Clinical Validation Research Articles

Clinical validation of C12FDG as a marker associated with senescence and osteoarthritic phenotypes.

Chronic conditions associated with aging have proven difficult to prevent or treat. Senescence is a cell fate defined by loss of proliferative capacity and the development of a pro-inflammatory senescence-associated secretory phenotype comprised of cytokines/chemokines, proteases, and other factors that promotes age-related diseases. Specifically, an increase in senescent peripheral blood mononuclear cells (PBMCs), including T cells, is associated with conditions like frailty, rheumatoid arthritis, and bone loss. However, it is unknown if the percentage of senescent PBMCs associated with age-associated orthopedic decline could be used for potential diagnostic or prognostic use in orthopedics. Here, we report senescent cell detection using the fluorescent compound C12FDG to quantify PBMCs senescence across a large cohort of healthy and osteoarthritic patients. There is an increase in the percent of circulating C12FDG+ PBMCs that is commensurate with increases in age and senescence-related serum biomarkers. Interestingly, C12FDG+ PBMCs and T cells also were found to be elevated in patients with mild to moderate osteoarthritis, a progressive joint disease that is strongly associated with inflammation. The percent of C12FDG+ PBMCs and age-related serum biomarkers were decreased in a small subgroup of study participants taking the senolytic drug fisetin. These results demonstrate quantifiable measurements in a large group of participants that could create a composite score of healthy aging sensitive enough to detect changes following senolytic therapy and may predict age-related orthopedic decline. Detection of peripheral senescence in PBMCs and subsets using C12FDG may be clinically useful for quantifying cellular senescence and determining how and if it plays a pathological role in osteoarthritic progression.

Clinical Implementation and Initial Validation of Respiratory-Gated Stereotactic Body Radiotherapy for Thoracoabdominal Tumors Under Abdominal Compression Using an Anzai Laser-Based Gating Device With Visual Guidance on an Elekta Linear Accelerator

Clinical Implementation and Initial Validation of Respiratory-Gated Stereotactic Body Radiotherapy for Thoracoabdominal Tumors Under Abdominal Compression Using an Anzai Laser-Based Gating Device With Visual Guidance on an Elekta Linear Accelerator

Medical comorbidities in bipolar disorder (BIPCOM): clinical validation of risk factors and biomarkers to improve prevention and treatment. Study protocol

BackgroundBIPCOM aims to (1) identify medical comorbidities in people with bipolar disorder (BD); (2) examine risk factors and clinical profiles of Medical Comorbidities (MC) in this clinical group, with a special focus on Metabolic Syndrome (MetS); (3) develop a Clinical Support Tool (CST) for the personalized management of BD and medical comorbidities.MethodsThe BIPCOM project aims to investigate MC, specifically MetS, in individuals with BD using various approaches. Initially, prevalence rates, characteristics, genetic and non-genetic risk factors, and the natural progression of MetS among individuals with BD will be assessed by analysing Nordic registers, biobanks, and existing patient datasets from 11 European recruiting centres across 5 countries. Subsequently, a clinical study involving 400 participants from these sites will be conducted to examine the clinical profiles and incidence of specific MetS risk factors over 1 year. Baseline assessments, 1-year follow-ups, biomarker analyses, and physical activity measurements with wearable biosensors, and focus groups will be performed. Using this comprehensive data, a CST will be developed to enhance the prevention, early detection, and personalized treatment of MC in BD, by incorporating clinical, biological, sex and genetic information. This protocol will highlight the study's methodology.DiscussionBIPCOM's data collection will pave the way for tailored treatment and prevention approaches for individuals with BD. This approach has the potential to generate significant healthcare savings by preventing complications, hospitalizations, and emergency visits related to comorbidities and cardiovascular risks in BD. BIPCOM's data collection will enhance BD patient care through personalized strategies, resulting in improved quality of life and reduced costly interventions. The findings of the study will contribute to a better understanding of the relationship between medical comorbidities and BD, enabling accurate prediction and effective management of MetS and cardiovascular diseases.Trial registration: ISRCTN68010602 at https://www.isrctn.com/ISRCTN68010602. Registration date: 18/04/2023.

Clinical validity of the Italian adaptation of the Uniform Data Set Neuropsychological Test Battery (I-UDSNB) in Mild Cognitive Impairment and Alzheimer’s Disease

BackgroundThe identification and staging of Alzheimer’s Disease (AD) represent a challenge, especially in the prodromal stage of Mild Cognitive Impairment (MCI), when cognitive changes can be subtle. Worldwide efforts were dedicated to select and harmonize available neuropsychological instruments. In Italy, the Italian Network of Neuroscience and Neuro-Rehabilitation has promoted the adaptation of the Uniform Data Set Neuropsychological Test Battery (I-UDSNB), collecting normative data from 433 healthy controls (HC).Here, we aimed to explore the ability of I-UDSNB to differentiate between a) MCI and HC, b) AD and HC, c) MCI and AD.MethodsOne hundred thirty-seven patients (65 MCI, 72 AD) diagnosed after clinical-neuropsychological assessment, and 137 HC were included. We compared the I-UDSNB scores between a) MCI and HC, b) AD and HC, c) MCI and AD, with t-tests. To identify the test(s) most capable of differentiating between groups, significant scores were entered in binary logistic and in stepwise regressions, and then in Receiver Operating Characteristic curve analyses.ResultsTwo episodic memory tests (Craft Story and Five Words test) differentiated MCI from HC subjects; Five Words test, Semantic Fluency (vegetables), and TMT-part B differentiated AD from, respectively, HC and MCI.ConclusionsOur findings indicate that the I-UDSNB is a suitable tool for the harmonized and concise assessment of patients with cognitive decline, showing high sensitivity and specificity for the diagnosis of MCI and AD.

Rehabilitation Assessment System for Stroke Patients Based on Fusion-Type Optoelectronic Plethysmography Device and Multi-Modality Fusion Model: Design and Validation

This study aimed to propose a portable and intelligent rehabilitation evaluation system for digital stroke-patient rehabilitation assessment. Specifically, the study designed and developed a fusion device capable of emitting red, green, and infrared lights simultaneously for photoplethysmography (PPG) acquisition. Leveraging the different penetration depths and tissue reflection characteristics of these light wavelengths, the device can provide richer and more comprehensive physiological information. Furthermore, a Multi-Channel Convolutional Neural Network–Long Short-Term Memory–Attention (MCNN-LSTM-Attention) evaluation model was developed. This model, constructed based on multiple convolutional channels, facilitates the feature extraction and fusion of collected multi-modality data. Additionally, it incorporated an attention mechanism module capable of dynamically adjusting the importance weights of input information, thereby enhancing the accuracy of rehabilitation assessment. To validate the effectiveness of the proposed system, sixteen volunteers were recruited for clinical data collection and validation, comprising eight stroke patients and eight healthy subjects. Experimental results demonstrated the system’s promising performance metrics (accuracy: 0.9125, precision: 0.8980, recall: 0.8970, F1 score: 0.8949, and loss function: 0.1261). This rehabilitation evaluation system holds the potential for stroke diagnosis and identification, laying a solid foundation for wearable-based stroke risk assessment and stroke rehabilitation assistance.

Abstract PO5-15-11: A Novel Biosignature to Predict Radiation Therapy Response in Early-Stage Invasive Breast Cancer Treated with Breast-Conserving Surgery

Abstract Introduction: Radiation therapy (RT) is a standard component of treatment for most patients with early-stage, hormone receptor-positive, invasive breast cancer (BC) following breast-conserving surgery (BCS). RT recommendations are primarily based on clinicopathologic (CP) factors, such as tumor size, nodal status, and histological grade. While these factors provide prognostic information, their ability to accurately identify patients at a higher risk of ipsilateral breast recurrence (IBR) or those who will significantly benefit from RT is unclear. Therefore, incorporation of additional predictive markers beyond CP factors is warranted and there remains a need to refine the selection criteria for patients who will derive the greatest benefit from RT. In this study, we aimed to explore the role of immune and metabolic signaling axes as potential predictive markers for RT response in hormone receptor-positive breast cancer patients. Methods: A cohort of 939 women from Sweden were diagnosed with BC between 1987 and 2004. A subset of 471 patients was diagnosed with T1-T2N0M0 BC and underwent BCS with negative margins. Patients were treated with BCS plus/minus RT (median dose 50Gy) based upon physician preference. Biomarkers were assayed in formalin-fixed paraffin-embedded tissue micro arrays using multiplex immunofluorescence and multi-spectral imaging in a CLIA lab (Laguna Hills, CA). Immune and metabolic axes were defined employing biomarkers and used to classify patients into three risk groups: 1) Low Risk (lower baseline risk and no RT benefit), 2) Elevated Risk (higher baseline risk and significant RT benefit), and 3) Residual Risk (higher baseline risk and suboptimal RT benefit). Risk groups were analyzed for IBR rate using Kaplan-Meier and Cox proportional hazards analyses. Risk group prognosis and RT prediction was assessed for RT-risk group interaction in multivariable analysis, adjusting for CP factors. Results: Among the patients with complete biomarker data, 261 were hormone receptor-positive, and had a median follow-up of 140 months. Median age was 62 years (28-88 years), and median tumor size was 1.1 cm (0.1-4 cm). Some patients (21%) received endocrine therapy (ET), and most (76%) received RT, with 79% of patients who received ET also receiving RT. In the overall population, RT following BCS was associated with a reduction in HR for recurrence (HR=0.3, 95%CI [0.1, 0.5], p< 0.001). Patients classified as Low Risk (41%) did not have a significant benefit from RT (HR=0.9, 95%CI [0.1, 8.3], p=0.95) in multivariable analysis adjusted for CP factors. For patients who were classified as Elevated Risk (31%), multivariable analysis showed a higher IBR rate without RT (HR=5.7, 95%CI [1.3, 25.1], p< 0.021) relative to the Low Risk group who had a significant benefit from RT (HR=0.1, 95%CI [0.02, 0.2], p< 0.001). Patients classified into the Residual Risk group (28%) had no significant reduction from RT (HR=0.7, 95%CI [0.2, 2.0], p=0.61). However, in multivariable analysis adjusted for CP factors, patients treated with RT in the Residual Risk group had higher IBR rates (HR=3.7, 95%CI [1.6, 8.7], p=0.0026) compared to RT-treated patients in other risk groups. The biosignature was predictive for RT benefit in multivariable analysis adjusted for CP factors (p-interaction < 0.001). Conclusions:The biosignature used in this study demonstrated the potential to identify patients who may derive limited benefit from RT, allowing for tailored therapeutic approaches. These findings underscore the importance of individualized treatment decisions based on risk stratification. Further clinical validation and refinement of risk stratification strategies are warranted to optimize outcomes in hormone receptor-positive, BC patients undergoing BCS. Citation Format: Troy Bremer, Karuna Mittal, Chirag Shah, Frank Vicini, Steven Shivers, Charlotta Wadsten. A Novel Biosignature to Predict Radiation Therapy Response in Early-Stage Invasive Breast Cancer Treated with Breast-Conserving Surgery [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-15-11.

Abstract PO1-28-06: Development of a plasma-based real-time qPCR gene expression assay for targeted screening and diagnosis of early-stage breast cancer

Abstract Background Over 12 million US women undergo follow-up breast imaging and exams annually due to breast cancer suspicions, incurring an $8 billion total cost, of which $2.18 billion is attributed to false-positive breast biopsies. Recent updates published to the Mammography Quality Standards Act (MQSA) now recommend supplemental imaging for women with heterogeneously dense and extremely dense breasts, regardless of suspicious findings. Given sensitivity limitations associated with mammography in dense-breasted women, as low as 30-47%, and non-specificity associated with ultrasound and MRI, these recent recommendations are likely to further strain the healthcare system and place substantial burden on patients and providers. With an estimated 25 million dense-breasted women who regularly screen for breast cancer now considered at-risk, there is a critical need for affordable and highly accurate testing, offering better specificity without sacrificing sensitivity. To address this gap, we developed an affordable and accurate plasma-based gene expression assay using real-time qPCR for targeted screening and diagnosis of early-stage breast cancer. Methods In previous studies, 26 cross-correlated mRNA gene targets were discovered and independently validated for non-invasive breast cancer detection across five independent patient cohorts using microarray gene expression profiling, comprised of peripheral blood mononuclear cells (n=337) and saliva (n=20), demonstrating strong efficacy, reproducibility, and concordance across bio-fluids and assay platforms in all studies. A sixth independent validation cohort comprised of 203 plasma samples was subsequently obtained for real-time qPCR clinical assay development and validation in our CLIA laboratory. The CLIA validation cohort was designed to be representative of different diagnosis status, stage, and ethnicity, and included Caucasian, Black/African American, Hispanic, and Asian women. After normalizing mRNA expression and clustering analysis, the signature was refined to 8 target genes and was assessed in a cohort of 87 plasma samples [Table 1] to verify the assay’s diagnostic performance for stage I breast cancer detection, under locked laboratory protocols. We leveraged machine learning methods derived from XGBoost classification, a supervised-learning algorithm that uses sequentially built shallow decision trees to provide accurate results and avoidance of overfitting. Results The XGBoost model, selecting from the 8 mRNA gene targets and patient age, achieved >99% sensitivity, 89% specificity for stage I breast cancer detection in the held-out test set, with an overall diagnostic accuracy of 94.5%. Normalized data showed significant differences in gene expression between healthy controls and stage I patients, and distinct clustering was observed for the 8 mRNA gene signature, including patient age. Conclusions Our plasma-based real-time qPCR gene expression clinical assay, with machine learning, is positioned as a highly accurate non-invasive tool for targeted screening and diagnosis of early-stage breast cancer. Furthermore, a clinically validated assay in this space addresses the need for more targeted diagnostics, while serving as a promising tool for clinicians as they make critical care decisions across the breast health continuum, accelerating time to earlier and more precise intervention and treatment, mitigating unnecessary healthcare expenditures, and reducing the mental, emotional, and financial burden on patients and their families. Citation Format: Martin Keiser, Elizabeth Cormier-May, Matthew Alderdice, Joy Kavanagh, William Guesdon, Heather Healy, Nathalie Jean-Charles, Jay Harness. Development of a plasma-based real-time qPCR gene expression assay for targeted screening and diagnosis of early-stage breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-28-06.

Abstract PS10-07: A second-generation polygenic risk score (PRS) based on genetic ancestry improves breast cancer (BC) risk prediction for all ancestries

Abstract Background: Common genetic variants, mainly single-nucleotide polymorphisms (SNPs) explain substantial genetic susceptibility to BC. PRS have been developed to quantify the combined effects of BC-associated SNPs, providing important information about BC risk. Historically, genome-wide association studies have been conducted in predominantly European populations, resulting in miscalibrated and inaccurate PRS for non-Europeans. We previously described a multiple-ancestry PRS (MA-PRS-149) based on 56 ancestry-informative and 93 BC-associated SNPs. The MA-PRS-149 achieved accuracy for all women by characterizing the genetic ancestry of each BC-associated SNP in terms of three reference ancestries (African, East Asian, and European), applying ancestry-specific SNP risks and frequencies, and combining the results as a weighted sum of three ancestry-specific PRS. Here, we aimed to improve the predictive accuracy of MA-PRS-149, particularly for non-Europeans, through the inclusion of additional BC-associated SNPs. Methods: Women referred for hereditary cancer testing and negative for pathogenic variants in BC-associated genes were divided into consecutive study cohorts to (1) quantify ancestry-specific SNP risks, (2) combine the three ancestry-specific PRS, and (3) pre-specified clinical validation. To select an optimal set of BC-associated SNPs, we developed a novel synthetic stepwise regression methodology that accounts for linkage disequilibrium. Ancestry-specific SNP risks were determined from meta-analyses of literature with clinical cohorts of 57,827 Black/African and 26,992 East Asian women. Ancestry-specific PRS were combined based on a diverse cohort of 157,740 women. Clinical validation was conducted in an independent cohort of 77,774 women. We used multivariable logistic regression adjusted for age, ancestry, and cancer history to test for improved BC risk prediction over clinical factors. We tested for improvement over the MA-PRS-149, and a European PRS, by including additional PRS as covariates. Calibration was assessed through goodness-of-fit tests. All analyses were conducted within the full cohort and ancestral subpopulations. Odds ratios (ORs) and 95% confidence intervals (CIs) are reported per standard deviation within the corresponding population. Results: An optimal set of 383 SNPs (56 ancestry-informative and 327 BC-associated) was included in the final PRS (MA-PRS-383). MA-PRS-383 added significant predictive information to clinical factors in the full cohort and within each ancestry (Table 1). MA-PRS-383 had greater predictive accuracy than MA-PRS-149 or a 383-SNP PRS with European weights. Goodness-of-fit tests showed that MA-PRS-383 was well-calibrated and predicted risk accurately in the tails of the distribution for both European and non-European women. Conclusion: MA-PRS-383 was well-calibrated and substantially improved upon existing PRS in all tested ancestral populations. Incorporation of MA-PRS-383 into BC risk assessment may lead to more accurate identification of women who are most likely to benefit from screening and preventive medications. Citation Format: Timothy Simmons, Elisha Hughes, Dmitry Pruss, Matthew Kucera, Benjamin Roa, Thaddeus Judkins, Thomas Slavin, Victor Abkevich, Ryan Hoff, Srikanth Jammulapati, Susanne Wagner, Dale Muzzey, Jerry Lanchbury, Alexander Gutin. A second-generation polygenic risk score (PRS) based on genetic ancestry improves breast cancer (BC) risk prediction for all ancestries [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS10-07.

Abstract PO2-16-02: RBsig gene-expression signature in patients with endocrine resistant metastatic breast cancer treated with palbociclib and fulvestrant in the PYTHIA trial

Abstract Background: We have previously identified RBsig, a gene-expression signature of resistance to CDK4/6 inhibitors composed of E2F1/E2F2 dependent genes, which is associated with genetic loss of RB1. Although RBsig has been previously tested in vitro and in clinical trials with CDK4/6 inhibitors, further clinical validation is needed. Additionally, data on the correlation of RBsig with other prognostic/predictive biomarkers for CDK4/6 inhibitors such as serum Thymidine Kinase activity (sTKa), is lacking. Here, we aimed to investigate RBsig in patients (pts) treated with palbociclib (P) + fulvestrant (F) within the PYTHIA trial (IBCSG 53- 14/BIG 14-04; NCT02536742), a downstream trial of the AURORA molecular program (BIG 14-01; NCT02102165). Methods: PYTHIA is a biomarker discovery phase II trial which enrolled 122 pts with endocrine-resistant ER+ and HER2 - metastatic breast cancer (MBC) who received P+F at standard schedule and dose. We analyzed a subset of 38 pts enrolled in PYTHIA and successfully included in AURORA who had available RNAseq from tumor tissue samples (24 primary, 24 metastatic, 10 both). Ki67 by IHC was also available. The PYTHIA biomarker assessment included sTKa at baseline and during treatment (D15 and D28) measured using DiviTumÒ TKa (Biovica International). Evaluation of the prognostic role of RBsig was a pre-specified primary objective of PYTHIA. RBsig is composed of 87 genes and was calculated as previously described. Wilcoxon rank sum test was used to compare continuous RBsig values between groups defined by sTKa. Median progression free survival (mPFS) was estimated using Kaplan-Meier method for groups defined by dichotomizing RBsig at zero. Results: Median RBsig was higher in metastatic than in primary samples; in paired samples, RBsig increased in metastatic for 7/10 pts vs primary. When evaluated as continuous variables, RBsig correlated with tissue Ki67 both in primary (Spearman R 0.8) and in metastatic (R 0.68) samples. Additionally, RBsig was higher in Basal and Luminal B intrinsic subtypes in metastatic samples. RBsig was significantly higher in pts with high baseline sTKa (median cut-off) (Wilcoxon test, p=0.04 and p=0.03 in primary and metastatic samples respectively), with a similar direction, albeit not statistically significant, observed in the group of pts with no sTKa clearance at D15 (below limit of detection cut-off). When evaluated in the combined cohort, where RBSig values were taken from metastatic samples when available and from primary samples otherwise, higher RBsig values were associated with poorer PFS as compared to lower RBsig values [mPFS 5.5 months (95% CI 3.7, 14) vs 13 months (95% CI 8.6, 28); Log-rank p-value=0.004]. Conclusions: Higher RBsig was associated with higher tumor proliferation (measured by both tissue ki67 and sTKa), luminal B status, and shorter PFS on treatment with P+F. This analysis reinforces prior data on RBsig prognostic significance in pts treated with CDK4/6 inhibitors. Further investigation on the added value of RBsig as a novel biomarker for patients’ stratification, including in the early setting, is warranted. Citation Format: Matteo Benelli, Svitlana Tyekucheva, Thayane Crestani, Michail Ignatiadis, Patrick Neven, Marco Colleoni, Stéphanie Henry, Konstantinos Papadimitriou, Antonio Bernardo, Elena Seles, François Duhoux, Iain Macpherson, Alastair Thomson, David Mark Davies, Ilenia Migliaccio, Gabriele Zoppoli, Roswitha Kammler, Heidi De Swert, Barbara Ruepp, Philippe Aftimos, Jose Seoane, Dario Romagnoli, David Venet, Theodora Goulioti, David Cameron, Sherene Loi, Martine Piccart, Meredith Regan, Luca Malorni. RBsig gene-expression signature in patients with endocrine resistant metastatic breast cancer treated with palbociclib and fulvestrant in the PYTHIA trial [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-16-02.

Abstract PO2-06-10: Prevalence of genomic alterations in Xerna tumor microenvironment subtypes in triple negative breast cancer patients

Abstract Introduction: Immune checkpoint inhibitors (ICI) are an important therapeutic option for patients with triple negative breast cancer (TNBC). However, identification of patients most likely to respond is challenging. PD-L1 positivity by immunohistochemistry is the standard biomarker used for ICI therapy selection in TNBC. However, other biomarkers, such as analysis of the tumor microenvironment (TME) may be more accurate in predicting response. The XernaTM TME Panel uses RNA sequencing data and machine learning to analyze the TME, utilizing the angiogenic and immunogenic biology of the TME to classify tumors into four TME subtypes. In this study, the distribution of Xerna TME subtypes and associated genomic alterations in TNBC were investigated for their potential use in therapy selection. Methods: A total of 203 TNBC patient samples underwent tumor-normal whole-exome, whole-transcriptome sequencing testing with the OncoExTraTM assay. The whole-transcriptome expression data were analyzed using the Xerna TME Panel to assign each sample to one of four subtypes: Immune Active (IA), Immune Suppressed (IS), Immune Desert (ID) and Angiogenic (A). The IA and IS subtypes both have high immune scores that may be particularly sensitive to ICI therapy. Actionable alterations, defined as those with FDA-approved matched therapies in any cancer, with matched clinical trials, or with evidence in cancer guidelines or the literature for possible matched therapies, were also identified and associations across Xerna subtypes were explored. Results: Approximately half (100 of 203; 49.3%) of the patient samples had high (IA+IS) immune subtypes (Table 1). Targetable alterations associated with an FDA-approved therapy were present in 114 (56.2%) patients. No biomarkers were significantly associated (p < 0.05) with high (IA+IS) versus low (ID+A) immune scores. Biomarkers associated with ICI response, namely mismatch repair gene alterations (MSH2/3/6, MLH1/3, PMS1/2), high tumor mutational burden (TMB-high) and microsatellite instability were detected in only 6 (3.0%), 3 (1.5%) and 1 (0.5%) patient samples respectively, and all but 1, an MSH6 alteration, were in high immune subtype samples. Conclusions: The Xerna TME Panel classified 49.3% of TNBC patient tumors to IA or IS, suggesting they may respond to ICI therapy. Many (56.2%) patient tumors harbored alterations associated with FDA-approved therapies, providing the potential for novel combination therapies. These findings warrant further study and clinical validation in TNBC patients treated with ICI therapy. Table 1. Frequency of actionable biomarkers that were present in at least 10 (5%) TNBC patient samples. Citation Format: Gargi Basu, Janine Lobello, Snehal Thakkar, Jessica Aldrich, Matthew Halbert, Patrick Eimerman, Cynthia Flannery, Nishitha Therala, David Hall, Daniel Pointing, Lea Vohar, Roman Luštrik, Luka Ausec, Mark Uhlik, Seema Iyer, Laura Benjamin, Frederick Baehner. Prevalence of genomic alterations in Xerna tumor microenvironment subtypes in triple negative breast cancer patients [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-06-10.

Abstract PO5-04-05: Clinical validation of the concordance performance of ERBB2 status by single-cells genomics ERBB2 (HER2) amplification Assay in Circulating Tumor Cells (CTCs) from patients with matched HER2 status from metastatic tissue biopsies

Abstract Background Liquid biopsies are a non-invasive diagnostic approach for detecting CTCs that may provide clinically actionable information for treatment decisions for metastatic breast cancer (MBC) patients when a conventional biopsy is infeasible. Blood-based liquid biopsy has the potential advantage that it allows broad sampling of metastatic tumor clones because it is capable of and likely to obtain cellular and genomic contributions from more than one metastatic site. Here we report the concordance performance of HER2 positivity by single-cell CTC genomics ERBB2 (HER2) amplification in patients with matched HER2 status from metastatic tissue biopsies. Methods A prospective Clinical Experience Program for CTC ERBB2 (HER2) Assay across 25 different United States-based cancer centers, was used to recruit 128 MBC patients with documented standard-of-care metastatic tissue biopsies performed within the last 5 years (Days from tissue biopsy to liquid biopsy, Median: 19.5; interquartile range: 40.75) CTCs from blood collection at each respective clinical site were isolated from plasma, nucleated cells were plated, & slides were bio-banked for Immunofluorescent staining & subsequent imaging in a CAP CLIA lab. CTCs were identified using Epic Sciences digital imaging & machine learning algorithms. Individual CTCs were sequenced for genomic quantification of chromosomal instability (LSTs) and ERBB2 (HER2) amplification. Results Within the clinical validation cohort of 128 MBC patients, 30% (38/128) were HER2-positive by standard pathologic criteria (IHC 3+ or IHC2+/ISH+) in metastatic tissue biopsies. Among the 64 out of 128 cases with reportable results by the CTC ERBB2 (HER2) amplification on chromosomally unstable CTCs, 31% (20/64) of patients had ERBB2 amplification. Concordance between CTC single cell genomics and available tissue results for HER2 positivity showed a sensitivity of 69% and specificity of 78%. We then addressed the effects of two known biological confounders to the use of tissue biopsy for HER2 status determination: 1) tissue biopsies performed on bone metastases, 2) tumor evolution driven by treatment pressure over time leading to potential HER2 conversion and tumor heterogeneity. Thus, we performed subgroup analysis including only validation set patients with tissue biopsies performed on non-bone tissues, which showed slightly improved concordance to the tissue comparator with a Sensitivity of 86%, and Specificity of 75%. Next, to exclude the possibility of tumor evolution we selected only patients with the HER2 status comparator performed on non-bone tissue biopsies contemporaneous (Days from tissue biopsy to liquid biopsy, Median: 14; interquartile range: 22) to the blood draw for the CTC ERBB2 (HER2) amplification assay. This showed dramatically improved concordance performance with a Sensitivity of 100%, and a Specificity of 75%. Conclusions Blood and tissue are two biologically distinct sample types where variations in HER2 and other biomarker results might be expected due to analytic, temporal and clinical differences. These data indicate that determination of HER2 status by single cell genomic ERBB2 (HER2) amplification assay can aid in the clinical assessment of ERBB2 status in patients with metastatic breast cancer (MBC) for whom tissue biopsy for HER2 status is not available, or infeasible. Additionally, these results provide evidence of the contribution of site differences (bone) of metastatic biopsies and also tumor evolution to discordance between the CTC ERBB2 (HER2) Assay and tumor biopsy. Citation Format: Giuseppe Di Caro, Ernest Lam, Megan Slade, Shuguang Huang, Rick Wenstrup, Lee Schwartzberg. Clinical validation of the concordance performance of ERBB2 status by single-cells genomics ERBB2 (HER2) amplification Assay in Circulating Tumor Cells (CTCs) from patients with matched HER2 status from metastatic tissue biopsies [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-04-05.

Abstract PO5-18-05: Assessment of the Potential of Photon mini-GRID Therapy for Pre-operative Partial Breast Cancer Treatment

Abstract Purpose We propose a new spatially fractionated radiation therapy (SFRT) technique, called mini-GRID therapy, to reduce normal tissue toxicities in pre-operative partial breast cancer radiotherapy. Mini-GRID therapy, an optimized implementation of GRID therapy, utilizes very narrow beams (widths ~ 1-2 mm) which significantly increase normal tissue tolerances. The concept, proposed by our team in 2017 [1], has been successfully implemented at Hospital de Santiago de Compostela (Spain) showing excellent results in terms of brain toxicity reduction [2]. Profiting from this remarkable increase in tolerance, this study aims to evaluate the feasibility and efficacy of photon mini-GRID therapy for pre-operative partial breast cancer treatment. Methods Ten unbiased clinical cases were selected for the study, comparing photon mini-GRID therapy with conventional broad beam radiotherapy. Monte Carlo simulations (TOPAS v.3.6) were performed based on the mini-GRID therapy implementation (megavoltage X-rays, Varian flattening-filter-free LINAC) realized at the University Hospital in Santiago de Compostela. A mean dose of 24 Gy was delivered in the PTV using three photon mini-GRID arrays with mini beams of approximately 1.8x1.8 mm² at the isocenter. The study aimed to achieve better tumor coverage than conventional pre-operative radiotherapy while ensuring comparable skin toxicity. Valley doses, determining tissue-sparing effects, were compared against organs-at-risk (OARs) tolerance limits. Results The mini-GRID therapy treatments ensured partial tumor coverage and met the dose tolerance limits for all the considered OARs, the most challenging of all being the skin [3]. Peak-to-valley dose ratio (PVDR) in the skin was higher than 3.5 for all the considered patients. Compared to standard radiotherapy (SRT) plans, mini-GRID therapy significantly reduced mean doses to the lungs, heart, and chest wall in the studied cases. Conclusion In this proof-of-concept study, we evaluated photon mini-GRID therapy with megavoltage X-rays as an alternative treatment approach for pre-operative partial breast cancer irradiation, demonstrating its feasibility and straightforward implementation. Because of its high skin and normal tissue-sparing capacities, photon mini-GRID therapy has great potential to improve pre-operative treatment outcomes. Complementary studies and clinical validation are needed to fully establish the optimal protocols and assess long-term effects, but the results encourage the integration of photon mini-GRID therapy as a promising modality for breast cancer therapy.

Abstract PO3-11-09: Cross-cultural adaptation and psychometric validation of lymphedema patient-reported outcome measurements in Chinese breast cancer patients

Abstract Background: Lymphedema is one of the most common complains reported by breast cancer survivors, which negatively affects the quality of life (QoL). Various internationally recognized patient-reported outcome measurements (PROMs), such as the Upper Limb Lymphedema 27 (ULL-27), Lymphoedema Functioning, Disability and Health questionnaire (Lymph-ICF), Lymphedema Quality of Life Questionnaire for upper limbs (LYMQOL-ARM), and Lymphedema Life Impact Scale Version 2 (LLIS), have been translated into different languages and widely used to assess QoL in patients with breast cancer-related lymphedema (BCRL). However, limited evidence has proved the efficiency of these tools in Chinese patients. In this study, we performed cross-cultural adaptation and psychometric validation of these four PROMs within the Chinese context. Additionally, we also compared the accuracy in detecting BCRL for early diagnosis. Methods: A process of forward–backward translation and cultural adaptation was conducted based on international standards. Content validity was assessed through patient interviews. Internal consistency was evaluated by Cronbach’s alpha coefficient. The correlation analysis of the MOS item short from health survey (SF-36) and each BCRL QoL questionnaires was utilized to indicate the convergent and divergent validity. Clinical validity was assessed by comparing scores among patients with different BCRL stages. Furthermore, questionnaires scores and the known group of lymphedema or not were compared to generate receiver operator characteristic (ROC) curves of these instruments. Results: A total of 204 female breast cancer patients were recruited in this study. Among them, 104 patients (61 with lymphedema and 43 without lymphedema) finished LYMQOL-ARM and ULL-27 questionnaire, while 100 patients (59 with lymphedema and 41 without lymphedema) were assessed by Lymph-ICF and LLIS. The internal consistency was high, with Cronbach's alpha coefficients above 0.9 for all subscales and global scales. The content validity was generally good, except for a few issues reported by the patients. Convergent and divergent validity confirmed the hypotheses of correlation between the tested questionnaire and SF-36 to varying degrees: 100% for ULL-27, 80% for Lymph-ICF, nearly 70% for LLIS, and only 50% for LYMQOL-ARM. LLIS and Lymph-ICF demonstrated strong clinical validity, with significant differences found in total and subscale scores among participants in different BCRL stages (P< 0.05). There were no significant differences between stages for the mood dimension (P=0.132) of LYMQOL-ARM, psychological dimension (P=0.506), social dimension (P=0.083), and total score (P=0.057) of ULL-27. Only the physical concerns and global scale of LLIS and the symptom subscale of Lymph-ICF exhibited acceptable AUC values near 0.7 (AUC 0.725, 95% CI 0.625-0.825, p< 0.029; AUC 0.680, 95% CI 0.573-0.787, p< 0.002; AUC 0.734, 95% CI 0.633-0.834, p< 0.001, respectively). In contrast, other subscales and questionnaires displayed lower AUC values. Conclusion: Lymphedema is a chronic disease that has a relatively low mortality risk but significantly affects the quality of life. Therefore, it is crucial to prioritize patient-reported outcomes in the assessment of lymphedema. Among the four PROMs, LLIS not only showed high internal consistency and content validity more suitable for Chinese patients, but also exhibited better performance in reflecting the presence and severity of lymphedema patients. Consequently, the validated and reliable Chinese version of LLIS is recommended for clinical and scientific practices in the assessment of BCRL, which will help to provide better interventions with a patient-centered focus. Citation Format: Yue Zhou, Bingqiu Xiu, Qi Zhang, Jiong Wu. Cross-cultural adaptation and psychometric validation of lymphedema patient-reported outcome measurements in Chinese breast cancer patients [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-11-09.

Clinical validation of automated depth camera-based measurement of the Fugl-Meyer assessment for upper extremity.

Depth camera-based measurement has demonstrated efficacy in automated assessment of upper limb Fugl-Meyer Assessment for paralysis rehabilitation. However, there is a lack of adequately sized studies to provide clinical support. Thus, we developed an automated system utilizing depth camera and machine learning, and assessed its feasibility and validity in a clinical setting. Validation and feasibility study of a measurement instrument based on single cross-sectional data. Rehabilitation unit in a general hospital. Ninety-five patients with hemiparesis admitted for inpatient rehabilitation unit (2021-2023). Scores for each item, excluding those related to reflexes, were computed utilizing machine learning models trained on participant videos and readouts from force test devices, while the remaining reflex scores were derived through regression algorithms. Concurrent criterion validity was evaluated using sensitivity, specificity, percent agreement and Cohen's Kappa coefficient for ordinal scores of individual items, as well as correlations and intraclass correlation coefficients for total scores. Video-based manual assessment was also conducted and compared to the automated tools. The majority of patients completed the assessment without therapist intervention. The automated scoring models demonstrated superior validity compared to video-based manual assessment across most items. The total scores derived from the automated assessment exhibited a high coefficient of 0.960. However, the validity of force test items utilizing force sensing resistors was relatively low. The integration of depth camera technology and machine learning models for automated Fugl-Meyer Assessment demonstrated acceptable validity and feasibility, suggesting its potential as a valuable tool in rehabilitation assessment.

Abstract PO5-14-08: External Reproducibility of PD-L1 IHC 22C3 pharmDx on Breast Cancer Specimens at CPS ≥ 1 and CPS ≥ 10 Cutoffs

Abstract Background: PD-L1 IHC 22C3 pharmDx is an approved companion diagnostic for pembrolizumab treatment in triple-negative breast cancer (TNBC). Clinical evidence for pembrolizumab approval in TNBC was demonstrated by KEYNOTE-355 (NCT02819518), which investigated the clinical validity of PD-L1 IHC 22C3 pharmDx in identifying patients with PD-L1 expressing Combined Positive Score (CPS) ≥ 10 in previously untreated locally recurrent unresectable or metastatic TNBC.1 The role of immune checkpoint inhibitors for breast cancer as a whole is not as well defined as it is for TNBC. In this study, we show evidence of the reproducibility of PD-L1 IHC 22C3 pharmDx in PD-L1 expression determination in breast cancer specimens at the CPS ≥ 1 and CPS ≥ 10 cutoffs documenting robust assay performance in breast cancer. Methods: A blinded, randomized study was conducted at three external sites using formalin-fixed, paraffin-embedded breast cancer specimens. Reproducibility of PD-L1 IHC 22C3 pharmDx was assessed at the inter-site, intra-site/inter-day, inter-observer, and intra-observer endpoints at the CPS ≥ 1 and CPS ≥ 10 cutoffs. To assess inter-site and intra-site reproducibility, five replicate sets of pre-qualified unstained specimens were stained and evaluated at each of the three external sites. To assess inter-observer and intra-observer reproducibility, one set of pre-stained specimens was evaluated three times by each of three external pathologists. Negative percent agreement (NPA), positive percent agreement (PPA), and overall percent agreement (OA), based on comparisons to consensus diagnostic outcome, were computed with corresponding two-sided 95% percentile bootstrap confidence intervals. Acceptance criteria required at least 85.0% for the lower-bound (LB) of the two-sided 95% confidence interval (CI) on NPA, PPA, and OA for all endpoints. Results: All but one endpoint in the external reproducibility study met pre-determined acceptance criteria. Both the inter- and intra-site reproducibility endpoints had 95% CI LB values of 98.8% for NPA, 95.9% for PPA, and 97.9% OA at the CPS ≥ 1 cutoff. At the CPS ≥ 10 cutoff, the inter-site reproducibility endpoint had 95% CI LB values of 88.0% for NPA, 87.1% for PPA, and 89.5% OA, and the intra-site reproducibility endpoint had 95% CI LB values of 96.7% for NPA, 92.2% for PPA, and 95.6% OA. The inter-observer reproducibility endpoint had 95% CI LB values of 95.3% for NPA, 92.3% for PPA, and 94.7% OA at the CPS ≥ 1 cutoff. At the CPS ≥ 10 cutoff, the inter-observer reproducibility endpoint had 95% CI LB values of 84.9% for NPA, 86.8% for PPA, and 87.3% OA. The intra-observer reproducibility endpoint had 95% CI LB values of 97.0% for NPA, 95.0% for PPA, and 96.6% OA at the CPS ≥ 1 cutoff. At the CPS ≥ 10 cutoff, the intra-observer reproducibility endpoint had 95% CI LB values of 92.2% for NPA, 89.2% for PPA, and 91.7% OA. Conclusions: Acceptance criteria were met for all endpoints in the breast cancer external reproducibility study of PD-L1 IHC 22C3 pharmDx at the CPS ≥ 1 and CPS ≥ 10 cutoffs, except for inter-observer at the CPS ≥ 10 cutoff, which had a 95% CI LB value of 84.9% for NPA.

Speech Biomarkers for Dysarthria in Parkinson's Disease — Clinical Validation Across Spanish and Czech Languages

Speech Biomarkers for Dysarthria in Parkinson's Disease — Clinical Validation Across Spanish and Czech Languages

Exosome secretion related gene signature predicts chemoresistance in patients with colorectal cancer

BackgroundColorectal cancer (CRC) is a highly heterogeneous malignancy, and patients often have different responses to treatment. In this study, the genetic characteristics related to exosome formation and secretion procedure were used to predict chemoresistance and guide the individualized treatment of patients. MethodsFirstly, seven microarray datasets in Gene Expression Omnibus (GEO) and RNA-Seq dataset from the Cancer Genome Atlas (TCGA) were used to analysis the transcriptome profiles and associated characteristics of CRC patients. Then, a predictive model based on gene features linked to exosome formation and secretion was created and validated using Least Absolute Shrinkage and Selection Operator (LASSO) regression analysis and Support Vector Machine-Recursive Feature Elimination (SVM-RFE) machine learning. Finally, we evaluated the model using chemoresistant/chemosensitive cells and tissues by immunofluorescence (IF), western blot (WB), quantitative real-time PCR (qRT-PCR) and immunocytochemistry (IHC) experiments, and the predictive value of integrated model in the clinical validation cohort were performed by Receiver Operating Characteristic (ROC) and Kaplan-Meier (K-M) curves analyses. ResultsWe established a risk score signature based on three genes related to exosome secretion in CRC. Better Overall Survival (OS) and greater chemosensitivity were seen in the low-risk group, whereas the high-risk group exhibited chemoresistance and a subpar response to immune checkpoint blockade (ICB) therapy. Higher expression of the model genes EXOC2, EXOC3 and STX4 were observed in chemoresistant cells and specimens. The AUC of 5-year disease-free survival (DFS) was 0.804. Compared with that in the low-risk group, patients’ DFS was found to be significantly worse in the high-risk group. ConclusionsIn summary, the gene signature related to exosome formation and secretion could reliably predict patients' chemosensitivity and ICB treatment response, which providing new independent biomarkers for the treatment of CRC.

Anticancer Drug Development: Evaluative Architecture

Abstract: Anticancer drug development is complex and multi-factorial, demanding robust drug evaluative architecture in experimental and preclinical studies. To increase the number of drug licensing, biochemical, pharmacological, technical and economic changes (balance and integration) in evaluative systems should be focused in labs throughout the world. Despite great progress, treatment breakthroughs and drug industry need new ideas and more competitive technology (new generation of in vitro test systems). Overall, drug evaluative systems comprise anticancer drug development (medicinal chemistry and pharmacology) from initial screening to clinical validity. Its advances greatly impact the outcomes of drug production (rates of final drug licensing and efficacy of therapeutics in the clinic). In this regard, creative ideas and new techniques will change the norm and convention of drug screening and mechanic exploration in the future. This article provides multi-disciplinary approaches for experimental and preclinical anticancer drug evaluations, selections and combinations (chemistry and biomedicine). Future trends for drug evaluative systems are especially highlighted in in-depth, multilateral and multidisciplinary approaches.

Biomechanical comparisons of F.E.R.I. techniques with different type of intramedullary screws fixation for Jones fractures

Biomechanical comparisons of F.E.R.I. techniques with different type of intramedullary screws fixation for Jones fractures

Assessing personalized responses to anti-PD-1 treatment using patient-derived lung tumor-on-chip.

There is a compelling need for approaches to predict the efficacy of immunotherapy drugs. Tumor-on-chip technology exploits microfluidics to generate 3D cell co-cultures embedded in hydrogels that recapitulate simplified tumor ecosystems. Here, we present the development and validation of lung tumor-on-chip platforms to quickly and precisely measure exvivo the effects of immune checkpoint inhibitors on Tcell-mediated cancer cell death by exploiting the power of live imaging and advanced image analysis algorithms. The integration of autologous immunosuppressive FAP+ cancer-associated fibroblasts impaired the response to anti-PD-1, indicating that tumors-on-chips are capable of recapitulating stroma-dependent mechanisms of immunotherapy resistance. For a small cohort of non-small cell lung cancer patients, we generated personalized tumors-on-chips with their autologous primary cells isolated from fresh tumor samples, and we measured the responses to anti-PD-1 treatment. These results support the power of tumor-on-chip technology in immuno-oncology research and open a path to future clinical validations.