PP2A B55α Inhibits Epithelial-Mesenchymal Transition Via Regulation of Slug Expression in Non-Small Cell Lung Cancer.

PP2A B55α, encoded by PPP2R2A, acts as a regulatory subunit of the serine/threonine phosphatase PP2A. Despite a frequent loss of heterozygosity of PPP2R2A in cases of non-small cell lung cancer (NSCLC), research on PP2A B55α's functions remains limited and controversial. To investigate the biological roles of PP2A B55α, we conducted bulk RNA-sequencing to assess the impact of PPP2R2A knockdown using two shRNAs in a NSCLC cell line. Gene set enrichment analysis (GSEA) of the RNA-sequencing data revealed significant enrichment of the epithelial-mesenchymal transition (EMT) pathway, with SNAI2 (the gene encoding Slug) emerging as one of the top candidates. Our findings demonstrate that PP2A B55α suppresses EMT, as PPP2R2A deficiency through knockdown or homozygous or hemizygous depletion promotes EMT and metastatic behavior in NSCLC cells, as evidenced by changes in EMT biomarkers, invasion and migration abilities, as well as metastasis in a tail vein assay. Mechanistically, PP2A B55α inhibits EMT by downregulating SNAI2 expression via the GSK3ß-ß-catenin pathway. Importantly, PPP2R2A deficiency also slows cell proliferation by disrupting DNA replication, particularly in PPP2R2A-/- cells. Furthermore, PPP2R2A deficiency, especially PPP2R2A-/- cells, leads to an increase in the cancer stem cell population, which correlates with enhanced resistance to chemotherapy. Overall, the decrease in PP2A B55α levels due to hemizygous/homozygous depletion heightens EMT and the metastatic or stemness/drug resistance potential of NSCLC cells despite their proliferation disadvantage. Our study highlights the significance of PP2A B55α in EMT and metastasis and suggests that targeting EMT/stemness could be a potential therapeutic strategy for treating PPP2R2A-deficient NSCLC.

Fumarate Hydratase-Deficient Abdominal Wall Leiomyoma Presenting a Decade Post-Fumarate Hydratase-Deficient Uterine Leiomyoma Excision: An Incidental or Syndromic Association?

Background. The occurrence of fumarate hydratase-deficient leiomyoma of the abdominal wall is exceptionally rare. Case Presentation. A 50-year-old female patient with a past medical history of fumarate hydratase-deficient uterine leiomyoma presented with a left lower quadrant abdominal mass that has been present for the past 2 years. An ultrasound revealed a 3.5 cm oval hypoechoic mass. A subsequent CT scan showed a 3.5 cm hyperdense mass within the left internal oblique musculature. No family history is noted. A biopsy of the mass exhibited bundles of spindle cell neoplasm exhibiting bizarre ovoid nuclei and eosinophilic cytoplasm. No evidence of mitotic figures or tumor necrosis was noted. Immunohistochemical staining showed positive staining for desmin and smooth muscle actin (SMA), but negative staining for MART-1, S100, and CD34. Lesional cells showed expression of 2-succinocysteine and loss of fumarate hydratase expression. A diagnosis of fumarate hydratase-deficient leiomyoma was rendered. Conclusion. This report reinforces the importance of considering genetic testing for fumarate hydratase mutations in the evaluation of extra-uterine leiomyomatous lesions. Comprehensive follow-up and clinical screening in individuals with new lesions and a known history of fumarate hydratase-deficient neoplasms is mandatory. Recent recommendations support the integration of morphology-based evaluation along with immunohistochemical staining and genetic testing as a part of the standard evaluation for all uterine leiomyomas.

The current state of digital cytology and artificial intelligence (AI): global survey results from the American Society of Cytopathology Digital Cytology Task Force.

INTRODUCTION: The integration of whole slide imaging (WSI) and artificial intelligence (AI) with digital cytology has been growing gradually. Therefore, there is a need to evaluate the current state of digital cytology. This study aimed to determine the current landscape of digital cytology via a survey conducted as part of the American Society of Cytopathology (ASC) Digital Cytology White Paper Task Force. MATERIALS AND METHODS: A survey with 43 questions pertaining to the current practices and experiences of WSI and AI in both surgical pathology and cytology was created. The survey was sent to members of the ASC, the International Academy of Cytology (IAC), and the Papanicolaou Society of Cytopathology (PSC). Responses were recorded and analyzed. RESULTS: In total, 327 individuals participated in the survey, spanning a diverse array of practice settings, roles, and experiences around the globe. The majority of responses indicated there was routine scanning of surgical pathology slides (n = 134; 61%) with fewer respondents scanning cytology slides (n = 150; 46%). The primary challenge for surgical WSI is the need for faster scanning and cost minimization, whereas image quality is the top issue for cytology WSI. AI tools are not widely utilized, with only 16% of participants using AI for surgical pathology samples and 13% for cytology practice. CONCLUSIONS: Utilization of digital pathology is limited in cytology laboratories as compared to surgical pathology. However, as more laboratories are willing to implement digital cytology in the near future, the establishment of practical clinical guidelines is needed.

Performance Analysis of Leica Biosystems Monoclonal Antibody Programmed Cell Death Ligand 1 Clone 73-10 on Breast, Colorectal, and Hepatocellular Carcinomas.

Programmed cell death receptor 1/Programmed cell death ligand 1 (PD-L1) checkpoint pathway is responsible for the control of immune cell responses. Immunotherapy using checkpoint inhibitors, such as anti-PD-L1 therapy, aids disease management and potentiates clinical outcomes. This study aimed to analyze the performance of the Leica Biosystems (LBS) USA FDA class I in vitro diagnostic monoclonal antibody (clone 73-10) to detect PD-L1 expression in breast, colorectal, and hepatocellular carcinomas compared with the class III FDA-approved PD-L1 detecting antibodies [SP263 (Ventana), 22C3 (Dako), and 28-8 (Dako)] using 208 unique tissue microarray-based cases for each tumor type. The interassay concordances between LBS 73-10 clone and other PD-L1 antibodies ranged from 0.59 to 0.95 Cohen kappa coefficient (K) and from 0.66 to 0.90 (K) for cutoff values of 1% and 50% tumor proportion score (TPS), respectively. The 73-10 clones showed inter-pathologist agreements ranging from 0.53 to 1.0 (K) and 0.34 to 0.94 (K) for cutoff values of 1% and 50% TPS, respectively. For the immune cell proportion score (IPS) using a cutoff of 1%, the Kappa coefficient of interassay concordances and inter-pathologist agreements ranged from 0.34 to 0.94. The 73-10 clone assay's sensitivity ranged from 78.3% to 100% (TPS ≥1%), 100% (TPS ≥50%), and 77.4% to 93.5% (IPS ≥1%), while its specificity was 97.9% to 100% (TPS ≥1%), 99.5% to 99.8% (TPS ≥50%), and 97.9% to 100% (IPS ≥1%). This exploratory evaluation of LBS 73-10 monoclonal antibody on a large set of breast, colorectal, and hepatocellular carcinomas showed the assay's technical performance is comparable to the FDA-approved companion/complementary diagnostics PD-L1 detection assays.

The thioredoxin system determines CHK1 inhibitor sensitivity via redox-mediated regulation of ribonucleotide reductase activity.

Checkpoint kinase 1 (CHK1) is critical for cell survival under replication stress (RS). CHK1 inhibitors (CHK1i's) in combination with chemotherapy have shown promising results in preclinical studies but have displayed minimal efficacy with substantial toxicity in clinical trials. To explore combinatorial strategies that can overcome these limitations, we perform an unbiased high-throughput screen in a non-small cell lung cancer (NSCLC) cell line and identify thioredoxin1 (Trx1), a major component of the mammalian antioxidant-system, as a determinant of CHK1i sensitivity. We establish a role for redox recycling of RRM1, the larger subunit of ribonucleotide reductase (RNR), and a depletion of the deoxynucleotide pool in this Trx1-mediated CHK1i sensitivity. Further, the TrxR inhibitor auranofin, an approved anti-rheumatoid arthritis drug, shows a synergistic interaction with CHK1i via interruption of the deoxynucleotide pool. Together, we show a pharmacological combination to treat NSCLC that relies on a redox regulatory link between the Trx system and mammalian RNR activity.

Breast Cancer With HER2 Immunohistochemical Score 2 and Average HER2 Signals/Cell 6 or More and HER2/CEP17 Ratio Less Than 2 ('ISH Group 3'): A Multiinstitutional Cohort Analysis Emphasizing Outcome and Molecular Subtype.

Breast cancer (BC) with average human epidermal growth factor receptor 2 (HER2) signals/cell ≥6 and HER2/chromosome enumeration probe 17 (CEP17) ratio <2 (in situ hybridization [ISH] group 3) is very rare, accounting for 0.4% to 3.0% of cases sent for the dual-probe ISH assay. Although such patients are currently eligible for treatment with HER2-targeted therapy, their characteristics and outcomes remain poorly understood. Sixty-two BCs with equivocal HER2 immunohistochemical score (2+) and reflex ISH group 3 results were identified across 4 institutions. Available clinicopathologic characteristics, MammaPrint and BluePrint molecular results, and follow-up information were retrospectively analyzed. Most BCs with HER2 equivocal immunohistochemical and ISH group 3 results were histologic grade 2 or 3 (100%), estrogen receptor (ER) positive (90.3%), with an average HER2 signals/cell of 7.3. Molecular profiles revealed that 80% (16/20) of tumors were luminal subtypes, and HER2 molecular subtype was identified in 10% of tumors (2/20). Twelve (19.4%) out of 62 patients developed local recurrence and/or distant metastasis with a median follow-up of 50 months. One (10%) of 10 patients achieved pathologic complete response after neoadjuvant chemotherapy. Forty-nine (79%) out of 62 patients completed anti-HER2 agents, and exploratory analysis showed no statistically significant difference in disease outcomes between patients who completed anti-HER2 treatment and those who did not. Univariate analysis revealed advanced clinical stage, and ER/progesterone receptor negativity was associated with unfavorable disease outcomes, and exploratory multivariate analysis demonstrated that clinical stage was the most significant factor associated with disease outcomes in the studied population. These findings increase our understanding of this rare, but clinically important HER2 category. Large-scale prospective randomized studies are needed to further evaluate the role of perioperative HER2-targeted therapy in this patient population.

Automated imaging analysis of Ki-67 immunohistochemistry on whole slide images of cell blocks from pancreatic neuroendocrine neoplasms.

INTRODUCTION: Accurate grading of pancreatic neuroendocrine tumors (PanNETs) relies on the assessment of Ki-67 immunohistochemistry (IHC). While digital imaging analysis (DIA) has been employed for Ki-67 IHC assessment in surgical specimens, its applicability to cytologic specimens remains underexplored. This study aimed to evaluate an automated DIA for assessing Ki-67 IHC on PanNET cell blocks. MATERIALS AND METHODS: The study included 61 consecutive PanNETs and 5 pancreatic neuroendocrine carcinomas. Ki-67 IHC slides from cell blocks were digitally scanned into whole slide images using Philips IntelliSite Scanners and analyzed in batches using the Visiopharm Ki-67 App in a digital workflow. Ki-67 scores obtained through DIA were compared to pathologists' manual scores. RESULTS: The Pearson correlation coefficient of the percentage of Ki-67-stained nuclei between DIA reads and the originally reported reads was 0.9681. Concordance between DIA Ki-67 grades and pathologists' Ki-67 grades was observed in 92.4% (61/66) of cases with the calculated Cohen's Kappa coefficient of 0.862 (almost perfect agreement). Discordance between DIA and pathologists' consensus reads occurred in 5 PanNET cases which were upgraded from G1 to G2 by DIA due to contaminated Ki-67-stained inflammatory cells. CONCLUSIONS: DIA demonstrated excellent concordance with pathologists' assessments, with only minor grading discrepancies. However, the essential role of pathologists in confirming results is emphasized to enhance overall accuracy.

Artificial intelligence's impact on breast cancer pathology: a literature review.

This review discusses the profound impact of artificial intelligence (AI) on breast cancer (BC) diagnosis and management within the field of pathology. It examines the various applications of AI across diverse aspects of BC pathology, highlighting key findings from multiple studies. Integrating AI into routine pathology practice stands to improve diagnostic accuracy, thereby contributing to reducing avoidable errors. Additionally, AI has excelled in identifying invasive breast tumors and lymph node metastasis through its capacity to process large whole-slide images adeptly. Adaptive sampling techniques and powerful convolutional neural networks mark these achievements. The evaluation of hormonal status, which is imperative for BC treatment choices, has also been enhanced by AI quantitative analysis, aiding interobserver concordance and reliability. Breast cancer grading and mitotic count evaluation also benefit from AI intervention. AI-based frameworks effectively classify breast carcinomas, even for moderately graded cases that traditional methods struggle with. Moreover, AI-assisted mitotic figures quantification surpasses manual counting in precision and sensitivity, fostering improved prognosis. The assessment of tumor-infiltrating lymphocytes in triple-negative breast cancer using AI yields insights into patient survival prognosis. Furthermore, AI-powered predictions of neoadjuvant chemotherapy response demonstrate potential for streamlining treatment strategies. Addressing limitations, such as preanalytical variables, annotation demands, and differentiation challenges, is pivotal for realizing AI's full potential in BC pathology. Despite the existing hurdles, AI's multifaceted contributions to BC pathology hold great promise, providing enhanced accuracy, efficiency, and standardization. Continued research and innovation are crucial for overcoming obstacles and fully harnessing AI's transformative capabilities in breast cancer diagnosis and assessment.

Comparing Accuracy of Helicobacter pylori Identification Using Traditional Hematoxylin and Eosin-Stained Glass Slides With Digital Whole Slide Imaging.

With advancements in the field of digital pathology, there has been a growing need to compare the diagnostic abilities of pathologists using digitized whole slide images against those when using traditional hematoxylin and eosin (H&E)-stained glass slides for primary diagnosis. One of the most common specimens received in pathology practices is an endoscopic gastric biopsy with a request to rule out Helicobacter pylori (H. pylori) infection. The current standard of care is the identification of the organisms on H&E-stained slides. Immunohistochemical or histochemical stains are used selectively. However, due to their small size (2-4 µm in length by 0.5-1 µm in width), visualization of the organisms can present a diagnostic challenge. The goal of the study was to compare the ability of pathologists to identify H. pylori on H&E slides using a digital platform against the gold standard of H&E glass slides using routine light microscopy. Diagnostic accuracy rates using glass slides vs digital slides were 81% vs 72% (P = .0142) based on H&E slides alone. When H. pylori immunohistochemical slides were provided, the diagnostic accuracy was significantly improved to comparable rates (96% glass vs 99% digital, P = 0.2199). Furthermore, differences in practice settings (academic/subspecialized vs community/general) and the duration of sign-out experience did not significantly impact the accuracy of detecting H. pylori on digital slides. We concluded that digital whole slide images, although amenable in different practice settings and teaching environments, does present some shortcomings in accuracy and precision, especially in certain circumstances and thus is not yet fully capable of completely replacing glass slide review for identification of H. pylori. We specifically recommend reviewing glass slides and/or performing ancillary stains, especially when there is a discrepancy between the degree of inflammation and the presence of microorganisms on digital images.

Digital cytology part 1: digital cytology implementation for practice: a concept paper with review and recommendations from the American Society of Cytopathology Digital Cytology Task Force.

Digital cytology and artificial intelligence (AI) are gaining greater adoption in the cytopathology laboratory. However, peer-reviewed real-world data and literature are lacking regarding the current clinical landscape. The American Society of Cytopathology in conjunction with the International Academy of Cytology and the Digital Pathology Association established a special task force comprising 20 members with expertise and/or interest in digital cytology. The aim of the group was to investigate the feasibility of incorporating digital cytology, specifically cytology whole slide scanning and AI applications, into the workflow of the laboratory. In turn, the impact on cytopathologists, cytologists (cytotechnologists), and cytology departments were also assessed. The task force reviewed existing literature on digital cytology, conducted a worldwide survey, and held a virtual roundtable discussion on digital cytology and AI with multiple industry corporate representatives. This white paper, presented in 2 parts, summarizes the current state of digital cytology and AI practice in global cytology practice. Part 1 of the white paper presented herein is a review and offers best practice recommendations for incorporating digital cytology into practice. Part 2 of the white paper provides a comprehensive review of AI in cytology practice along with best practice recommendations and legal considerations. Additionally, the results of a global survey regarding digital cytology are highlighted.

Digital cytology part 2: artificial intelligence in cytology: a concept paper with review and recommendations from the American Society of Cytopathology Digital Cytology Task Force.

Digital cytology and artificial intelligence (AI) are gaining greater adoption in the cytology laboratory. However, peer-reviewed real-world data and literature are lacking in regard to the current clinical landscape. The American Society of Cytopathology in conjunction with the International Academy of Cytology and the Digital Pathology Association established a special task force comprising 20 members with expertise and/or interest in digital cytology. The aim of the group was to investigate the feasibility of incorporating digital cytology, specifically cytology whole slide scanning and AI applications, into the workflow of the laboratory. In turn, the impact on cytopathologists, cytologists (cytotechnologists), and cytology departments were also assessed. The task force reviewed existing literature on digital cytology, conducted a worldwide survey, and held a virtual roundtable discussion on digital cytology and AI with multiple industry corporate representatives. This white paper, presented in 2 parts, summarizes the current state of digital cytology and AI practice in global cytology practice. Part 1 of the white paper is presented as a separate paper which details a review and best practice recommendations for incorporating digital cytology into practice. Part 2 of the white paper presented here provides a comprehensive review of AI in cytology practice along with best practice recommendations and legal considerations. Additionally, the cytology global survey results highlighting current AI practices by various laboratories, as well as current attitudes, are reported.

Computational pathology in the identification of HER2-low breast cancer: Opportunities and challenges.

For the past 2 decades, pathologists have been accustomed to reporting the HER2 status of breast cancer as either positive or negative, based on HER2 IHC. Today, however, there is a clinical imperative to employ a 3-tier approach to interpreting HER2 IHC that can also identify tumours categorised as HER2-low. Meeting this need for a finer degree of discrimination may be challenging, and in this article, we consider the potential for the integration of computational approaches to support pathologists in achieving accurate and reproducible HER2 IHC scoring as well as outlining some of the practicalities involved.

Comparison of four different displays for identification of select pathologic features extracted from whole slide images of surgical pathology cases.

BACKGROUND: The establishment of minimum standards for display selection for the whole slide image (WSI) interpretation has not been fully defined. Recently, pathologists have increasingly preferred using remote displays for clinical diagnostics. Our study aims to assess and compare the performance of three fixed work displays and one remote personal display in accurately identifying ten selected pathologic features integrated into WSIs. DESIGN: Hematoxylin and eosin-stained glass slides were digitized using Philips scanners. Seven practicing pathologists and three residents reviewed ninety WSIs to identify ten pathologic features using the LG, Dell, and Samsung and an optional consumer-grade display. Ten pathologic features included eosinophils, neutrophils, plasma cells, granulomas, necrosis, mucin, hemosiderin, crystals, nucleoli, and mitoses. RESULTS: The accuracy of the identification of ten features on different types of displays did not significantly differ among the three types of "fixed" workplace displays. The highest accuracy was observed for the identification of neutrophils, eosinophils, plasma cells, granuloma, and mucin. On the other hand, a lower accuracy was observed for the identification of crystals, mitoses, necrosis, hemosiderin, and nucleoli. Participant pathologists and residents preferred the use of larger displays (>30″) with a higher pixel count, resolution, and luminance. CONCLUSION: Most features can be identified using any display. However, certain features posed more challenges across the three fixed display types. Furthermore, the use of a remote personal consumer-grade display chosen according to the pathologists' preference showed similar feature identification accuracy. Several factors of display characteristics seemed to influence pathologists' display preferences such as the display size, color, contrast ratio, pixel count, and luminance calibration. This study supports the use of standard "unlocked" vendor-agnostic displays for clinical digital pathology workflow rather than purchasing "locked" and more expensive displays that are part of a digital pathology system.

Unlocking Translational Potential: Conditionally Reprogrammed Cells in Advancing Breast Cancer Research.

Preclinical in vitro models play an important role in studying cancer cell biology and facilitating translational research, especially in the identification of drug targets and drug discovery studies. This is particularly relevant in breast cancer, where the global burden of disease is quite high based on prevalence and a relatively high rate of lethality. Predictive tools to select patients who will be responsive to invasive or morbid therapies (radiotherapy, chemotherapy, immunotherapy, and/or surgery) are relatively lacking. To be clinically relevant, a model must accurately replicate the biology and cellular heterogeneity of the primary tumor. Addressing these requirements and overcoming the limitations of most existing cancer cell lines, which are typically derived from a single clone, we have recently developed conditional reprogramming (CR) technology. The CR technology refers to a co-culture system of primary human normal or tumor cells with irradiated murine fibroblasts in the presence of a Rho-associated kinase inhibitor to allow the primary cells to acquire stem cell properties and the ability to proliferate indefinitely in vitro without any exogenous gene or viral transfection. This innovative approach fulfills many of these needs and offers an alternative that surpasses the deficiencies associated with traditional cancer cell lines. These CR cells (CRCs) can be reprogrammed to maintain a highly proliferative state and reproduce the genomic and histological characteristics of the parental tissue. Therefore, CR technology may be a clinically relevant model to test and predict drug sensitivity, conduct gene profile analysis and xenograft research, and undertake personalized medicine. This review discusses studies that have applied CR technology to conduct breast cancer research.

Pancreaticobiliary Cytology Practice in 2021: Results of a College of American Pathologists Survey.

CONTEXT.­: The College of American Pathologists (CAP) surveys provide national benchmarks of pathology practice. OBJECTIVE.­: To investigate pancreaticobiliary cytology practice in domestic and international laboratories in 2021. DESIGN.­: We analyzed data from the CAP Pancreaticobiliary Cytology Practice Supplemental Questionnaire that was distributed to laboratories participating in the 2021 CAP Nongynecologic Cytopathology Education Program. RESULTS.­: Ninety-three percent (567 of 612) of respondent laboratories routinely evaluated pancreaticobiliary cytology specimens. Biliary brushing (85%) was the most common pancreaticobiliary cytology specimen evaluated, followed by pancreatic fine-needle aspiration (79%). The most used sampling methods reported by 235 laboratories were 22-gauge needle for fine-needle aspiration (62%) and SharkCore needle for fine-needle biopsy (27%). Cell block was the most used slide preparation method (76%), followed by liquid-based cytology (59%) for pancreatic cystic lesions. Up to 95% (303 of 320) of laboratories performed rapid on-site evaluation (ROSE) on pancreatic solid lesions, while 56% (180 of 320) performed ROSE for cystic lesions. Thirty-six percent (193 of 530) of laboratories used the Papanicolaou Society of Cytopathology System for Reporting Pancreaticobiliary Cytology in 2021. Among all institution types, significant differences in specimen volume, specimen type, ROSE practice, and case sign-out were identified. Additionally, significant differences in specimen type, slide preparation, and ROSE practice were found. CONCLUSIONS.­: This is the first survey from the CAP to investigate pancreaticobiliary cytology practice. The findings reveal significant differences among institution types and between domestic and international laboratories. These data provide a baseline for future studies in a variety of practice settings.

HER2 Intratumoral Heterogeneity in Breast Cancer, an Evolving Concept.

Amplification and/or overexpression of human epidermal growth factor receptor 2 (HER2) in breast cancer is associated with an adverse prognosis. The introduction of anti-HER2 targeted therapy has dramatically improved the clinical outcomes of patients with HER2-positive breast cancer. Unfortunately, a significant number of patients eventually relapse and develop distant metastasis. HER2 intratumoral heterogeneity (ITH) has been reported to be associated with poor prognosis in patients with anti-HER2 targeted therapies and was proposed to be a potential mechanism for anti-HER2 resistance. In this review, we described the current definition, common types of HER2 ITH in breast cancer, the challenge in interpretation of HER2 status in cases showing ITH and the clinical applications of anti-HER2 agents in breast cancer showing heterogeneous HER2 expression. Digital image analysis has emerged as an objective and reproducible scoring method and its role in the assessment of HER2 status with ITH remains to be demonstrated.

Triple-negative Breast Carcinoma With Apocrine and Histiocytoid Features: A Clinicopathologic and Molecular Study of 18 Cases.

Triple-negative breast cancer (TNBC) is a heterogenous group of tumors. Most TNBCs are high-grade aggressive tumors, but a minority of TNBCs are not high grade, with relatively indolent behavior and specific morphologic and molecular features. We performed a clinicopathologic and molecular assessment of 18 non-high-grade TNBCs with apocrine and/or histiocytoid features. All were grade I or II with low Ki-67 (≤20%). Thirteen (72%) showed apocrine features, and 5 (28%) showed histiocytoid and lobular features. In all, 17/18 expressed the androgen receptor, and 13/13 expressed gross cystic disease fluid protein 15. Four (22.2%) patients were treated with neoadjuvant chemotherapy, but none achieved a pathologic complete response. In all, 2/18 patients (11%) had lymph node metastasis at the time of surgery. None of the cases had a recurrence or disease-specific death, with an average follow-up time of 38 months. Thirteen cases were profiled by targeted capture-based next-generation DNA sequencing. Genomic alterations (GAs) were most significant for PI3K-PKB/Akt pathway (69%) genes, including PIK3R1 (23%), PIK3CA (38%), and PTEN (23%), and RTK-RAS pathway (62%) including FGFR4 (46%) and ERBB2 (15%). TP53 GA was seen in only 31% of patients. Our findings support those on high-grade TNBCs with apocrine and/or histiocytoid features as a clinicopathologic and genetically distinct subgroup of TNBC. They can be defined by features including tubule formation, rare mitosis, low Ki-67 (≤20%), triple-negative status, expression of androgen receptor and/or gross cystic disease fluid protein 15, and GA in the PI3K-PKB/Akt and/or RTK-RAS pathway. These tumors are not sensitive to chemotherapy but have favorable clinical behavior. Tumor subtype definitions are the first step to implementing future trial designs to select these patients.

The thioredoxin system determines CHK1 inhibitor sensitivity via redox-mediated regulation of ribonucleotide reductase activity.

Checkpoint kinase 1 (CHK1) is critical for cell survival under replication stress (RS). CHK1 inhibitors (CHK1i's) in combination with chemotherapy have shown promising results in preclinical studies but minimal efficacy with substantial toxicity in clinical trials. To explore novel combinational strategies that can overcome these limitations, we performed an unbiased high-throughput screen in a non-small cell lung cancer (NSCLC) cell line and identified thioredoxin1 (Trx1), a major component of the mammalian antioxidant-system, as a novel determinant of CHK1i sensitivity. We established a role for redox recycling of RRM1, the larger subunit of ribonucleotide reductase (RNR), and a depletion of the deoxynucleotide pool in this Trx1-mediated CHK1i sensitivity. Further, the TrxR1 inhibitor auronafin, an anti-rheumatoid arthritis drug, shows a synergistic interaction with CHK1i via interruption of the deoxynucleotide pool. Together, these findings identify a new pharmacological combination to treat NSCLC that relies on a redox regulatory link between the Trx system and mammalian RNR activity.