platinum in combination with PARPis [30]. Kummar et al. conducted a randomized phase II trial of veliparib in combination with cyclophosphamide or cyclophosphamide alone [31]. Three other studies assessed the efficacies of monotherapy with veliparib or olaparib. Additionally, we included a recently published phase III randomized trial: the OlympiA trial [32], which is a large international randomized trial that evaluated 1-year adjuvant therapy with olaparib vs placebo after chemotherapy and local treatment in germline BRCA mutation (gBRCAm) carriers with human epidermal growth factor receptor 2 (HER2)-negative breast cancer. In March 2022, the European Society of Medical Oncology (ESMO) virtual plenary was updated with a significant overall survival benefit (hazard ratio [HR] 0.68, p = 0.009) [33]. The risk of bias for each study was assessed according to the Cochrane Manual 5.1.0 assessment criteria ( Figure 2). Randomization was unclear in four studies, and two studies had other unclear risk of bias, although the risk of bias is generally low.

Primary endpoint: PFS
Progression-free survival (PFS) was the primary endpoint of most studies, although this cannot be obtained in one study (OlympiA) [32]. As disease-free survival (DFS) was the primary endpoint of the present study, we excluded this study and pooled the results of five other studies. Due to the moderate heterogeneity (I 2 = 29%, p = 0.23), we chose the fixed-effects model. From the forest map, we found that PARPis were closely related to the improvement of PFS, with HR 0.63 ([95% CI, 0.55-0.73], p < 0.00001; Figure 3a). Additionally, PARPis, either as a single-agent or in combination, significantly prolonged PFS in patients compared with control groups (HR 0.56 [95%CI, 0.46 to 0.68], p < 0.00001; HR 0.71 [95%CI, 0.59 to 0.85], p = 0.00002, respectively), with no significant difference in the benefit of PFS between monotherapy and combination therapy (p = 0.09; Figure 3b). Additionally, we compared the median PFS data provided in the selected studies (Table 2). Median survival was significantly longer in the PARPi arms than in the placebo or chemotherapy arms.
In breast cancer, both olaparib and talazoparib have been approved for the treatment of gBRCAm carriers with metastatic HER2-negative breast cancer [25,34]. On 11 March 2022, the FDA approved olaparib as the an adjunctive treatment of HER2-negative in adults with gBRCAm who are at high risk for early breast cancer and who have previously received neoadjuvant or adjuvant chemotherapy [35]. Veliparib, although not approved by the FDA, has shown promising results in several clinical trials in gBRCAm carriers with metastatic HER2negative breast cancer [36,37]. Therefore, we focused on the PFS benefits for HER2-negative breast cancer in the subgroup analysis. In a subgroup analysis based on hormone receptor status, we found an exciting correlation between PARPis and improved PFS in patients with both triple negative breast cancer (TNBC) and HER2-negative hormone receptor (HR)positive breast cancer (HR 0.59 [95%CI, 0.49 to 0.72], p < 0.00001; HR 0.66 [95%CI, 0.54 to 0.80], p < 0.00001, respectively; Figure 3c) . Furthermore, there is no significant heterogeneity (I [2] = 41%, p = 0.15; I 2 = 48%, p = 0.14).
Patients with breast cancer with BRCA1/2 mutations are more sensitive to platinum drugs such as cisplatin and carboplatin than the wild type [38,39]. The effect of prior use of platinum therapy was also analyzed. However, PARPis significantly improved PFS in patients not receiving platinumbased therapy with an HR of 0.64 ([95% CI, 0.55 to 0.75] p < 0.00001; Figure 3d). In patients who had received platinum therapy, the risk of disease progression was also statistically significant in the PARP inhibitor group (HR 0.70 [95%CI, 0.53 to 0.91]). As for BRCA mutation status, PARPis provided benefits uniformly in BRCA mutations 1 or 2 (Supplementary Figure 1a).

Secondary endpoint: OS and ORR
We included three single-agent studies and two combination chemotherapy studies to analyze overall survival (OS) without significant heterogeneity (I 2 = 0%, p = 0.51). The pooled HR    Overall, five studies provided data for objective response rate (ORR) analysis. The pooled results showed a significant correlation between ORR and the experimental arms (RR 1.55 [95% CI, 1.02 to 2.34], p = 0.04, I 2 = 90%;), with high heterogeneity (Figure 4c). Subgroup analysis revealed that monotherapy (RR 2.21 [95% CI, 1.73 to 2.84]) had a higher ORR than combination therapy (RR 1.11 [95% CI, 0.93 to 1.33]); Figure 4d). Surprisingly, subgroup analysis based on hormone receptor status indicated that ORR was significantly different between patients with hormone-receptor positive and triple negative breast cancers (RR 7. 10 FigureS1d). Although the sample size was limited, the ORR was higher in patients with TNBC than in HR-positive breast cancer, and there was no significant subgroup difference between the two (p = 0.10, I 2 = 62.6%).

Safety
Apart from efficacy, we also examined the possible adverse reactions of PARPis, thus, safety analysis. The comparative safety profile of the (adverse events) AEs of interest is shown in Table 3 and Supplementary Figure 1. Overall, forest plot results showed there is no difference in the probability of AEs between the PARPi arms and placebo or chemotherapy arms, regardless of whether it was AEs of any grade with RR 1.03  Figures 5a, b). RE model was used due to high heterogeneity (I [2] = 97%, respectively). In the subgroup analyses of AEs of any grade, the incidence of AEs with monotherapy and combination therapy was similar and there was no difference between each and their control groups (RR 1.04 [95% CI, 0.96 to 1.12], p = 0.34 and RR 1.02 [95% CI, 0.92 to 1.14], p = 0.68; Figure 5c). The results of the subgroup with AE grade ≥ 3 were the same as indicated above (Figure 5d). In the subgroup analysis, patients with early-stage breast cancer had lower grade ≥3 AEs than did those with metastatic breast cancer (RR 0.83 [95% CI, 0.57 to 1.21], p =      FigureS2a). There was a statistically significant difference between the two groups, which may be due to the small sample size (p < 0.00001). Furthermore, we assessed the safety of different PARPis shown in Table 4. Olaparib has a higher risk than the other two drugs for any type of AEs of grade

Sensitivity analyses and publication bias
Sensitivity analysis was used to assess the impact of individual studies on the overall results. The results showed that except for NCT02163694 for ORR and NCT02032823 for AEs, a single study did not significantly change the overall results of HRs (for PFS and OS) and RRs (for ORR and AEs), which demonstrated the robustness of the analysis (Supplementary Figure 2). We evaluated the publication bias of the included literatures using Begg's Funnel Plot and Egger's Test, and the results showed that there was no publication bias in PFS,OS, ORR and AEs (Begg's funnel plot p = 0.806 for PFS, p = 0.806 for OS, p = 0.806 for ORR, p = 0.452 for AEs; Egger's test p = 0.410 for PFS, p = 0.665 for OS, p = 0.186 for ORR, p = 0.12 for AEs; Supplementary Figure 3).

Discussion
In conclusion and to the best of our knowledge, this is the most recent and comprehensive meta-analysis known to evaluate the efficacy and safety of PARPis in BRCA-mutated breast cancer. Our results suggest that PARPis, both as a monotherapy or combination therapy, demonstrates a strong and excellent therapeutic effect, significantly improves survival, and is well tolerated by patients with possible toxicity.
As olaparib entered clinical trials for the first time in 2009, PARPis have gradually become emerging targets for cancer treatment in the past decade [40]. In 2018, olaparib and talazoparib were approved for the treatment of with HER2-negative advanced or metastatic breast cancer patients with gBRCAm. As research continues, PARPis are also being evaluated in combination with other drugs. In the Phase III BROCADE3 trial, the addition of veliparib to carboplatin and paclitaxel significantly improved median PFS compared to placebo added to carboplatin and paclitaxel, with encouraging results. In two previously reported meta-analyses, monotherapy with PARPis improved PFS but not OS [41,42]. By contrast, our meta-analysis showed significant improvement in both OS and PFS in monotherapy with PARPis. Our metaanalysis also evaluated the efficacy of PARPis in combination with chemotherapeutic agents. Although OS and ORR did not improve, PFS had significant benefits (HR 0.71 [95%CI, 0.59 to 0.85], p = 0.00002), which is consistent with the stratified results of our PARPis combination therapy [43]. Meanwhile, our analysis also noted that there was no significant difference in the benefit between monotherapy and combination therapy, which may provide strong evidence for future clinical drug combination.
Additionally, our meta-analysis provides perspectives on the patients who may benefit more from PARPis. In previous meta-analyses, only patients with TNBC achieved statistically significant improvement in PFS based on a subgroup analysis of the hormone receptor status [41]. However, our results suggest that patients with hormone-receptor-positive tumors also benefit in the form of better PFS (HR 0.66 [95%CI, 0.54 to 0.80], p < 0.00001) and ORR (RR 2.99 [95%CI, 1.81 to 4.93], p < 0.0001). According to the guidelines, platinum-based chemotherapy is the preferred option for patients with advanced breast cancer associated with gBRCAm [44]. It is important to note that PFS results were significantly improved compared to PARPis and chemotherapy in patients with or without prior platinum-based therapy, which also differs from what has been reported in previous meta-analyses [41,42].
As PARPis are gradually approved for clinical use, their safety and tolerability in patients are of great value and indispensable significance. In general, PARPis appear to be safe and well tolerated in patients with breast cancer despite the hematologic gastrointestinal adverse effects, such as anemia, leukopenia, nausea, and vomiting, consistent with previous reports; especially in patients with early-stage breast cancer, adverse effects are relatively mild. Evaluation of three different PARPis for safety showed that olaparib had the most severe adverse reactions, whereas talazoparib and veliparib had less severe adverse reactions. This may be because the control group for olaparib in the latest trial we included was placebo, so there was an increased risk of adverse effects. The reactions may also be related to different doses of PARPis. The current approved dose of olaparib is 300 mg twice daily, whereas talazoparib, the strongest PARP trapper, has the lowest recommended dose [45]. This meta-analysis has several limitations. As with other reported meta-analyses, the data obtained are based on research-level evidence rather than individual patient data results and lacks some raw data. Second, ORR and AEs have high data heterogeneity, which may introduce selection bias associated with the results. Finally, single-arm and Phase I trials were excluded from this meta-analysis, which may lead to some limitations in the assessment of safety.
There have been many clinical trials exploring the efficacy of different PARPis in breast cancer. These included phase III trials of niraparib in patients with HER2-negative and gBRCAm breast cancer (BRAVO) [46]. Additionally, the role of PARP inhibitors in neoadjuvant therapy for patients with breast cancer has been evaluated; for example, BrighTNess and GeparOLA studies [47,48]. It is believed that as clinical trials get concluded, PARPis in the treatment of breast cancer will be more outstanding in performance.

Conclusions
Our results confirm and strengthen the efficacy and safety of PARPis in patients with BRCA-mutated breast cancer, and more specifically clarify the efficacy of PARPis alone or in combination with other chemotherapy drugs. However, beyond the approved indications, the therapeutic value of PARPis for patients with other types of breast cancer needs to be further evaluated in future studies.