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Although circulating tumour DNA (ctDNA)-based next-generation sequencing (NGS) is a less invasive method for assessing ESR1 mutations that are essential mechanisms of endocrine therapy resistance in patients with oestrogen receptor-positive breast cancer, adequate amounts of DNA are required to assess polyclonal ESR1 mutations. By combining a peptide nucleic acid and locked nucleic acid polymerase chain reaction (PNA-LNA PCR) clamping assay, we have developed a novel detection system to screen for polyclonal ESR1 mutations in ctDNA. A validation assay was prospectively performed on clinical samples and compared with the NGS results. The PNA-LNA PCR clamp assay was validated using six and four blood samples in which ESR1 mutations were detected by NGS and no mutations were detected, respectively. The PNA-LNA assay results were comparable with those of NGS. We prospectively assessed the concordance between the PNA-LNA PCR clamp method and NGS. Using the PNA-LNA PCR clamp method, ESR1 mutations were detected in 5 out of 18 samples, including those in which mutations were not detected by NGS due to small amounts of ctDNA. The PNA-LNA PCR clamping method is a highly sensitive and minimally invasive assay for polyclonal ESR1 mutation detection in the ctDNA of patients with breast cancer.

Background: Trabectedin is used as a treatment for advanced-stage soft tissue sarcomas (STSs), particularly liposarcoma and leiomyosarcoma. Aside from its direct effect on tumor cells, trabectedin can affect the immune system in the tumor microenvironment. This study aimed to evaluate whether inflammatory biomarkers predict trabectedin efficacy in STSs. Methods: We retrospectively reviewed the clinical features and outcomes of patients with STS treated with trabectedin at our institution between 2016 and 2020. The neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), platelet-to-lymphocyte ratio (PLR), and systemic inflammation response index (SIRI = neutrophil × monocyte/lymphocyte) were calculated based on the blood samples obtained prior to trabectedin treatment initiation. Analyses of the overall survival (OS) and progression-free survival (PFS) were performed according to various factors. Results: Of the 101 patients identified, 54 had L-sarcoma (leiomyosarcoma: 30; liposarcoma: 24), and 47 had other types of STSs. Elevated SIRI, NLR, PLR, LMR, and C-reactive protein (CRP) were associated with worse PFS (P < 0.001, P = 0.008, P = 0.027, P = 0.013, and P < 0.001, respectively) according to the results of the univariate analysis. Multivariate analysis showed that elevated SIRI, other histology, and CRP were associated with poor PFS (P = 0.007, P = 0.008, and P = 0.029, respectively). In addition, the multivariate analysis of OS showed that SIRI was an independent prognostic factor (hazard ratio: 2.16, P = 0.006). Conclusion: Pretreatment SIRI can be considered a biomarker for the prognostic prediction of patients with STS treated with trabectedin.

Background: Abemaciclib is a drug approved for ER-positive and HER2-negative metastatic or recurrent breast cancer and those treatable in an early setting. Some patients with ER-positive and HER2-negative breast cancer pretreated with abemaciclib and endocrine therapy may still benefit from abemaciclib rechallenge, especially in case of acquired resistance to endocrine therapy. However, there is no evidence for abemaciclib rechallenge. To address this, a single-arm phase 2 trial is planned to determine whether abemaciclib rechallenge has clinical benefits. Translational research to evaluate the gene alteration and immunohistochemistry will be conducted as an accompanying research to determine the predictive biomarkers of resistance or sensitivity to abemaciclib. Patients and Methods: The eligible patients are histologically confirmed with ER-positive and HER2-negative invasive breast cancer, locally advanced or metastatic, and previously received no more than two lines of endocrine therapy. The patients who previously received abemaciclib in combination with endocrine therapy led to a clinical benefit, including complete response, partial response, or stable disease, during abemaciclib-based treatment (≥6 months). The patients who previously received chemotherapy, immune checkpoint inhibitors (excluding perioperative systemic treatment), everolimus, olaparib, and palbociclib for advanced or metastatic diseases are excluded. The patients will be treated with abemaciclib 150 mg orally q12h on days 1 to 28 of a 28-day cycle. The other endocrine drug prescribed was not used in previous treatment. We will evaluate the gene mutations and/or amplification of ESR1, PIK3CA, CCNE1, FGFR1, RB1, TP53, NF1, MYC, AR, and MET at the time of inclusion and three months after day 1 and ctDNA and immunostaining levels of HER3, PTEN, CCNE1, FGFR1/2, and AR at the time of inclusion. The primary endpoint is progression-free survival (PFS), while the secondary endpoints are overall response rate, clinical benefit rate, chemotherapy-free interval, overall survival (OS), safety, and evaluation of the mechanism of sensitivity and resistance to abemaciclib. The expected median PFS for the study treatment arm is 5.0 months compared to the previous data of 3.0 months. The number of necessary cases is 59, with a significance level of 5% (two-sided) and power of 80% during the two-year recruitment period and one-year follow-up period. In the planned total of 65 cases, 10% is considered inappropriate. The planned duration of enrollment and follow-up is two years and one year, respectively. The planned total study duration is four years. Subgroup analysis will be performed on ESR1 and PIK3CA mutations, disease site (visceral/bone only/other), previous lines of therapy for advanced or metastatic disease (first and second), endocrine therapy for pretreatment (aromatase inhibitors/fuluvestrant), performance status (0/1), organs involved (1/2/≥3), age ( Citation Format: Meiko Nishimura, Takahiro Kogawa, Yuko Akaishi, Misato Ogata, Jun Masuda, Mitsuo Terada, Hitomi Sakai, Kazuki Nozawa, Sasagu Kurozumi, Takamichi Yokoe, Yukinori Ozaki Ozaki, Shu Yazaki, Mai Onishi, Tsutomu Iwasa, Takuma Onoe, Yuta Okumura, Sayaka Nakayama, Kanako Hagio, Yuko Takahashi, Hirokazu Tanino, Junji Tsurutani, Koji Matsumoto, Mototsugu Shimokawa, Toshimi Takano. Clinical evaluation of the efficacy and liquid molecular analysis of abemaciclib rechallenge upon progression to abemaciclib combination therapies for ER-positive HER2-negative metastatic breast cancer patients [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr OT2-19-02.

Objective Given the rarity of cutaneous/subcutaneous Ewing sarcoma, their clinical characteristics remain poorly understood. In this study, we aimed to analyse the clinical characteristics of patients with cutaneous/subcutaneous Ewing sarcoma and review the treatment strategy. Methods We reviewed the clinical data of 154 patients with Ewing sarcoma who were treated at our hospital between 2005 and 2019. Amongst these patients, 21 patients with cutaneous/subcutaneous Ewing sarcoma were analysed. As a basic strategy, patients with localized disease received intensive chemotherapy (vincristine–doxorubicin–cyclophosphamide/ifosfamide–etoposide), followed by definitive surgery with or without radiotherapy. In total, 15 patients underwent pre-diagnostic resection with macroscopic residue (seven patients) or non-macroscopic residue (eight patients) before intensive chemotherapy. Results The median tumour length of the measurable lesions was 3.2 cm, and the ratio of metastasis was significantly lower than the Ewing sarcoma of other anatomical sites (10% vs. 37%, P = 0.013). Despite the pre-diagnostic resection, local recurrence after additional resection and/or adjuvant radiotherapy did not occur in any of the patients with localized disease. Overall survival was significantly higher in patients with cutaneous/subcutaneous Ewing sarcoma than in patients with Ewing sarcoma of other anatomical sites (hazard ratio = 0.33, P = 0.013). The event-free survival rate of cutaneous/subcutaneous Ewing sarcoma was also superior to that of Ewing sarcoma of other anatomical sites (hazard ratio = 0.35, P = 0.01). Conclusions Patients with cutaneous/subcutaneous Ewing sarcoma may have better prognosis than those with Ewing sarcoma at other anatomical sites. Although pre-diagnostic resection without appropriate investigations is not recommended, local control may be recovered by using a combination of additional resection, chemotherapy and radiotherapy.

Background Human epidermal growth factor receptor-3 (HER3) is a member of the epidermal growth factor receptor family of receptor tyrosine kinases, and its overexpression is associated with inferior prognosis in several cancers. However, it is unclear whether HER3 expression status changes in tumor tissue at recurrence. Therefore, this study aimed to evaluate the changes in HER3 expression between primary and recurrent status in gynecological cancers. Methods This retrospective study used matched-pair tissues of gynecological cancer patients at initial diagnosis and at recurrence. Immunohistochemical (IHC) scores of 3 + or 2 + were termed “HER3-high”, while IHC scores of 1 + or 0 were designated as “HER3-low/zero”. Results A total of 86 patients (40 with ovarian cancers, 32 with endometrial cancers, and 14 with cervical cancers) were included in this study. In ovarian cancer, 67.5% and 80.0% of the patients received a HER3-high at initial and recurrent diagnosis, respectively. The H-score was significantly increased at recurrence (p = 0.004). The proportion of HER3-high endometrial cancer patients increased from 46.9% at initial diagnosis to 68.8% at recurrence, and the H-score tended to increase at recurrence (p = 0.08). The fraction of HER3-high-rated cervical cancer patients remained unchanged at 85.7% both at initial and recurrent diagnosis. The discordance rate of HER3 expression detection in initial and recurrent diagnosis samples was 27.5%, 53.1%, and 14.3% for ovarian, endometrial, and cervical cancers, respectively. Ovarian and endometrial cancers with a HER3-high recurrent score tended to show shorter median survival time than those with a HER3-low/zero recurrent rating. Conclusion Our findings suggest that, in main types of gynecological cancers, the proportion of patients having a HER3-high score increased from initial to recurrent diagnosis.

Introduction: Synovial sarcoma (SS) predominantly affects adolescents and young adults. Doxorubicin with or without ifosfamide therapy is the standard first-line treatment for unresectable or metastatic SS. However, there is no standard second-line chemotherapy regimen. The purpose of the current study was to evaluate the outcomes of second-line chemotherapy for patients with SS. Methods: We retrospectively evaluated the outcomes of 61 patients with unresectable or metastatic SS who had received first-line chemotherapy at our institution between 1997 and 2017. Patients who received second-line chemotherapy were included in the analysis. Outcomes of the chemotherapy were evaluated. Results: Among the 61 patients treated with first-line chemotherapy, we identified 32 patients who received second-line chemotherapy. Most patients (62.5%) were under 40 years of age. Regarding second-line chemotherapy regimens, 6 (18.8%) patients were treated with doxorubicin with/without ifosfamide, 6 (18.8%) with ifosfamide and etoposide, 4 (12.5%) with docetaxel and gemcitabine, 5 (15.6%) with pazopanib, 2 (6.2%) with trabectedin, and 1 (3.1%) with eribulin. The overall response rate according to the Response Evaluation Criteria in Solid Tumors for all patients was 9.4%. Eleven patients (34.3%) achieved disease-control for >6 months. The median follow-up duration was 15.2 months. The 1-year progression-free and overall survival rates were 33.1% and 67.1%, respectively. Conclusion: Our exploratory study revealed that the response rate of second-line chemotherapy regimens for patients with SS was 9.4%. Therefore, there is an urgent need to develop more active therapeutic regimens for SSs.

Purpose Stromal tumor-infiltrating lymphocytes (TILs) are independent prognostic factors in systemically untreated early-stage triple-negative breast cancer (TNBC). Other immune biomarkers including CD8, CD20, programmed cell death-ligand 1 (PD-L1), and tertiary lymphoid structures (TLS) are also reported to be associated with prognosis. However, whether combining other immune biomarkers with TILs would allow for further prognostic stratification is unknown. Methods We retrospectively analyzed 125 patients with early-stage TNBC not receiving perioperative chemotherapy. Stromal TILs and TLS were evaluated on hematoxylin–eosin slides. PD-L1 expression was evaluated using the SP142 assay. CD8 and CD20 were assessed by immunohistochemistry and counted by digital pathology. Results Immune biomarker levels were positively correlated (p 2 = 9.24, p = 0.01). In Cox regression analysis, high CD8 was significantly associated with better prognosis [hazard ratio (HR) 0.69, 95% confidence interval (CI) 0.48–0.98, p = 0.04], and PD-L1 positivity was significantly associated with worse prognosis (HR 4.33, 95%CI 1.57–11.99, p = 0.005). Patients with high CD8/PD–L1 (–) tumors had the most favorable prognosis [5 year invasive disease-free survival (iDFS), 100%], while patients with low CD8/PD-L1( +) tumors had the worst prognosis (5 year iDFS, 33.3%). Conclusion CD8 and PD-L1 levels add prognostic information beyond TILs for early-stage TNBC not receiving perioperative chemotherapy. CD8–positive T cells and PD-L1 may be useful for prognostic stratification and in designing future clinical trials of TNBC.

Background Chemotherapy and radiotherapy were postulated to induce an inflamed tumour microenvironment. We aimed to evaluate the effects of adjuvant chemotherapy/radiotherapy on tumour-infiltrating lymphocytes (TILs) and programmed death-ligand 1 (PD-L1) expression in metastatic breast cancer. Methods We identified paired primary and metastatic tumours in 85 patients with breast cancer. Stromal TILs were assessed according to international guidelines. PD-L1 expression was evaluated using the VENTANA SP142 assay. Results TILs were significantly lower in metastatic tumours than in primary tumours (12.2 vs. 8.3%, p = 0.049). PD-L1 positivity was similar between primary and metastatic tumours (21.2 vs. 14.1%, p = 0.23). TILs were significantly lower in patients who received adjuvant chemotherapy than in those who did not (−9.07 vs. 1.19%, p = 0.01). However, radiotherapy had no significant effect on TILs (p = 0.44). Decreased TILs predicted worse post-recurrence survival (hazard ratio, 2.94; 95% confidence interval [CI]: 1.41–6.13, p = 0.003), while increased TILs was associated with a better prognosis (HR, 0.12; 95% CI: 0.02–0.08, p = 0.04). Conclusions TILs decreased in metastatic tumours, particularly in patients who relapsed after adjuvant chemotherapy. Changes in TILs from primary to metastatic sites could be a prognostic factor after recurrence.

Background Mesothelin (MSLN) is a cell-surface glycoprotein found in various solid tumours. Cancer therapies targeting MSLN have been developed in recent years; however, the available information on MSLN expression in cervical cancer is limited. This study aimed to evaluate MSLN expression in various histological types of cervical cancer and examine its relationship with prognosis. Methods This retrospective study included patients with cervical cancer who underwent primary surgery between January 2000 and December 2020 at our institution. MSLN expression was evaluated by immunohistochemistry using clone SP74 and defined as positive if MSLN was expressed at any intensity. High MSLN expression was defined as an intensity of ≥ 2 + in ≥ 30% of tumour cells. The association between MSLN expression and clinicopathological factors was evaluated. Results Overall, 123 patients were identified, and 140 tumour samples, including 17 paired primary and metastatic samples, were evaluated. Concerning histological type, 67 patients had squamous cell carcinoma (SCC), whereas 56 had non-SCC. MSLN expression was observed in 98.4% (121/123) of primary tumours. High MSLN expression was observed in 63.4% of samples (78/123), but it differed between the histological types (49.2% for SCC vs. 80.4% for non-SCC, p p = 0.015). In patients with common histological types, overall survival (OS) was shorter in the high MSLN expression group than in the low MSLN expression group (hazard ratio, 3.53; 95% confidence interval, 1.16–15.3, p = 0.03). Conclusions MSLN was highly expressed in patients with cervical cancer, especially in those with non-SCC. High MSLN expression in the primary lesion was significantly associated with poor OS, and its expression was maintained in metastatic lesions. Our findings indicate that MSLN may be an attractive therapeutic target for cervical cancer. Trial registration Retrospectively registered. 2014-393. 1 June 2015

Background Trastuzumab emtansine (T-DM1) treatment for human epidermal growth factor receptor-2 (HER2)-positive metastatic breast cancer after taxane with trastuzumab and pertuzumab is standard therapy. However, treatment strategies beyond T-DM1 are still in development with insufficient evidence of their effectiveness. Here, we aimed to evaluate real-world treatment choice and efficacy of treatments after T-DM1 for HER2-positive metastatic breast cancer. Methods In this multi-centre retrospective cohort study involving 17 hospitals, 325 female HER2-positive metastatic breast cancer patients whose post-T-DM1 treatment began between April 15, 2014 and December 31, 2018 were enrolled. The primary end point was the objective response rate (ORR) of post-T-DM1 treatments. Secondary end points included disease control rate (DCR), progression-free survival (PFS), time to treatment failure (TTF), and overall survival (OS). Results The median number of prior treatments of post-T-DM1 treatment was four. The types of post-T-DM1 treatments included (1) chemotherapy in combination with trastuzumab and pertuzumab (n = 102; 31.4%), (2) chemotherapy concomitant with trastuzumab (n = 78; 24.0%), (3), lapatinib with capecitabine (n = 63; 19.4%), and (4) others (n = 82; 25.2%). ORR was 22.8% [95% confidence interval (CI): 18.1–28.0], DCR = 66.6% (95% CI 60.8–72.0), median PFS = 6.1 months (95% CI 5.3–6.7), median TTF = 5.1 months (95% CI 4.4–5.6), and median OS = 23.7 months (95% CI 20.7–27.4). Conclusion The benefits of treatments after T-DM1 are limited. Further investigation of new treatment strategies beyond T-DM1 is awaited for HER2-positive metastatic breast cancer patients.

Tumor human epidermal growth factor receptor 2 (HER2) status is defined by either protein expression using immunohistochemistry (IHC) or gene amplification using fluorescence in situ hybridization (FISH). Approximately 20% of HER2-positive breast cancer is HER2 IHC 2+/FISH-positive. Unlike trastuzumab, it has not been studied whether the response to trastuzumab emtansine (T-DM1) differs according to HER2-positive status. We retrospectively identified and reviewed medical records of all patients with HER2-positive advanced breast cancer (ABC) who received T-DM1 in our hospital from October 2013 to December 2016. We compared the objective response rate (ORR) and progression-free survival (PFS) between patients in the HER2 IHC 3+ group and those in the HER2 IHC 2+/FISH-positive group. A total of 39 patients (IHC 3+: n = 32; IHC 2+/FISH-positive: n = 7) were analyzed. Nineteen (48.7%), 13 (33.3%), and 29 (74.4%) patients had received at least one prior chemotherapy, more than three lines of chemotherapy, and prior pertuzumab for ABC, respectively. ORR was significantly higher in the IHC 3+ group than in the IHC 2+/FISH-positive group (53.3% vs. 0%, P = 0.024). Median PFS was 7.9 months in the IHC 3+ group versus 3.9 months in the IHC 2+/FISH-positive group (hazard ratio 0.68; 95% confidence interval 0.28-1.69, P = 0.408). Among the HER2-positive ABC patients treated with T-DM1, ORR was significantly worse in HER2 IHC 2+/FISH-positive than in HER2 IHC 3+ patients. Median PFS tended to be shorter in patients with HER2 IHC 2+/FISH-positive.

Background Routine measurement of tumor markers is not recommended in daily clinical practice for patients with cancer of unknown primary (CUP). We evaluated the diagnostic value of tumor markers in identifying favorable or unfavorable subsets in patients with CUP. Methods We retrospectively reviewed the medical records of patients who were diagnosed with CUP between October 2010 and July 2015 at the National Cancer Center Hospital. The tumor markers of the patients were examined, including squamous cell carcinoma antigen, cytokeratin fraction, carcinoembryonic antigen, sialyl Lewis X, neuron-specific enolase, pro-gastrin-releasing peptide, α-fetoprotein, protein induced by vitamin K absence or antagonist II, prostate-specific antigen, soluble interleukin-2 receptor, carbohydrate antigen 19–9, cancer antigen 125, cancer antigen 15–3, NCC-ST-439 (ST439), elastase-1, human chorionic gonadotropin, and sialyl-Tn (STN). Results Among 199 patients with suspected CUP, 90 were diagnosed with confirmed CUP (12 in the favorable subset and 78 in the unfavorable subset). No tumor markers showed 100% sensitivity for unfavorable subsets. ST439 (p = 0.03) and STN (p = 0.049) showed 100% specificity for unfavorable subsets. Conclusions For patients with suspected CUP who show elevated ST439 or STN levels, the treatment strategy should be based on the premise that the patient is likely to be placed in the unfavorable subset.

Background: ESR1 gene mutations are an essential mechanism of resistance to endocrine therapy in patients with oestrogen receptor-positive breast cancer. ESR1 mutations can exist at multiple sites and are typically detected polyclonally after endocrine therapy. The use of circulating tumour DNA (ctDNA) is a less invasive and less expensive method for assessing ESR1 mutations. Adequate amounts of DNA are required to assess polyclonal ESR1 mutations using next-generation sequencing (NGS). The aim of this study was to develop a method to assess polyclonal ESR1 mutations in blood samples.Methods: We developed a novel detection system to screen for ESR1-activated mutations in ctDNA by combining a peptide nucleic acid and locked nucleic acid polymerase chain reaction (PNA-LNA PCR) clamping assay for detection of the most common mutations: E380Q, Y537S, and D538G. Other mutations in codons 536-538 were detected by direct sequencing of the amplified product following clamp PCR. A validation assay was prospectively performed on clinical samples and compared with the NGS results. Results: The PNA-LNA PCR clamp assay was validated using six blood samples in which ESR1 mutations were detected by NGS and four samples in which no mutations were detected. Results of the PNA-LNA assay were comparable with those of NGS. We prospectively assessed the concordance between the PNA-LNA PCR clamp method and NGS. Using the PNA-LNA PCR clamp method, ESR1 mutations were detected in 5 of 18 samples, including those in which mutations were not detected by NGS due to the small amount of ctDNA.Conclusions: The PNA-LNA PCR clamping method is a highly sensitive and minimally invasive assay for the detection of polyclonal ESR1 mutations in the ctDNA of patients with breast cancer.

Background: The prognostic value of stromal tumor-infiltrating lymphocytes (TILs) as a biomarker for triple-negative breast cancer (TNBC) has been extensively demonstrated in patients (pts) receiving (neo)adjuvant systemic therapy. In addition, several small studies suggest that a subset of pts with early-stage TNBC and high TILs have excellent long-term outcomes, even in the absence of systemic therapy [1-3]. However, data on the absolute risk of TNBC recurrence according to TIL levels in the absence of systemic therapy are limited and critical to inform the design of future systemic therapy de-escalation clinical trials. Methods: We conducted an individual patient data pooled analysis of 12 international cohorts of pts with TNBC treated with locoregional therapy but no systemic therapy. TNBC was defined as tumors with estrogen and progesterone receptor of < 1% and HER2 negative (IHC 0, 1+ or IHC 2+ and FISH negative) per local evaluation. TILs were locally assessed in hematoxylin & eosin-stained slides according to the International Immuno-Oncology Biomarker Working Group guidelines (www.tilsinbreastcancer.org). We used the Kaplan-Meier method to assess survival outcomes according to prespecified TIL thresholds: 30% and 50%. Confidence intervals (CI) for survival probabilities were calculated using a percentile bootstrap method. The primary endpoint was invasive disease-free survival (iDFS, STEEP 2.0 definition). Key secondary outcomes included recurrence-free survival (RFS), distant disease-free survival (DDFS) and overall survival (OS). Results: 1,835 pts diagnosed with TNBC between 1982 and 2017 who did not receive systemic therapy were included. The median age at diagnosis was 56 (IQR 38-71). Menopausal status was known in 1,184 women, of whom 78% were post-menopausal. The median tumor size was 2.0 cm (IQR 1.2-2.6). Most pts (87%) had no axillary lymph node involvement (N0). Most tumors were invasive ductal carcinoma (74%) and grade 3 (70%). The median level of TILs was 15% (IQR 5-40). The median duration of follow-up was 30.4 years (95% CI 29.9, 31.1). A total of 950 (52%) iDFS, 828 (45%) RFS, 767 (42%) DDFS events, and 604 (33%) deaths were observed. In multivariable analyses, higher TILs were independently associated with improved iDFS, RFS, DDFS, and OS beyond clinicopathological factors (likelihood ratio p< 10e-6). Each 10% increment in stromal TILs was associated with an 8% (95% CI: 6-11), 10% (95% CI: 7-13), and 13% (95% CI: 10-15) reduction in the risk of experiencing an iDFS, RFS or DDFS event, and with a 12% (95% CI: 9-15) reduction in the risk of death. iDFS, RFS, DDFS and OS rates according to different TIL thresholds and nodal status are shown in the Table. Of note, the RFS estimates (which exclude second non-breast primaries and contralateral breast cancers) were consistently higher than the iDFS counterparts (which include both), consistent with a high rate of contralateral breast cancers and second primary tumors in this cohort. Notably, patients with node-negative—and especially stage I—TNBC with high TILs had excellent survival rates at 10-year follow-up. Conclusion: TILs are highly prognostic in pts with systemically untreated early-stage TNBC. Pts with pN0 (and especially stage I) TNBC with high TILs exhibited very favorable long-term outcomes even in the absence of systemic therapy. These data define the natural history of TIL-rich TNBC pts and are crucial to identifying the optimal patient population for future chemotherapy and immunotherapy de-escalation clinical trials. References: [1] Leon-Ferre et al, 2017, PMID: 28913760 [2] Park et al, 2019, PMID: 31566659 [3] de Jong et al, 2022, PMID: 35353548 Table 5 and 10-year survival endpoints according TIL level, nodal status, and stage Citation Format: Roberto A. Leon-Ferre, Sarah Flora Jonas, Roberto Salgado, Sherene Loi, Vincent De Jong, Jodi M. Carter, Torsten Nielson, Samuel Leung, Nazia Riaz, Giuseppe Curigliano, Carmen Criscitiello, Vincent Cockenpot, Matteo Lambertini, Vera Suman, Barbro Linderholm, John WM Martens, Carolien HM van Deurzen, Mieke Timmermans, Tatsunori Shimoi, Shu Yazaki, Masayuki Yoshida, Sung-Bae Kim, Hee Jin Lee, Maria Vittoria Dieci, Guillaume Bataillon, Anne Salomon, Fabrice Andre, Marleen Kok, Sabine Linn, Matthew P. Goetz, Stefan Michiels. Stromal tumor-infiltrating lymphocytes identify early-stage triple-negative breast cancer patients with favorable outcomes at 10-year follow-up in the absence of systemic therapy: a pooled analysis of 1835 patients [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD9-05.

Background Eribulin or capecitabine monotherapy is the next cytotoxic chemotherapy option for patients with metastatic or recurrent breast cancer who have previously received an anthracycline or a taxane. However, it is unclear what factors can guide the selection of eribulin or capecitabine in this setting, and prognostic factors are needed to guide appropriate treatment selection. The neutrophil-to-lymphocyte ratio (NLR) is a prognostic factor for eribulin-treated patients, although it is unclear whether it is a prognostic factor for capecitabine-treated patients. Therefore, we analysed the ability of the NLR to predict oncological outcomes among patients who received capecitabine after previous anthracycline or taxane treatment for breast cancer. Methods We retrospectively reviewed the medical records of patients with metastatic or recurrent breast cancer who had previously received anthracycline or taxane treatment at the National Cancer Center Hospital between 2007 and 2015. Patients were included if they received eribulin or capecitabine monotherapy as first-line, second-line, or third-line chemotherapy. Analyses of overall survival (OS) and progression-free survival (PFS) were performed according to various factors. Results Between 2007 and 2015, we identified 125 eligible patients, including 46 patients who received only eribulin, 34 patients who received only capecitabine, and 45 patients who received eribulin and capecitabine. The median follow-up period was 19.1 months. Among eribulin-treated patients, an NLR of Conclusions To the best of our knowledge, this is the first study to evaluate whether the NLR is a prognostic factor for capecitabine-treated patients with metastatic or recurrent breast cancer. However, the NLR only independently predicted PFS in this setting, despite it being a useful prognostic factor for other chemotherapies.

Folate receptor α (FRα) is a membrane protein expressed in various solid tumors but has limited expression in normal cells. Therefore, FRα is an attractive target for cancer treatment. This study aimed to investigate the relationship between FRα expression and the clinicopathological characteristics and survivals of cervical cancer.This retrospective study included patients with cervical cancer who underwent primary surgery between 2000 and 2020 at our institution. Immunohistochemical staining of FRα was performed using an anti-folate-binding protein/FBP antibody. FRα-positive staining was defined as ≥5% of tumor staining and FRα-high as ≥50% tumor staining with ≥2+ intensity. The association between FRα expression and survival was assessed using multivariate Cox regression analysis, adjusting for established prognostic factors.Overall, 123 patients were identified, and 140 tumor samples, including 17 paired primary and metastatic samples, were evaluated. As histological types, 67 patients had squamous cell carcinoma (SCC), and 56 patients had non-SCC. All primary tumors were FRα-positive. High FRα expression was observed in 25% of the cases and differed according to histology (SCC vs. non-SCC, 14.9% vs. 37.5%, p=0.004). FRα expression was significantly higher in metastatic tumors than in primary (170 [IQR, 140-205] vs. 125 [IQR, 110-150], p=0.0006). High FRα expression was significantly associated with worse overall survival (hazard ratio, 6.73; 95% confidence interval, 2.21-20.53; p=0.001).In cervical cancer, FRα expression was elevated in metastatic tumors and high expression was associated with a worse prognosis. Our study supports the development of FRα-targeted therapy for advanced cervical cancer.

IMPORTANCE: Patients with cancer and health care workers (HCWs) are at high risk of SARS-CoV-2 infection. Assessing the antibody status of patients with cancer and HCWs can help understand the spread of COVID-19 in cancer care. OBJECTIVE: To evaluate serum SARS-CoV-2 antibody status in patients with cancer and HCWs during the COVID-19 pandemic in Japan. DESIGN, SETTING, AND PARTICIPANTS: Participants were enrolled for this prospective cross-sectional study between August 3 and October 30, 2020, from 2 comprehensive cancer centers in the epidemic area around Tokyo, Japan. Patients with cancer aged 16 years or older and employees were enrolled. Participants with suspected COVID-19 infection at the time of enrollment were excluded. EXPOSURES: Cancer of any type and cancer treatment, including chemotherapy, surgery, immune checkpoint inhibitors, radiotherapy, and targeted molecular therapy. MAIN OUTCOMES AND MEASURES: Seroprevalence and antibody levels in patients with cancer and HCWs. Seropositivity was defined as positivity to nucleocapsid IgG (N-IgG) and/or spike IgG (S-IgG). Serum levels of SARS-CoV-2 IgM and IgG antibodies against the nucleocapsid and spike proteins were measured by chemiluminescent enzyme immunoassay. RESULTS: A total of 500 patients with cancer (median age, 62.5 years [range, 21-88 years]; 265 men [55.4%]) and 1190 HCWs (median age, 40 years [range, 20-70 years]; 382 men [25.4%]) were enrolled. In patients with cancer, 489 (97.8%) had solid tumors, and 355 (71.0%) had received anticancer treatment within 1 month. Among HCWs, 385 (32.3%) were nurses or assistant nurses, 266 (22.4%) were administrative officers, 197 (16.6%) were researchers, 179 (15.0%) were physicians, 113 (9.5%) were technicians, and 50 (4.2%) were pharmacists. The seroprevalence was 1.0% (95% CI, 0.33%-2.32%) in patients and 0.67% (95% CI, 0.29%-1.32%) in HCWs (P = .48). However, the N-IgG and S-IgG antibody levels were significantly lower in patients than in HCWs (N-IgG: β, −0.38; 95% CI, −0.55 to −0.21; P

Background: HER3 (ErbB-3) is a member of the epidermal growth factor receptor family of receptor tyrosine kinases, and its overexpression is associated with poorer prognosis in several cancer types. It is unclear whether HER3 expression status changes in tumor tissue at relapse. The purpose of this study is to evaluate changes in HER3 expression between initial diagnosis and recurrence in gynecologic cancers. Methods: This retrospective study included gynecologic cancer patients with matched paired tissues at the time of initial diagnosis and at the time of recurrence between 1999 and 2019 at National Cancer Center Hospital, Japan. Immunohistochemical (IHC) staining for HER3 was performed using formalin-fixed paraffin-embedded specimens. The primary antibody was HER3/ErbB3 (D22C5) XP Rabbit mAb (Cell Signaling Technology). HER3-positive was defined as an IHC score of 2+ or 3+, and HER3-negative was an IHC score of 0 or 1+, scored according to HER2 testing guidelines for gastroesophageal cancer. The H-score (range, 0-300) was calculated using the following formula: 3X+2Y+Z, where X, Y, and Z are the percentage of tumor cells showing strong, moderate, and weak staining intensity. The difference in HER3 expression between initial diagnosis and recurrence was evaluated using the χ2 test. Results: Eighty-six patients with gynecologic cancers were included (40 ovarian; 32 endometrial; 14 cervical). In ovarian cancer, 67.5% and 80.0% of the patients were HER3-positive at initial diagnosis and recurrence, respectively. There was a statistically significant increase in H-Score at recurrence (p=0.004). The HER3-positive rate in patients with endometrial cancer increased from 46.9% at initial diagnosis to 68.8% at recurrence, and H-Score tended to increase at recurrence (p=0.08). Twelve of the 14 patients (85.7%) with cervical cancer were HER3-positive, both at initial diagnosis and at recurrence, and H-Score tended to increase at recurrence (p=0.19). The discordance rate of HER3 expression determination in samples at initial diagnosis and recurrence was 27.5%, 46.9%, and 14.3% for ovarian, endometrial, and cervical cancers, respectively. Conclusion: Our findings suggest that HER3 expression may increase at recurrence in patients with gynecologic cancers. HER3-targeted therapy may represent a promising option to overcome treatment failure and improve patient outcomes. Citation Format: Yuki Kojima, Kazuki Sudo, Hiroshi Yoshida, Shu Yazaki, Momoko Tokura, Shosuke Kita, Kasumi Yamamoto, Chiharu Mizoguchi, Hitomi S Okuma, Tadaaki Nishikawa, Emi Noguchi, Tatsunori Shimoi, Yasuhito Tanase, Masaya Uno, Mitsuya Ishikawa, Tomoyasu Kato, Kumiko Koyama, Maki Kobayashi, Tomoya Kakegawa, Yasuhiro Fujiwara, Kan Yonemori. Changes in HER3 expression profiles between initial diagnosis and recurrence in gynecologic cancers [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 5083.

Supplemental Digital Content is available in the text, Since 2013, trastuzumab emtansine (T-DM1) has been widely used in Japan to treat patients with human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer (MBC) who were previously administered trastuzumab and a taxane. However, there is no information about the treatment outcomes after exposure to T-DM1 in Japanese patients with HER2-positive MBC. In this study, we aimed to describe the survival outcomes of patients with HER2-positive MBC who received a treatment following T-DM1 and clarify the predictive factors of their prognosis. We retrospectively identified patients with HER2-positive MBC who received T-DM1 between April 1, 2014, and December 31, 2018, at the National Cancer Center Hospital, and focused on the population that received another line of therapy following T-DM1 discontinuation. Thirty patients were available for the outcome analysis. Median progression-free survival (PFS) of the first subsequent therapy was 6.0 months [95% confidence interval (95% CI) 4.1–6.4], whereas the median overall survival (OS) from the first subsequent therapy was 20.6 months (95% CI 13.5 months to not reached). We divided the patients into 2 groups according to their PFS with T-DM1 treatment and compared their PFS with the subsequent therapy. The results revealed a significant difference in the median PFS with the first subsequent treatment between patients with the PFS of less than and more than 3 months [5.1 (95% CI 1.7–6.2) vs 6.2 (95% CI 4.0–11.3) months, P = .03]. This is the first study to evaluate the survival outcomes of post-T-DM1 therapy in Japanese patients with HER2-positive MBC. A short PFS with T-DM1 might affect the PFS with a treatment after T-DM1.

Background: Mesothelin (MSLN) is a cell-surface glycoprotein highly expressed in many cancers, while its expression is limited to normal mesothelial cells. Aberrant MSLN expression is considered to play an essential role in promoting proliferation, cell migration, and invasion. Early phase clinical trial of the antibody-drug conjugate of anti-MSLN-antibody demonstrated patients who responded to treatment had high MSLN expression. Hence, MSLN is an attractive target for cancer therapy. However, the association between MSLN expression and clinicopathologic features in cervical cancer has not been clarified. Methods: This retrospective study included patients diagnosed with cervical cancer who underwent primary surgery between 2000 and 2020. The VENTANA MSLN (SP74) IHC assay was used to assess MSLN expression. The MSLN positive was defined as the tumors expressing MSLN at any intensity. Tumors were considered high MSLN expression if MSLN was detected at a 2+ or 3+ intensity on ≥30% of tumor cells. The association of MSLN expression with overall survival was assessed by multivariate cox regression analysis, adjusting for established prognostic factors. Results: Overall, 123 patients were enrolled, and 140 tumor samples (17 paired primary and metastatic samples) were evaluated. The histopathologic classification was squamous cell carcinoma (Sq)/adenocarcinoma (Ad)/adenosquamous carcinoma (Adsq)/adenocarcinoma, gastric type (Ga)/neuroendocrine carcinoma=67/27/13/12/4. MSLN-positive was found in 95.7% (134/140), and MSLN-high was 48% (36/75) of Sq and 86.5% (45/52) of Ad/Adsq/Ga. An analysis of the Cancer Genome Atlas (TCGA) cervical cancer dataset (N=280) confirmed that a higher level of MSLN mRNA expression in Ad compared to Sq (log fold-change 3.13, p Conclusion: MSLN was highly expressed in cervical adenocarcinoma, including adenosquamous carcinoma and adenocarcinoma, gastric type. Despite the MSLN-high in the primary tumor, MSLN expression was reduced in metastatic tumors in 37.5% of patients. The MSLN may be a potential target for cervical adenocarcinoma, and reassessment of MSLN expression at metastatic sites is desirable to maximize the therapeutic effects. Citation Format: Shu Yazaki, Yuki Kojima, Hiroshi Yoshida, Rui Kitadai, Shigemasa Takamizawa, Tatsunori Shimoi, Tadaaki Nishikawa, Kazuki Sudo, Tomoyasu Kato, Kan Yonemori. Mesothelin expression varies by histological type in patients with cervical cancer and is reduced in metastatic tumors [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 361.

Background: The folate receptor alpha (FRα) is a glycosyl-phosphatidylinositol-anchored membrane protein that binds and internalizes folate for purine and thymidine DNA biosynthesis, repair, and methylation. FRα have a role in cellular migration and invasion, and overexpression of this receptor is associated with tumor progression. FRα is known to be over-expressed in numerous epithelial malignancies and considered a potential target for anticancer treatment. However, the association between FRα expression and clinicopathologic features and prognosis in cervical cancer remains unclear. Methods: This retrospective study included patients diagnosed with cervical cancer who underwent primary surgery between 2000 and 2020. Immunohistochemical staining of FRα was performed using Anti-Folate Binding Protein/FBP antibody (abcam, ab67422). The evaluation of FRα expression was performed by an experienced pathologist. FRα-positive was defined as > 5% tumor staining. FRα-high was defined as ≥ 50% tumor staining with 2+ or 3+ intensity. A possible influence of FRα expression on survival was studied by multivariate Cox regression adjusting for established risk factors for postoperative recurrence. Results: Overall, 123 patients were enrolled, and 136 tumor samples (13 paired primary and metastatic samples) were assessed. The histopathologic classification was squamous cell carcinoma, 67; adenocarcinoma, 27; adenosquamous carcinoma, 13; other, 16. FRα-positive was found in 100% (136/136), and FRα-high was 20.8% (15/72) of squamous cell carcinoma and 33.3% (10/30) of adenocarcinoma. The FRα expression of the metastatic tumor was significantly higher than the primary tumor (H-score, 180 vs. 105, p=0.006). In 69.2% (9/13) of cases with paired samples of primary and metastatic, FRα expression was elevated in metastatic tumors than primary.In a multivariate analysis, FRα-high was associated with poor recurrence-free survival (hazard ratio [HR] 2.30, 95% confidence interval [CI] 1.17-4.52, p=0.016) and overall survival (HR 5.45, 95% CI 1.91-15.5, p=0.002). Conclusion: FRα was widely expressed regardless of histological subtypes. FRα expression was elevated in metastatic tumors, and its expression was independently associated with postoperative recurrence and decreased survival in patients with cervical cancer. FRα may be a potential target for therapy of cervical cancer. Citation Format: Shigemasa Takamizawa, Shu Yazaki, Yuki Kojima, Hiroshi Yoshida, Rui Kitadai, Tatsunori Shimoi, Tadaaki Nishikawa, Kazuki Sudo, Tomoyasu Kato, Kan Yonemori. Folate receptor alpha expression is associated with poor prognosis in patients with cervical cancer [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 657.

e13019 Background: Since trastuzumab emtansine (T-DM1) was established as a standard treatment option, the treatment strategy of metastatic HER2-positive breast cancer has markedly changed. However, clinical evidence regarding the treatments beyond T-DM1 is insufficient. In this study, we attempted to describe real-world selection and efficacy of treatments following T-DM1 for metastatic HER2-positive breast cancer. Methods: This multi-center retrospective cohort study was conducted in 17 hospitals in Japan. Consecutive patients with metastatic HER2-positive breast cancer who had received T-DM1 and started post-T-DM1 treatments between January 2014 and December 2018 were enrolled. Patients treated with investigational drugs were excluded. The primary endpoint was objective response rate (ORR) of post-T-DM1 treatments. Secondary endpoints included disease control rate (DCR), progression-free survival (PFS), time to treatment failure (TTF), and overall survival (OS). Results: In this study, 325 patients were eligible, of which 182 (56.0%) cases were estrogen receptor-positive and 61 (18.8%) had brain metastases. T-DM1 had been used as the first to 11th (median third) line treatment for metastatic disease. The types of post-T-DM1 treatment were as follows: 1) chemotherapy concomitant with trastuzumab and pertuzumab (n = 102; 31.4%), 2) chemotherapy concomitant with trastuzumab only (n = 78; 24.0%), 3) lapatinib with capecitabine (n = 63; 19.4%), and 4) others (n = 82; 25.2%). ORR of post-T-DM1 treatments was 22.8% (95% confidence interval [CI]: 18.1 to 28.0) and DCR was 66.6% (95% CI: 60.8 to 72.0), calculated with 290 eligible cases with the target lesion. Median PFS was 6.1 months (95%CI: 5.3 to 6.7), median TTF was 5.1 months (95%CI: 4.4 to 5.6), and median OS was 23.7 months (95%CI: 20.7 to 27.4). Conclusions: This real-world study showed that post-T-DM1 treatments had modest anti-tumor activity. Development of more effective treatments beyond T-DM1 is needed for metastatic HER2-positive breast cancer. Clinical trial information: UMIN000037747 .

Background: Procalcitonin (PCT) is known as a useful diagnostic marker signifying bacterial infection and its serum levels correlate with disease severity measured by scoring systems such as pneumonia severity index (PSI) or Age, Dehydration, Respiratory failure, Orientation disturbance, Pressure (A-DROP) score, a Japanese prognostic scale for community-acquired pneumonia (CAP). However, for aspiration pneumonia, a major cause of death among elderly patients, little is known about the role of PCT. Objective: We investigated whether PCT levels would be useful for predicting severity and clinical course of aspiration pneumonia. Method: We conducted a prospective study in 49 patients with aspiration pneumonia who were admitted to our hospital from September 2013 and April 2014. Patients with immunosuppression or on treatment with chemotherapy were excluded. We analyzed the association between PCT levels on admission and PSI and A-DROP scores. In addition, correlation between PCT levels and the duration of administration of intravenous antimicrobial agents as an indicator of clinical course was examined. Results: The mean age of patients was 85 years and 25 (51%) were male. PCT levels did not correlate with PSI risk classification (correlation coefficient (R):0.25, p=0.08) nor A-DROP scores (R: 0.20, p=0.17). A multiple regression analysis showed no association between PCT levels on admission and the number of days of intravenous antimicrobial agent administration (regression coefficient (β): 1.59, 95%CI -0.8 to 4.0, p=0.19). Conclusions: PCT levels were not useful for predicting severity and clinical course of aspiration pneumonia, in contrast to previous reports of CAP.