BACKGROUND:The introduction of Trastuzumab deruxtecan (T-Dxd) has exposed clinically significant limitations in accurately detecting HER2-low expression testing when using immunohistochemistry (IHC) assays originally developed to detect HER2 over-expression. While HER2 testing is widely used to determine T-Dxd eligibility, no HER2-low assay was ever validated against HER2 protein expression. METHODS:To address this pressing need, the Consortium for Analytic Standardization in Immunohistochemistry (CASI) conducted the CASI-01 study, involving 54 IHC laboratories across Europe and the U.S. The study aimed to identify optimal assay conditions for accurate HER2 testing, differentiating between HER2 overexpression (3+) for Trastuzumab eligibility and HER2-low expression (1+ or ultra-low) for T-Dxd eligibility. The conventional FDA-cleared HER2 assay ("predicate") was compared with higher-sensitivity assays using pathologist versus image analysis readouts. HER2 overexpression was validated against HER2 gene amplification via in situ hybridisation (ISH), while HER2-low accuracy was evaluated using newly introduced HER2 reference standards and a novel IHC parameter-dynamic range. FINDINGS:CASI-01 revealed variability in predicate HER2 assays, with detection thresholds ranging from 30,000 to 60,000 among laboratories. Despite this variability, these assays demonstrated high accuracy for identifying HER2 overexpression (3+), with 85.7% (18/21) sensitivity (95% confidence limits 63.66-96.95%) and 100% (49/49) specificity (95% confidence limits 92.75-100%), though sensitivity may have been limited by the use of older tissue specimens, with loss or reduced expression levels of the HER2 protein. However, these same assays exhibited poor dynamic range for detecting HER2-low scores. Enhanced analytic sensitivity of IHC assays combined with image analysis overcame this limitation with HER2-low scores, achieving a six-fold improvement (p = 0.0017). INTERPRETATION:IHC assays with detection thresholds in the range of 30,000-60,000 HER2 molecules per cell yield accurate results for determination of Trastuzumab eligibility (HER2 3+) but fail to demonstrate the dynamic range for accurate HER2-low scores. Enhanced analytic sensitivity of HER2 assays combined with image analysis addresses this critical gap in HER2-low testing. More generally, CASI-01 introduces pivotal advancements in precision medicine: (a) the importance of reporting IHC analytic sensitivity and ability to demonstrate an assay dynamic range, and (b) image analysis can surpass pathologist readout accuracy in specific clinical contexts. FUNDING:This work was supported by the National Cancer Institute of the National Institutes of Health under Award Number R44CA268484.

Background/Objectives: The NavDx+Gyn blood test detects and quantifies fourteen HPV types in various sample types to provide a reliable means of detecting and monitoring HPV-driven gynecologic cancers. NavDx+Gyn is an extension of the NavDx assay, which identifies five high-risk HPV types. NavDx has been clinically validated in multiple independent studies for the surveillance of HPV-driven oropharyngeal cancer and has been integrated into clinical practice by over 1300 healthcare providers at over 500 medical sites in the US. The NavDx+Gyn assay incorporates an analysis of nine additional high-risk HPV types. Here, we report a detailed analytical validation of the NavDx+Gyn assay for use in cervical, vaginal, and vulvar cancer patients to detect fourteen high-risk HPV types related to HPV-driven gynecologic cancers. Methods: Parameters include specificity as measured by limits of blank (LoBs) and sensitivity illustrated via limits of detection and quantitation (LoDs and LoQs). Results: The LoBs were between 0 and 0.0926 copies/μL, LoDs were 0.1009 to 0.3147 copies/μL, and LoQs were 0.1009 to 0.3147 copies/μL, demonstrating the high analytic sensitivity and specificity provided by NavDx+Gyn. In-depth evaluations, including accuracy and intra- and inter-assay precision studies, were shown to be within acceptable ranges. Regression analysis revealed a high degree of correlation between expected and effective concentrations, demonstrating excellent linearity (R2 > 0.99) across a broad range of analyte concentrations. Conclusions: These results demonstrate that NavDx+Gyn accurately and reproducibly detects fourteen types of high-risk HPV, which aids in the diagnosis and surveillance of the vast majority of HPV-driven gynecologic cancers.

PDF file - 59KB, The knockdown of NDY1/KDM2B suppresses the activity of the Wnt pathway, despite the upregulation of Wnt1.

Supplementary Figure 3 from Systemic Stromal Effects of Estrogen Promote the Growth of Estrogen Receptor–Negative Cancers

Supplementary Data from Chemotherapy-Induced Collagen IV Drives Cancer Cell Motility through Activation of Src and Focal Adhesion Kinase

PDF file - 3801KB, The knockdown of NDY1/KDM2B is not rescued by exogenous EZH2.

Supplementary Material, Methods and Figure Legend from Akt1 Ablation Inhibits, whereas Akt2 Ablation Accelerates, the Development of Mammary Adenocarcinomas in Mouse Mammary Tumor Virus (MMTV)-ErbB2/Neu and MMTV-Polyoma Middle T Transgenic Mice

The NavDx® blood test analyzes tumor tissue modified viral (TTMV)-HPV DNA to provide a reliable means of detecting and monitoring HPV-driven cancers. The test has been clinically validated in a large number of independent studies and has been integrated into clinical practice by over 1000 healthcare providers at over 400 medical sites in the US. This Clinical Laboratory Improvement Amendments (CLIA), high complexity laboratory developed test, has also been accredited by the College of American Pathologists (CAP) and the New York State Department of Health. Here, we report a detailed analytical validation of the NavDx assay, including sample stability, specificity as measured by limits of blank (LOBs), and sensitivity illustrated via limits of detection and quantitation (LODs and LOQs). LOBs were 0–0.32 copies/μL, LODs were 0–1.10 copies/μL, and LOQs were <1.20–4.11 copies/μL, demonstrating the high sensitivity and specificity of data provided by NavDx. In-depth evaluations including accuracy and intra- and inter-assay precision studies were shown to be well within acceptable ranges. Regression analysis revealed a high degree of correlation between expected and effective concentrations, demonstrating excellent linearity (R2 = 1) across a broad range of analyte concentrations. These results demonstrate that NavDx accurately and reproducibly detects circulating TTMV-HPV DNA, which has been shown to aid in the diagnosis and surveillance of HPV-driven cancers.

PDF file, 945K, Adipose stromal cells synthesize lipid droplets in response to adipogenic media.

PDF file - 78KB, The NDY1/KDM2B knockdown inhibits anchorage-dependent and independent growth of cancer cells.

Supplementary Figure 1 from Akt1 Ablation Inhibits, whereas Akt2 Ablation Accelerates, the Development of Mammary Adenocarcinomas in Mouse Mammary Tumor Virus (MMTV)-ErbB2/Neu and MMTV-Polyoma Middle T Transgenic Mice

PDF file, 440K, Characterization of SVF cell lines and effects of treatment with diphtheria toxin in vivo.

PDF file, 1009K, Characterization of SVF/EV and SVF/CCL2-derived end stage tumors.

PDF file - 154KB, Table S1. Cell lines, media. Table S2. Pre-miRs and anti-miRs. Table S3. Primers. Table S4. Antibodies. Table S5. Clinical characteristics of the tumor-samples used in Figure 7.

Supplementary Figure 4 from Systemic Stromal Effects of Estrogen Promote the Growth of Estrogen Receptor–Negative Cancers

Supplementary Figure 2 from Systemic Stromal Effects of Estrogen Promote the Growth of Estrogen Receptor–Negative Cancers

Supplementary Figure 1 from Systemic Stromal Effects of Estrogen Promote the Growth of Estrogen Receptor–Negative Cancers

PDF file - 91KB, The knockdown of NDY1/KDM2B is not rescued by exogenous BMI1.

PDF file - 139KB, The knockdown of NDY1/KDM2B interferes with the replating efficiency of mammospheres from both basal and luminal mammary adenocarcinoma cell lines.

PDF file - 3478KB, Knockdown of NDY1/KDM2B promotes differentiation of human mammary adenocarcinoma cell lines in culture.