High prevalence of CD44 and its ligand low molecular weight hyaluronan in plasma of HNSCC patients: clinical significance.

BACKGROUND: This study aims to evaluate the role of cancer stem cell marker, CD44, and its ligand HA as potential molecular biomarker for early detection of HNSCC. METHODS AND RESULTS: The expression profile (mRNA/Protein) of CD44 variants were analysed in primary HNSCC lesions and plasma of the patients. Then, prevalence of HA variants was analysed in plasma of the patients. The mRNA expression of CD44 variants, CD44S and CD44v3, were significantly high in both early (stage I/II) and late (stage III/IV) invasive lesions, with predominant expression of CD44v3 in the late-stage lesions. In plasma of HNSCC patients, increased levels of SolCD44, CD44-ICD and unique 62 KD CD44 variants with respect to standard CD44S were seen, in comparison to their prevalence in plasma of normal individuals. The abundance of CD44-ICD and 62 KD variants were significantly high in plasma of late stage HNSCC patients. Interestingly, significantly high level of low molecular weight HA(LMW HA) with respect to high molecular weight HA(HMW HA) was seen in plasma of HNSCC patients irrespective of clinical stages. On the contrary, high HMW HA level in plasma of normal individuals was seen. The high level of LMW HA in plasma of HNSCC patients might be due to combinatorial effect of increased mRNA expression of HA synthesizing enzyme HAS1/2/3 and HA degrading enzyme HYAL1/2, as seen in the primary HNSCC samples. CONCLUSION: Thus, our data revealed the importance of specific CD44 and HA variants in plasma of HNSCC patients during its development as potential non-invasive molecular biomarker of the disease.

Visual detection of serum asialohaptoglobin by plasmonic sandwich ELLSA--a new platform for cirrhosis diagnosis.

The cirrhotic condition of the liver has long been acknowledged as the preface to liver cancer. The desialylation status of the serum acute phase protein, haptoglobin, has been introduced as a new diagnostic analyte for liver cirrhosis. The reliability of this new diagnostic molecule has been evaluated in 30 liver cirrhosis patients having a history of earlier viral hepatitis C (HCV-LC). A novel enzyme linked lectinosorbent assay has been developed coupled with the plasmon mechanism of gold nanoparticle aggregation as the colorimetric read out which can visually distinguish the cirrhotic liver patients from the normal healthy and hepatitis C controls. The assay can be useful for rapid point-of-care detection, and even an untrained person can execute it without a specialized instrument. This method employs Sambucus nigra agglutinin (SNA) to detect the extent of α-2,6 sialylation of serum haptoglobin, the new diagnostic molecule for liver cirrhosis.

Alterations of glycan branching and differential expression of sialic acid on alpha fetoprotein among hepatitis patients.

The level of serum glycoproteins and their glycosylation pattern change in liver diseases including hepatocellular carcinoma (HCC). Some of them, especially alpha fetoprotein (AFP), serve as useful biomarkers for HCC. The present investigation showed high level of AFP in hepatitis B cirrhosis (HBV-LC) and hepatitis C cirrhosis (HCV-LC) patients. However, increase of AFP level was not significantly high in chronic hepatitis B (HBV-CH) as determined by ELISA using monoclonal anti-human AFP (mAb-AFP). The differential expression of sialic acid linkage was observed in HBV-CH and HCV-LC by ELISA; the former bound strongly with Sambucus nigra agglutinin (SNA), which has exclusive binding specificity for NeuAcα2-6-, whereas HCV-LC reacted preferably with Maackia amurensis agglutinin (MAA) which recognizes NeuAcα2-3-. There was significantly high glycan branching in HBV-LC and HCV-LC in comparison to controls as illustrated by concanavalin A. This was further confirmed by Phaseolus vulgaris erythroagglutinin (E-PHA) and Datura stramonium agglutinin (DSA). Enhanced fucosylation of AFP was observed in HBV-LC, HCV-LC and HCC patients by ELISA using fucose binding Aleuria aurantia lectin; however, maximum binding was found in HCC. Fucosylation with α1-6 linkage was further confirmed by Lens culinaris agglutinin (LCA). From the above results it is concluded that the changes in concentration of AFP, differential expression of sialic acid, increase of glycan branching and fucosylation have a prognostic value of hepatitis and it could be possible that lectin-based assay with AFP can aid in diagnosis of hepatitis diseases besides clinical examination and routine laboratory investigation.

Purification and molecular characterization of a sialic acid specific lectin from the phytopathogenic fungus Macrophomina phaseolina.

A lectin was isolated and purified from the culture filtrate of the plant pathogenic fungus Macrophomina phaseolina by a combination of ammonium sulfate precipitation, affinity chromatography on fetuin-Sepharose 4B and ion-exchange chromatography on DEAE-A 50. The lectin designated MPL was homogeneous by PAGE and HPLC and a monomeric protein with a molecular weight of approximately 34 kDa as demonstrated by SDS-PAGE. It is a glycoprotein and agglutinated human erythrocytes regardless of the human blood type. Neuraminidase treatment of erythrocytes reduced the agglutination activity of the lectin. It is thermally stable and exhibits maximum activity between pH 6 and 7.2. Its carbohydrate binding specificity was investigated both by hapten inhibition of hemagglutination and by enzyme-conjugated lectin inhibition assay. Although, M. phaseolina lectin bound sialic acid, it exhibited binding affinity towards neuraminyl oligosaccharides of N-linked glycoproteins, alpha-Neu5Ac-(2-->3)-beta-Gal-(1-->4)-GlcNAc being maximum.