N-acetylneuraminic Acid Levels Research Articles

Urinary Metabolite Diagnostic and Prognostic Liquid Biopsy Biomarkers of Lung Cancer in Non-smokers and Tobacco Smokers.

Non-smokers account for 10-13% of all lung cancer cases in the United States. Etiology is attributed to multiple risk factors including exposure to secondhand smoking, asbestos, environmental pollution, and radon, but these exposures are not within the current eligibility criteria for early lung cancer screening by low-dose computed tomography (LDCT). Urine samples were collected from two independent cohorts comprising 846 participants (exploratory cohort) and 505 participants (validation cohort). The cancer urinary biomarkers, creatine riboside (CR) and N-acetylneuraminic acid (NANA) were analyzed and quantified using liquid chromatography-mass spectrometry to determine if non-smoker cases can be distinguished from sex and age-matched controls in comparison to tobacco smoker cases and controls, potentially leading to more precise eligibility criteria for LDCT screening. Urinary levels of CR and NANA were significantly higher and comparable in non-smokers and tobacco smoker cases as compared to population controls in both cohorts. Receiver Operating Characteristics (ROC) analysis for combined CR and NANA levels in non-smokers of the exploratory cohort resulted in better predictive performance with the area under the curve (AUC) of 0.94, whereas the validation cohort non-smokers had an AUC of 0.80. Kaplan-Meier survival curves showed that high levels of CR and NANA were associated with increased cancer-specific death in non-smokers as well as tobacco smoker cases in both cohorts. Measuring CR and NANA in urine liquid biopsies could identify non-smokers at high risk for lung cancer as candidates for LDCT screening and warrant prospective studies of these biomarkers.

Abstract 1001: Urinary metabolite diagnostic and prognostic liquid biopsy biomarkers of lung cancer - never smokers versus tobacco smokers

Abstract Introduction: Never smokers account for 10-13% of all lung cancer cases in the United States. The etiology is attributed to multiple causes including exposure to passive smoking, parental smoking, asbestos and radon. However, these lung cancer cases do not fit the current United States Government-funded criterion of low-dose computed tomography that identifies early stages of lung cancer with higher survival following surgical removal. Methods: The cancer urinary biomarkers, creatine riboside (CR) and N-acetylneuraminic acid (NANA), were analyzed and quantified by liquid chromatography-mass spectrometry to determine if never smokers with non-small cell lung cancer (NSCLC) can also be identified alongside tobacco smoker NSCLC cases as compared with gender and age-matched non-cancer population controls. Results: These cancer biomarkers were significantly higher and comparable in never-smokers and tobacco smokers cases compared to population controls. Receiver Operating Characteristics (ROC) analysis in never smokers was equivalent for CR and NANA levels area under the curve (AUC) 0.91. In contrast, combined CR and NANA levels resulted in better predictive performance (AUC 0.94). Kaplan-Meier survival curves showed that high levels of CR and NANA were associated (p<0.05) with increased cancer-specific death in never-smokers and tobacco smokers. Therefore these metabolites are independent of Tobacco smoking. NANA metabolite category factor, histology and stage were independent predictors of survival (p<0.05) in multivariate analyses for never smokers as shown in Table 1. Conclusions: These results indicate that measuring CR and NANA in liquid biopsy urine could identify never-smoking lung cancer candidates for LDCT screening and warrant prospective studies of these biomarkers in high-risk environments. Table 1. Univariate and multivariate regression analysis for factors associated with survival in the never smokers population Univariate Analysis Multivariate analysis Factors Levels N HR 95%CI P-value HR 95%CI P-value Stage Early Stage (I & II); Late Stage (III & IV) 145 3.52 2.3, 5.3 <0.001 3.94 2.2, 6.8 <0.001 Histology Adenocarcinoma; Squamous cell carcinoma 114 1.62 0.7, 3.5 0.26 3.42 1.4, 7.9 0.01 CR Category Low; High 166 1.34 0.8-2.1 0.19 1.67 0.9, 3.0 0.08 NANA Category Low; High 166 2.11 1.1-3.9 0.009 2.31 1.0, 5.0 0.02 Age N=166 166 1.00 0.9, 1.0 0.50 0.99 0.9, 1.0 0.4 Sex Male; Female 166 0.87 0.5, 1.2 0.49 0.90 0.5, 1.5 0.7 Race African American; European American 166 0.82 0.2, 2.5 0.75 0.40 0.1, 1.3 0.2 Citation Format: Bhavik Dalal, Takeshi Tada, Daxesh P. Patel, Mohammed Khan, Takahiro Oike, Yasuyuki Kanke, Amelia Parker, Majda Haznadar, Leila Toulabi, Kristopher Krausz, Ana Robles, Elise Bowman, Frank Gonzalez, Curtis Harris. Urinary metabolite diagnostic and prognostic liquid biopsy biomarkers of lung cancer - never smokers versus tobacco smokers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1001.

Correlation between plasma glutathione peroxidase 4 and N-acetylneuraminic acid levels with clinical risk stratification and prognosis of patients with acute coronary syndrome.

To investigate the correlation between plasma glutathione peroxidase 4 (GPX4) and N-acetyl-neuraminic acid (Neu5Ac) with clinical risk stratification and outcomes of acute coronary syndrome (ACS) patients. Between October 2018 and July 2019, 413 patients that were scheduled for coronary angiography were enrolled in this prospective study at the First Affiliated Hospital of Bengbu Medical College, Bengbu, China. Patients were divided into control and ACS groups. Patients with ACS were divided into 3 risk levels based on their thrombolysis in myocardial infarction risk score. After discharge, ACS patients were followed for the incidence of major adverse cardiac events (MACEs). For the analysis of cumulative endpoint event occurrences, the Kaplan-Meier method was applied. The ACS group had lower plasma GPX4 but higher Neu5Ac levels than the control group. There was a greater increase in plasma Neu5Ac in the high-risk group when compared with the medium-risk and low-risk groups, while GPX4 levels were higher in the low-risk group. The MACEs group had higher plasma Neu5Ac but lower GPX4 levels than the non-MACEs group. The plasma Neu5Ac was an independent risk factor but GPX4 was a protective factor for MACEs. Glutathione peroxidase 4 and Neu5Ac levels in plasma can be used to diagnose, stratify risks, and predict long-term outcomes in patients with ACS.

A rapid and sensitive procedure for determination of 5- N-acetyl neuraminic acid in lipopolysaccharides of Haemophilus influenzae: a survey of 24 non-typeable H. influenzae strains

In view of the importance of 5- N-acetyl neuraminic acid in bacterial pathogenesis, a sensitive, reproducible and reliable method for the determination of 5- N-acetyl neuraminic acid levels in lipopolysaccharide (LPS) is described and applied to 24 different non-typeable Haemophilus influenzae (NTHi) strains. The method involves analysis by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) of terminal 5- N-acetyl neuraminic acid residues released by neuraminidase treatment of O-deacylated LPS. The procedure is relatively fast and the instrumental effort is moderate. The results of the procedure were compared with data obtained by 1H NMR and electrospray ionisation-mass spectrometry (ESI-MS). The analysis of LPS from 24 NTHi strains showed that 5- N-acetyl neuraminic acid was found to be a common constituent of LPS in NTHi. Only one strain (NTHi 432) did not show any sialylation. Molar ratios (LPS/5- N-acetyl neuraminic acid) ranged between 5/1 and 500/1. Several strains in which no 5- N-acetyl neuraminic acid could be determined by other methods including 1H NMR and ESI-MS were shown to contain 5- N-acetyl neuraminic acid by this HPAEC-PAD procedure. The method was applied to determine levels of terminal 5- N-acetyl neuraminic acid in LPS from NTHi strains grown under different conditions and mutant strains containing inactive LPS biosynthetic genes.

Implication of altered levels of plasma alpha(1)-acid glycoprotein and its derived sialic acid on plasma protein binding of trimethoprim in pigs in physiological and pathological states.

In growing pigs (0 to 158 days after birth), pregnant sows (0 to 90 days), and in pigs suffering from respiratory disease (Actinobacillus pleuropneumoniae), the plasma levels of alpha(1)-acid glycoprotein (AGP) and its derived sialic acid were determined, as was the meaning of their altered levels on the plasma protein binding of trimethoprim. The AGP level was very high immediately after birth (more than 10,000 mu g/ml), decreased markedly during 2 weeks after birth (about 700 mu g/ml) and thereafter stayed at a constant level (about 400 mu g/ml). Pregnant sows had a low AGP level with a narrow variation throughout pregnancy (about 190 to 260 mu g/ml). Pigs infected with A. pleuropneumoniae showed an increased AGP level (mean value; 732 mu g/ml) with a wide variation (range: 170-1,840 mu g/ml). N-acetylneuraminic acid (NANA) and N-glycolylneuraminic acid (NGNA) were sialic acid subtypes detected in porcine plasma. In growing pigs, the time course of changes in NANA concentrations was consistent with that of AGP, whereas that of NGNA was different, implying that NANA is a sialic acid subtype derived from porcine AGP, in contrast to NGNA. The relationship between AGP and NANA levels in growing pigs could be expressed by the following equation: NANA=0.14 x AGP + 159 mu g/ml, whereas that in pigs with respiratory disease could be expressed by NANA=0.O67 x AGP + 357 mu g/ml, indicating a low fraction of NANA in AGP in diseased pigs. The regression lines between the AGP level and the plasma protein binding of trimethoprim < or = 2,000 mu g of AGP/ml were similar as follows: binding(%)=0.O23 x GP + 34 in growing pigs and binding(%)=0.O22 x GP+29 in diseased pigs, implying a minor role of sialic acid residues in the binding of basic drugs to AGP. In conclusion, the wide change in plasma AGP levels in diseased pigs as well as during the initial growth phase can alter the plasma protein binding of basic drugs such as trimethoprim, probably leading to a change in drug disposition. The low sialylation of AGP in diseased pigs may not have a great influence on the binding of basic drugs to AGP, implying the quantitative importance of AGP.

Hypothalamic opioids and the acute-phase glycoprotein response in guinea pigs

Endogenous opioids (EO) probably do not modulate endotoxin (LPS)- or interleukin 1 (IL1)-induced fever because naloxone does not prevent its development. Yet, increases in CSF and hypothalamic levels of β-endorphin have been reported during LPS-and IL1-induced fevers. Since IL1 also reduces the specific binding of opioids to their receptors in guinea pig brain, the opioids could be involved in modulating nonfebrile effects of IL1. To determine whether EO might have a role in the IL1-induced acute-phase glycoprotein response of guinea pigs, (1) naloxone (5 and 10 mg/kg, SC) was injected prior to LPS ( S. enteritidis 2 μg/kg, IV; N = 5), and (2) morphine (MOR, 10 μg/μl), [D-ala 2]-met-enkephalinamide (DAME, 5 μg/μl), or dynorphin A (DYN, 5 μg/μl) was injected into the preoptic area (1 μl, bilaterally; N = 8/treatment) or into the 3rd ventricle (N = 4/treatment); pyrogen-free saline was the control injection. Measurements were: core temperature (T co) and, as indices of acute-phase glycoproteins, plasma levels of copper (Cu) and N-acetylneuraminic acid (NANA). Naloxone did not prevent the fever or the increases in plasma Cu and NANA levels evoked by LPS. The intracerebral administration of opioid agonists by either route induced variable rises in T co, each with a different pattern, but no increases in plasma Cu and NANA levels. Thus, EO do not participate in the central modulation of acute-phase glycoprotein synthesis, but may have a role in influencing other nonthermal IL1 effects in the CNS.

Analysis of mammary tumour cell metastasis and release of bound n-acetylneuraminic acid.

A tumour model consisting of the highly metastatic mammary 13762 parental line, the non-metastatic and 6-thioguanine-resistant (TgR) variant line, and two TgR revertant lines (TgRrev, TgRrevM) that were occasionally metastatic, were used to determine whether the release of N-acetylneuraminic acid (NANA) was related to tumour metastasis. For comparative purposes, the occasionally metastatic R3230AC and the nonmetastatic DMBA8 tumour lines were studied. The NANA was considered to be in bound form, because acid hydrolysis was required to release it for high-pressure liquid chromatographic analyses. Sera of animals bearing the 13762 and R3230AC tumours had high levels of bound NANA. No differences were found in serum NANA levels in animals bearing metastatic or non-metastatic R3230AC tumours. Low levels of bound NANA were found in animals bearing the other tumour lines regardless of whether metastasis occurred or not. The experiments in vitro substantiated the in vivo findings. The phenotypic expression of bound NANA shedding did not correlate well with metastatic potential of the mammary tumour line. Our analyses suggest that this phenotypic marker cannot be used as a reliable indicator of metastasis.

Activation of acute-phase responses by intrapreoptic injections of endogenous pyrogen in guinea pigs

The acute-phase reaction (APR) is the concatenation of events that develops in response to infectious or other acute inflammatory stimuli. It includes fever and changes in plasma trace metal and glycoprotein levels. Endogenous pyrogen (EP) is believed to be the mediator of the APR. It acts within the preoptic-anterior hypothalamus (PO) to initiate fever; prostaglandins E (PGE) may modulate this action. To determine whether the nonfebrile responses to EP also are mediated by the PO and through PGE, guinea pigs were injected bilaterally intra-PO (iPO) with homologous EP (1 microliter) or PGE2 (0.1 microgram), and their colonic temperatures (Tco) and plasma iron (Fe), zinc (Zn), copper (Cu), and N-acetylneuraminic acid (NANA) levels were measured. For comparison, EP (2 ml) also was injected intraperitoneally (IP). Heat-denatured EP (delta EP) or pyrogen-free saline (PFS) was the corresponding control. Fevers were induced by IP EP (1.0 +/- 0.1 degrees C [mean +/- SD]), iPO EP (1.1 +/- 0.2 degrees C), and iPO PGE2 (1.4 +/- 0.2 degrees C); neither delta EP nor PFS was pyrogenic. Plasma Fe and Zn levels were decreased significantly after IP EP, but unchanged after iPO EP and PGE2. Plasma Cu and NANA levels were elevated significantly following both IP and iPO EP, but not after iPO PGE2. delta EP or PFS did not cause any changes, by either route. It appears, therefore, that EP-induced fever and rises in plasma Cu and NANA are mediated by the PO, while the decreases of plasma Fe and Zn are direct, peripheral effects. On the other hand, PGE2 appears to be involved only in the central febrile response. Indeed, guinea pigs, pretreated with indomethacin (5 mg/kg, IP), and injected iPO with EP or IP with S. enteritidis endotoxin (2 micrograms/kg), did not develop fever, but exhibited the rise in plasma Cu and NANA.

Elimination of 2-deoxyribose interference in the thiobarbituric acid determination of N-acetylneuraminic acid in tumor cells by pH-dependent extraction with cyclohexanone

The widely used thiobarbituric acid technique for the quantitation of N-acetylneuraminic acid (NANA) was improved to eliminate the interference of the ubiquitous 2-deoxyribose. The 2-deoxyribose chromogen was completely removed by cyclohexanone extraction at pH 5.6–6.0. After readjusting the pH to 1.7–1.9, the chromogen representative of NANA was quantitatively extracted with cyclohexanone. All other aspects of the original technique (L. Warren 1959 J. Biol. Chem. 534, 1971–1975) remained unchanged. The technique has been applied to determine total as well as neuraminidase-susceptible NANA in the preparation of the immunogen (neuraminidase-treated myeloblasts) utilized to stimulate specific immunity in patients with myeloblastic leukemia and certain solid tumors; NANA levels significantly affect immunogenicity. Data obtained from a variety of tumors using pH-dependent extraction as compared to the thiobarbituric acid method, isoamyl alcohol extraction, and ion-exchange purification showed that 2-deoxyribose interference may cause as much a two- to threefold error in the quantitation of NANA.

Correlation between perchloric-acid-soluble serum proteins, cellular immunity and tumor-cell burden.

AbstractSeveral in vitro‐cultured neoplastic cells, particularly human mammary carcinoma and malignant melanoma cells, shed or secrete glycoproteins which are tumorspecific. These compounds contain N‐acetylneuraminic acid (NANA), and they differ from the bulk of serum proteins and glycoproteins in being soluble in perchloric acid. The present studies examined the relation between perchloric acid soluble serum proteins (PA‐proteins), and N‐acetylneuraminic acid (PA‐NANA) and the number of R3230 AdCa implanted into syngeneic Fischer rats. A graded number of R3230 AdCa cells from a mammary rat adenocarcinoma was implanted into separate groups (20 rats/group) of imbred 12‐ to 16‐week‐old female Fischer rats. At 0, 24,48, 72, 196 h, and at 6, 9, 12, 15 and 21 days after tumor cell implantation, blood was examined for protein and NANA levels, and spleen cells for migration inhibition. Vibrio cholerae neuraminidase (VCN)‐treated formalinized tumor cells were used as the antigen. Animals which received 100 R3230 AdCa cells/animal showed a progressive increase in PA‐proteins and PA‐NANA 48 h after implanting the tumor cells. The serum PA‐protein and PA‐NANA levels were time‐dependent and increased with the number of implanted tumor cells. Maximum levels were found in sera from blood drawn 196 h or later from animals which received 1,000 tumor cells/rat or more. At 72 h or more after tumor cell implantation, spleen cells from animals which received 100 or more R3230 AdCa cells per animal were inhibited on contact with VCN‐treated formalinized R3230 AdCa cells. The magnitude of inhibition increased with time and with number of implanted tumor cells. A maximum of 85% inhibition was attained with spleens from animals 196 h after implantation of 100 R3230 AdCa cell/animal, or from animals 72 h after implantation of 103 or 104 R3230 AdCa cells/animal.

Significance of protein-bound neuraminic acid levels in patients with prostatic and bladder carcinoma*

The role of plasma N-acetyl neuraminic acid (NANA) level in detection and staging prostatic and bladder carcinomas was evaluated. Multiple determinations were obtained in 73 male subjects, including normal controls, patients with prostatic or bladder carcinoma or with benign prostatic hyperplasia. Patients with prostatic or bladder carcinoma had significantly higher NANA levels than normal individuals. Further, in patients with prostatic carcinoma, NANA values correlated well with both the stage of the disease and the response to therapy.

Serum Sialic Acid Levels in Lung Cancer Patients

The levels of N-acetyl neuraminic acid (sialic acid) in normal men and pre-and post-menopausal women were determined. Smoking post-menopausal estrogen therapy, oral contraceptives, and refreezing had no effects on sialic acid levels. Pre-treatment values from patients with lung carcinoma showed markedly elevated levels of sialic acid (0.697 +/- 0.149 muM/ml) as compared to those from normal controls (0.432 +/- 0.067 muM/ml). The potential usefulness of sialic acid as a biological marker is discussed.

Bronchorrhoea.

Bronchorrohea has been defined as a condition in which more than 100 ml of sputum is produced within 24 hours, an amount in excess of that seen in chronic lung diseases. The rheological and chemical characteristics of the sputum are here described. Levels of viscosity, dry weight, N-acetyl neuraminic acid (NANA), fucose, and sulphate fall between those in saliva and mucoid sputum from chronic lung diseases. These levels were always higher in bronchorrhoea sputum than in saliva and therefore may be used in the differential diagnosis of bronchorrhoea and hypersalivation. Bronchorrhoea sputum has the constituents of a bronchial secretion but is low in acid glycoprotein. Certain other features are commonly found - a large amount of froth, increase in viscosity with time, and separation into two phases. Some cases respond to steroids, particularly when the levels of NANA in the sputum are low.

Effect of pneumococcal neuraminidase on canine cerebral cortex in experimental meningitis.

We examined the effect of experimental meningeal infection with a neuraminidase-producing strain of Diplococcus pneumoniae on the N-acetylneuraminic acid (NANA) content of canine cerebral cortical gray matter. The Warren thiobarbituric acid assay for NANA was used. There was no significant difference between the free and bound NANA levels of the tissue from control animals and from those with meningitis. This finding suggests that the neurological impairment so frequent in clinical pneumococcal meningitis is not a result of the activity of this bacterial enzyme upon cortical glycolipids.