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Introduction: The epithelial cells of the endometrium undergo rapid proliferation and shedding each month under the influence of estrogen and progesterone. Recent studies have demonstrated pervasive driver mutations in the endometrium with ageing. However, previous observations have been limited to the microscopic scale, and it is still unclear how clonal expansion occurs over the entire endometrium and what driver mutations are responsible therein. To address these issues, we performed multisampling of the endometrium with extensive spatial mapping. Methods: We collected multiple endometrial glands from normal endometrium in a 5mm lattice in 19 patients who underwent hysterectomy (3-31 samples per case), followed by DNA extraction, detected somatic mutations by whole exome sequencing, to identify large clones that spread across samples based on shared mutations. Results: In total, 216 bulk endometrial glands were analyzed. The number of detected somatic mutations per sample ranged from 6 to 98 (mean±SD: 37.8±16.0), and most of the samples had at least one known driver gene mutation (mean±SD: 4.38±2.27). The mutational signature analysis revealed the predominance of the age-related SBS1 and SBS5 signatures. dN/dS analysis identified 21 genes that were positively selected in normal endometrium, including PIK3CA, PIK3R1, KRAS, PPP2R1A, ARHGAP35, and FBXW7, most of which are also known as driver genes for endometrial cancer. The number of mutations and their frequency for each driver gene did not differ by background diseases: endometriosis, endometriosis-associated ovarian cancer and other non-endometrial related diseases. We found clones showing a large expansion involving multiple samples in 8 cases, most of which harbored one or more driver mutations, such as PIK3CA and KRAS. The size of the largest clone carrying a KRAS mutation, which was found in a 32-year-old woman with cervical cancer, spanned as long as 10 mm in diameter. This suggests that clonal expansion in the endometrium could be very rapid even in healthy women, which contrasts to the observations that clonal expansion in other organs is usually seen in aged individuals. Conclusion: We confirmed the high frequency of driver mutations in the normal endometrium, most of which overlapped to those found in endometrial cancers, suggesting that these driver genes are involved in the early phase of the development in the endometrial cancer. Some of the driver-mutated clones expanded in macroscopic size. Our findings provide an interesting insight into the clonal structure and underlying genetic variation in the normal endometrium. Citation Format: Koichi Watanabe, Nobuyuki Kakiuchi, Shiro Takamatsu, Sachiko Kitamura, Mana Taki, Koji Yamanoi, Ryusuke Murakami, Ken Yamaguchi, Junzo Hamanishi, Hiroko Tanaka, Satoru Miyano, Masaki Mandai, Seishi Ogawa. Clonal expansion with driver mutations in normal human endometrium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 128.

Background: Since the advent of high-throughput sequencing technologies, multi-omics data along with large-scale drug sensitivity screening data on cancer cell lines from the NCI-60 panel and the CCLE (Cancer Cell Line Encyclopedia) project have been established and widely used for cancer research. Although we previously reported the utility of homologous recombination deficiency (HRD), commonly found in ovarian and breast cancer, as a clinical biomarker across cancer types (PMID:35613413), whether HRD status in cancer cell lines can be an indicator of drug sensitivity to platinum agents or PARP inhibitors has not been fully investigated. Methods: From CCLE publicly available datasets, BRCA1/2 gene mutations and their loss of heterozygosity (LOH) status, HRD score, mutational signature 3 (SBS3), and BRCA1 methylation silencing were analyzed. In vitro drug screening data were obtained from the GDSC (Genomics of Drug Sensitivity in Cancer), CTRP (Cancer Therapeutic Response Portal), and PRISM (Profiling Relative Inhibition Simultaneously in Mixtures) databases. We additionally analyzed anticancer drug response in cell line-derived xenograft models using the dataset of ROADMAPS (Responses to Oncology Agents and Dosing in Models to Aid Preclinical Studies). Results: A total of 1260 cell lines from the CCLE were analyzed. HRD score and SBS3 were positively correlated (ρ=0.36). These scores were higher in samples with BRCA1 methylation and BRCA1/2 mutations involving locus-specific LOH (referred to collectively as BRCA alteration), but not in samples with BRCA1/2 mutations lacking locus-specific LOH. Per-drug correlation analysis between cell line HRD scores and the areas under the drug response curve (AUCs) showed statistically significant (p Conclusion: When comparing in vitro drug sensitivity among cancer cell lines, HRD correlates with resistance rather than with response to platinum or PARP inhibitors. Thus, the association between HRD status and drug sensitivity of cancer cells is substantially different between tumors in dish culture and in clinical settings. This study provides critical insights that should be considered when using cancer cell lines and underscores the need for experimental models that better reflect efficacy in clinical tumors. Citation Format: Shiro Takamatsu, Noriomi Matsumura. Homologous recombination deficiency in the CCLE cell lines correlates with in vitro resistance to platinum agents and PARP inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2773.

The improvement of the quality of life (QOL) of gynecological cancer patients is a critical purpose in cancer treatment. At present, there is no other methodology to evaluate QOL other than patients reported outcomes using QOL questionnaires. Therefore, management of QOL is not performed clinically because there is no objective and easy method to evaluate QOL. In this study, we extracted the worst symptom of QOL in gynecological cancer patients and examined whether QOL can be evaluated by lifelogs objectively. Lifelogs (diets, sleep duration, the number of steps, pulse rate, voice, and so on) and QOL questionnaires (EORTC-qlq-c30, PHQ9, and so on) were collected from 139 gynecological cancer patients at our hospital using mobile applications. Scales of symptoms and functions were calculated from EORTC-qlq-c30. Logistic regression analyses and calculation of feature impact in the predictive model depicting receiver operating characteristic (ROC) curve were performed to extract symptoms and functions that contributed most to global health status (GHS) indicating a general QOL status. Fatigue-related metabolites in patient serum were measured by ELISA. Correlations between each lifelog, fatigue scale, and fatigue-related metabolites were examined. A computational model to detect high fatigue scale status was developed using lifelogs. The fatigue scale was the highest among all treatment periods. The fatigue scale, appetite loss scale, and physical functioning scale were symptoms that deteriorate QOL frequently in gynecologic cancer patients (odds ratio: 0.9708, 0.9797, and 1.0374, respectively; 95% CI: 0.9493 to 0.9927, 0.9677 to 0.9919, and 1.0155 to 1.0597, respectively). ROC curve indicated that an accurate prediction of GHS using scales of symptoms and functions (area under the curve, AUC: 0.85). From the scales of symptoms and functions, the fatigue scale showed the highest feature impact to predict GSH. Several lifelogs were significantly correlated with fatigue scores (p In summary, fatigue is a significant problem contributing to the decline in QOL of gynecological cancer patients. Fatigue can be monitored and diagnosed using digital data using lifelogs. These findings lead to the development of future management strategies of QOL for cancer patients. Citation Format: Ken Yamaguchi, Nozomi higashiyama, Akihiko Ueda, Shiro Takamatsu, Masayo Ukita, Mana Taki, Koji Yamanoi, Junzo Hamanishi, Masaki Mandai. Development of a computational algorithm to detect fatigue of gynecologic cancer patients using lifelogs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5261.

Objective: Tertiary lymphoid structures (TLSs) are known to be a marker of peripheral inflammation. While high densities of TLSs were reported to be associated with longer patient survival in several cancer types, there is no direct evidence where B cells in TLSs(TLBs) are derived from and whether these clones are consistent with tumor infiltrating B cells (TIBs), in endometrial cancer. Methods: We analyzed the pathological distribution of TLSs and TIBs in human endometrial cancer samples using immunohistochemistry of a series of B cell markers and evaluated the relationship with patients’ clinical outcomes. Additionally, we micro-dissected six pairs of tumor nests and their neighbor TLSs and analyzed the clonal consistency between TLBs and TIBs using the sequencing of complementarity-determining region (CDR) of B cell receptor (BCR). Results: In 104 patients with endometrial cancer, TLSs were detected in 81 patients (78%), and the absence of TLS was related to worse progression-free survival (PFS) (HR, 0.15, P Conclusion: TLSs and TIBs are associated with better prognosis in endometrial cancer. Common B cell clones were found in the advanced stage cases, suggesting that B cells may have gradually acquired immunospecificity in TLSs. Citation Format: Haruka Suzuki, Junzo Hamanishi, Meng Qin, Masayo Ukita, Akihiko Ueda, Shunsuke Kawahara, Koji Yamanoi, Taito Miyamoto, Shiro Takamatsu, Mana Taki, Yoko Hosoe, Takashi Kobayashi, Ken Yamaguchi, Masaki Mandai. Clonal relationship of B cells between tertiary lymphoid structures (TLSs) and tumor infiltrating lymphocytes (TILs) in advanced endometrial cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6109.

[Background] Genomic alterations in BRCA1/2 and the ‘genomic scar' signatures are associated with homologous recombination DNA repair deficiency (HRD) and serve as therapeutic biomarkers for sensitivity to platinum and PARP inhibitors in breast and ovarian cancers. However, the clinical significance of these biomarkers in other homologous recombination repair (HRR) pathway genes or in other cancer types is not fully understood. [Method] From the TCGA and the AACR GENIE project databases, we obtained clinical and genomic data of patients with multiple solid cancers, evaluated both locus-specific LOH in HRR pathway gene mutations and two genomic scar signature scores (HRD score and mutational signature 3), identified potential HRD cases, and examined the efficacy of DNA-damaging drugs in these HRD cases. [Results] Germline and somatic gene mutations in the HRR pathway genes with locus-specific LOH were significantly enriched in cases having high genomic scar scores, whereas those without the LOH were not enriched. Somatic mutations without the LOH were remarkably enriched in hypermutated cases and considered to be passenger mutations. The mutation rate and frequency with LOH in the HRR pathway genes varied widely by cancer type. When stratifying patients by gender and presence of TP53 mutation, the correlation between the two genomic scar scores and the resulting optimal cutoff values for biallelic HRR pathway gene alterations differed greatly. The potential HRD cases that were selected by the two genomic scar cutoffs and biallelic HRR pathway gene alterations showed significantly high gene expression-based HRD scores. In Cox proportional hazard multivariate analyses with covariates of age, stage, gender, HRD, and TP53 mutation, HRD was a good prognostic factor in the group with DNA-damaging agents, but a poor one in the group without the agents. [Conclusion] Increased attention on locus-specific LOH in HRR pathway gene alterations and the differences in genomic scar signatures stratified by gender and TP53 mutation is newly warranted in order to properly assess HRD in a pan-cancer manner. The results in the present study proved the clinical significance of HRD across cancer types and may contribute to the development of personalized medicine based on HRD. Citation Format: Shiro Takamatsu, JB Brown, Koji Yamanoi, Ken Yamaguchi, Junzo Hamanishi, Takashi Kobayashi, Masaki Mandai, Noriomi Matsumura. Identification of a clinically significant pan-cancer biomarker for homologous recombination deficiency [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 366.