The presence of circulating genetically abnormal cells in blood predicts risk of lung cancer in individuals with indeterminate pulmonary nodules.

PURPOSE: Computed tomography is the standard method by which pulmonary nodules are detected. Greater than 40% of pulmonary biopsies are not lung cancer and therefore not necessary, suggesting that improved diagnostic tools are needed. The LungLB™ blood test was developed to aid the clinical assessment of indeterminate nodules suspicious for lung cancer. LungLB™ identifies circulating genetically abnormal cells (CGACs) that are present early in lung cancer pathogenesis. METHODS: LungLB™ is a 4-color fluorescence in-situ hybridization assay for detecting CGACs from peripheral blood. A prospective correlational study was performed on 151 participants scheduled for a pulmonary nodule biopsy. Mann-Whitney, Fisher's Exact and Chi-Square tests were used to assess participant demographics and correlation of LungLB™ with biopsy results, and sensitivity and specificity were also evaluated. RESULTS: Participants from Mount Sinai Hospital (n = 83) and MD Anderson (n = 68), scheduled for a pulmonary biopsy were enrolled to have a LungLB™ test. Additional clinical variables including smoking history, previous cancer, lesion size, and nodule appearance were also collected. LungLB™ achieved 77% sensitivity and 72% specificity with an AUC of 0.78 for predicting lung cancer in the associated needle biopsy. Multivariate analysis found that clinical and radiological factors commonly used in malignancy prediction models did not impact the test performance. High test performance was observed across all participant characteristics, including clinical categories where other tests perform poorly (Mayo Clinic Model, AUC = 0.52). CONCLUSION: Early clinical performance of the LungLB™ test supports a role in the discrimination of benign from malignant pulmonary nodules. Extended studies are underway.

PathogenDx DetectX Combined Demonstrates Equivalent Performance in Comparison to Four AOAC Certified Methods for the Detection of Aspergillus Species, Salmonella Species, and STEC in Dried Hemp Flower.

BACKGROUND: The PathogenDx DetectX Combined method is a certified Performance Tested MethodSM (012201) that is enrichment-free and utilizes a DNA microarray-based end point PCR method for the simultaneous detection of Aspergillus (A. flavus, A. fumigatus, A. niger, and A. terreus), Salmonella spp., and a broad range of Shiga toxin-producing Esherichia coli (STEC) from hemp and cannabis flower, edibles, and concentrates. OBJECTIVE: This study aimed to compare the PathogenDx DetectX Combined enrichment-free method to four AOAC INTERNATIONAL certified molecular methods that utilize enrichment prior to quantitative PCR (qPCR) amplification in hemp flower for the detection of Aspergillus (A. flavus), S. enterica, and Escherichia coli 026. METHODS: In this method comparison study, each method was evaluated according to the AOAC validated instructions for use (IFU) and the AOAC Appendix J validation guidelines. A total of 16 samples at three levels of contamination (0, 0.7, and 2 CFU/10g test portion) were analyzed by each method. The results for all methods were evaluated by using the probability of detection statistical model (POD). RESULTS: Results of the validation study demonstrate that the PathogenDx DetectX Combined enrichment-free method is equivalent in performance to the three proprietary methods evaluated in this study. CONCLUSION: The method comparison study indicated that the PathogenDx DetectX Combined enrichment-free method provides equivalent detection of the target analytes (A. flavus, Salmonella, and a broad range of STEC) in hemp flower. HIGHLIGHTS: The performance of The PathogenDx DetectX Combined method is significantly faster and possesses a higher or equivalent degree of sensitivity and specificity. Implementation of this method for routine microbial pathogen analysis in laboratories would save significant time and resources.

Rapid detection of Salmonella enterica in leafy greens by a novel DNA microarray-based PathogenDx system.

The diverse matrices pose great challenges for rapid detection of low Salmonella level (<10 CFU) in fresh produce. The applicability of microarray-based PathogenDx system for detecting low contamination of Salmonella Newport from leafy greens was evaluated. A pre-PCR preparation protocol including enrichment in universal pre-enrichment broth for 3 h followed by sample concentration using an InnovaPrep bio-concentrator or 6 h enrichment without a concentration step was used for detecting S. Newport from leafy greens with initial inoculum level at ∼6 CFU/25 g. Among 205 samples tested, 98%, 93%, 76%, and 60% of Romaine lettuce, Iceberg lettuce, kale, and spinach samples were tested positive after 3 h of enrichment with sample concentration. After 6 h of enrichment, 100%, 98%, 90%, and 82% of Romaine lettuce, Iceberg lettuce, kale, and spinach samples were positive. The samples were parallelly tested by the FDA bacterial analytical manual (BAM) method and 100% of spiked produce samples were tested positive. The overall analysis time of this methodology was between 8 and 11 h, including all pre-enrichment and concentration steps, in contrast to 4-5 days required for BAM method. The system correctly differentiated all 108 Salmonella strains and 35 non-Salmonella strains used in the study. This novel microarray approach provides a rapid method for detecting Salmonella in leafy greens.

Validation Study for PathogenDx EnviroX-F Assay for the Detection of Listeria, L. monocytogenes, and Salmonella in Environmental Surface Samples: AOAC Performance Tested MethodSM 092001.

BACKGROUND: The PathogenDx EnviroX-F uses microarray technology to simultaneously detect Listeria species, Listeria monocytogenes, and Salmonella species from environmental samples without the need for enrichment. OBJECTIVE: The validation study included a matrix study of four matrices (stainless steel, plastic, rubber, and sealed concrete) comparing the PathogenDx EnviroX-F assay to the US Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) Chapter 10, "Detection of L. monocytogenes in Foods and Environmental Samples, and Enumeration of L. monocytogenes in Foods" (revised March 2017), and FDA BAM Chapter 5,"Salmonella" (revised February 2020). Other required Performance Tested MethodSM parameters evaluated included: inclusivity and exclusivity, robustness, instrument variation, and product consistency and stability. METHODS: The PathogenDx EnviroX-F assay was evaluated with 30 unpaired replicate surface areas for each environmental surface. The candidate method was evaluated without an enrichment step. RESULTS: In the inclusivity and exclusivity study 50 out of 50 Listeria isolates were detected, 50 out of 50 L. monocytogenes strains were detected, 108 out of 108 Salmonella strains were detected, and 95 out of 95 exclusivity strains were correctly excluded. In the matrix study, the PathogenDx EnviroX-F assay showed no significant differences between confirmed results or between candidate and reference method results for 4" × 4" environmental surface areas for each matrix. CONCLUSION: The PathogenDx EnviroX-F assay is an effective method for the qualitative detection of Listeria spp., L. monocytogenes, and Salmonella spp. from environmental surface swabs. HIGHLIGHTS: The PathogenDx EnviroX-F assay will be the first PTM-approved multiplex assay for Listeria spp., L. monocytogenes, and Salmonella without the need for an environment step.

Independent Study for the Detectx Combined Assay for the Detection of Aspergillus, Salmonella, and STEC (stx1 and/or 2) in Dried Cannabis Flower and Dried Hemp Flower: Level 3 Modification Study 012201.

BACKGROUND: The PathogenDx family of assays uses microarray technology to simultaneously detect the presence of bacterial and fungal pathogens in food products, environmental surfaces, and cannabis products. OBJECTIVE: The Detectx Combined assay was validated for the detection of Aspergillus, (Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, and Aspergillus terreus), Salmonella, and a broad range of STEC (stx1 and/or 2) species. The validation consisted of two matrix studies in dried hemp flower and dried cannabis flower (>0.3% delta-9 tetrahydrocannabinol) flower, product consistency, stability, robustness, and inclusivity and exclusivity for two targets: Aspergillus and STEC. METHOD: The PathogenDx Detectx Combined assay was evaluated with 30 replicates in each matrix and confirmed according to the instructions outlined in this study. RESULTS: Results of the validation study met the requirements of AOAC Standard Method Performance Requirement (SMPR®) 2020.002 and 2020.012. In the inclusivity and exclusivity study, all target isolates (Aspergillus and STEC) were correctly detected. For the exclusivity study, 26 out of 30 Aspergillus and 30 out of 30 STEC non-target strains were correctly excluded. In the matrix study, the PathogenDx Detectx Combined assay showed no significant statistical differences between confirmed results for dried hemp and cannabis flower. Robustness testing indicated that small changes to the method parameters did not impact the performance of the assay. Stability and consistency studies verified that the assay's shelf-life claims were appropriate, and manufacturing of the assay was consistent. CONCLUSIONS: The validation study indicated that the PathogenDx DetectX Combined assay was successful in detection of the new target analytes (Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, and Aspergillus terreus and STEC containing stx1 and/or 2) and could successfully recover these organisms and Salmonella from dried hemp flower and dried cannabis flower (>0.3% delta-9 tetrahydrocannabinol). HIGHLIGHTS: The PathogenDx DetectX Combined Assay will be the first PTM approved multiplex assay for Aspergillus, E. coli and Salmonella that does not require an enrichment step.

Validation Study for PathogenDx EnviroX-Rv Assay for the Detection of SARS-CoV-2 from Stainless Steel Environmental Surface Swabs Performance Tested MethodSM 122003.

BACKGROUND: The PathogenDx EnviroX-Rv uses endpoint PCR + DNA microarray technology to detect SARS-CoV-2, the causative agent of COVID-19, from stainless-steel environmental sample swabs. OBJECTIVE: To validate the PathogenDx EnviroX-Rv assay as part of the Emergency Response Validation (ERV) Performance Tested Method(s)SM (PTM) program. METHOD: The PathogenDx EnviroX-Rv assay was evaluated for specificity using in silico analysis of ≥41 000 SARS-CoV-2 sequences and over 50 exclusivity organisms (both near neighbors and background organisms). The candidate method was evaluated in an unpaired study design for one environmental surface (stainless steel) and compared to the US Centers for Disease Control and Prevention (CDC) 2019-Novel Coronavirus (2019-nCoV) Real-Time-Polymerase Chain Reaction (RT-PCR) Diagnostic Panel, Instructions for Use (Revision 4, Effective 6/12/2020). RESULTS: Results of the in silico analysis demonstrated the high specificity of the method in being able to detect target SARS-CoV-2 sequences and discriminate them from near neighbors and environmental background organisms. In the matrix study, the candidate method demonstrated a statistically significant difference when compared to the results of the CDC method utilized in this study, with the candidate method resulting in more positive replicates as it only requires one target to be present for a positive sample. CONCLUSIONS: The EnviroX-Rv assay rapidly and accurately detected SARS-CoV-2 RNA on environmental swabs from stainless-steel surfaces at a concentration of 2000 genomic copies per 2 × 2" test area. HIGHLIGHTS: The EnviroX-Rv assay employs dual PCR and hybridization techniques to provide highly accurate results when detecting SARS-CoV-2 from surfaces.

Accessing analytes in biofluids for peripheral biochemical monitoring.

Peripheral biochemical monitoring involves the use of wearable devices for minimally invasive or noninvasive measurement of analytes in biofluids such as interstitial fluid, saliva, tears and sweat. The goal in most cases is to obtain measurements that serve as surrogates for circulating analyte concentrations in blood. Key technological developments to date include continuous glucose monitors, which use an indwelling sensor needle to measure glucose in interstitial fluid, and device-integrated sweat stimulation for continuous access to analytes in sweat. Further development of continuous sensing technologies through new electrochemical sensing modalities will be a major focus of future research. While there has been much investment in wearable technologies to sense analytes, less effort has been directed to understanding the physiology of biofluid secretion. Elucidating the underlying biology is crucial for accelerating technological progress, as the biofluid itself often presents the greatest challenge in terms of sample volumes, secretion rates, filtration, active analyte channels, variable pH and salinity, analyte breakdown and other confounding factors.

Correction to: Expression of quiescin sulfhydryl oxidase 1 is associated with a highly invasive phenotype and correlates with a poor prognosis in luminal B breast cancer.

After the publication of this work [1], an error was noticed in Fig. 4a. The micrograph image sh528 was accidentally duplicated.

A compact, low-cost, quantitative and multiplexed fluorescence detection platform for point-of-care applications.

An effective method of combating infectious diseases is the deployment of hand-held devices at the point-of-care (POC) for screening or self-monitoring applications. There is a need for very sensitive, low-cost and quantitative diagnostic devices. In this study, we present a low-cost, multiplexed fluorescence detection platform that has a high sensitivity and wide dynamic range. Our system features inexpensive 3 × 3 mm interference filters with a high stopband rejection, sharp transition edges, and greater than 90% transmission in the passband. In addition to the filters, we improve signal-to-noise ratio by leveraging time for accuracy using a charge-integration-based readout. The fluorescence sensing platform provides a sensitivity to photon flux of ∼1×104photons/mm2sec and has the potential for 2-3 orders of magnitude improvement in sensitivity over standard colorimetric detection that uses colored latex microspheres. We also detail the design, development, and characterization of our low-cost fluorescence detection platform and demonstrate 100% and 97.96% reduction in crosstalk probability and filter cost, respectively. This is achieved by reducing filter dimensions and ensuring appropriate channel isolation in a 2 × 2 array configuration. Practical considerations with low-cost interference filter system design, analysis, and system performance are also discussed. The performance of our platform is compared to that of a standard laboratory array scanner. We also demonstrate the detection of antibodies to human papillomavirus (HPV16) E7 protein, as a potential biomarker for early cervical cancer detection in human plasma.

Eccrine Sweat as a Biofluid for Profiling Immune Biomarkers.

PURPOSE: Sweat is a relatively unexplored biofluid for diagnosis and monitoring of disease states. In this study, the proteomic profiling of immune-related biomarkers from healthy individuals are presented. EXPERIMENTAL DESIGN: Eccrine sweat samples are collected from 50 healthy individuals. LC-MS/MS is performed on two pools of sweat samples from five male and female participants. Individual sweat samples are analyzed by antibody isotyping microarrays (n = 49), human cytokine arrays (n = 30), and quantitative ELISAs for interleukin-1α (n = 16), epidermal growth factor (n = 6), and angiogenin (n = 7). RESULTS: In sweat, 220 unique proteins are identified by shotgun analysis. Detectable antibody isotypes include IgA (100% positive; median 1230 ± 28 700 pg mL-1 ), IgD (18%; 22.0 ± 119 pg mL-1 ), IgG1 (96%; 1640 ± 6750 pg mL-1 ), IgG2 (37%; 292 ± 6810 pg mL-1 ), IgG3 (71%; 74.0 ± 119 pg mL-1 ), IgG4 (69%; 43.0 ± 42.0 pg mL-1 ), and IgM (41%; 69.0 ± 1630 pg mL-1 ). Of 42 cytokines, three are readily detected in all sweat samples (p < 0.01). The median concentration for interleukin-1α is 352 ± 521 pg mL-1 , epidermal growth factor is 86.5 ± 147 pg mL-1 , and angiogenin is 38.3 ± 96.3 pg mL-1 . Multiple other cytokines are detected at lower levels. CONCLUSIONS AND CLINICAL RELEVANCE: Sweat can be used for profiling antibodies and innate immune biomarkers.

Autoantibody biomarkers for the detection of serous ovarian cancer.

Objective The purpose of this study was to identify a panel of novel serum tumor antigen-associated autoantibody (TAAb) biomarkers for the diagnosis of high-grade serous ovarian cancer. METHODS: To detect TAAb we probed high-density programmable protein microarrays (NAPPA) containing 10,247 antigens with sera from patients with serous ovarian cancer (n=30 cases/30 healthy controls) and measured bound IgG. We identified 735 promising tumor antigens and evaluated these with an independent set of serous ovarian cancer sera (n=30 cases/30 benign disease controls/30 healthy controls). Thirty-nine potential tumor autoantigens were identified and evaluated using an orthogonal programmable ELISA platform against a total of 153 sera samples (n=63 cases/30 benign disease controls/60 healthy controls). Sensitivities at 95% specificity were calculated and a classifier for the detection of high-grade serous ovarian cancer was constructed. RESULTS: We identified 11-TAAbs (ICAM3, CTAG2, p53, STYXL1, PVR, POMC, NUDT11, TRIM39, UHMK1, KSR1, and NXF3) that distinguished high-grade serous ovarian cancer cases from healthy controls with a combined 45% sensitivity at 98% specificity. CONCLUSION: These are potential circulating biomarkers for the detection of serous ovarian cancer, and warrant confirmation in larger clinical cohorts.

Application of flat panel OLED display technology for the point-of-care detection of circulating cancer biomarkers.

Point-of-care molecular diagnostics can provide efficient and cost-effective medical care, and they have the potential to fundamentally change our approach to global health. However, most existing approaches are not scalable to include multiple biomarkers. As a solution, we have combined commercial flat panel OLED display technology with protein microarray technology to enable high-density fluorescent, programmable, multiplexed biorecognition in a compact and disposable configuration with clinical-level sensitivity. Our approach leverages advances in commercial display technology to reduce pre-functionalized biosensor substrate costs to pennies per cm(2). Here, we demonstrate quantitative detection of IgG antibodies to multiple viral antigens in patient serum samples with detection limits for human IgG in the 10 pg/mL range. We also demonstrate multiplexed detection of antibodies to the HPV16 proteins E2, E6, and E7, which are circulating biomarkers for cervical as well as head and neck cancers.

Proteomic mapping of p53 immunogenicity in pancreatic, ovarian, and breast cancers.

PURPOSE: Mutations in TP53 induce autoantibody immune responses in a subset of cancer patients, which have been proposed as biomarkers for early detection. Here, we investigate the association of p53-specific autoantibodies with multiple tumor subtypes and determine the association with p53 mutation status and epitope specificity. EXPERIMENTAL DESIGN: IgG p53 autoantibodies (p53-AAb), were quantified in 412 serum samples using a programmable ELISA assay from patients with serous ovarian, pancreatic adenocarcinoma, and breast cancer. To determine if patients generated mutation-specific autoantibodies we designed a panel of the most relevant 51 p53 point mutant proteins, to be displayed on custom programmable protein microarrays. To determine the epitope specificity we displayed 12 overlapping tiling fragments and 38 N- and C-terminal deletions spanning the length of the wild-type p53 protein. RESULTS: We detected p53-AAb with sensitivities of 58.8% (ovarian), 22% (pancreatic), 32% (triple negative breast cancer), and 10.2% (HER2+ breast cancer) at 94% specificity. Sera with p53-AAb contained broadly reactive autoantibodies to 51 displayed p53 mutant proteins, demonstrating a polyclonal response to common epitopes. All p53-AAb displayed broad polyclonal immune response to both continuous and discontinuous epitopes at the N- and C-terminus as well as the DNA-binding domain. CONCLUSION AND CLINICAL RELEVANCE: In this comprehensive analysis, mutations in tumor p53 induce strong, polyclonal autoantibodies with broadly reactive epitope specificity.

Programmable protein arrays for immunoprofiling HPV-associated cancers.

Over 600,000 cancers each year are attributed to the human papillomavirus (HPV), including cervical, anogenital and oropharyngeal cancers (OPC). A key challenge in understanding HPV immunobiology is the diversity of oncogenic HPV types and the need for multiplexed display of HPV antigens to measure antibody responses. We have generated custom HPV protein microarrays displaying 98 proteins as C-terminal GST fusion proteins, representing eight antigens of two low-risk HPV types (HPV6 and 11) and ten oncogenic high-risk HPV types (HPV16, 18, 31, 33, 35, 39, 45, 51, 52 and 58). We demonstrate robust and reproducible protein expression of 96/98 of the antigens using a human cell lysate expression system. The target epitopes and specificities of four monoclonal antibodies were identified. Using sera from ten patients with newly diagnosed OPC and ten controls, we demonstrate specific IgG seroreactivity to HPV16 E1, E2, and E7 (a fold increase of 1.52, 2.19 and 1.35 in cases vs. controls, respectively, all p < 0.005), confirming our prior data on an ELISA platform. We also detect HPV52 E7 Abs in serum from a patient with cervical cancer. The HPV protein array has potential for rapid identification of serologic responses to 12 HPV types.

Ebselen inhibits QSOX1 enzymatic activity and suppresses invasion of pancreatic and renal cancer cell lines.

Quiescin sulfhydryl oxidase 1 (QSOX1) is a highly conserved disulfide bond-generating enzyme that is overexpressed in diverse tumor types. Its enzymatic activity promotes the growth and invasion of tumor cells and alters extracellular matrix composition. In a nude mouse-human tumor xenograft model, tumors containing shRNA for QSOX1 grew significantly more slowly than controls, suggesting that QSOX1 supports a proliferative phenotype in vivo. High throughput screening experiments identified ebselen as an in vitro inhibitor of QSOX1 enzymatic activity. Ebselen treatment of pancreatic and renal cancer cell lines stalled tumor growth and inhibited invasion through Matrigel in vitro. Daily oral treatment with ebselen resulted in a 58% reduction in tumor growth in mice bearing human pancreatic tumor xenografts compared to controls. Mass spectrometric analysis of ebselen-treated QSOX1 mechanistically revealed that C165 and C237 of QSOX1 covalently bound to ebselen. This report details the anti-neoplastic properties of ebselen in pancreatic and renal cancer cell lines. The results here offer a "proof-of-principle" that enzymatic inhibition of QSOX1 may have clinical relevancy.

Expression of quiescin sulfhydryl oxidase 1 is associated with a highly invasive phenotype and correlates with a poor prognosis in Luminal B breast cancer.

INTRODUCTION: Quiescin sulfhydryl oxidase 1 (QSOX1) oxidizes sulfhydryl groups to form disulfide bonds in proteins. Tumor specific expression of QSOX1 has been reported for numerous tumor types. In this study, we investigate QSOX1 as a marker of breast tumor progression and evaluate the role of QSOX1 as it relates to breast tumor growth and metastasis. METHODS: Correlation of QSOX1 expression with breast tumor grade, subtype and estrogen receptor (ER) status was gathered through informatic analysis using the "Gene expression based Outcome for Breast cancer Online" (GOBO) web-based tool. Expression of QSOX1 protein in breast tumors tissue microarray (TMA) and in a panel of breast cancer cell lines was used to confirm our informatics analysis. To investigate malignant cell mechanisms for which QSOX1 might play a key role, we suppressed QSOX1 protein expression using short hairpin (sh) RNA in ER+ Luminal A-like MCF7, ER+ Luminal B-like BT474 and ER- Basal-like BT549 breast cancer cell lines. RESULTS: GOBO analysis revealed high levels of QSOX1 RNA expression in ER+ subtypes of breast cancer. In addition, Kaplan Meyer analyses revealed QSOX1 RNA as a highly significant predictive marker for both relapse and poor overall survival in Luminal B tumors. We confirmed this finding by evaluation of QSOX1 protein expression in breast tumors and in a panel of breast cancer cell lines. Expression of QSOX1 in breast tumors correlates with increasing tumor grade and high Ki-67 expression. Suppression of QSOX1 protein slowed cell proliferation as well as dramatic inhibition of MCF7, BT474 and BT549 breast tumor cells from invading through Matrigel™ in a modified Boyden chamber assay. Inhibition of invasion could be rescued by the exogenous addition of recombinant QSOX1. Gelatin zymography indicated that QSOX1 plays an important role in the function of MMP-9, a key mediator of breast cancer invasive behavior. CONCLUSIONS: Taken together, our results suggest that QSOX1 is a novel biomarker for risk of relapse and poor survival in Luminal B breast cancer, and has a pro-proliferative and pro-invasive role in malignant progression partly mediated through a decrease in MMP-9 functional activity.

Quiescin sulfhydryl oxidase 1 promotes invasion of pancreatic tumor cells mediated by matrix metalloproteinases.

Quiescin sulfhydryl oxidase 1 (QSOX1) oxidizes sulfhydryl groups to form disulfide bonds in proteins. We previously mapped a peptide in plasma from pancreatic ductal adenocarcinoma (PDA) patients back to an overexpressed QSOX1 parent protein. In addition to overexpression in pancreatic cancer cell lines, 29 of 37 patients diagnosed with PDA expressed QSOX1 protein in tumor cells, but QSOX1 was not detected in normal adjacent tissues or in a transformed, but nontumorigenic cell line. To begin to evaluate the advantage QSOX1 might provide to tumors, we suppressed QSOX1 protein expression using short hairpin (sh) RNA in two pancreatic cancer cell lines. Growth, cell cycle, apoptosis, invasion, and matrix metalloproteinase (MMP) activity were evaluated. QSOX1 shRNA suppressed both short and long isoforms of the protein, showing a significant effect on cell growth, cell cycle, and apoptosis. However, QSOX1 shRNA dramatically inhibited the abilities of BxPC-3 and Panc-1 pancreatic tumor cells to invade through Matrigel in a modified Boyden chamber assay. Mechanistically, gelatin zymography indicated that QSOX1 plays an important role in activation of MMP-2 and MMP-9. Taken together, our results suggest that the mechanism of QSOX1-mediated tumor cell invasion is by activation of MMP-2 and MMP-9.

Analysis of the plasma peptidome from pancreas cancer patients connects a peptide in plasma to overexpression of the parent protein in tumors.

Blood circulates through nearly every organ including tumors. Therefore, plasma is a logical source to search for tumor-derived proteins and peptides. The challenge with plasma is that it is a complex bodily fluid composed of high concentrations of normal host proteins that obscure identification of tumor-derived molecules. To simplify plasma, we examined a low molecular weight (LMW) fraction (plasma peptidome) using liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods. In the plasma peptidome of patients with ductal adenocarcinoma of the pancreas (DAP), a prominent peptide was identified from the QSOX1 parent protein. This peptide is stable in whole blood over 24 h and was present in 16 of 23 DAP patients and 4 of 5 patients with intraductal papillary mucinous neoplasm (IPMN). QSOX1 peptides were never identified in the plasma peptidome from 42 normal healthy donors using the same methods. Immunohistochemical staining of DAP tissue sections with anti-QSOX1 antibody shows overexpression of QSOX1 in tumor but not in adjacent stroma or normal ducts. Three of four pancreas tumor cell lines also express QSOX1 protein by Western blot analysis. This is the first report of QSOX1 peptides in plasma from DAP patients and makes the rare connection between a peptide in plasma from cancer patients and overexpression of the parent protein in tumors.