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Proteomics of Sentinel Lymph Nodes in Early Breast Cancer for Identification of Thymidylate Synthase as a Potential Biomarker to Flag Metastasis: A Preliminary Study

Authors Pathania S, Khan MI, Kumar A, Gupta AK, Rani K, Ramesh Parashar T, Jayaram J, Ranjan Mishra P, Srivastava A, Mathur S, Hari S, Hariprasad G

Received 26 March 2020

Accepted for publication 21 May 2020

Published 23 June 2020 Volume 2020:12 Pages 4841—4854


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Dr Sanjeev Srivastava

Video abstract presented by Gururao Hariprasad.

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Sheetal Pathania,1,* Mohd Imran Khan,1,* Abhishek Kumar,1 Ashish Kumar Gupta,1 Komal Rani,1 Tanvi Ramesh Parashar,1 Jnaneshwari Jayaram,2 Piyush Ranjan Mishra,2 Anurag Srivastava,2 Sandeep Mathur,3 Smriti Hari,4 Gururao Hariprasad1

1Department of Biophysics, All India Institute of Medical Sciences, New Delhi 110029, India; 2Department of Surgery, All India Institute of Medical Sciences, New Delhi 110029, India; 3Department of Pathology, All India Institute of Medical Sciences, New Delhi 110029, India; 4Department of Radiology, All India Institute of Medical Sciences, New Delhi 110029, India

*These authors contributed equally to this work

Correspondence: Gururao Hariprasad Tel +91-11-26594240
Fax +91-11-26588663

Introduction: Breast cancer is the second most common cancer in women across the world. Some of the patients who present in the early stage of disease are affected by metastasis to the axillary group of lymph nodes. The first among this group that is affected is called as sentinel lymph node, and its diagnosis is crucial for the staging of cancer thereby dictating the type of surgical therapy. Therefore, the sentinel lymph node status provides the most relevant information to the surgeon and patient prognosis. The expanded utilization of breast conservation surgery has declined the morbidity associated with mastectomy and axillary lymph node surgery. Recent interest is, therefore, centered on techniques that allow accurate assessment of the sentinel lymph node metastasis. A current procedure such as sentinel lymph node biopsy (SLNB) that is used to assess axillary lymph node metastasis is neither specific nor sensitive, and besides, it is time-consuming.
Objective: To compare the protein profiles between metastatic and non-metastatic lymph nodes to identify a biomarker that can flag lymph node metastasis.
Materials and Methods: Women with early breast cancer were screened using mammography imaging and recruited to the study. Surgical resection was done to remove the breast tissue, and sentinel lymph node was identified using fluorescein and methylene blue tracer. Lymph node was sliced, and one set was sent for histopathology, which was considered the gold standard to assess the metastatic status of the lymph node. One set of slices was taken for proteomic experiments. Proteins were labelled with fluorescent cyanine tags and were subjected to difference gel electrophoresis experiment. Differentially expressed spots that had at least a twofold relative ratio and consistent pattern across three sets of biological replicate experiments were marked. Gel spots were trypsin digested and identified on mass spectrometry machine. Validation study was done by Western blot experiment on the same set of samples.
Results: Thymidylate synthase has a twofold higher expression in the metastatic sentinel lymph nodes as compared to non-metastatic lymph nodes in early breast cancer patients.
Conclusion: Differential in gel expression proteomics is an ideal platform for the identification of potential protein biomarker candidates that can differentiate metastatic from non-metastatic lymph nodes in early breast cancer. The identification of thymidylate synthase offers a scope to develop an on-table diagnostic kit to assess the status of sentinel lymph nodes during mastectomy procedure to guide surgical management of axillary lymph nodes in early breast cancer.

Keywords: early breast cancer, sentinel lymph node metastasis, gel-based proteomics, difference gel expression, biomarkers, thymidylate synthase, surgical intervention

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