Androgen Receptor Splice Variant 7 in Asian Patients With Metastatic Castration-Resistant Prostate Cancer.

PURPOSE: Androgen receptor splice variant 7 (ARV-7) is a resistance mechanism to hormonal therapy in metastatic castrate-resistant prostate cancer (mCRPC). It has been associated with poor outcomes. On progression to castrate resistance, ARV-7 positivity has been identified in global populations at an incidence of 17.8%-28.8%. Here, we characterize the incidence of ARV-7 positivity in Asian patients with mCRPC in a prospective fashion and evaluate its implications on treatment outcomes. METHODS: Patients with mCRPC from multiple centers in Southeast and East Asia were enrolled in a prospective manner before initiation of androgen receptor signaling inhibitors or docetaxel. ARV-7 status was evaluated at baseline with three commercially available assays: AdnaTest Prostate Cancer platform, Clearbridge method, and IBN method. Clinical outcomes at progression were assessed. The primary end point of this study was prevalence of ARV-7 positivity; secondary end points were incidence of ARV-7 positivity, prostate specific antigen (PSA) response rate, PSA progression-free survival (PFS), and overall survival (OS). RESULTS: A total of 102 patients with a median age of 72 years at enrollment participated. Overall, an incidence of ARV-7 positivity of between 14.3% and 33.7% in Asian patients with mCRPC was demonstrated depending on the assay used. Patients found to have ARV-7 positivity at enrollment had a numerically worse PSA PFS compared with ARV-7 negative patients. CONCLUSION: In this study, the incidence of ARV-7 positivity in Asian patients with mCRPC was shown to be similar to the global population. Patients with ARV-7 positivity appear to have more aggressive disease with numerically worse PSA PFS and OS. Further prospective studies are needed to fully characterize the relationship that ARV-7 positivity has on prognosis of Asian patients with mCRPC.

Machine learning unveils an immune-related DNA methylation profile in germline DNA from breast cancer patients.

BACKGROUND: There is an unmet need for precise biomarkers for early non-invasive breast cancer detection. Here, we aimed to identify blood-based DNA methylation biomarkers that are associated with breast cancer. METHODS: DNA methylation profiling was performed for 524 Asian Chinese individuals, comprising 256 breast cancer patients and 268 age-matched healthy controls, using the Infinium MethylationEPIC array. Feature selection was applied to 649,688 CpG sites in the training set. Predictive models were built by training three machine learning models, with performance evaluated on an independent test set. Enrichment analysis to identify transcription factors binding to regions associated with the selected CpG sites and pathway analysis for genes located nearby were conducted. RESULTS: A methylation profile comprising 51 CpGs was identified that effectively distinguishes breast cancer patients from healthy controls achieving an AUC of 0.823 on an independent test set. Notably, it outperformed all four previously reported breast cancer-associated methylation profiles. Enrichment analysis revealed enrichment of genomic loci associated with the binding of immune modulating AP-1 transcription factors, while pathway analysis of nearby genes showed an overrepresentation of immune-related pathways. CONCLUSION: This study has identified a breast cancer-associated methylation profile that is immune-related to potential for early cancer detection.

Landscape of germline pathogenic variants in patients with dual primary breast and lung cancer.

BACKGROUND: Cancer predisposition is most often studied in the context of single cancers. However, inherited cancer predispositions can also give rise to multiple primary cancers. Yet, there is a paucity of studies on genetic predisposition in multiple primary cancers, especially those outside of well-defined cancer predisposition syndromes. This study aimed to identify germline variants associated with dual primary cancers of the breast and lung. METHODS: Exome sequencing was performed on germline DNA from 55 Singapore patients (52 [95%] never-smokers) with dual primaries in the breast and lung, confirmed by histopathology. Using two large control cohorts: the local SG10K_Health (n = 9770) and gnomAD non-cancer East Asians (n = 9626); and two additional local case cohorts of early-onset or familial breast cancer (n = 290), and lung cancer (n = 209), variants were assessed for pathogenicity in accordance with ACMG/AMP guidelines. In particular, comparisons were made with known pathogenic or likely pathogenic variants in the ClinVar database, pathogenicity predictions were obtained from in silico prediction software, and case-control association analyses were performed. RESULTS: Altogether, we identified 19 pathogenic or likely pathogenic variants from 16 genes, detected in 17 of 55 (31%) patients. Six of the 19 variants were identified using ClinVar, while 13 variants were classified pathogenic or likely pathogenic using ACMG/AMP guidelines. The 16 genes include well-known cancer predisposition genes such as BRCA2, TP53, and RAD51D; but also lesser known cancer genes EXT2, WWOX, GATA2, and GPC3. Most of these genes are involved in DNA damage repair, reaffirming the role of impaired DNA repair mechanisms in the development of multiple malignancies. These variants warrant further investigations in additional populations. CONCLUSIONS: We have identified both known and novel variants significantly enriched in patients with primary breast and lung malignancies, expanding the body of known cancer predisposition variants for both breast and lung cancer. These variants are mostly from genes involved in DNA repair, affirming the role of impaired DNA repair in the predisposition and development of multiple cancers.

An approach to genetic testing in patients with metastatic castration-resistant prostate cancer in Singapore.

Introduction: There has been a rapid evolution in the treatment strategies for metastatic castration-resistant prostate cancer (mCRPC) following the identification of targetable mutations, making genetic testing essential for patient selection. Although several international guidelines recommend genetic testing for patients with mCRPC, there is a lack of locally endorsed clinical practice guidelines in Singapore. Method: A multidisciplinary specialist panel with representation from medical and radiation oncology, urology, pathology, interventional radiology, and medical genetics discussed the challenges associated with patient selection, genetic counselling and sample processing in mCRPC. Results: A clinical model for incorporating genetic testing into routine clinical practice in Singapore was formulated. Tumour testing with an assay that is able to detect both somatic and germline mutations should be utilised. The panel also recommended the "mainstreaming" approach for genetic counselling in which pre-test counselling is conducted by the managing clinician and post-test discussion with a genetic counsellor, to alleviate the bottlenecks at genetic counselling stage in Singapore. The need for training of clinicians to provide pre-test genetic counselling and educating the laboratory personnel for appropriate sample processing that facilitates downstream genetic testing was recognised. Molecular tumour boards and multidisciplinary discussions are recommended to guide therapeutic decisions in mCRPC. The panel also highlighted the issue of reimbursement for genetic testing to reduce patient-borne costs and increase the reach of genetic testing among this patient population. Conclusion: This article aims to provide strategic and implementable recommendations to overcome the challenges in genetic testing for patients with mCRPC in Singapore.

Bone marrow-targetable Green Tea Catechin-Based Micellar Nanocomplex for synergistic therapy of Acute myeloid leukemia.

BACKGROUND: Currently available anti-leukemia drugs have shown limited success in the treatment of acute myeloid leukemia (AML) due to their poor access to bone marrow niche supporting leukemic cell proliferation. RESULTS: Herein, we report a bone marrow-targetable green tea catechin-based micellar nanocomplex for synergistic AML therapy. The nanocomplex was found to synergistically amplify the anti-leukemic potency of sorafenib via selective disruption of pro-survival mTOR signaling. In vivo biodistribution study demonstrated about 11-fold greater bone marrow accumulation of the nanocomplex compared to free sorafenib. In AML patient-derived xenograft (AML-PDX) mouse model, administration of the nanocomplex effectively eradicated bone marrow-residing leukemic blasts and improved survival rates without noticeable off-target toxicity. CONCLUSION: This study may provide insights into the rational design of nanomedicine platforms enabling bone marrow-targeted delivery of therapeutic agents for the treatment of AML and other bone marrow diseases.

Whole-exome sequencing of BRCA-negative breast cancer patients and case-control analyses identify variants associated with breast cancer susceptibility.

BACKGROUND: For the majority of individuals with early-onset or familial breast cancer referred for genetic testing, the genetic basis of their familial breast cancer remains unexplained. To identify novel germline variants associated with breast cancer predisposition, whole-exome sequencing (WES) was performed. METHODS: WES on 290 BRCA1/BRCA2-negative Singaporeans with early-onset breast cancer and/or a family history of breast cancer was done. Case-control analysis against the East-Asian subpopulation (EAS) from the Genome Aggregation Database (gnomAD) identified variants enriched in cases, which were further selected by occurrence in cancer gene databases. Variants were further evaluated in repeated case-control analyses using a second case cohort from the database of Genotypes and Phenotypes (dbGaP) comprising 466 early-onset breast cancer patients from the United States, and a Singapore SG10K_Health control cohort. RESULTS: Forty-nine breast cancer-associated germline pathogenic variants in 37 genes were identified in Singapore cases versus gnomAD (EAS). Compared against SG10K_Health controls, 13 of 49 variants remain significantly enriched (False Discovery Rate (FDR)-adjusted p < 0.05). Comparing these 49 variants in dbGaP cases against gnomAD (EAS) and SG10K_Health controls revealed 23 concordant variants that were significantly enriched (FDR-adjusted p < 0.05). Fourteen variants were consistently enriched in breast cancer cases across all comparisons (FDR-adjusted p < 0.05). Seven variants in GPRIN2, NRG1, MYO5A, CLIP1, CUX1, GNAS and MGA were confirmed by Sanger sequencing. CONCLUSIONS: In conclusion, we have identified pathogenic variants in genes associated with breast cancer predisposition. Importantly, many of these variants were significant in a second case cohort from dbGaP, suggesting that the strategy of using case-control analysis to select variants could potentially be utilized for identifying variants associated with cancer susceptibility.

A comprehensive next generation sequencing tissue assay for Asian-prevalent cancers-Analytical validation and performance evaluation with clinical samples.

Introduction: A well-validated diagnostic assay with curated biomarkers complements clinicopathological factors to facilitate early diagnosis and ensure timely treatment delivery. This study focuses on an Asian-centric cancer diagnostic assay designed and thoroughly validated against commercially available standard references and a cohort of over 200 clinical specimens spanning 12 diverse Asian-centric cancer types. Methods: The assay uses hybrid-capture probes capable of profiling DNA aberrations from 572 cancer-related genes and 91 RNA fusion partners. The panel can detect clinically-tractable biomarkers such as microsatellite instability (MSI) and tumor mutation burden (TMB). Results: Analytical evaluation demonstrated 100% specificity and 99.9% sensitivity within a ≥5% VAF limit of detection (LoD) for SNV/Indels. RNA-based fusion features an LoD of ≥5 copies per nanogram input when evaluated against commercial references. Excellent linearity and concordance were observed when benchmarking against orthogonal methods in identifying MSI status, TMB scores and RNA fusions. Actionable genetic alterations were identified in 65% of the clinical samples. Conclusion: These results demonstrate a molecular diagnostic assay that accurately detects genomic alterations and complex biomarkers. The data also supports an excellent performance of this assay for making critical diagnoses and well-informed therapeutic decisions in Asian prevalent cancers.

Longitudinal Evaluation of Circulating Tumor DNA Using Sensitive Amplicon-Based Next-Generation Sequencing to Identify Resistance Mechanisms to Immune Checkpoint Inhibitors for Advanced Urothelial Carcinoma.

Serial evaluation of circulating tumor DNA may allow noninvasive assessment of drivers of resistance to immune checkpoint inhibitors (ICIs) in advanced urothelial cancer (aUC). We used a novel, amplicon-based next-generation sequencing assay to identify genomic alterations (GAs) pre- and post-therapy in 39 patients with aUC receiving ICI and 6 receiving platinum-based chemotherapy (PBC). One or more GA was seen in 95% and 100% of pre- and post-ICI samples, respectively, commonly in TP53 (54% and 54%), TERT (49% and 59%), and BRCA1/BRCA2 (33% and 33%). Clearance of ≥1 GA was seen in 7 of 9 patients responding to ICI, commonly in TP53 (n = 4), PIK3CA (n = 2), and BRCA1/BRCA2 (n = 2). A new GA was seen in 17 of 20 patients progressing on ICI, frequently in BRCA1/BRCA2 (n = 6), PIK3CA (n = 3), and TP53 (n = 3), which seldom emerged in patients receiving PBC. These findings highlight the potential for longitudinal circulating tumor DNA evaluation in tracking response and resistance to therapy.

Complementing Tissue Testing With Plasma Mutation Profiling Improves Therapeutic Decision-Making for Patients With Lung Cancer.

BACKGROUND: Tissue biopsy is an integral part of the diagnostic approach to lung cancer. It is however invasive and limited by heterogeneity. Liquid biopsies may complement tissue testing by providing additional molecular information and may be particularly helpful in patients from whom obtaining sufficient tissue for genomic profiling is challenging. METHODS: Patients with suspected lung cancer (n = 71) were prospectively recruited. Blood and diagnostic tissue samples were collected within 48 h of each other. Plasma cell-free DNA (cfDNA) testing was done using an ultrasensitive amplicon-based next-generation sequencing (NGS) panel (plasma NGS testing). For cases diagnosed as non-small cell lung carcinoma (NSCLC) via histology or cytology, targeted testing for epidermal growth factor receptor (EGFR) mutations was performed using tissue biopsy samples (tissue EGFR testing), where available. Concordance of clinically actionable mutations between methods and sample types was assessed. RESULTS: For confirmed NSCLC cases (n = 54), tissue EGFR test results were available only for 70.3% (38/54) due to sample inadequacies, compared to blood samples for 98.1% (53/54) cases. Tissue EGFR testing identified sensitizing EGFR (L858R or exon 19 deletion) mutation in 31.6% (12/38) of cases. Plasma NGS identified clinically actionable mutations in 37.7% (20/53) of cases, including EGFR mutations in two cases with no tissue EGFR results, and mutations in KRAS, BRAF, and MET. The overall sensitivity of sensitizing EGFR mutation detection by plasma NGS was 75% (9/12), and specificity was 100% (25/25) in patients tested in both tissue EGFR and plasma NGS (n = 37). In this cohort of patients, tissue EGFR testing alone informed clinical decisions in 22.2% (12/54) of cases. Adding plasma NGS to tissue EGFR testing increased the detection rate of actionable mutations to 42.6% (23/54), representing a 1.9-fold increase in clinically relevant findings. The average turnaround time of plasma NGS was shorter than standard tissue testing (10 vs. 29.9 days, p < 0.05). CONCLUSIONS: In the first-line setting, plasma NGS was highly concordant with tissue EGFR testing. Plasma NGS increases the detection of actionable findings with a shorter time to results. This study outlines the clinical utility of complementary plasma mutation profiling in the routine management of lung cancer patients.

Regional registration of whole slide image stacks containing major histological artifacts.

BACKGROUND: High resolution 2D whole slide imaging provides rich information about the tissue structure. This information can be a lot richer if these 2D images can be stacked into a 3D tissue volume. A 3D analysis, however, requires accurate reconstruction of the tissue volume from the 2D image stack. This task is not trivial due to the distortions such as tissue tearing, folding and missing at each slide. Performing registration for the whole tissue slices may be adversely affected by distorted tissue regions. Consequently, regional registration is found to be more effective. In this paper, we propose a new approach to an accurate and robust registration of regions of interest for whole slide images. We introduce the idea of multi-scale attention for registration. RESULTS: Using mean similarity index as the metric, the proposed algorithm (mean ± SD [Formula: see text]) followed by a fine registration algorithm ([Formula: see text]) outperformed the state-of-the-art linear whole tissue registration algorithm ([Formula: see text]) and the regional version of this algorithm ([Formula: see text]). The proposed algorithm also outperforms the state-of-the-art nonlinear registration algorithm (original: [Formula: see text], regional: [Formula: see text]) for whole slide images and a recently proposed patch-based registration algorithm (patch size 256: [Formula: see text] , patch size 512: [Formula: see text]) for medical images. CONCLUSION: Using multi-scale attention mechanism leads to a more robust and accurate solution to the problem of regional registration of whole slide images corrupted in some parts by major histological artifacts in the imaged tissue.

Computerized Image Analysis of Tumor Cell Nuclear Morphology Can Improve Patient Selection for Clinical Trials in Localized Clear Cell Renal Cell Carcinoma.

BACKGROUND: Clinicopathological scores are used to predict the likelihood of recurrence-free survival for patients with clear cell renal cell carcinoma (ccRCC) after surgery. These are fallible, particularly in the middle range. This inevitably means that a significant proportion of ccRCC patients who will not develop recurrent disease enroll into clinical trials. As an exemplar of using digital pathology, we sought to improve the predictive power of "recurrence free" designation in localized ccRCC patients, by precise measurement of ccRCC nuclear morphological features using computational image analysis, thereby replacing manual nuclear grade assessment. MATERIALS AND METHODS: TNM 8 UICC pathological stage pT1-pT3 ccRCC cases were recruited in Scotland and in Singapore. A Leibovich score (LS) was calculated. Definiens Tissue studio® (Definiens GmbH, Munich) image analysis platform was used to measure tumor nuclear morphological features in digitized hematoxylin and eosin (H&E) images. RESULTS: Replacing human-defined nuclear grade with computer-defined mean perimeter generated a modified Leibovich algorithm, improved overall specificity 0.86 from 0.76 in the training cohort. The greatest increase in specificity was seen in LS 5 and 6, which went from 0 to 0.57 and 0.40, respectively. The modified Leibovich algorithm increased the specificity from 0.84 to 0.94 in the validation cohort. CONCLUSIONS: CcRCC nuclear mean perimeter, measured by computational image analysis, together with tumor stage and size, node status and necrosis improved the accuracy of predicting recurrence-free in the localized ccRCC patients. This finding was validated in an ethnically different Singaporean cohort, despite the different H and E staining protocol and scanner used. This may be a useful patient selection tool for recruitment to multicenter studies, preventing some patients from receiving unnecessary additional treatment while reducing the number of patients required to achieve adequate power within neoadjuvant and adjuvant clinical studies.

Baseline Plasma EGFR Circulating Tumour DNA Levels in a Pilot Cohort of EGFR-Mutant Limited-Stage Lung Adenocarcinoma Patients Undergoing Radical Lung Radiotherapy.

The use of circulating cell-free tumour DNA (ctDNA) is established in metastatic lung adenocarcinoma to detect and monitor sensitising EGFR mutations. In early-stage disease, there is very little data supporting its role as a potential biomarker. We report on a prospective cohort of 9 limited-stage EGFR mutant lung cancer patients who were treated with radical radiotherapy. We looked at baseline plasma EGFR ctDNA and noted the detection rates to be higher in locally advanced disease. At a median follow-up of 13.5 months, an association between a detectable pre-radiotherapy plasma EGFR ctDNA and early tumour relapse (155 days vs. NR, p = 0.004) was noted. One patient with persistent plasma EGFR ctDNA predated radiological progression. The role of ctDNA in early-stage lung cancer is developing. Plasma EGFR ctDNA could be a useful biomarker in lung cancer patients undergoing radical treatments for staging, prognostication, and follow-up. These preliminary findings should be explored in larger studies.

Single Cell Hydrodynamic Stretching and Microsieve Filtration Reveal Genetic, Phenotypic and Treatment-Related Links to Cellular Deformability.

Deformability is shown to correlate with the invasiveness and metastasis of cancer cells. Recent studies suggest epithelial-to-mesenchymal transition (EMT) might enable cancer metastasis. However, the correlation of EMT with cancer cell deformability has not been well elucidated. Cellular deformability could also help evaluate the drug response of cancer cells. Here, we combine hydrodynamic stretching and microsieve filtration to study cellular deformability in several cellular models. Hydrodynamic stretching uses extensional flow to rapidly quantify cellular deformability and size with high throughput at the single cell level. Microsieve filtration can rapidly estimate relative deformability in cellular populations. We show that colorectal cancer cell line RKO with the mesenchymal-like feature is more flexible than the epithelial-like HCT116. In another model, the breast epithelial cells MCF10A with deletion of the TP53 gene are also significantly more deformable compared to their isogenic wildtype counterpart, indicating a potential genetic link to cellular deformability. We also find that the drug docetaxel leads to an increase in the size of A549 lung cancer cells. The ability to associate mechanical properties of cancer cells with their phenotypes and genetics using single cell hydrodynamic stretching or the microsieve may help to deepen our understanding of the basic properties of cancer progression.

Incidence of breast cancer attributable to breast density, modifiable and non-modifiable breast cancer risk factors in Singapore.

Incidence of breast cancer is rising rapidly in Asia. Some breast cancer risk factors are modifiable. We examined the impact of known breast cancer risk factors, including body mass index (BMI), reproductive and hormonal risk factors, and breast density on the incidence of breast cancer, in Singapore. The study population was a population-based prospective trial of screening mammography - Singapore Breast Cancer Screening Project. Population attributable risk and absolute risks of breast cancer due to various risk factors were calculated. Among 28,130 women, 474 women (1.7%) developed breast cancer. The population attributable risk was highest for ethnicity (49.4%) and lowest for family history of breast cancer (3.8%). The proportion of breast cancers that is attributable to modifiable risk factor BMI was 16.2%. The proportion of breast cancers that is attributable to reproductive risk factors were low; 9.2% for age at menarche and 4.2% for number of live births. Up to 45.9% of all breast cancers could be avoided if all women had breast density <12% and BMI <25 kg/m2. Notably, sixty percent of women with the lowest risk based on non-modifiable risk factors will never reach the risk level recommended for mammography screening. A combination of easily assessable breast cancer risk factors can help to identify women at high risk of developing breast cancer for targeted screening. A large number of high-risk women could benefit from risk-reduction and risk stratification strategies.

Mechanically and chemically defined hydrogel matrices for patient-derived colorectal tumor organoid culture.

Patient-derived tumor organoids offer potentially useful models of cancer tissue physiology. Yet, conventional organoid cultures utilize generic matrices that are difficult to tailor for various unique tumor microenvironments. Here, we employ synthetic, enzymatically crosslinked hydrogels to define mechanical and biochemical properties hypothesized to be relevant for maintaining these organoids. We show that a single extracellular matrix component, gelatin, suffices to support colorectal cancer patient-derived xenograft (CRC-PDX) organoid survival, and that high matrix stiffness synergizes with hypoxia to increase organoid growth and metabolism in a majority of CRC-PDX lines tested. Moreover, we demonstrate that defined gelatin-based hydrogels support CRC-PDX tumor growth in vivo and organoid sensitivity to various CRC therapeutic drugs in vitro in a largely comparable fashion to a conventional reconstituted basement membrane matrix. Based on our findings, we propose that enzymatically crosslinked hydrogels potentially provide a platform for designing mechanically and biochemically defined matrices for various types of patient-derived tumor organoids.

Carrier-Enhanced Anticancer Efficacy of Sunitinib-Loaded Green Tea-Based Micellar Nanocomplex beyond Tumor-Targeted Delivery.

Although a few nanomedicines have been approved for clinical use in cancer treatment, that recognizes improved patient safety through targeted delivery, their improved efficacy over conventional drugs has remained marginal. One of the typical drawbacks of nanocarriers for cancer therapy is a low drug-loading capacity that leads to insufficient efficacy and requires an increase in dosage and/or frequency of administration, which in turn increases carrier toxicity. In contrast, elevating drug-loading would cause the risk of nanocarrier instability, resulting in low efficacy and off-target toxicity. This intractable drug-to-carrier ratio has imposed constraints on the design and development of nanocarriers. However, if the nanocarrier has intrinsic therapeutic effects, the efficacy would be synergistically augmented with less concern for the drug-to-carrier ratio. Sunitinib-loaded micellar nanocomplex (SU-MNC) was formed using poly(ethylene glycol)-conjugated epigallocatechin-3-O-gallate (PEG-EGCG) as such a carrier. SU-MNC specifically inhibited the vascular endothelial growth factor-induced proliferation of endothelial cells, exhibiting minimal cytotoxicity to normal renal cells. SU-MNC showed enhanced anticancer effects and less toxicity than SU administered orally/intravenously on human renal cell carcinoma-xenografted mice, demonstrating more efficient effects on anti-angiogenesis, apoptosis induction, and proliferation inhibition against tumors. In comparison, a conventional nanocarrier, SU-loaded polymeric micelle (SU-PM) comprised of PEG-b-poly(lactic acid) (PEG-PLA) copolymer, only reduced toxicity with no elevated efficacy, despite comparable drug-loading and tumor-targeting efficiency to SU-MNC. Improved efficacy of SU-MNC was ascribed to the carrier-drug synergies with the high-performance carrier of PEG-EGCG besides tumor-targeted delivery.

Factors associated with false-positive mammography at first screen in an Asian population.

INTRODUCTION: False-positive recall is an issue in national screening programmes. The aim of this study is to investigate the recall rate at first screen and to identify potential predictors of false-positive recall in a multi-ethnic Asian population-based breast cancer screening programme. METHODS: Women aged 50-64 years attending screening mammography for the first time (n = 25,318) were included in this study. The associations between potential predictors (sociodemographic, lifestyle and reproductive) and false-positive recall were evaluated using multivariable logistic regression models. RESULTS: The recall rate was 7.6% (n = 1,923), of which with 93.8% were false-positive. Factors independently associated with higher false-positive recall included Indian ethnicity (odds ratio [95% confidence interval]: 1.52 [1.25 to 1.84]), premenopause (1.23 [1.04 to 1.44]), nulliparity (1.85 [1.57 to 2.17]), recent breast symptoms (1.72 [1.31 to 2.23]) and history of breast lump excision (1.87 [1.53 to 2.26]). Factors associated with lower risk of false-positive recall included older age at screen (0.84 [0.73 to 0.97]) and use of oral contraceptives (0.87 [0.78 to 0.97]). After further adjustment of percent mammographic density, associations with older age at screening (0.97 [0.84 to 1.11]) and menopausal status (1.12 [0.95 to 1.32]) were attenuated and no longer significant. CONCLUSION: For every breast cancer identified, 15 women without cancer were subjected to further testing. Efforts to educate Asian women on what it means to be recalled will be useful in reducing unnecessary stress and anxiety.

Generation of mature kupffer cells from human induced pluripotent stem cells.

Liver macrophages, Kupffer cells (KCs), play a critical role in drug-induced liver injury (DILI) and liver diseases including cholestasis, liver fibrosis and viral hepatitis. Application of KCs in in vitro models of DILI and liver diseases is hindered due to limited source of human KCs. In vivo, KCs originate from MYB-independent macrophage progenitors, which differentiate into liver-specific macrophages in response to hepatic cues in the liver. Here, we recapitulated KCs ontogeny by differentiation of MYB-independent iPSCs to macrophage-precursors and exposing them to hepatic cues to generate iPSC-derived KCs (iKCs). iKCs expressed macrophage markers (CD11/CD14/CD68/CD163/CD32) at 0.3-5 folds of primary adult human KCs (pKCs) and KC-specific CLEC-4F, ID1 and ID3. iKCs phagocytosed and secreted IL-6 and TNFα upon stimulation at levels similar to pKCs but different from non-liver macrophages. Hepatocyte-iKCs co-culture model was more sensitive in detecting hepatotoxicity induced by inflammation-associated drugs, Acetaminophen and Trovafloxacin, and Chlorpromazine-induced cholestasis when compared to hepatocytes alone. Overall, iKCs were mature, liver-specific and functional. Furthermore, donor-matched iKCs and iPSC-hepatocyte co-culture exhibited minimal non-specific background response compared to donor-mismatched counterpart. iKCs offer a mature renewable human cell source for liver-specific macrophages, useful in developing in vitro model to study DILI and liver diseases such as cholestasis.