<p>Glycosylation alterations have emerged as significant indicators in the landscape of cancer diagnosis, with aberrant N-glycans standing out as potential biomarkers. The existing methods for purifying N-glycans mainly rely on their hydrophilicity, affinity, and size. These methods exhibit relatively weak binding forces and low specificity based on non-covalent interactions. Moreover, the current liquid chromatography-based N-glycan purification and detection techniques must be bettered for clinical settings' rapid diagnostic requirements. Here, we have developed a method involving the covalent binding of graphene modified with photocleavable amino groups to N-glycans in hepatocellular carcinoma (HCC) patients' serum glycoproteins. The N-glycans are identified using MALDI-TOF/TOF MS, catering to the requirements of biological research and the diagnosis of liver cancer disease. Specifically, amino-functionalized graphene is linked with photocleavable groups bearing hydrazide functional group, then undergoes acyl hydrazone covalent reaction with aldehyde groups on the side chains of oxidized monosaccharides to capture the N-glycans. Finally, under UV irradiation at a wavelength of 356&#xa0;nm for 15&#xa0;s, the ortho-nitrobenzyl photocleavable groups undergo cleavage, completing the release of captured N-glycans, which are then subjected to MALDI-TOF/TOF MS mass spectrometry for detection. Our findings demonstrate significant differences in the N-glycan profiles between serum samples of liver cancer patients and healthy individuals. These differences are characterized by increased abundance and molecular weight of N-glycans. Through biomarker screening, one N-glycan with an m/z ratio of 1847.01 shows the potential to serve as a diagnostic biomarker for liver cancer (P &lt; 0.05). Our research underscores the potential of photocleavable amino-modified graphene for detecting aberrant N-glycans in HCC patients' serum to elucidate the intricate between aberrant N-glycan alterations and HCC. This research paves the way for developing novel biomarkers, diagnostic strategies, and therapeutic interventions to combat this complex disease.</p> Graphical Abstract <p></p>

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A novel photocleavable amino-modified graphene for covalent purification of N-glycans from hepatocellular carcinoma patients’ serum for potential biomarkers discovery

  • Xuyuan Chao,
  • Shengjie Yang,
  • Baoying Zhang,
  • Xin Zang,
  • Jingyi Zhang,
  • Xizi Liu,
  • Lu Chen,
  • Lu Qi,
  • Xiaofeng Xue,
  • Han Hu,
  • Xinghe Wang

摘要

Glycosylation alterations have emerged as significant indicators in the landscape of cancer diagnosis, with aberrant N-glycans standing out as potential biomarkers. The existing methods for purifying N-glycans mainly rely on their hydrophilicity, affinity, and size. These methods exhibit relatively weak binding forces and low specificity based on non-covalent interactions. Moreover, the current liquid chromatography-based N-glycan purification and detection techniques must be bettered for clinical settings' rapid diagnostic requirements. Here, we have developed a method involving the covalent binding of graphene modified with photocleavable amino groups to N-glycans in hepatocellular carcinoma (HCC) patients' serum glycoproteins. The N-glycans are identified using MALDI-TOF/TOF MS, catering to the requirements of biological research and the diagnosis of liver cancer disease. Specifically, amino-functionalized graphene is linked with photocleavable groups bearing hydrazide functional group, then undergoes acyl hydrazone covalent reaction with aldehyde groups on the side chains of oxidized monosaccharides to capture the N-glycans. Finally, under UV irradiation at a wavelength of 356 nm for 15 s, the ortho-nitrobenzyl photocleavable groups undergo cleavage, completing the release of captured N-glycans, which are then subjected to MALDI-TOF/TOF MS mass spectrometry for detection. Our findings demonstrate significant differences in the N-glycan profiles between serum samples of liver cancer patients and healthy individuals. These differences are characterized by increased abundance and molecular weight of N-glycans. Through biomarker screening, one N-glycan with an m/z ratio of 1847.01 shows the potential to serve as a diagnostic biomarker for liver cancer (P < 0.05). Our research underscores the potential of photocleavable amino-modified graphene for detecting aberrant N-glycans in HCC patients' serum to elucidate the intricate between aberrant N-glycan alterations and HCC. This research paves the way for developing novel biomarkers, diagnostic strategies, and therapeutic interventions to combat this complex disease.

Graphical Abstract