Monitoring Of Therapeutic Response Research Articles

Leukocyte Heterogeneity in Pancreatic Ductal Adenocarcinoma: Phenotypic and Spatial Features Associated with Clinical Outcome.

Immunotherapies targeting aspects of T cell functionality are efficacious in many solid tumors, but pancreatic ductal adenocarcinoma (PDAC) remains refractory to these treatments. Deeper understanding of the PDAC immune ecosystem is needed to identify additional therapeutic targets and predictive biomarkers for therapeutic response and resistance monitoring. To address these needs, we quantitatively evaluated leukocyte contexture in 135 human PDACs at single-cell resolution by profiling density and spatial distribution of myeloid and lymphoid cells within histopathologically defined regions of surgical resections from treatment-naive and presurgically (neoadjuvant)-treated patients and biopsy specimens from metastatic PDAC. Resultant data establish an immune atlas of PDAC heterogeneity, identify leukocyte features correlating with clinical outcomes, and, through an in silico study, provide guidance for use of PDAC tissue microarrays to optimally measure intratumoral immune heterogeneity. Atlas data have direct applicability as a reference for evaluating immune responses to investigational neoadjuvant PDAC therapeutics where pretherapy baseline specimens are not available. SIGNIFICANCE: We provide a phenotypic and spatial immune atlas of human PDAC identifying leukocyte composition at steady state and following standard neoadjuvant therapies. These data have broad utility as a resource that can inform on leukocyte responses to emerging therapies where baseline tissues were not acquired.This article is highlighted in the In This Issue feature, p. 1861.

Diffusion histology imaging differentiates distinct pediatric brain tumor histology

High-grade pediatric brain tumors exhibit the highest cancer mortality rates in children. While conventional MRI has been widely adopted for examining pediatric high-grade brain tumors clinically, accurate neuroimaging detection and differentiation of tumor histopathology for improved diagnosis, surgical planning, and treatment evaluation, remains an unmet need in their clinical management. We employed a novel Diffusion Histology Imaging (DHI) approach employing diffusion basis spectrum imaging (DBSI) derived metrics as the input classifiers for deep neural network analysis. DHI aims to detect, differentiate, and quantify heterogeneous areas in pediatric high-grade brain tumors, which include normal white matter (WM), densely cellular tumor, less densely cellular tumor, infiltrating edge, necrosis, and hemorrhage. Distinct diffusion metric combination would thus indicate the unique distributions of each distinct tumor histology features. DHI, by incorporating DBSI metrics and the deep neural network algorithm, classified pediatric tumor histology with an overall accuracy of 85.8%. Receiver operating analysis (ROC) analysis suggested DHI’s great capability in distinguishing individual tumor histology with AUC values (95% CI) of 0.984 (0.982–0.986), 0.960 (0.956–0.963), 0.991 (0.990–0.993), 0.950 (0.944–0.956), 0.977 (0.973–0.981) and 0.976 (0.972–0.979) for normal WM, densely cellular tumor, less densely cellular tumor, infiltrating edge, necrosis and hemorrhage, respectively. Our results suggest that DBSI-DNN, or DHI, accurately characterized and classified multiple tumor histologic features in pediatric high-grade brain tumors. If these results could be further validated in patients, the novel DHI might emerge as a favorable alternative to the current neuroimaging techniques to better guide biopsy and resection as well as monitor therapeutic response in patients with high-grade brain tumors.

Landscape of circulating tumor DNA (ctDNA) abnormalities in advanced prostate cancer (aPCa): Distinctions in African American (AA) versus Caucasian (Ca) patients.

156 Background: AA have substantially higher prostate cancer incidence rates, are diagnosed at a younger age and with a more advanced stage as compared to Ca. However, after adjusting for known prognostic factors, AA have an increased overall survival. We hypothesized that these differences might be due to the underlying changes in the genomic landscape which can be revealed by liquid biopsy. Methods: Real world comprehensive genomic profiling of ctDNA from aPCa patients from two institutions. The first ctDNA results as reported by Guardant 360 panel (Redwood City, CA) were included. Association between genetic mutation and gene were tested using Barnard’s test. To account for multiple testing, we used Benjamini-Hochberg’s False Discovery Rate adjustment across all tests to determine thresholds for false discovery rates. Same analysis was performed using a Bayesian Network Machine learning approach. Results: Overall, 361 patients with aPCa (81 AA and 280 Ca) were included in the analysis. Pathogenic genomic alterations were found in 87.0% of the cases, more frequently TP53 (42.4%), AR (34.1%), PIK3CA (13.9%), BRAF (12.7%), NF1 (10.8%) and MYC (10.0%). Targetable alterations of interest included DNA repair genes [BRCA 2 (7.8%), BRCA 1 (4.4%), ATM (6.4%), CDK12 (2.2%)], PIK3CA/mTOR/AKT (19.1%), PTEN (3.3%) and NTRK (1.9%). MSI-high was found in 4 patients. AA as compared to Ca had a significantly higher prevalence of CDK12 (20.7% vs. 3.8%, p=0.016) and GNA11 mutations (3.7% vs. 0.4%, p=0.0225). BayesNet analysis also supported these results (table). Conclusions: In this dataset, liquid biopsy of ctDNA was useful for genetic characterization of aPCa and reveal differences in the molecular phenotype of AA and Ca in aPCa with potential clinical implications. These findings support ongoing research on the clinical utility of non-invasive genotyping and therapeutic response monitoring with a focus on AA population. [Table: see text]

Histiocytosis and Neoplasms of Macrophage-Dendritic Cell Lineages: Multimodality Imaging with Emphasis on PET/CT.

Histiocytosis is a rare inflammatory process characterized by pathologic infiltration and accumulation of cells derived from the monocytic lineage in normal tissue. It encompasses more than 100 different subtypes of disorders that were recently classified into five main groups: (a) Langerhans-related histiocytosis, (b) Rosai-Dorfman histiocytosis, (c) cutaneous and mucocutaneous histiocytosis, (d) malignant histiocytosis, and (e) hemophagocytic lymphohistiocytosis and macrophage activation syndrome. Langerhans cell histiocytosis is the most common histiocytic disorder. Less common types include Erdheim-Chester disease, Rosai-Dorfman disease, adult and juvenile xanthogranuloma, necrobiotic xanthogranuloma, histiocytic sarcoma, interdigitating dendritic cell sarcoma, Langerhans cell sarcoma, and hemophagocytic lymphohistiocytosis. Although the pathogenesis of these disorders may be attributable to mutations in the oncogenic driver, recent discoveries have shown that inflammation and fibrosis secondary to mutated histiocytes, rather than a proliferative cell mechanism, result in manifestation of the disease. Diagnosis, which relies on a multidisciplinary approach, is challenging and often delayed because clinical findings are nonspecific and may mimic malignant processes at radiologic evaluation. Compared with conventional imaging, PET/CT allows detection of the increased metabolic activity in histiocytes. Diagnostic algorithms for histiocytic disorders should include functional imaging with fluorine 18 (18F) fluorodeoxyglucose (FDG) PET/CT, which provides a comprehensive whole-body evaluation of their potential involvement with multiple organ systems and allows monitoring of therapeutic response. The most recent revised classification, pathophysiologic and clinical manifestations, sites of involvement, and imaging features of histiocytosis are described in this review and a multimodality approach is used, with emphasis on 18F-FDG PET/CT evaluation. ©RSNA, 2021.

Epigenetic Landscape of Liquid Biopsy in Colorectal Cancer.

Colorectal cancer (CRC) is one of the most common malignancies and is a major cause of cancer-related deaths worldwide. Thus, there is a clinical need to improve early detection of CRC and personalize therapy for patients with this disease. In the era of precision oncology, liquid biopsy has emerged as a major approach to characterize the circulating tumor elements present in body fluids, including cell-free DNA and RNA, circulating tumor cells, and extracellular vesicles. This non-invasive tool has allowed the identification of relevant molecular alterations in CRC patients, including some indicating the disruption of epigenetic mechanisms. Epigenetic alterations found in solid and liquid biopsies have shown great utility as biomarkers for early detection, prognosis, monitoring, and evaluation of therapeutic response in CRC patients. Here, we summarize current knowledge of the most relevant epigenetic mechanisms associated with cancer development and progression, and the implications of their deregulation in cancer cells and liquid biopsy of CRC patients. In particular, we describe the methodologies used to analyze these epigenetic alterations in circulating tumor material, and we focus on the clinical utility of epigenetic marks in liquid biopsy as tumor biomarkers for CRC patients. We also discuss the great challenges and emerging opportunities of this field for the diagnosis and personalized management of CRC patients.

Real-Time Quantification of Cartilage Degeneration by GAG-Targeted Cationic Nanoparticles for Efficient Therapeutic Monitoring in Living Mice.

One of the characterizations of degenerative cartilage disease is the progressive loss of glycosaminoglycans (GAGs). The real-time imaging method to quantify GAGs is of great significance for the biochemical analysis of cartilage and diagnosis and therapeutic monitoring of cartilage degeneration in vivo. To this end, a cationic photoacoustic (PA) contrast agent, poly-l-lysine melanin nanoparticles (PLL-MNPs), specifically targeting anionic GAGs was developed in this study to investigate whether it can image cartilage degeneration. PLL-MNP assessed GAG depletion by Chondroitinase ABC in vitro rat cartilage and intact ex vivo mouse knee joint. A papain-induced cartilage degenerative mice model was used for in vivo photoacoustic imaging (PAI). Oral cartilage supplement glucosamine sulfate was intragastrically administered for mice cartilage repair and the therapeutic efficacy was monitored by PLL-MNP-enhanced PAI. Histologic findings were used to further confirm PAI results. In vitro results revealed that the PLL-MNPs not only had a high binding ability with GAGs but also sensitively monitored GAG content changes by PAI. The PA signal was gradually weakened along with the depletion of GAGs in cartilage. Particularly, PLL-MNPs depicted the cartilage structure and the distribution of GAGs was demonstrated in PA images in ex vivo joints. Compared with the normal joint, a lower signal intensity was detected from degenerative joint at 3 weeks after papain injection, suggesting an early diagnosis of cartilage lesion by PLL-MNPs. Importantly, this PA-enhanced nanoprobe was suitable for monitoring in vivo efficacy of glucosamine sulfate, which effectively blocked cartilage degradation in a high dose manner. In vivo imaging findings correlated well with histological examinations. PLL-MNPs provided sensitive visualization of cartilage degeneration and promising monitoring of therapeutic response in living subjects.

What Is Known about Theragnostic Strategies in Colorectal Cancer.

Despite the paradigmatic shift occurred in recent years for defined molecular subtypes in the metastatic setting treatment, colorectal cancer (CRC) still remains an incurable disease in most of the cases. Therefore, there is an urgent need for new tools and biomarkers for both early tumor diagnosis and to improve personalized treatment. Thus, liquid biopsy has emerged as a minimally invasive tool that is capable of detecting genomic alterations from primary or metastatic tumors, allowing the prognostic stratification of patients, the detection of the minimal residual disease after surgical or systemic treatments, the monitoring of therapeutic response, and the development of resistance, establishing an opportunity for early intervention before imaging detection or worsening of clinical symptoms. On the other hand, preclinical and clinical evidence demonstrated the role of gut microbiota dysbiosis in promoting inflammatory responses and cancer initiation. Altered gut microbiota is associated with resistance to chemo drugs and immune checkpoint inhibitors, whereas the use of microbe-targeted therapies including antibiotics, pre-probiotics, and fecal microbiota transplantation can restore response to anticancer drugs, promote immune response, and therefore support current treatment strategies in CRC. In this review, we aim to summarize preclinical and clinical evidence for the utilization of liquid biopsy and gut microbiota in CRC.

Circulating free DNA in the plasma of individuals with neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is an autosomal dominant syndrome whose characteristic manifestations include benign neurofibromas, yet NF1 is also associated with a high risk of cancer. Measurements of circulating free plasma DNA (cfDNA) are gaining wider applicability in cancer diagnostics, targeting of therapy, and monitoring of therapeutic response. Individuals with NF1 are likely to be followed up using this method, but the effects of NF1 and neurofibromas on cfDNA levels are not known. We studied peripheral blood samples from 19 adults with NF1 and 12 healthy controls. The cfDNA was isolated from plasma with QIAamp Circulating Nucleic Acid Kit and quantified using the Qubit 2.0 Fluorometer. The cfDNA concentration of each sample was normalized relative to the plasma protein concentration. The normalized median concentration of cfDNA in plasma was 19.3 ng/ml (range 6.6-78.6) among individuals with NF1 and 15.9 ng/ml (range 4.8-47.0) among controls (p = .369). Individuals with NF1 who also had plexiform neurofibroma (pNF) showed non-significantly elevated cfDNA concentration compared to individuals with NF1 and without known pNF (median 25.4 vs. 18.8 ng/ml, p = .122). The effect of NF1 on cfDNA seems to be relatively small and NF1 is therefore unlikely to hamper the use of cfDNA-based assays.

Rational Design of Albumin Theranostic Conjugates for Gold Nanoparticles Anticancer Drugs: Where the Seed Meets the Soil?

Multifunctional gold nanoparticles (AuNPs) may serve as a scaffold to integrate diagnostic and therapeutic functions into one theranostic system, thereby simultaneously facilitating diagnosis and therapy and monitoring therapeutic responses. Herein, albumin-AuNP theranostic agents have been obtained by conjugation of an anticancer nucleotide trifluorothymidine (TFT) or a boron-neutron capture therapy drug undecahydro-closo-dodecaborate (B12H12) to bimodal human serum albumin (HSA) followed by reacting of the albumin conjugates with AuNPs. In vitro studies have revealed a stronger cytotoxicity by the AuNPs decorated with the TFT-tagged bimodal HSA than by the boronated albumin conjugates. Despite long circulation time, lack of the significant accumulation in the tumor was observed for the AuNP theranostic conjugates. Our unique labelling strategy allows for monitoring of spatial distribution of the AuNPs theranostic in vivo in real time with high sensitivity, thus reducing the number of animals required for testing and optimizing new nanosystems as chemotherapeutic agents and boron-neutron capture therapy drug candidates.

Sleep Disturbances Associated with Neurological Autoimmunity.

Associations between sleep disorders and neurological autoimmunity have been notably expanding recently. Potential immune-mediated etiopathogenesis has been proposed for various sleep disorders including narcolepsy, Kleine-Levin syndrome, and Morvan syndrome. Sleep manifestations are also common in various autoimmune neurological syndromes, but may be underestimated as overriding presenting (and potentially dangerous) neurological symptoms often require more urgent attention. Even so, sleep dysfunction has been described with various neural-specific antibody biomarkers, including IgLON5; leucine-rich, glioma-inactivated protein 1 (LGI1); contactin-associated protein 2 (CASPR2); N-methyl-D-aspartate (NMDA)-receptor; Ma2; dipeptidyl-peptidase-like protein-6 (DPPX); alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPA-R); anti-neuronal nuclear antibody type-1 (ANNA-1, i.e., Hu); anti-neuronal nuclear antibody type-2 (ANNA-2, i.e., Ri); gamma-aminobutyric acid (GABA)-B-receptor (GABA-B-R); metabotropic glutamate receptor 5 (mGluR5); and aquaporin-4 (AQP-4). Given potentially distinctive findings, it is possible that sleep testing could potentially provide objective biomarkers (polysomnography, quantitative muscle activity during REM sleep, cerebrospinal fluid hypocretin-1) to support an autoimmune diagnosis, monitor therapeutic response, or disease progression/relapse. However, more comprehensive characterization of sleep manifestations is needed to better understand the underlying sleep disruption with neurological autoimmunity.

Renal Cell Carcinoma Mimicking with Peritoneal Carcinomatosis and Krukenberg Tumor: Diagnosis Seen on Fluorodeoxyglucose Positron Emission Tomography-Computed Tomography.

Krukenberg tumor (KT) is a rare clinical entity with a mysterious origin. It originates most commonly from adenocarcinoma of the stomach. We present an interestingly rare case of this entity in renal cell carcinoma, revealed by 18F-fluorodeoxyglucose-positron emission tomography/computed tomography (PET/CT) scan. Ovarian cancers with diffuse peritoneal metastasis were considered the differential diagnosis of the disease, based on PET/CT. The potential efficacy of this functional imaging for KT is still in the exploratory phase, but its applications in diagnosis, disease prognostication, therapeutic response monitoring, and follow-up recurrence detection are superior than other imaging modalities.

Synergic combination of paclitaxel and irinotecan loaded poly(lactic-co-glycolic acid) nanocomplexes for MRI and the treatment of endometrial cancer

We have established dual paclitaxel (PTX)/superparamagnetic iron oxide (SPIO)-loaded PLGA-based nanoparticles for theranostic purposes. The fabricated nanoparticles exhibited a spherical morphology with a size of ∼240 nm. The PTX and iron loading were 1.84 ± 0.4 and 10.4 ± 1.93 mg/100 mg, respectively. The relaxometry studies and phantom magnetic resonance imaging demonstrated their efficacy as T2 contrast agents. A significant cellular uptake of nanoparticles by the endometrial Ishikawa cells was demonstrated using confocal laser scanning microscopy. While SPIO did not show any toxicity in endometrial Ishikawa cells, PTX-loaded nanoparticles indicated cytotoxic activity. Together, these advantages of multifunctional nanoparticles may be considered as future nanomedicines for simultaneous molecular imaging, drug delivery, and real-time monitoring of therapeutic response.

A combination of surgery, theranostics, and liquid biopsy - a personalised oncologic approach to treatment of patients with advanced metastatic neuroendocrine neoplasms.

Rationale: Neuroendocrine neoplasia (NEN) of small bowel (SBNEN) frequently present with metastatic disease. Theranostics (molecular imaging followed by targeting therapy) allow for personalised medicine. Liquid biopsies enable precise identification of residual disease and real-time monitoring of therapeutic response. Our aim was to determine the clinical utility of a combination of surgery, theranostics, and a multigene blood measurement in metastasised SBNEN.Methods: Inclusion criteria were SBNEN, G1/G2 NEN, initial tumour diagnosis, stage IV NEN, positivity on 68Ga somatostatin analogue PET/CT, eligible for surgery, and 177Lu peptide receptor radionuclide therapy (PRRT). Blood samples for NETest were collected longitudinally. Progression-free survival (PFS) and overall survival (OS) were calculated. NETest results were assessed prior to surgery and during clinical follow-up.Results: A surgical cohort of 39 SBNEN patients met eligibility criteria. Thirty-two patients underwent ileal resection and 7 right hemicolectomy. The mean number of 177Lu PRRT cycles was 4. Mortality was nil. Surgical morbidity was 10.3%. Transient grade 1/2 toxicity occurred in 41% (PRRT). NETest scores (n=9 patients) decreased in 100% following treatment and correlated with diminished tumour volume and disease stabilization following surgery and PRRT. Median follow-up: 78 months. Median PFS and OS: 42.7 and 110 months, respectively. Progression-free survival at 1-, 3-, and 5-years was 79.4%, 57.1% and 40.5%, respectively. Overall survival at 1-, 3-, and 5-years was 97.4%, 97.4%, and 94.1%, respectively.Conclusions: Surgery combined with 177Lu PRRT is safe and provides favourable PFS and OS in selected patients with advanced SBNEN. Liquid biopsy (NETest) has the potential to accurately delineate disease status.

Therapeutic response monitoring after targeted therapy in an orthotopic rat model of hepatocellular carcinoma using contrast-enhanced ultrasound: Focusing on inter-scanner, and inter-operator reproducibility.

PurposeTo assess therapeutic response monitoring after targeted therapy in an orthotopic rat model of hepatocellular carcinoma (HCC) using CEUS with focusing on inter-scanner and inter-operator reproducibility.Materials and methodsFor reproducibility, CEUS was performed using two different US scanners by two operators in sixteen rat models of HCC. Using perfusion analysis software (VueBox ®), eleven parameters were collected, and intra-class correlation coefficient (ICC) was used to analyze reproducibility. Then seventeen rat models of HCC were divided into treatment group (n = 8, 30 mg/kg/day sorafenib for five days) and control group (n = 9). CEUS was performed at baseline and 14 days after first treatment, and changes of perfusion parameters were analyzed.ResultsIn treatment group, CEUS perfusion parameters showed a significant change. The peak enhancement (PE, 2.50 x103±1.68 x103 vs 5.55x102±4.65x102, p = 0.010) and wash-in and wash out AUC (WiWoAUC, 1.07x105±6.48 x104 vs 2.65x104±2.25x104, p = 0.009) had significantly decreased two weeks after treatment. On the contrary, control group did not show a significant change, including PE (1.15 x103±7.53x102 vs 9.43x102± 7.81 x102, p = 0.632) and WiWoAUC (5.09 x104±3.25x104 vs 5.92 x104±3.20x104, p = 0.646). For reproducibility, the various degrees of inter-scanner reproducibility were from poor to good (ICC: <0.01–0.63). However, inter-operator reproducibility of important perfusion parameters, including WiAUC, WoAUC, and WiWoAUC, ranged from fair to excellent (ICC: 0.59–0.93) in a different scanner.ConclusionOur results suggest that CEUS is useful for assessment of the treatment response after targeted therapy and with fair to excellent inter-operator reproducibility.

Transcranial detection of amyloid‐beta at single plaque resolution in vivo with large‐field multifocal illumination fluorescence microscopy

AbstractBackgroundThe abnormal deposition of beta‐amyloid proteins in the brain is one of the major histopathological hallmarks of Alzheimer’s disease. Currently available intravital microscopy techniques for high‐resolution plaque visualization commonly involve highly invasive procedures and are limited to small field‐of‐views within the rodent brain.MethodWe devised a large‐field multi‐focal illumination (LMI) fluorescence microscopy method that provides a 20 × 20 mm field‐of‐view and an axial resolution of ∼15 mm. In vivo and ex vivo transcranial LMI fluorescence microscopy and wide‐field fluorescence microscopy for amyloid deposits were performed in APP/PS1 and arcAb mouse models of Alzheimer’s disease amyloidosis using luminescent conjugated oligothiophene probe HS‐169. Immunohistochemical staining with HS‐169, anti‐Ab antibody 6E10, and conformation antibodies OC (fibrillar) of brain tissue sectionsResultLMI fluorescence microscopy detected amyloid‐beta deposits at a single plaque level in APP/PS1 and arcAb mice with high sensitivity and specificity. The contrast‐to‐noise ratio was approximately 24 times higher in LMI fluorescence microscopy of amyloid deposits compared to wide‐field fluorescence microscopy. Immunohistochemical staining showed HS‐169 co‐localized with 6E10 and OC stained compact parenchymal and vessel‐associated amyloid deposits.ConclusionThe novel LMI in vivo amyloid imaging platform offers new prospects for studies into Alzheimer’s disease mechanisms in animal models as well as longitudinal monitoring of therapeutic responses at a single plaque level.

Dynamic analysis of circulating tumor DNA to predict prognosis and monitor therapeutic response in metastatic relapsed cervical cancer.

Limited and inefficient treatment options exist for metastatic relapsed cervical cancer (MRCC), and there are currently no reliable indicators to guide therapeutic selection. We performed deep sequencing analyses targeting 322 cancer-related genes in plasma cell-free DNA and matched white blood cells in 173 serial blood samples from 82 locally advanced CC (LACC) or MRCC patients and when possible during treatment. We identified five notable nonsynonymous mutant genes (PIK3CA, BRAF, GNA11, FBXW7 and CDH1) in the MRCC samples as the metastatic relapse significantly mutated (MSG) genes and found that MRCC patients with any detectable MSG mutations had significantly shorter progression-free survival (PFS) (P = .005) and overall survival (OS) (P = .007) times than those without detectable MSG mutations. Additionally, analyses of matched prechemotherapy and postchemotherapy plasma revealed that a reduction in the number of MSG mutations after chemotherapy was significantly associated with partial remission (PR) and stable disease (SD) (P = .007). Among the patients included in the longitudinal tracking ctDNA analysis, an increase in MSG mutations was observed earlier in response to disease progression than radiological imaging. Our results outline the mutation profiles of MRCC. We show how longitudinal monitoring with ctDNA in liquid biopsy samples provides both predictive and prognostic information during treatment.

Pictorial review of the clinical applications of MIBG in neuroblastoma: current practices

Neuroblastoma is the most common extra-cranial solid tumor in pediatrics. The objective of this manuscript is to present an image-rich review of the presentation, diagnosis, staging and response assessment in individuals with neuroblastoma with special attention to the role of I-123 mIBG (meta-iodobenzylguanidine). A literature search was performed in the PubMed and Scopus databases to identity articles published between 1980 and March 2020 regarding different aspects of neuroblastoma (presentation, diagnosis, pathologic classification, staging, and response monitoring). Different combinations of search terms were used including “neuroblastoma”, “presentation”, “diagnosis”, “staging”, “imaging”, “pathology”, “MIBG”, “FDG”, “norepinephrine transporter”, and “18F DOPA”. The references of the resultant articles were also used to identify additional manuscripts. A total of 101 key articles were identified that discussed the spectrum of presentation, diagnosis, staging and response assessment in individuals with neuroblastoma. These articles form the basis of this manuscript. Iodine-123-labeled meta-iodobenzylguanidine (mIBG) is the cornerstone of functional imaging for staging, therapeutic response monitoring, prognostication, and the detection of recurrence in patients with neuroblastoma. MIBG images are interpreted both qualitatively and semiquantitatively. The International Neuroblastoma Risk Group Staging System and the International Neuroblastoma Staging System are used to stage the disease extent before and after surgical resection, respectively. The Revised International Neuroblastoma Response Criteria, updated in 2017, stratify the treatment response based on findings on functional (with mIBG or if mIBG non-avid with [18F]-fluorodeoxyglucose) and anatomical (computed tomography or magnetic resonance) imaging and bone marrow aspirates.

Ultrasensitive Measurable Residual Disease (MRD) Detection in Acute Myeloid Leukemia (AML) Using a Targeted Next Generation Sequencing (NGS) Panel

INTRODUCTION: Assessment of MRD in AML is emerging as an important tool for refining risk classification and improving patient management and outcomes. MRD also shows promise as a surrogate endpoint to accelerate therapeutic development and may facilitate more robust post-transplant surveillance. Highly sensitive digital PCR (dPCR) methods such as BEAMing - which are 50-100X more sensitive than "pan-heme" NGS assays - have generated compelling data for therapeutic response monitoring and clearance of molecular MRD in clinical trials: for example, for both relapsed/refractory as well as newly diagnosed patients, declining levels and eventual clearance of IDH1 mutations correlated with more robust hematologic response (DiNardo CD et al. N Engl J Med. 2018, 378(25):2386-98; Daigle S et al. Presented at ASH 2019). Nevertheless, the complex clonal dynamics of AML demand broader coverage across more genomic regions than are currently able to be interrogated using dPCR. At the same time, NGS pan-heme panels that offer broad genomic coverage lack sufficient sensitivity to detect MRD, as their limits of detection (LoD) are between 1-5% mutant allele frequency (MAF). To overcome these limitations, we developed a highly sensitive assay based on the Safe-SeqS technology (SafeSEQ) to detect de novo mutations across genomic regions with established and emerging clinical validity for AML MRD. This highly sensitive error-corrected NGS-based workflow detects molecular MRD with an LoD comparable to dPCR assays such as BEAMing. METHODS: A multiplexed enrichment panel targeting 68 highly relevant genomic regions was developed for ultra-sensitive mutation detection across 57 exons spanning 20 genes: BCOR, BRAF, CEBPA, FLT3, GATA2, IDH1, IDH2, JAK2, KIT, KRAS, NPM1, NRAS, PRPF8, PTPN11, SETBP1, SF3B1, SRSF2, TP53, U2AF1, and ZRSR2. Initial analytical performance demonstration for the SafeSEQ AML MRD test examined limit of blank (LoB) as well as LoD at the 95% confidence level (LoD95). LoB was measured using wildtype (wt) gDNA prepared from two NIST cell lines run in replicate; LoD was determined through replicate testing of contrived materials containing 30 characterized mutations across different mutant molecule (MM)/ MAF tiers ranging from 20 MM down to &amp;lt;1 MM. Low and high DNA input levels were also tested, from 1,000 genomic equivalents (GE) up to a subset of tiers run at 20,000 GE (3.3-66 ng) to demonstrate performance irrespective of wt DNA amount. Direct evaluation of analytical and clinical performance between the SafeSEQ AML MRD panel and a subset of mutations detected using BEAMing is currently underway. RESULTS: A rank-based approach was used to evaluate the distribution of background signals observed in the wt cell line DNA across all interrogated base positions. Consistent with previous observations of LoB across different SafeSEQ panels, a very low level of background noise was observed, with the vast majority of bases exhibiting signal &amp;lt;1 MM, indicating robust analytical specificity. Importantly, there were no positions affected by elevated noise for which clinically relevant mutations have been characterized for AML. For the variant-positive contrived materials run in replicate across different mutation levels, hit rates were consistent with those observed in other SafeSEQ assays and yielded 95% detection at 5 MM; this corresponds to 0.025% and 0.5% MAF for 20,000 and 1,000 GE inputs, respectively. Hit rates for the different MM tiers were also consistent between low and high DNA input levels, indicating robust performance for mutation detection across a wide range of background wt DNA. CONCLUSIONS: The SafeSEQ AML MRD assay demonstrates ultra-sensitive detection of low frequency mutations, with an LoD95 of 5 MM (0.025% MAF for 20,000 GE DNA input), while specificity remains very high. Similar to other SafeSEQ platform configurations, this performance is comparable with data obtained using the BEAMing dPCR technology; it is also orders of magnitude more sensitive than broad pan-heme NGS tests. With this combination of ultra-high analytical sensitivity paired with clinically-informed genomic coverage, we anticipate the SafeSEQ AML MRD test will play a fundamental role in accelerating therapeutic development. It should also provide clinicians with more reliable MRD information at important decision points, such as pre-transplant, or when considering novel maintenance therapies. Disclosures Sloane: Sysmex Inostics, Inc: Current Employment.

Rising Circulating Tumor DNA As a Molecular Biomarker of Early Disease Progression in Metastatic Breast Cancer

Accurate monitoring of therapeutic response remains an important unmet need for patients with metastatic breast cancer (MBC). Analysis of tumor genomics obtained via circulating tumor DNA (ctDNA) can provide a comprehensive overview of tumor evolution. Here, we evaluated ctDNA change as an early prognostic biomarker of subsequent radiologic progression and survival in MBC. Paired blood samples from patients with MBC were analyzed for levels of ctDNA, carcinoembryonic antigen, and cancer antigen 15-3 at baseline and during treatment. A Clinical Laboratory Improvement Amendments-certified sequencing panel of 73 genes was used to quantify tumor-specific point mutations in ctDNA. Multivariable logistic regression analysis was conducted to evaluate the association between ctDNA rise from baseline to during-treatment (genomic progression) and subsequent radiologic progression and progression-free survival (PFS). Somatic mutations were detected in 76 baseline samples (90.5%) and 71 during-treatment samples (84.5%). Patients with genomic progression were more than twice as likely to have subsequent radiologic progression (odds ratio, 2.04; 95% CI, 1.74 to 2.41; P < .0001), with a mean lead time of 5.8 weeks. Genomic assessment provided a high positive predictive value of 81.8% and a negative predictive value of 89.7%. The subset of patients with genomic progression also had shorter PFS (median, 4.2 v 8.3 months; hazard ratio, 2.97; 95% CI, 1.75 to 5.04; log-rank P < .0001) compared with those without genomic progression. Genomic progression, as assessed by early rise in ctDNA, is an independent biomarker of disease progression before overt radiologic or clinical progression becomes evident in patients with MBC.

Screen-Printed Sensor for Low-Cost Chloride Analysis in Sweat for Rapid Diagnosis and Monitoring of Cystic Fibrosis.

Analysis of sweat chloride levels in cystic fibrosis (CF) patients is essential not only for diagnosis but also for the monitoring of therapeutic responses to new drugs, such as cystic fibrosis transmembrane conductance regulator (CFTR) modulators and potentiators. Using iontophoresis as the gold standard can cause complications like burns, is uncomfortable, and requires repetitive hospital visits, which can be particularly problematic during a pandemic, where distancing and hygiene requirements are increased; therefore, it is necessary to develop fast and simple measures for the diagnosis and monitoring of CF. A screen-printed, low-cost chloride sensor was developed to remotely monitor CF patients. Using potentiometric measurements, the performance of the sensor was tested. It showed good sensitivity and a detection limit of 2.7 × 10−5 mol/L, which covered more than the complete concentration range of interest for CF diagnosis. Due to its fast response of 30 s, it competes well with standard sensor systems. It also offers significantly reduced costs and can be used as a portable device. The analysis of real sweat samples from healthy subjects, as well as CF patients, demonstrates a proper distinction using the screen-printed sensor. This approach presents an attractive remote measurement alternative for fast, simple, and low-cost CF diagnosis and monitoring