TP53 Missense Mutations Research Articles

TP53 mutations and survival in ovarian carcinoma patients receiving first-line chemotherapy plus bevacizumab: Results of the MITO16A/MaNGO OV-2 study.

To date, there are no biomarkers that define a patient subpopulation responsive to bevacizumab (BEV), an effective treatment option for advanced ovarian carcinoma (OC). In the context of the MITO16A/MaNGO OV-2 trial, a Phase IV study of chemotherapy combined with BEV in first-line treatment of advanced OC, we evaluated TP53 mutations by next-generation sequencing and p53 expression by immunohistochemistry (IHC) on 202 and 311 cases, respectively. We further correlated TP53 mutations in terms of type, function, and site, and IHC data with patients' clinicopathological characteristics and survival. TP53 missense mutations of unknown function (named unclassified) represented the majority of variants in our population (44.4%) and were associated with a significantly improved overall survival (OS) both in univariable (hazard ratio [HR] = 0.43, 95% confidence interval [CI] = 0.20-0.92, p = .03) and multivariable analysis (HR = 0.39, 95% CI = 0.18-0.86, p = .02). Concordance between TP53 mutational analysis and IHC was 91%. We observed an HR of 0.70 for OS in patients with p53 IHC overexpression compared to p53 wild-type, which however did not reach statistical significance (p = .31, 95% CI = 0.36-1.38). Our results indicate that the presence of unclassified TP53 mutations has favorable prognostic significance in patients with OC receiving upfront BEV plus chemotherapy. In particular, unclassified missense TP53 mutations characterize a subpopulation of patients with a significant survival advantage, independently of clinicopathological characteristics. Our findings warrant future investigations to confirm the prognostic impact of TP53 mutations in BEV-treated OC patients and deserve to be assessed for their potential predictive role in future randomized clinical studies.

1338P TP53 truncating and missense mutations are linked to differential response to checkpoint blockade in patients with advanced non-small cell lung cancer (NSCLC)

1338P TP53 truncating and missense mutations are linked to differential response to checkpoint blockade in patients with advanced non-small cell lung cancer (NSCLC)

Age and Sex Affects the Frequency and Mutation Type of FGFR2 Alterations in Cholangiocarcinoma

PURPOSE FGFR2 alterations are infrequent but currently represent the most common targetable mutation in cholangiocarcinoma. We performed a large-scale genomic analysis to assess associations between FGFR2 and other driver gene alterations in cholangiocarcinoma with patient demographics. METHODS Clinical and molecular data for patients with intrahepatic cholangiocarcinoma were abstracted from the Foundation Medicine (FMI) and Genomics, Evidence, Neoplasia, Information, Exchange (GENIE) genomic databases. The Chi-squared test was used to assess the association between sex and alterations in driver genes. FGFR2 fusion partners were analyzed using descriptive statistics. RESULTS Across both FMI (N = 7,859) and GENIE (N = 1,395) cohorts, FGFR2 rearrangements were more frequent in female than male patients with cholangiocarcinoma (odds ratio, 1.69 and 2.45). By contrast, FGFR2 missense mutations, TP53 mutations, and KRAS mutations were not associated with sex. Stratifying patients by age groups showed that FGFR2 rearrangements were specifically enriched in younger ages (<45 years). Although the most common FGFR2 fusion partners were shared across both sexes, a subset of fusion partners was recurrent and uniquely associated with female or male patients. CONCLUSION FGFR2 rearrangements but not other driver alterations in cholangiocarcinoma are concentrated in young female patients, where the prevalence reached nearly 20% across two large patient cohorts. These results suggest that the molecular basis of FGFR2 rearrangements may be distinct between sexes. These results indicate the importance of genetic testing in cholangiocarcinoma, particularly for younger patients, and suggest that patient recruitment strategies in trials for FGFR2-targeting therapies should seek to reduce barriers to enhance the enrollment of young female patients.

Identify truly high-risk TP53-mutated diffuse large B cell lymphoma patients and explore the underlying biological mechanisms

TP53 mutation (TP53-mut) correlates with inferior survival in many cancers, whereas its prognostic role in diffuse large B-cell lymphoma (DLBCL) is still in controversy. Therefore, more precise risk stratification needs to be further explored for TP53-mut DLBCL patients. A set of 2637 DLBCL cases from multiple cohorts, was enrolled in our analysis. Among the 2637 DLBCL patients, 14.0% patients (370/2637) had TP53-mut. Since missense mutations account for the vast majority of TP53-mut DLBCL patients, and most non-missense mutations affect the function of the P53 protein, leading to worse survival rates, we distinguished patients with missense mutations. A TP53 missense mutation risk model was constructed based on a 150-combination machine learning computational framework, demonstrating excellent performance in predicting prognosis. Further analysis revealed that patients with high-risk missense mutations are significantly associated with early progression and exhibit dysregulation of multiple immune and metabolic pathways at the transcriptional level. Additionally, the high-risk group showed an absolutely suppressed immune microenvironment. To stratify the entire cohort of TP53-mut DLBCL, we combined clinical characteristics and ultimately constructed the TP53 Prognostic Index (TP53PI) model. In summary, we identified the truly high-risk TP53-mut DLBCL patients and explained this difference at the mutation and transcriptional levels.

P53R172H and p53R245W Hotspot Mutations Drive Distinct Transcriptomes in Mouse Mammary Tumors Through a Convergent Transcriptional Mediator.

Our findings implicate NR5A2 as a novel mediator of mutant p53 transcriptional activity in breast cancer. NR5A2 may be an important therapeutic target in hard-to-treat breast cancers such as endocrine-resistant tumors and metastatic triple-negative breast cancers harboring TP53 missense mutations.

TP53 Mutations in Acute Leukemias and Myelodysplastic Syndromes: Insights and Treatment Updates.

TP53 mutations are found in 5%-10% of de novo myelodysplastic syndrome (MDS) and AML cases. By contrast, in therapy related MDS and AML, mutations in TP53 are found in up to 30%-40% of patients. The majority of inactivating mutations observed in MDS and AML are missense mutations localized in a few prevalent hotspots. TP53 missense mutations together with truncating mutations or chromosomal loss of TP53 determine a loss-of-function effect on normal p53 function. Clonal expansion of TP53-mutant clones is observed under the selection pressure of chemotherapy or MDM2 inhibitor therapy. TP53-mutant clones are resistant to current chemotherapy, and when responses to treatment have been observed, they have correlated poorly with overall survival. The most heavily investigated and targeted agent for patients with TP53-mutant MDS and AML has been APR-246 (eprenetapopt) a p53 reactivator, in combination with azacitidine, but also in triplets with venetoclax. Despite positive results in phase II trials, a phase III trial did not confirm superior response or improved survival. Other agents, like magrolimab (anti-CD47 antibody), failed to demonstrate improved activity in TP53-mutant MDS and AML. Agents whose activity is not dependent on a functional apoptosis system like anti-CD123 antibodies or cellular therapies are in development and may hold promises. Delivering prognostic information in a dismal disease like TP53-mutated MDS and AML is particularly challenging. The physician should balance hope and realism, describing the trajectory of possible treatments and at the same time indicating the poor outcome, together with promoting adaptive coping in patients and elaborating on the nature of the disease.

KDM7A-DT induces genotoxic stress, tumorigenesis, and progression of p53 missense mutation-associated invasive breast cancer.

Stress-induced promoter-associated and antisense lncRNAs (si-paancRNAs) originate from a reservoir of oxidative stress (OS)-specific promoters via RNAPII pausing-mediated divergent antisense transcription. Several studies have shown that the KDM7A divergent transcript gene (KDM7A-DT), which encodes a si-paancRNA, is overexpressed in some cancer types. However, the mechanisms of this overexpression and its corresponding roles in oncogenesis and cancer progression are poorly understood. We found that KDM7A-DT expression is correlated with highly aggressive cancer types and specific inherently determined subtypes (such as ductal invasive breast carcinoma (BRCA) basal subtype). Its regulation is determined by missense TP53 mutations in a subtype-specific context. KDM7A-DT transcribes several intermediate-sized ncRNAs and a full-length transcript, exhibiting distinct expression and localization patterns. Overexpression of KDM7A-DT upregulates TP53 protein expression and H2AX phosphorylation in nonmalignant fibroblasts, while in semi-transformed fibroblasts, OS superinduces KDM7A-DT expression in a TP53-dependent manner. KDM7A-DT knockdown and gene expression profiling in TP53-missense mutated luminal A BRCA variant, where it is abundantly expressed, indicate its significant role in cancer pathways. Endogenous over-expression of KDM7A-DT inhibits DNA damage response/repair (DDR/R) via the TP53BP1-mediated pathway, reducing apoptosis and promoting G2/M checkpoint arrest. Higher KDM7A-DT expression in BRCA is associated with KDM7A-DT locus gain/amplification, higher histologic grade, aneuploidy, hypoxia, immune modulation scores, and activation of the c-myc pathway. Higher KDM7A-DT expression is associated with relatively poor survival outcomes in patients with luminal A or Basal subtypes. In contrast, it is associated with favorable outcomes in patients with HER2+ER- or luminal B subtypes. KDM7A-DT levels are coregulated with critical transcripts and proteins aberrantly expressed in BRCA, including those involved in DNA repair via non-homologous end joining and epithelial-to-mesenchymal transition pathway. In summary, KDM7A-DT and its si-lncRNA exhibit several intrinsic biological and clinical characteristics that suggest important roles in invasive BRCA and its subtypes. KDM7A-DT-defined mRNA and protein subnetworks offer resources for identifying clinically relevant RNA-based signatures and prospective targets for therapeutic intervention.

Longitudinal Comparative Analysis of Circulating Tumor DNA and Matched Tumor Tissue DNA in Patients with Metastatic Colorectal Cancer Receiving Palliative First-Line Systemic Anti-Cancer Therapy.

This study aimed to compare tumor tissue DNA (ttDNA) and circulating tumor DNA (ctDNA) to explore the clinical applicability of ctDNA and to better understand clonal evolution in patients with metastatic colorectal cancer undergoing palliative first-line systemic therapy. We performed targeted sequencing analysis of 88 cancer-associated genes using germline DNA, ctDNA at baseline (baseline-ctDNA), and ctDNA at progressive disease (PD-ctDNA). The results were compared with ttDNA data. Among 208 consecutively enrolled patients, we selected 84 (41 males; median age, 59 years; range, 35 to 90 years) with all four sample types available. A total of 202 driver mutations were found in 34 genes. ttDNA exhibited the highest mutation frequency (n=232), followed by baseline-ctDNA (n=155) and PD-ctDNA (n=117). Sequencing ctDNA alongside ttDNA revealed additional mutations in 40 patients (47.6%). PD-ctDNA detected 13 novel mutations in 10 patients (11.9%) compared to ttDNA and baseline-ctDNA. Notably, seven mutations in five patients (6.0%) were missense or nonsense mutations in APC, TP53, SMAD4, and CDH1 genes. In baseline-ctDNA, higher maximal variant allele frequency (VAF) values (p=0.010) and higher VAF values of APC (p=0.012), TP53 (p=0.012), and KRAS (p=0.005) mutations were significantly associated with worse overall survival. While ttDNA remains more sensitive than ctDNA, our ctDNA platform demonstrated validity and potential value when ttDNA was unavailable. Post-treatment analysis of PD-ctDNA unveiled new pathogenic mutations, signifying cancer's clonal evolution. Additionally, baseline-ctDNA's VAF values were prognostic after treatment.

Abstract 6143: Association of polygenic risk score and prostate tumor biomarkers with lethal prostate cancer

Abstract Prostate cancer (PCa) is a highly heritable cancer. While polygenic risk scores (PRS) stratify overall PCa risk among cancer-free men, they have not been reported to predict lethal outcomes in PCa patients. We aimed to examine to what extent PRS modifies associations between tumor biomarkers with cancer prognosis after diagnosis. We included men with primary, non-metastatic PCa in the Health Professionals Follow-up Study (HPFS) and Physicians’ Health Study (PHS). The PRS was constructed with 451 risk variants identified from prior PCa GWAS, measured on DNA from blood or buccal, standardized by adjusting for mean and standard deviation (SD) based on men without PCa with array-based genotyping data in HPFS/PHS. Prostate tumor biomarkers of molecular subtypes (TMPRSS2:ERG fusion, PTEN loss, and TP53 missense mutations from validated immunohistochemistry assays), insulin/lipid signaling pathways (FASN, IR, and IGF1-R), and cellular proliferation and tumor microenvironment (Ki-67, apoptosis rates, and angiogenesis) were measured on tumor tissue from radical prostatectomy or TURMP. Sample sizes varied by tumor biomarkers, ranging from 214 for angiogenesis markers to 598 for ERG protein expression. We first used either logistic or negative binomial regression to evaluate the association between the PRS and the protein expression (levels) of each individual biomarker, and then evaluated the addition of multiplicative interaction terms between PRS and biomarkers to logistic regression models for lethal PCa, defined as metastases or death from PCa. Lastly, we evaluated the predictive performance for lethal PCa by area under the curve (AUC). All models adjusted for age at diagnosis and Gleason score. Among all patients included in the analysis, the median age at diagnosis was 66 (interquartile range: 62-70) years, with 20% having a Gleason score≥8 and 10% having lethal PCa. PRS showed no association with lethal PCa (OR per SD increase PRS=0.95, 95%CI 0.85-1.06) or most tumor biomarkers, except for the apoptosis index (per SD in PRS corresponding to 0.22 units increase, 95%CI 0.02-0.42). Statistically significant interactions were found between PRS and p53 overexpression (p interaction = 1.6x10-5), PTEN loss (p=3.7x10-3), FASN (p=0.03), and Ki-67 (p=1.6x10-3), suggesting that compared to men with a PRS higher than the median, TP53 missense mutation, PTEN loss, higher FASN intensity, and higher Ki-67 indices were more strongly associated with lethal PCa in patients with lower PRS. Incorporating PRS with tumor biomarkers into predictive models enhanced the AUC beyond a model with tumor biomarker and age. The largest improvements were observed when combining the PRS with Ki-67 and apoptosis, showing absolute increases of 5% and 7% respectively. In conclusion, PRS integration with tumor biomarkers at diagnosis potentially improves the prediction of lethal PCa, thereby informing more targeted management strategies. Citation Format: Anqi Wang, Anna Plym, Konrad H. Stopsack, Lorelei A. Mucci, Kathryn L. Penney. Association of polygenic risk score and prostate tumor biomarkers with lethal prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6143.

Abstract 6463: Highly frequent gene mutations and co-occurring genes analysis in colorectal cancer

Abstract Background: Colorectal cancer (CRC) is the second most leading cause of cancer-related deaths worldwide. Understanding the molecular alterations that drive CRC is critical for improving patient outcomes. Next-generation sequencing (NGS) has revolutionized CRC molecular profiling by simultaneous analysis of multiple genes with high accuracy and sensitivity. In this study, we analyzed a cohort of CRC formalin-fixed paraffin-embedded (FFPE) samples to identify highly frequent gene mutations and co-occurring genes in CRC. Methods: FFPE samples from CRC patients tested from January 2020 to May 2023 were analyzed using NeoTYPE® Colorectal Tumor Profile. This profile analyzes 44 biomarkers through a combination of NGS focusing on a panel of 36 genes known to be frequently mutated in CRC, as well as 3 FISH and 3 IHC biomarkers. Gene mutation frequency, co-occurring mutation genes and clinical significance were explored and analyzed. In addition, immunotherapy markers, Microsatellite Instability (MSI) and Tumor Mutation Burden (TMB) were assessed. Results: A total of 2,920 patients (1,577 men and 1,343 women) with CRC were included in this study. Average sequence coverage was 2386X with mean minor allelic frequency of 30%. A higher average number of variant allele frequency were observed in metastatic versus primary colorectal cancer (P < .001). We found that APC, TP53, KRAS, PIK3CA and ARID1A were the most frequently mutated genes. APC truncated mutation p. R1450* were the most common mutations. KRAS, PIK3CA and TP53 missense mutations are the prevalent consequence. Notably, a subset of patients exhibited high microsatellite instability (MSI-H) or TMB high, indicating potential eligibility for immune checkpoint inhibitor therapy. In addition, we found APC mutations co-occurring with TP53 or KRAS mutations (p <.001) in our cohort. Conclusion: Our results provide insight into the genetic landscape of CRC and highlight the importance of understanding co-occurring mutations in this disease. Specifically, APC mutations frequently coincide with TP53 mutations, jointly contributing to the onset of CRC. TP53 often play a role in promoting genomic instability and evading mechanisms that regulate cell growth. The identification of highly frequent mutations and co-occurring genes may have important implications for the development of targeted therapies and personalized treatment approaches for CRC patients. Citation Format: Qinqin Zha, Frank Scarpa, Madhu Rengaraj, Anna Juncker-Jensen. Highly frequent gene mutations and co-occurring genes analysis in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6463.

Abstract 966: Longitudinal comparative analysis of circulating tumor DNA and matched tumor tissue DNA in patients with metastatic colorectal cancer receiving palliative first-line systemic anti-cancer therapy

Abstract Background: Plasma circulating tumor DNA (ctDNA) analysis has become an attractive method for precision oncology. This study compared mutational landscapes of ctDNA with matched tumor tissue DNA (ttDNA) to explore clinical applicability and to better understand clonal evolution in patients with metastatic colorectal cancer receiving palliative first-line systemic anti-cancer therapy (SACT). Methods: We performed unique molecular identifier-based targeted sequencing of 88 cancer-associated genes using the following samples: ttDNA, germline DNA, ctDNA at baseline (baseline-ctDNA), and ctDNA at progressive disease (PD-ctDNA). Results: Of the 208 consecutively enrolled patients, 84 were selected (41 males; median age 59, ranging from 35 to 90) with access to all four samples. Excluding duplicated mutations, a total of 202 driver mutations were discovered in 34 genes. ttDNA exhibited the highest occurrence of mutations (N=232), followed by baseline-ctDNA (N=155) and PD-ctDNA (N=117). Genotype concordance for the ten most mutated genes was highest between baseline-ctDNA and PD-ctDNA, at 94.3%, while the concordance rates for ttDNA were 85.5% with baseline-ctDNA and 83.6% with PD-ctDNA. Overall, 40 of the 84 patients (47.6%) benefited from sequencing ctDNA in addition to ttDNA as it identified additional mutations. Compared to ttDNA and baseline-ctDNA, PD-ctDNA identified 13 novel mutations in ten patients (11.9%). Notably, 7 mutations in five patients (6.0%) were missense or nonsense mutations in APC, TP53, SMAD4, and CDH1 genes. In baseline-ctDNA, patients with maximal variant allele frequency (VAF) values of >0.059 showed significantly worse progression-free survival (median 10.5 vs. 17.1 months, P=0.036) and overall survival (median 18.1 vs. 40.3 months, P=0.010). Moreover, overall survival was significantly worse in those with higher VAF values of APC (median 18.1 vs. 36.8 months, P=0.012), TP53 (median 24.7 vs. 40.3 months, P=0.012), and KRAS (median 13.7 vs. 27.6 months, P=0.005) mutations. Conclusions: While ttDNA remains more sensitive than ctDNA, our unique molecular identifier-based ctDNA platform demonstrated the validity and potential value when ttDNA was unavailable. Furthermore, post-SACT analysis of PD-ctDNA revealed new pathogenic mutations, indicating the clonal evolution of cancer. In addition, the VAF values of baseline-ctDNA predicted prognosis after SACT. Citation Format: Ji-Won Kim, Seung-been Lee, Hong-Geun Kim, Sung-Hyun Hwang, Kui-Jin Kim, Ju Hyun Lee, Jeongmin Seo, Minsu Kang, Eun Hee Jung, Koung Jin Suh, Se Hyun Kim, Jin Won Kim, Yu Jung Kim, Jee Hyun Kim, Nak-Jung Kwon, Keun-Wook Lee. Longitudinal comparative analysis of circulating tumor DNA and matched tumor tissue DNA in patients with metastatic colorectal cancer receiving palliative first-line systemic anti-cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 966.

Evolutionary trajectories of small cell lung cancer under therapy

The evolutionary processes that underlie the marked sensitivity of small cell lung cancer (SCLC) to chemotherapy and rapid relapse are unknown1–3. Here we determined tumour phylogenies at diagnosis and throughout chemotherapy and immunotherapy by multiregion sequencing of 160 tumours from 65 patients. Treatment-naive SCLC exhibited clonal homogeneity at distinct tumour sites, whereas first-line platinum-based chemotherapy led to a burst in genomic intratumour heterogeneity and spatial clonal diversity. We observed branched evolution and a shift to ancestral clones underlying tumour relapse. Effective radio- or immunotherapy induced a re-expansion of founder clones with acquired genomic damage from first-line chemotherapy. Whereas TP53 and RB1 alterations were exclusively part of the common ancestor, MYC family amplifications were frequently not constituents of the founder clone. At relapse, emerging subclonal mutations affected key genes associated with SCLC biology, and tumours harbouring clonal CREBBP/EP300 alterations underwent genome duplications. Gene-damaging TP53 alterations and co-alterations of TP53 missense mutations with TP73, CREBBP/EP300 or FMN2 were significantly associated with shorter disease relapse following chemotherapy. In summary, we uncover key processes of the genomic evolution of SCLC under therapy, identify the common ancestor as the source of clonal diversity at relapse and show central genomic patterns associated with sensitivity and resistance to chemotherapy.

The spectrum of TP53 mutations in Rwandan patients with gastric cancer

BackgroundGastric cancer is the sixth most frequently diagnosed cancer and third in causing cancer-related death globally. The most frequently mutated gene in human cancers is TP53, which plays a pivotal role in cancer initiation and progression. In Africa, particularly in Rwanda, data on TP53 mutations are lacking. Therefore, this study intended to obtain TP53 mutation status in Rwandan patients with gastric cancer.ResultsFormalin-fixed paraffin-embedded tissue blocks of 95 Rwandan patients with histopathologically proven gastric carcinoma were obtained from the University Teaching Hospital of Kigali. After DNA extraction, all coding regions of the TP53 gene and the exon–intron boundary region of TP53 were sequenced using the Sanger sequencing. Mutated TP53 were observed in 24 (25.3%) of the 95 cases, and a total of 29 mutations were identified. These TP53 mutations were distributed between exon 4 and 8 and most of them were missense mutations (19/29; 65.5%). Immunohistochemical analysis for TP53 revealed that most of the TP53 missense mutations were associated with TP53 protein accumulation. Among the 29 mutations, one was novel (c.459_477delCGGCACCCGCGTCCGCGCC). This 19-bp deletion mutation in exon 5 caused the production of truncated TP53 protein (p.G154Wfs*10). Regarding the spectrum of TP53 mutations, G:C > A:T at CpG sites was the most prevalent (10/29; 34.5%) and G:C > T:A was the second most prevalent (7/29; 24.1%). Interestingly, when the mutation spectrum of TP53 was compared to three previous TP53 mutational studies on non-Rwandan patients with gastric cancer, G:C > T:A mutations were significantly more frequent in this study than in our previous study (p = 0.013), the TCGA database (p = 0.017), and a previous study on patients from Hong Kong (p = 0.006). Even after correcting for false discovery, statistical significance was observed.ConclusionsOur results suggested that TP53 G:C > T:A transversion mutation in Rwandan patients with gastric cancer is more frequent than in non-Rwandan patients with gastric cancer, indicating at an alternative etiological and carcinogenic progression of gastric cancer in Rwanda.

Targeted DNA sequencing of high-grade serous ovarian carcinoma reveals association of TP53 mutations with platinum resistance when combined with gene expression.

High-grade serous ovarian carcinoma (HGSC) is the most common subtype of ovarian cancer and is among the most fatal gynecological malignancies worldwide, due to late diagnosis at advanced stages and frequent therapy resistance. In 47 HGSC patients, we assessed somatic and germline genetic variability of a custom panel of 144 known or suspected HGSC-related genes by high-coverage targeted DNA sequencing to identify the genetic determinants associated with resistance to platinum-based therapy. In the germline, the most mutated genes were DNAH14 (17%), RAD51B (17%), CFTR (13%), BRCA1 (11%), and RAD51 (11%). Somatically, the most mutated gene was TP53 (98%), followed by CSMD1/2/3 (19/19/36%), and CFTR (23%). Results were compared with those from whole exome sequencing of a similar set of 35 HGSC patients. Somatic variants in TP53 were also validated using GENIE data of 1287 HGSC samples. Our approach showed increased prevalence of high impact somatic and germline mutations, especially those affecting splice sites of TP53, compared to validation datasets. Furthermore, nonsense TP53 somatic mutations were negatively associated with patient survival. Elevated TP53 transcript levels were associated with platinum resistance and presence of TP53 missense mutations, while decreased TP53 levels were found in tumors carrying mutations with predicted high impact, which was confirmed in The Cancer Genome Atlas data (n = 260). Targeted DNA sequencing of TP53 combined with transcript quantification may contribute to the concept of precision oncology of HGSC. Future studies should explore targeting the p53 pathway based on specific mutation types and co-analyze the expression and mutational profiles of other key cancer genes.

Abstract A052: ROS in ovulatory follicular fluid exert de novo mutagenic activities through activation-induced cytidine deaminase acting on the TP53 gene in the fallopian tube epithelial cell

Abstract Objective: Ovulation is a major risk factor of epithelial ovarian cancer (EOC), presumably acts from the cancer initiation stage. High-grade serous carcinoma (HGSC) is the most prevalent and fatal EOC, mainly arising from fallopian tube fimbrial epithelium (FTE). TP53 missense mutations are considered the earliest event of carcinogenesis since they form “p53 signatures” in FTE and were found in virtually all HGSC. Previously, we discovered that a subset of ovulatory follicular fluids (FF) harbored high levels of ROS, which were both mutagenic and oncogenic, causing DNA double-strand breaks (DSB) and tumorigenesis. However, DSB is unlikely the cause of TP53 mutations which are mostly CG: TA base substitutions involving deamination. We hypothesized that during ovulation FF exposes the fallopian tube fimbria and causes TP53 mutations by activating the activation-induced cytidine deaminase (AID). Materials and Methods: By conducting an HPRT gene mutation assay, we determined the mutagenic activity of FF on FTE cells with or without AID-knockdown. AID gene activation and protein nuclear translocation were analyzed in human FTE tissues and cells with different degrees of transformation, as well as in the oviducts of mice under controlled ovulation. The binding of AID to the TP53 gene and the subsequent deamination were determined by ChIP-PCR and uracil-qPCR, respectively. Results: After FF exposure, the mutation frequency of HPRT in FTE cells increased by 4.1 folds, at levels 40% higher than the ethyl methane sulfonate control and could be diminished by antioxidants or knockdown of the AID gene. AID activation, including mRNA and protein inductions and nuclear translocation, was evident in FTE cells and tissue after FF exposure and in mouse oviducts after ovulation. Two waves of AID activations were noted, peaking at 45 minutes (early activity) and 8.5 hours (late activity) after ovulation or FF exposure. TNF-α and ROS were found to be responsible for the activities in FF. Mechanistically, the early activity worked through the MAPK, while the late activity worked through the NF-κB pathway. The induced AID in both activities physically binds to and biochemically deaminates the TP53 gene irrelative to the known mutational hotspots of HGSC. However, it did not act on PTEN, the other prevalent tumor suppressor gene in HGSC. Meanwhile, ovulatory FF also activates the putative tumor-initiating and stemness gene LGR6 in FTE at locations where AID was also activated. Conclusion: The serial studies characterized the mutagenic activities of ovulatory FF, which acts on the LGR6-expressing cells in the fallopian tube fimbria and targets the TP53 gene. The study also identified ROS as the mutagen and pointed to strategies of ovulation inhibition and antioxidation for the prevention of EOC. Citation Format: Tang-Yuan Chu, Sanchana Subramani, Hsuan-Shun Huang. ROS in ovulatory follicular fluid exert de novo mutagenic activities through activation-induced cytidine deaminase acting on the TP53 gene in the fallopian tube epithelial cell [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A052.

Relevance of mutations in protein deubiquitinases genes and TP53 in corticotroph pituitary tumors.

Corticotroph pituitary neuroendocrine tumors (PitNETs) develop from ACTH-producing cells. They commonly cause Cushing's disease (CD), however, some remain clinically silent. Recurrent USP8, USP48, BRAF and TP53 mutations occur in corticotroph PitNETs. The aim of our study was to determine frequency and relevance of these mutations in a possibly large series of corticotroph PitNETs. Study included 147 patients (100 CD and 47 silent tumors) that were screened for hot-spot mutations in USP8, USP48 and BRAF with Sanger sequencing, while 128 of these patients were screened for TP53 mutations with next generation sequencing and immunohistochemistry. USP8 mutations were found in 41% CD and 8,5% silent tumors, while USP48 mutations were found in 6% CD patients only. Both were more prevalent in women. They were related to higher rate of biochemical remission, non-invasive tumor growth, its smaller size and densely granulated histology, suggesting that these mutation may be favorable clinical features. Multivariate survival analyses did not confirm possible prognostic value of mutation in protein deubiquitinases. No BRAF mutations were found. Four TP53 mutations were identified (2 in CD, 2 in silent tumors) in tumors with size >10mm including 3 invasive ones. They were found in Crooke's cell and sparsely granulated tumors. Tumors with missense TP53 mutations had higher TP53 immunoreactivity score than wild-type tumors. Tumor with frameshift TP53 variant had low protein expression. TP53 mutation was a poor prognostic factor in CD according to uni- and multivariate survival analyses in spite of low mutations frequency. We confirmed high prevalence of USP8 mutations and low incidence of USP48 and TP53 mutations. Changes in protein deubiquitinases genes appear to be favorable prognostic factors in CD. TP53 mutations are rare, occur in both functioning and silent tumors and are related to poor clinical outcome in CD.

EGFR amplification indicated poor prognosis in EGFR‐mutated lung cancer with leptomeningeal metastases

AbstractBackgroundLeptomeningeal metastasis (LM) is a lethal complication of advanced lung cancer, and due to limited access to the leptomeningeal lesion, we explored the potential role of cerebrospinal fluid (CSF) as a source for liquid biopsy in LM patients of lung cancer with EGFR mutation.Materials and MethodsFrom August 2018 to June 2021, we collected CSF samples of lung cancer patients at First Affiliated Hospital of Zhengzhou University. Next‐generation sequencing was performed to detect the mutations in EGFR genes, and 38 patients detected with EGFR mutations were finally enrolled in further clinical analyses.ResultsTP53 missense mutation (50%) was the most frequently detected concurrent gene in CSF. In those 10 patients whose CSF was obtained upon resistance to first TKI, TP53 missense mutation (40%, n = 4) and EGFR copy number amplification (40%, n = 4) was detected with high frequency; meanwhile, T790M mutation was found only in two patients. Known mechanisms of acquired resistance to third EGFR‐TKIs were found in 31.8% of cases. C797S or C797G mutation was identified in three patients. Possible EGFR‐independent resistant mechanism included MET amplification (4.5%), RET gene fusion (4.5%), PIK3CA missense mutation (4.5%) and CDK4 amplification (4.5%). The median OS was 55 months, the median OSLM was 38 months. EGFR amplification was associated with shortened OS in these EGFR‐mutated lung cancer with leptomeningeal metastases.ConclusionEGFR amplification indicated poor prognosis in EGFR‐mutated lung cancer with leptomeningeal metastases, providing a novel pathogenesis and treatment direction of these patients.

Abstract B012: TP53 missense mutations and keratin 17 are negative prognostic biomarkers in pancreatic ductal adenocarcinoma

Abstract Background: Cancer patients with similar clinical and pathologic characteristics may have different responses to treatments and survival, which may be attributable to distinct molecular properties of their tumors. In pancreatic ductal adenocarcinomas (PDACs), there is a pressing need to identify prognostic biomarkers that could help us to predict disease progression and patient survival. Our group previously reported that keratin17 (K17) expression is a hallmark of PDACs of patients with the shortest patient survival. Furthermore, there is some evidence suggesting that missense mutations in TP53 are associated with worse survival in patients with PDAC. To date, however, no studies have addressed the differential prognostic value of molecular characterization of TP53 coupled with K17 expression for predicting survival in patients with PDAC. Objective and methods: To fill this gap, we assessed the effect of different TP53 mutations and K17 expression on patient survival, using a clinically annotated cohort of 128 patients containing molecular profiling on patient tumors. K17 Immunohistochemistry (IHC) staining on patient’s formalin-fixed, paraffin-embedded (FFPE) tissue blocks were performed to assess the expression of this marker in tumor cells. K17 IHC scoring was conducted based on a subjective assessment of the overall proportion of stained cells relative to the overall number of tumor cells present in a single representative section from each case. Kaplan-Meier and Cox proportional hazards regression models were analyzed to determine survival differences of patients with PDACs harboring different TP53 mutations and levels of K17 IHC expression, adjusting for patient demographics (age at diagnosis, sex, race/ethnicity) and tumor characteristics (presence of distant metastasis at diagnosis). Results: Tumors were classified as high for K17 when ≥90% of tumor cells showed strong staining. High-K17 cases (n=16;12.50%) had shorter survival compared with low-K17 cases (n=112; 87.50%) (26.80 vs. 16.77 months; P=0.035). The majority of cases harbored missense mutations in TP53 (42.19%), while about one-third harbored truncating TP53 (27.34%) mutations. We found that PDACs with missense TP53 mutations and high K17 were associated with worse patient survival compared to tumors with missense TP53 and low K17 expression (HR=5.27; 95% CI, 1.76-15.82; P=0.003). Similarly, compared with patients with wild-type TP53 and low K17, patients with both missense TP53 mutations and high K17 had significantly worse survival (HR=3.51, 95% CI, 1.45-8.51; P=0.005). Conclusions: Our findings suggest that molecular characterization of specific TP53 mutations coupled with K17 IHC staining may help to determine differences in patient prognosis and identify subgroups with worse patient survival, therefore, potentially guiding future clinical management of patients. Citation Format: Carlos Mauricio Mejia Arbelaez, Gabriella Baraks, Lyanne Oblein, Michael Horowitz, Lynn Matrisian, Kenneth Shroyer, Luisa Fernanda Escobar-Hoyos. TP53 missense mutations and keratin 17 are negative prognostic biomarkers in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr B012.

TP53 missense mutation reveals gain-of-function properties in small-sized KRAS transformed pancreatic ductal adenocarcinoma

BackgroundAlthough the molecular features of pancreatic ductal adenocarcinoma (PDAC) have been well described, the impact of detailed gene mutation subtypes on disease progression remained unclear. This study aimed to evaluate the impact of different TP53 mutation subtypes on clinical characteristics and outcomes of patients with PDAC.MethodsWe included 639 patients treated with PDAC in Ruijin Hospital affiliated to Shanghai Jiaotong University School of Medicine between Jan 2019 and Jun 2021. The genomic alterations of PDAC were analyzed, and the association of TP53 mutation subtypes and other core gene pathway alterations with patients’ clinical characteristics were evaluated by Chi-squared test, Kaplan-Meier method and Cox regression model.ResultsTP53 missense mutation was significantly associated with poor differentiation in KRASmut PDAC (50.7% vs. 36.1%, P = 0.001). In small-sized (≤ 2 cm) KRASmut tumors, significantly higher LNs involvement (54.8% vs. 23.5%, P = 0.010) and distal metastic rate (20.5% vs. 2.9%, P = 0.030) were observed in those with TP53 missense mutation instead of truncating mutation. Compared with TP53 truncating mutation, missense mutation was significantly associated with reduced DFS (6.6 [5.6–7.6] vs. 9.2 [5.2–13.3] months, HR 0.368 [0.200–0.677], P = 0.005) and OS (9.6 [8.0-11.1] vs. 18.3 [6.7–30.0] months, HR 0.457 [0.248–0.842], P = 0.012) in patients who failed to receive chemotherapy, while higher OS (24.2 [20.8–27.7] vs. 23.8 [19.0–28.5] months, HR 1.461 [1.005–2.124], P = 0.047) was observed in TP53missense cases after chemotherapy.ConclusionsTP53 missense mutation was associated with poor tumor differentiation, and revealed gain-of-function properties in small-sized KRAS transformed PDAC. Nonetheless, it was not associated with insensitivity to chemotherapy, highlighting the neoadjuvant therapy before surgery as the potential optimized strategy for the treatment of a subset of patients.

Longitudinal Noninvasive Surveillance & Fragmentomic Characterization of Follicular Lymphoma

Introduction While cell-free DNA (cfDNA) plays an increasingly defined role in aggressive lymphomas, its characteristics in indolent lymphomas are less established. Many follicular lymphoma (FL) patients experience durable remissions and late relapses ( Fig A). We therefore explored circulating tumor DNA (ctDNA) kinetics during long-term blood-based surveillance in patients with FL. In addition to noninvasive detection using somatic mutations, we also evaluated gene expression inference from cfDNA utilizing a novel fragmentomic method to detect histological transformation (tFL). Methods We studied 121 serial samples in a cohort of 17 FL patients (median 7 per patient) with long term clinical and blood-based MRD monitoring (median 5.6 years, max 10.8). We considered serial ctDNA status at 4 milestones: baseline/pre-treatment, post-treatment, in radiographic remission (MRD), and at relapse. We also profiled pre-treatment cfDNA for 9 tFL cases. Cases were genotyped to identify somatic mutations from diagnostic tumor or baseline plasma, with matched PBMCs used to censor germline variants and clonal hematopoiesis. ctDNA status in subsequent samples was evaluated via CAPP-Seq (SNVs) and PhasED-Seq (PVs). The cfDNA fragmentation profiles of pre-treatment FL and control samples were evaluated for inferred gene expression via EPIC-seq (Esfahani Nat Biotech 2022) using a novel 1676-gene panel focused on lymphoid neoplasms, including classic and transformed FL. Results Treatment & Relapse Kinetics Most patients had advanced disease (88% stage III-IV) and Low to Intermediate risk FLIPI (71% 0-2). At first cfDNA evaluation, 9/17 (53%) were previously untreated. Median pre-treatment ctDNA levels were 73.4 hGE/mL in FL, as compared to 236 hGE/mL in DLBCL (Kurtz JCO 2018). Post-treatment, median ctDNA levels dropped to 1.2 hGE/mL. Depth of response was similar between rituximab monotherapy and cytotoxic regimens, with median -2.2 log10 fold change from baseline. MRD Detection PVs were observed in all tumors, consistent with expected aberrant somatic hypermutation in FL. To evaluate MRD detection via PhasED-Seq, we considered plasma timepoints with undetectable ctDNA by CAPP-Seq (n=26) from patients (n=9) with subsequent clinical relapse. MRD was detected in 11 additional specimens (42%) by PhasED-Seq. Among FL patients tested for MRD while in radiographic remission, low-level ctDNA was detectable in 62% of plasma specimens (8/13) obtained within 1 month of negative PET/CT, suggesting potential additional utility for surveillance. Long-term Genomic Stability Sequential biopsies of FL tumors have shown genomic heterogeneity between diagnosis and relapse. We therefore evaluated ctDNA concordance across variant classes & regions. Relative AF was calculated pre-treatment and at relapse (median 2 years between samples, range 1-8 years), normalized by total ctDNA levels. The most stable lesions included missense mutations in CREBBP and TP53, nonsense KMT2D mutations, and BCL2 5'-UTR variants. BCL6 intronic mutations and chr2/chr14 SHM varied between diagnosis/relapse. Plasma versus Cellular Concentrations In contrast to DLBCL, circulating FL cells are frequently detectable at low levels in blood. To compare relative FL concentrations in the cellular versus plasma compartments, we profiled 10 paired specimens from patients with radiographic disease. While tumor-confirmed variants were detected in 7 of 10 cellular specimens, mean allelic frequency (AF) was 5.3x higher in plasma. Fragmentomic Assessment of Transformation We compared gene expression profiles inferred from cfDNA from pre-treatment lymphoma (FL=15, tFL=9) versus control (n=20) samples ( Fig B). Considering genes associated with FL (Huet Lancet Onc 2018), FL patients had significantly higher inferred expression than healthy adults (p=0.036). Likewise, patients with tFL showed higher inferred expression of genes associated with histologic transformation (Gentles Blood 2009, p=0.025). Conclusions Sensitive methods are required for effective MRD assessment in FL given lower ctDNA shedding as compared with DLBCL. Frequent detectable MRD in radiographic remission suggests that integrating molecular surveillance may augment serial imaging in the long-term management of FL. Simultaneous inferred gene expression from FL cfDNA appears to hold promise, including for noninvasive detection of transformation.