ADP-ribose Research Articles

Niraparib enhances antitumor immunity and contributes to the efficacy of PD-L1 blockade in cervical cancer

PurposeWith the development of immunotherapy research, the role of immune checkpoint blockade (ICB) in the treatment of cervical cancer has been emphasized, but many patients still can’t receive long-term benefits from ICB. Poly ADP ribose polymerase inhibitor (PARPi) has been proved to exert significant antitumor effects in multiple solid tumors. Whether cervical cancer patients obtain better benefits from the treatment regimen of PARPi combined with ICB remains unclear.MethodsThe alteration of PD-L1 expression induced by niraparib in cervical cancer cells and its underlying mechanism were assessed by western blot and immunofluorescence and quantitative real-time polymerase chain reaction (qRT–PCR).The regulation of PTEN by KDM5A was confirmed using Chromatin immunoprecipitation (ChIP) assay and RNA interference. Analyzing the relationship between PD-L1 and immune effector molecules through searching online databases. Therapeutic efficacy of niraparib, PD-L1 blockade or combination was assessed in syngeneic tumor model. The changes of immune cells and cytokines in vivo was detected by immunohistochemistry (IHC) and qRT–PCR.ResultsWe found that niraparib upregulated PD-L1 expression and potentiated the antitumor effects of PD-L1 blockade in a murine cervical cancer model. Niraparib inhibited the Pten expression by increasing the abundance of KDM5A, which expanded PD-L1 abundance through activating the PI3K-AKT-S6K1 pathway. PD-L1 was positively correlated with immune effector molecules including TNF-α, IFN-γ, granzyme A and granzyme B based on biological information analysis. Niraparib increased the infiltration of CD8+ T cells and the level of IFN-γ, granzyme B in vivo.ConclusionOur findings demonstrates the regulation of niraparib on local immune microenvironment of cervical cancer, and provides theoretical basis for supporting the combination of PARPi and PD-L1 blockade as a potential treatment for cervical cancer.

Stratification of Homologous Recombination Deficiency-Negative High-Grade Ovarian Cancer by the Type of Peritoneal Spread into Two Groups with Distinct Survival Outcomes.

Homologous recombination deficiency (HRD) has evolved into a major diagnostic marker in high-grade ovarian cancer (HGOC), predicting the response to poly (adenosine diphosphate-ribose) polymerase inhibitors (PARPi) and also platinum-based therapy. In addition to HRD, the type of peritoneal tumor spread influences the treatment response and patient survival; miliary type tumor spread has a poorer predicted outcome than non-miliary type tumor spread. Known methods for HRD assessment were adapted for our technical requirements and the predictive-value integrated genomic instability score (PIGIS) for HRD assessment evolved as an outcome. PIGIS was validated in HGOC samples from 122 patients. We used PIGIS to analyze whether the type of tumor spread correlated with HRD status and whether this had an impact on survival. We demonstrated that PIGIS can discriminate HRD-positive from HRD-negative samples. Tumors with a miliary tumor spread are HRD-negative and have a very bad prognosis with a progression-free survival (PFS) of 15.6 months and an overall survival (OS) of 3.9 years. However, HRD-negative non-miliary spreading tumors in our cohort had a much better prognosis (PFS 35.4 months, OS 8.9 years); similar to HRD-positive tumors (PFS 34.7 months, OS 8.9 years). Our results indicate that in a predominantly PARPi naïve cohort, the type of tumor spread and concomitant cytoreduction efficiency is a better predictor of survival than HRD and that HRD may be an accidental surrogate marker for tumor spread and concomitant cytoreduction efficiency. It remains to be determined whether this also applies for sensitivity to PARPi.

Synergistic cytotoxicity of histone deacetylase and poly-ADP ribose polymerase inhibitors and decitabine in pancreatic cancer cells: Implications for novel therapy.

Histone deacetylase inhibitors (HDACi) can modulate the acetylation status of proteins, influencing the genomic instability exhibited by cancer cells. Poly (ADP ribose) polymerase (PARP) inhibitors (PARPi) have a direct effect on protein poly (ADP-ribosyl)ation, which is important for DNA repair. Decitabine is a nucleoside cytidine analogue, which when phosphorylated gets incorporated into the growing DNA strand, inhibiting methylation and inducing DNA damage by inactivating and trapping DNA methyltransferase on the DNA, thereby activating transcriptionally silenced DNA loci. We explored various combinations of HDACi and PARPi +/- decitabine (hypomethylating agent) in pancreatic cancer cell lines BxPC-3 and PL45 (wild-type BRCA1 and BRCA2) and Capan-1 (mutated BRCA2). The combination of HDACi (panobinostat or vorinostat) with PARPi (talazoparib or olaparib) resulted in synergistic cytotoxicity in all cell lines tested. The addition of decitabine further increased the synergistic cytotoxicity noted with HDACi and PARPi, triggering apoptosis (evidenced by increased cleavage of caspase 3 and PARP1). The 3-drug combination treatments (vorinostat, talazoparib, and decitabine; vorinostat, olaparib, and decitabine; panobinostat, talazoparib, and decitabine; panobinostat, olaparib, and decitabine) induced more DNA damage (increased phosphorylation of histone 2AX) than the individual drugs and impaired the DNA repair pathways (decreased levels of ATM, BRCA1, and ATRX proteins). The 3-drug combinations also altered the epigenetic regulation of gene expression (NuRD complex subunits, reduced levels). This is the first study to demonstrate synergistic interactions between the aforementioned agents in pancreatic cancer cell lines and provides preclinical data to design individualized therapeutic approaches with the potential to improve pancreatic cancer treatment outcomes.

Financial Toxicity of Withdrawn Poly (Adenosine Diphosphate Ribose) Polymerase Inhibitor Indications for Ovarian Cancer

ObjectivesWe sought to quantify exposure to and financial impacts of poly (adenosine diphosphate ribose) polymerase inhibitor (PARPi) treatments for eventually withdrawn ovarian cancer indications. MethodsWe identified in Optum’s deidentified Clinformatics® Data Mart database 1695 patients with ovarian cancer diagnoses who received olaparib, rucaparib, or niraparib between January 2015 and September 2021. We describe PARPi use and out-of-pocket, total healthcare, and PARPi spending among patients with ovarian cancer with 3 or more previous lines of therapy. ResultsOf the 1695 patients who received PARPi, 254 were estimated to have been heavily pretreated and exposed to eventually withdrawn indications. Cumulative total medical and pharmacy costs for these patients were $53 392 184; PARPi costs accounted for 34%. Median PARPi cost per patient was $43 347. Cumulative out-of-pocket costs totaled $533 281. ConclusionsPotential patient harm, including financial toxicity, might have been mitigated through more stringent drug approval requirements.

Association of homologous recombination repair mutation defined with a 15-gene panel with the prognosis of epithelial ovarian cancer: A Chinese multicenter retrospective study.

e17556 Background: There was no consensus regarding the optimal homologous recombination repair (HRR) gene panel to predict prognosis of epithelial ovarian cancer (EOC). Methods: The genetic testing results in a 15-gene panel from 308 EOC patients diagnosed between 2014 and 2022 from six centres were collected. The association of clinicopathologic characteristics, the use of poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitors and progression-free survival (PFS) with 15-gene panel mutation status was assessed. Results: 43.2% (133/308) of patients were determined to carry 144 deleterious HRR mutations (HRRm), among which 68.1% (98/144) were germline mutations and 32.8% (101/308) were BRCA1/2 gene lethal mutations. Patients who received PARP inhibitors (PARPis) were 29.1% (37/127) and 50.5% (46/91) in the HRR wild-type (HRRwt) group and HRRm group at IIIC-IV stages, respectively. A prognostic difference was observed only between the BRCA mutation group and the HRRwt group, not between the non-BRCA HRRm group and the HRRwt group. In univariate analysis, significant predictors of relapse included nonresidual status (HR, 0.62; 95% CI, 0.39-0.98; P = 0.0287), 15-gene panel mutation (HR, 0.42; 95% CI, 0.28-0.64; P = 0.0001), platinum sensitivity (HR, 0.07; 95% CI, 0.02-0.22; P < 0.001) and maintenance therapy (PARPi) (HR, 0.48; 95% CI, 0.31-0.74; P = 0.0028) (Figure 5). In multivariate Cox analysis, nonresidual (HR, 0.63; 95% CI, 0.40-0.99; P = 0.046), 15-gene panel mutation (HR, 0.49; 95% CI, 0.28-0.88; P = 0.016) and platinum sensitivity (HR, 0.02; 95% CI, 0.01-0.05; P < 0.001) remained independent prognostic factors. Conclusions: This study provides evidence that HRRm with a 15-gene panel can predict the prognosis of EOC, among which only BRCA1/2 mutations contribute to prognosis prediction. [Table: see text]

Clinical homologous recombination repair gene reversion analysis identifies mechanisms of resistance to PARP inhibitors and platinum‐chemotherapy

AbstractIdentifying mechanisms underlying cancer resistance to therapy is vital for advancing treatment strategies. Pathogenic mutations of homologous recombination repair (HRR) genes are known biomarkers for platinum (Pt)‐based chemotherapy and poly ADP ribose polymerase inhibitors (PARPi) effectiveness. Yet, the dynamics of HRR reversion mutations, which may herald therapy resistance, are not fully elucidated. Addressing this gap, our study analyzed secondary HRR gene mutations in a comprehensive pan‐cancer data set of approximately 13,000 patients who underwent targeted next‐generation sequencing. We identified a subset of patients harboring secondary mutations, which were further categorized into three tiers based on their nature, and occur in the presence of a primary pathogenic mutation, notably in BRCA1, BRCA2, PALB2, and RAD51D genes. Here we show that secondary BRCA2 mutations, indicative of adaptive resistance, emerge post‐Pt/Olaparib treatment. This challenges the prevailing notion that pathogenic HRR mutations uniformly predict therapeutic sensitivity, highlighting a nuanced genetic interplay that impacts treatment success. This investigation enriches our understanding of cancer's adaptive mechanisms against therapy, suggesting a pivotal shift towards more personalized, dynamic treatment strategies. It underscores the imperative of adapting to cancer's genetic evolution, aiming for a step ahead in the ongoing battle against this disease.

(CDK) 4/6 inhibitors: An effective treatment for homologous recombination deficient (HRD) luminal breast cancers (BC)?

e13100 Background: Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) with endocrine therapy (ET) still represents the best 1st-line choice for hormone receptor (HR) positive(+)/HER2 negative(-) metastatic breast cancer (MBC) patients (pts). Approximately 8% of pts with HR+ BC harbors germline BRCA mutation (g BRCAmt). The interaction between Cyclin/CDK pathway, endocrine receptor (ER) and Breast Cancer gene 1/2 (BRCA1/2) is complex and evidence reported so far, await confirmation in the context of future randomized clinical trials. So far, limited data exist about treatment effectiveness of CDK4/6i in homologous recombination deficiency (HRD) Luminal BC. A current frequent question in clinical practice is which additional adjuvant therapy to prioritize for luminal BRCA tumors (tu), considering the approval of both CDK4/6i andpoly ADP ribose polymerase (PARP) inhibitors in this scenario. Considering potential impacts of genetic instability in tu biology, such as potential loss of Rb protein related to Loss of Heterozygosity (LOH) of BRCA2 as a primary resistance mechanism in CDK4/6i, we sought to analyze the clinical results of that drug in tu with genetic instability. Methods: We retrospectively analyzed real-world outcomes of CDK4/6i performance in HR+/HER2- MBC treated in a single institution, with focus in clinical outcomes of HRD BC. germline HR genes status and somatic phosphatidylinositol 3-kinase (PIK3CA) status were also analyzed. Results: A total of 384 consecutive MBC pts treated with CDK4/6i were included in this study. The most used CDK4/6i was palbociclib (40,6%), followed by ribociclib (29,8%) and abemaciclib (29,2%). Considering the whole cohort PFSm 30months (95%IC 26-34) and OSm were 56 months (95%IC 52-60) No differences in PFS or OS was observed among the 3 CDK4/6i. Median age of the cohort was 60 years old. 38.5% were premenopause Pts and 59.9% were postmenopause. A total of 149 (39%) pts performed germline screening. Between 19 pts with gHR mt, 12 were BRCA1/2 mt (just one BRCA1). The other were: 2 PALB2, 2 CHEK2, 1 ATM, 1 RAD51D, 1 RAD51C mt. So far, no differences in PFS ou OS was observed in the presence of BRCA 1/2 or other gHR mt. Nevertheless, PIK3CA mt was tested in 142 pts, 51 were positives. That mt was a predictor of worse OS (55m; 95%IC 45-65) x 63m (95%IC 56-69), p 0,019. Conclusions: The results of this real-world study suggest that treatment outcomes with CDK4/6i may be worse in pts with HRD tu due to potential differences in tu biology. Future exploration in larger samples of patients who have had biomarker testing is ongoing. Patients with Pik3CA mt treated with CDK4/6i presented worse clinical outcomes.

TRPM2 channels are essential for regulation of cytokine production in lung interstitial macrophages.

Interstitial macrophages (IMs) are essential for organ homeostasis, inflammation, and autonomous immune response in lung tissues, which are achieved through polarization to a pro-inflammatory M1 and an M2 state for tissue repair. Their remote parenchymal localization and low counts, however, are limiting factors for their isolation and molecular characterization of their specific role during tissue inflammation. We isolated viable murine IMs in sufficient quantities by coculturing them with stromal cells and analyzed mRNA expression patterns of transient receptor potential (TRP) channels in naïve and M1 polarized IMs after application of lipopolysaccharide (LPS) and interferon γ. M-RNAs for the second member of the melastatin family of TRP channels, TRPM2, were upregulated in the M1 state and functional channels were identified by their characteristic currents induced by ADP-ribose, its specific activator. Most interestingly, cytokine production and secretion of interleukin-1α (IL-1α), IL-6 and tumor necrosis factor-α in M1 polarized but TRPM2-deficient IMs was significantly enhanced compared to WT cells. Activation of TRPM2 channels by ADP-ribose (ADPR) released from mitochondria by ROS-produced H2O2 significantly increases plasma membrane depolarization, which inhibits production of reactive oxygen species by NADPH oxidases and reduces cytokine production and secretion in a negative feedback loop. Therefore, TRPM2 channels are essential for the regulation of cytokine production in M1-polarized murine IMs. Specific activation of these channels may promote an anti-inflammatory phenotype and prevent a harmful cytokine storm often observed in COVID-19 patients.

Regulation of carbohydrate metabolism and insulin secretion in diabetic rats following treatment with Hypericum perforatum L. hairy root aqueous methanol extract

Hairy root (HR) cultures of Hypericum perforatum L. have shown promise in controlling hyperglycemia, regulating blood lipid and enzyme profiles, and improving metabolic function in vivo. These benefits are mainly attributed to the phenolic acids, flavonoids, and xanthones present in HR extracts. However, the specific mechanism underlying these effects remains unclear. This study was performed to elucidate the biochemical and molecular mechanisms driving HR the antihyperglycemic and antidiabetic effects of HR extracts. HR extract (200 mg/kg body weight) was administered daily for 14 days to healthy rats and rats with streptozotocin-induced diabetes, with glibenclamide serving as a positive control. The phenolic composition of the HR extracts was confirmed through high-performance liquid chromatography/diode array detection/electrospray ionization mass spectrometry (HPLC/DAD/ESI-MSn ) analysis. The results showed that HR extract treatment increased the plasma insulin level and pancreatic poly (adenosine diphosphate-ribose) polymerase (PARP) activity in diabetic rats, thus normalizing blood glucose levels. Additionally, it reduced the activity of gluconeogenic enzymes, increased the activity of glycolytic enzymes, and normalized the glycogen content in the liver. HR extract-treated rats also exhibited increased hepatic adenosine monophosphate-activated protein kinase (AMPK) mRNA expression and a decreased protein kinase Cε (PKCε) concentration. In conclusion, HR extract demonstrated insulinotropic effects and effectively regulated hepatic carbohydrate metabolism in diabetic rats by modulating AMPK expression and the PKCε concentration. These findings suggest the potential use of HR extract as an herbal medicine for diabetes treatment and a source of antidiabetic drug development.

Real-world drug treatment models of novel targeted drugs in Chinese patients with gynecological cancer from 2017 to 2021: A cross-sectional analysis

ObjectiveThe significance of novel anti-tumor pharmaceuticals in the treatment of gynecological tumors is growing, but there is no consensus regarding the optimal drug delivery strategy for gynecological tumors. This study seeks to investigate the treatment models of novel anti-tumor drugs in patients with gynecological cancer in China over the past five years, with a particular emphasis on the trend and rationality of their use. MethodWe conducted a cross-sectional analysis of data from a China Medical Association-supervised hospital prescription analysis cooperation initiative. The data was derived from prescriptions written for patients diagnosed with cancer between January 2017 and December 2021. The required information for patients was extracted. Our study included 2308 patients that were diagnosed as gynecological tumors which were treated with novel antineoplastic targeted drugs. Patients were categorized by age and region. Then, the selection, application, and indications of the most essential treatment pharmaceuticals were investigated. We evaluated anti-tumor prescription information based on the recommended drug labeling protocol and the most recent domestic and international guidelines.Excel 2013 and SPSS (version 25; SPSS Inc., Chicago, IL, United States) were utilized to conduct statistical analysis.In addition，we also used Sankey diagram to evalute the relation between novel antineoplastic targeted drugs and corresponding diagnoses. ResultThe top three cities for the 2308 patients included in this study were Guangzhou (28.51%), Hangzhou (21.79%), and Beijing (20.06%). In the past five years, the average age of medication patients was 55.61-year-old, with 37.86% of women aged of 51–60. Each patient's primary treatment regimens were statistically analyzed, yielding a total of 16 single-drug and combination-drug primary treatment regimens. Bevacizumab, Olaparib, Trastuzumab, Apatinib, and Arotinib were the top five treatment strategies. The maximum proportion, up to 0.74%, was attributed to the combination of human epidermal growth factor receptor-2 inhibitor (HER2i), including Trastuzumab and Parostuzumab. Vascular endothelial growth factor receptor inhibitor (VEGFRi), including Bevacizumab and Apatinib was the most frequently prescribed medication for outpatients in major cities across the country. According to the 5-year change in time, poly adenosine diphosphate ribose polymerase inhibitor (PARPi) rated first in terms of usage, with Olaparib ranking first with the highest concentration of 33.44% and Niraparib ranking second overall with the fastest growth in 2021. The quantity of VEGFRi variants utilized was the greatest, and their proportion of total usage increased annually. The top five drugs by total drug costs were Bevacizumab, Carelizumab, Olaparib, Trastuzumab, and Apatinib. However, the top five drugs by per capita drug cost were Olaparib + Bevacizumab, Bevacizumab + Sidilimab, Arrotinib + Olaparib, Olaparib, and Patuzumab + Trastuzumab. ConclusionThe incidence rate of gynecological tumor patients rises with age, and the cost of drug treatment has risen annually over the past five years, which is also related to the rising incidence rate of tumors in recent years. Bevacizumab rates first in the drug treatment scheme for the application of novel anti-tumor targeted drugs, which may be related to the widespread use of VEGFRi drugs in gynecological and reproductive tumors. Breast cancer and adenocarcinoma are at the top of the female cancer incidence spectrum, which may explain why HER2i multi-drug combination regimen rates highest among multi-drug combination regimens. Future research may concentrate on how novel anti-tumor targeted drugs can minimize the economic burden and maximize the benefits of patient treatment for patients with gynecological cancer.

Senaparib as first-line maintenance therapy in advanced ovarian cancer: a randomized phase 3 trial.

Poly(adenosine diphosphate-ribose) polymerase (PARP) inhibitors as maintenance therapy after first-line chemotherapy have improved progression-free survival in women with advanced ovarian cancer; however, not all PARP inhibitors can provide benefit for a biomarker-unselected population. Senaparib is a PARP inhibitor that demonstrated antitumor activity in patients with solid tumors, including ovarian cancer, in phase 1 studies. The multicenter, double-blind, phase 3 trial FLAMES randomized (2:1) 404 females with advanced ovarian cancer (International Federation of Gynecology and Obstetrics stage III-IV) and response to first-line platinum-based chemotherapy to senaparib 100 mg (n = 271) or placebo (n = 133) orally once daily for up to 2 years. The primary endpoint was progression-free survival assessed by blinded independent central review. At the prespecified interim analysis, the median progression-free survival was not reached with senaparib and was 13.6 months with placebo (hazard ratio 0.43, 95% confidence interval 0.32-0.58; P < 0.0001). The benefit with senaparib over placebo was consistent in the subgroups defined by BRCA1 and BRCA2 mutation or homologous recombination status. Grade ≥3 treatment-emergent adverse events occurred in 179 (66%) and 27 (20%) patients, respectively. Senaparib significantly improved progression-free survival versus placebo in patients with advanced ovarian cancer after response to first-line platinum-based chemotherapy, irrespective of BRCA1 and BRCA2 mutation status and with consistent benefits observed between homologous recombination subgroups, and was well tolerated. These results support senaparib as a maintenance treatment for patients with advanced ovarian cancer after a response to first-line chemotherapy. ClinicalTrials.gov identifier: NCT04169997 .

Outcomes of secondary cytoreductive surgery in patients with platinum-sensitive recurrent ovarian cancer progressed after prior poly (adenosine diphosphate-ribose) polymerase inhibitors: A retrospective cohort study

ObjectiveTo evaluate the impact of previous poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitor therapy on the effectiveness of secondary cytoreductive surgery (SCS) in patients with platinum-sensitive recurrent ovarian cancer (PSROC). MethodsWe identified patients with PSROC who underwent SCS at the Cancer Hospital, Chinese Academy of Medical Science, between January 2010 and December 2022. Postoperative complications within 30 days were categorized using the Accordion Severity Grading System. The Kaplan‒Meier method was used to estimate both overall survival (OS) and progression-free survival (PFS), and multivariate analysis was used to identify independent prognostic factors. ResultsOf the 265 patients included, 39 received prior PARP inhibitor therapy (Group A), and 226 did not (Group B). The rates of complete resection after SCS did not significantly differ between the two groups (79.5 % for Group A vs. 81.0 % for Group B; p = 0.766). As of December 2023, Group A exhibited a significantly shorter median PFS (14.2 months) than Group B (22.5 months; p = 0.002). Furthermore, the 3-year OS rate was lower in Group A (72.5 %) than in Group B (82.7 %; p = 0.015). The incidence of severe postoperative complications was comparable between Groups A and B (7.7 % vs. 1.8 %; p = 0.061). Multivariate analysis revealed that prior PARP inhibitor therapy significantly reduced the median PFS (hazard ratio (HR) = 4.434; p = 0.021) and OS (HR = 2.076; p = 0.010). ConclusionsSCS for PSROC demonstrated reduced efficacy in patients previously treated with PARP inhibitors compared to those without prior PARP inhibitor treatment.

Abstract PO3-09-04: Identification of germline pathogenic variants to guide systemic therapy: Prevalence and spectrum of PVs among Mexican patients with metastatic breast cancer

Abstract BACKGROUND Currently, international genetic cancer risk assessment (GCRA) guidelines recommend genetic testing for the identification of germline pathogenic variants (PVs) in patients with metastatic breast cancer (MBC) to guide systemic therapy. Patients with PVs in BRCA genes are candidates to receive poly(adenosine diphosphate–ribose) polymerase inhibitors (PARPi). The prevalence of PVs in patients with HER2-negative MBC has been reported to be 10%. The proportion of MBC in low- and middle-income countries (LMIC) is higher. However, access to genetic testing in LMIC is limited and contributes to the gap in genetic epidemiology knowledge. We aimed to describe the frequency of germline PVs in breast cancer associated genes and clinic-pathologic characteristics of Mexican women with MBC. METHODS This retrospective study included Mexican women from 7 centers in Mexico, who were diagnosed with MBC and had genetic testing. Pathogenic and likely pathogenic variants were reported. Demographic, histopathological characteristics, family history and access to treatment with PARPi, were obtained from chart review. RESULTS From August 2017 to May 2023, 114 Mexican women with MBC were included, median age at diagnosis was 47 years (range 24-80 y), 42% had de novo metastatic, and 58% recurrent disease. 86.8% had infiltrating ductal carcinoma, 51.3% had HR+HER2- (n=58), 4.4% HR+HER+ (n=5), 5.3% HR-HER2+ (n=6), and 38.6% triple negative tumors (n=44). Sites of metastatic disease were as follows: 61.4% non-regional nodes (n=70), 57% bone (n=65), 40.3% lung (n=46), 30.7% liver (n=35), 15.8% central nervous system (n=18) and 7% peritoneal carcinomatosis (n=8). The frequency of PVs was 22% (25/114); 84% of the PVs were heterozygous, mainly identified in BRCA genes (BRCA1 n=12, BRCA2 n=9), and in 4 patients in other genes (PALB2 n=2, PTEN n=1, MUTYH n=1). Among PARPi candidates, 40% (10/25) received Olaparib, the majority received it on the second or subsequent lines of treatment (7/10) with a median progression free survival of 6 months (3-13 m). BRCA carriers were younger at BC diagnosis (38.5 vs. 49 years, p=0.00694) and had a significant higher proportion of family history of ovarian cancer (p=0.00039), but not for breast (p=0.06), prostate (p=0.71) or pancreatic cancer (p=0.098), compared to non-carriers. CONCLUSION We found a high frequency of PVs in BRCA genes in Mexican women with MBC. Access to the genetic testing and targeted therapy should be improve in the Mexican population in order to decrease disparities in cancer care in the Mexican Population. In addition, genetic testing allows to identify other family members at risk with cascade testing and to recommend them risk reduction strategies. Citation Format: Jose Luis Rodriguez-Olivares, Maria Fernanda Esparza-Orozco, Jose Alfredo Ramirez-Gonzalez, Roberto Ivan Sanchez-Reyes, Enrique Isay Talamantes-Gamez, Gregorio Quintero-Beulo, Gilberto Lopez-Rosas, Karla Alicia Centelles-López, Brizio Moreno-Jaime, Daniela Vazquez-Juarez, Ana Maria Hernandez-Murillo, Ilana de la Puente Tawil, Jeffrey Weitzel, Cynthia Villareal Garza, Yanin Chavarri-Guerra. Identification of germline pathogenic variants to guide systemic therapy: Prevalence and spectrum of PVs among Mexican patients with metastatic breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-09-04.

Abstract PS05-02: PET Imaging of PARP Expression as a Biomarker of Response to Chemotherapy in Breast Cancer: A Nonrandomized Clinical Trial

Abstract Introduction: Poly–(adenosine diphosphate–ribose) polymerase (PARP) proteins are involved in double-stranded break repair, mediated through homologous recombination (HR) and non-homologous end joining (NHEJ). The majority of PARP activity in this context is attributed to PARP-1 (85-90%). PARP-1 binding to damaged DNA, catalytic activity, and subsequent release to allow access for other DNA repair proteins are necessary for efficient DNA repair. The critical function of PARP-1 in these processes makes this protein a unique potential biomarker of DNA repair capability. [18F]FluorThanatrace ([18F]FTT) is an 18F-labeled PARP inhibitor analog that binds to the same site as approved PARPi drugs. We hypothesized that the biologically relevant form of PARP in untreated breast cancer might provide an indication of functional tumor DNA repair capabilities and therefore predict chemotherapy response. Thus, this study aimed to determine whether a non-invasive quantitative measure of in vivo PARP expression correlated with response to chemotherapy in breast cancer. Methods: A single-arm prospective trial enriched for subjects with triple negative breast cancer (TNBC) was conducted at the University of Pennsylvania from May 2017 to March 2022 (NCT03083288 and NCT03846167). Inclusion criteria were primary breast cancer with tumor diameter of at least 1 cm on conventional imaging and willingness to undergo an [18F]FTT-PET scan prior to neoadjuvant chemotherapy. Participants provided written informed consent. [18F]FTT uptake in breast cancer was measured pre-therapy and tested for association with pathologic complete response (pCR). Subjects were scanned on an Ingenuity TF PET/CT (Philips Healthcare) following injection of [18F]FTT using a 20-minute static acquisition. Peak 1 cm3 average uptake (SUVpeak) were recorded from a spherical region-of-interest (ROI) covering the primary malignancy, guided by CT and prior imaging studies. All SUVs were partial volume corrected and normalized using muscle (NM). A Mann-Whitney test was used to determine if [18F]FTT uptake differed by pCR. Receiver operating characteristic (ROC) curves were constructed to test the performance of [18F]FTT uptake in predicting pCR. Statistical analyses utilized STATA v15.1 with significance based on a two-sided alpha level of ≤0.05. Participants: Twenty-six women with stage I-III breast cancer met inclusion criteria with age range of 29-74 years and self-reported race as Asian (1, 4%), Black (9, 37%) or White (16, 60%). One subject had bilateral tumors (ER+ stage IIA and HER2+ stage I) that were evaluated separately (27 total tumors). Breast tumors were pathologically confirmed and included 6 (22%) ER+/HER2-, 5 (19%) HER2+, and 16 (59%) TNBC. Germline sequencing was available for 21 participants. Results: Mean tumor diameter ranged from 15 to 91 mm (median 30 mm). A considerable range of [18F]FTT uptake was seen across subjects (SUVpeak/NM 0.66-6.5). [18F]FTT uptake was independent of subtype (ER+, HER2+, TN) (P=0.35), stage (P=0.39), and between mutations in genes associated with homologous recombination deficiency (i.e., BRCA1/2, PALB2) and wild-type (P=0.73). In the TNBC cohort, pre-treatment [18F]FTT uptake was higher in subjects who went on to have a pCR (n=16, P=0.05), while a trend toward higher uptake was seen when including all tumor types (n=27, P=0.11). ROC analysis of the value of [18F]FTT uptake for predicting pCR revealed an AUC of 0.79 and showed that a threshold SUV ratio &amp;lt; 2.47 predicted pCR in TNBC patients with a 100% specificity and 64% sensitivity. Conclusions: These early clinical results suggest a relationship between [18F]FTT uptake, a measure of PARP expression, and chemotherapy sensitivity in TNBC, warranting future studies to elucidate underlying mechanistic processes and potential clinical implications. Citation Format: Sarah Gitto, Anthony Young, Ira Bleiweiss, Amy Clark, Elizabeth McDonald. PET Imaging of PARP Expression as a Biomarker of Response to Chemotherapy in Breast Cancer: A Nonrandomized Clinical Trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS05-02.

Abstract PO2-09-03: Twenty-one gene recurrence scores in individuals with breast cancer associated with PALB2 germline pathogenic variants

Abstract Background: The 21-gene recurrence score assay (Oncotype DX, Genomic Health Inc., CA) is prognostic, quantifies predicted benefit of adjuvant chemotherapy, and informs clinical decisions for patients with early-stage, hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) breast cancer (BC). Differences in Oncotype recurrence scores (RS) in individuals with and without BRCA1 and BRCA2 (BRCA1/2) pathogenic variants (PV) have been demonstrated, but the results of this assay in patients with BC and germline PALB2 PV have not previously been described. This is of particular relevance as PALB2 BC has demonstrated biologic features similar to BRCA1/2 BC such as poly(adenosine diphosphate ribose) polymerase inhibitor (PARPi) sensitivity. Methods: Patients with early-stage, HR+, HER2-, invasive breast cancer and a germline PALB2 PV were ascertained from academic cancer center databases and the Prospective Registry of MultiPlex Testing (PROMPT), an online research registry for individuals who have had multigene panel testing for inherited cancer susceptibility. Study participants (pts) were categorized by age &amp;lt; 50 and ≥50. Oncotype DX RS were retrospectively reviewed and categorized as those that would and would not warrant consideration of chemotherapy (≥16 and &amp;lt; 16, respectively, for pts &amp;lt; 50; ≥26 and &amp;lt; 26, respectively, for pts ≥50). Yates corrected Chi-square tests were used to compare RS distributions between carriers and a previously published genetically unselected reference population. Results: Oncotype DX reports were reviewed for 20 pts with BC and germline PALB2 PV. Median RS was 19 (range 6-41) overall. Four patients were diagnosed under the age of 50 (range 33-48); median RS for these individuals was 19 (range 12-41). Sixteen patients were diagnosed at age 50 or greater (median 59, range 50-70); median RS for these individuals was also 19 (range 6-31). Compared to the genetically unselected reference population, Oncotype DX RS in pts with PALB2-associated BC demonstrated a numerical but not statistically significant shift in distribution towards categories in which chemotherapy would be considered. Among pts under the age of 50, 75% had Oncotype DX RS for which chemotherapy would be considered and 25% had Oncotype DX RS for which endocrine therapy alone would likely be recommended (p=0.726 compared to reference). Among pts aged 50 and older, 25% had Oncotype DX RS for which chemotherapy would be considered and 75% had Oncotype DX RS for which endocrine therapy alone would likely be recommended (p=0.579 compared to reference). Conclusions: Compared to data from the published genetically unselected commercial database, BC associated with PALB2 PV may have higher Oncotype DX RS reflective of intrinsically less favorable biology and greater therapeutic sensitivity. Larger datasets are forthcoming to confirm presence of significant differences in Oncotype DX RS distributions between BC with and without PALB2 PV. Table 1. Oncotype DX RS distribution by age among pts with gPALB2 PV and genetically unselected reference *Jakubowski et al., J Surg Oncol 2020. PMID 3249731 Citation Format: Payal Shah, Siddhartha Yadav, Jamie Brower, Jada Hamilton, Erica Simmons, Judy Garber, Kenneth Offit, Mark Robson, Fergus Couch, Susan Domchek. Twenty-one gene recurrence scores in individuals with breast cancer associated with PALB2 germline pathogenic variants [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-09-03.

Abstract PO5-08-08: Tumor Registry Guided Genetic Testing: Opportunities Unlocked

Abstract Introduction With the expansion of guidelines for germline genetic testing, more patients are eligible for testing. Unfortunately, many remain untested. Testing is critical, as patients with pathogenic variants (and their families) can benefit from more aggressive screening and preventative strategies, as well as targeted therapies (e.g., poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitors) if cancer does occur. Objectives To study the prevalence of Hereditary Breast Ovarian Cancer (HBOC) genetic testing eligibility and uptake among cancer patients from the tumor registry at the Medical University of South Carolina (MUSC) in order to identify a group of living patients who require testing now. Methods This Quality Improvement study included patients 18 years of age or older diagnosed with breast, ovarian/fallopian/primary peritoneal, pancreas and prostate cancers between 1991 and 2021 as reported to the MUSC Tumor Registry. Each patient was evaluated using the latest National Comprehensive Cancer Network (NCCN) criteria for breast-ovarian genetic testing (Excluding family history criteria, which were not available in the tumor registry). Using our genetic testing database, we also determined which had had testing. We describe the prevalence of genetic testing performed vs the number of patients that would benefit from testing under the newest guidelines. Results Patients identified as eligible for testing included 399 breast cancer patients, all 544 ovarian/fallopian/primary peritoneal cancer patients, all 1534 pancreas cancer patients and 872 prostate cancer patients. Reported testing of eligible patients was low (breast: 35%, Ovarian: 33%, pancreas: 4%, and prostate cancer: 6%). Of eligible patients not tested, 310 patients with breast cancer, 343 patients with ovarian/fallopian/primary peritoneal cancer, 717 patients with pancreas cancer and 763 patients with prostate cancer, are alive and eligible for testing (2133 patients) Discussion Using Tumor Registry data plus genetic testing data allowed us to detect a large number of living patients with cancer that would be eligible for HBOC genetic testing. This is despite the fact that this approach underestimates the number of patients that would benefit from further testing as it does not include family history and other factors. A similar approach for colorectal or other cancers will yield additional patients. Conclusion Most cancer patients eligible for breast ovarian genetic testing have not been tested. There is an opportunity to use Tumor Registry and genetic testing data to identify a large number of living patients who can be approached for testing. As we showed in our study, by adopting updated cancer genetic testing guidelines, there is increased opportunity to detect more patients that would benefit from such testing. In our study this benefit is shown especially in the prostate and pancreas cancer groups. Next steps will be to determine how best to notify and make testing available to these patients. Table 1. Testing Status Comparison Assessing Genetic Testing Rates Using Tumor Registry Among Eligible Patients with Breast, Ovarian/Fallopian/Primary Peritoneum, Pancreas and Prostate cancer as per the New Testing Guidelines Citation Format: Kevin Hughes, Antoine Finianos, Kiersten Meeder. Tumor Registry Guided Genetic Testing: Opportunities Unlocked [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-08-08.

Abstract PO4-08-02: Genetic testing among patients with triple-negative breast cancer at a single health system: a quality improvement project

Abstract Background: Breast cancer is the most common type and the second leading cause of cancer-related deaths in women. Approximately 15-20% of patients have triple-negative breast cancer (TNBC) at diagnosis. TNBC is aggressive, difficult to treat, prone to relapse, and associated with an increased mortality rate. The incidence of BRCA1/2 mutation is about 20% in patients with TNBC (Hartman et al. 2012). Despite the high prevalence of germline mutations in cancer susceptibility genes among TNBC patients, many do not received genetic counseling (GC) or genetic testing (GT). National Comprehensive Cancer Network (NCCN) guidelines now recommend GT for all patients with TNBC, regardless of age, especially after the approval of Poly Adenosine diphosphate Ribose Polymerase (PARP) inhibitors for targeted therapy against BRCA1/2 mutations. The aim of this quality improvement (QI) project was to identify patients with TNBC who did not receive GT and design an intervention to understand potential barriers to receiving GC and GT. Methods: We implemented a QI project to identify previously diagnosed adult patients with TNBC (18 years and older) of any stage (early or advanced/metastatic) without prior GT or family history of genetic disorders with active follow-up at the Henry Ford health breast cancer clinic. Eligible patients diagnosed between January 2015 and January 2020 from the Syapse Learning Health Network database were included. Baseline information as well as additional data including demographics, breast cancer treatment details, and whether GT and/or GC was offered or not was obtained through electronic health record review after institutional review board committee approval. Bi-monthly meetings were held amongst breast cancer providers, genetic counselors, and nurse navigators to develop an intervention to contact the patients who did not undergo GT and/or GC. After obtaining the list of patients who did not undergo GT and/or GC despite meeting eligibility criteria, they were contacted by their primary breast cancer providers via telephone regarding their willingness to obtain GT and/or GC at this time and participate in a questionnaire-based interview to understand barriers towards the same. Results: In the 5 years, a total of 123 patients were noted to have been diagnosed with triple-negative breast cancer. Of these, 93 patients were actively being followed at the Henry Ford breast clinic at the time of the QI project. 2 of the 9 patients who underwent GT and GC were positive for BRCA1/2 mutation. Of the remaining 83 patients that did not receive GC or GT, 65 (78.3%) had no clear reason explained in the chart, 27 patients (32.5%) chose not to get tested, 23 (27.7%) were not deemed to be eligible upon initial assessment and 4 (4.8%) had pending orders placed by breast surgeons upon initial diagnosis. Unfortunately, 29 of these 83 patients were noted to have been deceased when the intervention was being designed. Remaining 54 patients were contacted by their primary breast cancer oncologists to determine their willingness in participating in the questionnaire-based interview. Conclusion: Based on the results of our QI project, we have now developed a process to contact the identified patients and renew the offer for GT and GC as well as participation in a questionnaire-based interview to understand the potential barriers to testing. We also plan to improve physician education and awareness by conducting mini sessions during our annual symposium meetings as well as grand rounds and develop a protocol for ordering GT and GC in patients with TNBC at our institution. Citation Format: Manasi Godbole, Mary Nyhuis, Travis Washburn, Joann Hirth, Haythem Ali. Genetic testing among patients with triple-negative breast cancer at a single health system: a quality improvement project [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-08-02.

Evolutionary trajectory of TRPM2 channel activation by adenosine diphosphate ribose and calcium

Ion channel activation upon ligand gating triggers a myriad of biological events and, therefore, evolution of ligand gating mechanism is of fundamental importance. TRPM2, a typical ancient ion channel, is activated by adenosine diphosphate ribose (ADPR) and calcium and its activation has evolved from a simple mode in invertebrates to a more complex one in vertebrates, but the evolutionary process is still unknown. Molecular evolutionary analysis of TRPM2s from more than 280 different animal species has revealed that, the C-terminal NUDT9-H domain has evolved from an enzyme to a ligand binding site for activation, while the N-terminal MHR domain maintains a conserved ligand binding site. Calcium gating pattern has also evolved, from one Ca2+-binding site as in sea anemones to three sites as in human. Importantly, we identified a new group represented by olTRPM2, which has a novel gating mode and fills the missing link of the channel gating evolution. We conclude that the TRPM2 ligand binding or activation mode evolved through at least three identifiable stages in the past billion years from simple to complicated and coordinated. Such findings benefit the evolutionary investigations of other channels and proteins.

Prevalence of Homologous Recombination Deficiency Among Patients With Germline RAD51C/D Breast or Ovarian Cancer

RAD51C and RAD51D are involved in DNA repair by homologous recombination. Germline pathogenic variants (PVs) in these genes are associated with an increased risk of ovarian and breast cancer. Understanding the homologous recombination deficiency (HRD) status of tumors from patients with germline PVs in RAD51C/D could guide therapeutic decision-making and improve survival. To characterize the clinical and tumor characteristics of germline RAD51C/D PV carriers, including the evaluation of HRD status. This retrospective cohort study included 91 index patients plus 90 relatives carrying germline RAD51C/D PV (n = 181) in Spanish hospitals from January 1, 2014, to December 31, 2021. Genomic and functional HRD biomarkers were assessed in untreated breast and ovarian tumor samples (n = 45) from June 2022 to February 2023. Clinical and pathologic characteristics were assessed using descriptive statistics. Genomic HRD by genomic instability scores, functional HRD by RAD51, and gene-specific loss of heterozygosity were analyzed. Associations between HRD status and tumor subtype, age at diagnosis, and gene-specific loss of heterozygosity in RAD51C/D were investigated using logistic regression or the t test. A total of 9507 index patients were reviewed, and 91 patients (1.0%) were found to carry a PV in RAD51C/D; 90 family members with a germline PV in RAD51C/D were also included. A total of 157 of carriers (86.7%) were women and 181 (55.8%) had received a diagnosis of cancer, mainly breast cancer or ovarian cancer. The most prevalent PVs were c.1026+5_1026+7del (11 of 56 [19.6%]) and c.709C>T (9 of 56 [16.1%]) in RAD51C and c.694C>T (20 of 35 [57.1%]) in RAD51D. In untreated breast cancer and ovarian cancer, the prevalence of functional and genomic HRD was 55.2% (16 of 29) and 61.1% (11 of 18) for RAD51C, respectively, and 66.7% (6 of 9) and 90.0% (9 of 10) for RAD51D. The concordance between HRD biomarkers was 91%. Tumors with the same PV displayed contrasting HRD status, and age at diagnosis did not correlate with the occurrence of HRD. All breast cancers retaining the wild-type allele were estrogen receptor positive and lacked HRD. In this cohort study of germline RAD51C/D breast cancer and ovarian cancer, less than 70% of tumors displayed functional HRD, and half of those that did not display HRD were explained by retention of the wild-type allele, which was more frequent among estrogen receptor-positive breast cancers. Understanding which tumors are associated with RAD51C/D and HRD is key to identify patients who can benefit from targeted therapies, such as PARP (poly [adenosine diphosphate-ribose] polymerase) inhibitors.

Unexpected Noncovalent Off-Target Activity of Clinical BTK Inhibitors Leads to Discovery of a Dual NUDT5/14 Antagonist.

Cofactor mimicry represents an attractive strategy for the development of enzyme inhibitors but can lead to off-target effects due to the evolutionary conservation of binding sites across the proteome. Here, we uncover the ADP-ribose (ADPr) hydrolase NUDT5 as an unexpected, noncovalent, off-target of clinical BTK inhibitors. Using a combination of biochemical, biophysical, and intact cell NanoBRET assays as well as X-ray crystallography, we confirm catalytic inhibition and cellular target engagement of NUDT5 and reveal an unusual binding mode that is independent of the reactive acrylamide warhead. Further investigation of the prototypical BTK inhibitor ibrutinib also revealed potent inhibition of the largely unstudied NUDIX hydrolase family member NUDT14. By exploring structure-activity relationships (SARs) around the core scaffold, we identify a potent, noncovalent, and cell-active dual NUDT5/14 inhibitor. Cocrystallization experiments yielded new insights into the NUDT14 hydrolase active site architecture and inhibitor binding, thus providing a basis for future chemical probe design.