Pranav Patel (Presenter)
Imperial College London. London, UK.
Bio: I joined Professor Zoltan Takat’s research group in August 2015 as a clinical PhD Research Fellow at Imperial College London. I completed my undergraduate medical education (MBBS) honours degree at Imperial College London in 2008. During my undergraduate training, I intercalated my honours BSc in 2006, where my interest in surgical oncology and analytical sciences research began. In 2012, I commenced my postgraduate specialist residency program in General and Gastrointestinal Oncology Surgery. This enabled me to work in a nationally renowned cancer centre and commence my research. My current work is centred on the creation of a novel electrospray interface for analysis of aspiration cytology directly from the minimally invasive collection device. This has great potential in accurately diagnosing local disease spread more accurately to tailor oncological and surgical treatment.
Authorship: Pranav H Patel (1), Pamela Pruski (1), Nima Abbassi-Ghadi (1), Sacheen Kumar (1), Renata Soares (1), George B Hanna (1), Zoltan Takats (1)
(1) Imperial College London, London, UK.
| Short Abstract Oesophago-gastric cancer is the 5th most common cause of cancer in the world, however patients typically present late with advanced disease. Disease staging is crucial in Oesophago-gastric cancer, lymph node fluid cytology assessment is used to determine local disease spread. The accuracy of this technique can be variable. Here, we demonstrate a novel technique for the analysis of fluid cytology directly from the aspiration needle collection device using mass spectrometry. This method is based upon analysis of the cancer cell lipidome, and has proven to be rapid, easy and tissue type specific. Thereby being translational to alternative cancer cytology sub-types. |
Long Abstract
Introduction
Oesophago-gastric cancer is the 5th most common cause of cancer in the world, however due to the insidious nature of symptoms patients present late with advanced disease and local lymph node metastases. Only 38% of newly diagnosed patients are treated by curative intent as there is a paucity of symptoms in the early stages of the disease, with the majority of patients presenting with disease spread (1). Overall 5-year age-adjusted relative survival rates for oesophageal and gastric cancer being 15 and 20 per cent, respectively (2). Current staging protocols include CT imaging and Endoscopic Ultrasound (EUS), these modalities have good tumour specificity but poor for lymph node metastases. EUS with lymph node cytology had been used to bridge this diagnostic gap, however traditional analysis is cellular light microscopy based with some studies reporting a 0.54 negative predictive value (3). The clinical utility of newly developed techniques using mass spectrometry based analysis for fluid cytology, is still to be determined. Here, we demonstrate a quick, easy and selective method for the analysis of cytological fluid directly from the needle collection device using lipid-based cellular markers.
Methods
Methods:
The malignant cytology fluid was collected by direct needle aspiration of cellular constituents of lymph nodes dissected at time of surgical resection from ex-vivo surgical specimens. Samples were collected with matched lymph node tissue for classification by the current gold standard cellular histopathology. Needle analysis was performed using a novel direct analysis electrospray ionisation platform. The needle analysis stage enabled a direct 3.5kV current application to the needle with Methanol: Water (95:5) used as the solvent for sample elution and ionisation from the inner channel and analysed with a LTQ-Orbitrap Discovery (Thermo Fischer Scientific, Massachusetts, USA) high resolution mass spectrometer. Analysis was performed in both negative and positive ionisation modes in full scan mode to identify lipid structures. Raw data files were converted into mzXML format and subsequently peak aligned using a predefined chemometric toolbox programmed in Matlab. Supervised and unsupervised statistical modelling was performed with incorporation of clinical metadata. The lipid spectra acquired during the lymph nodal cytology analysis referenced to the gold standard cytopathology and tissue histopathology.
Troubleshooting steps:
Initial inconsistencies in sample elution from the needle inner channel were improved by developing an inner channel metal capillary to prevent solvent pooling in the base of the needle. Once applied a linear sample flow was established with uniform detection by the mass spectrometer during analysis.
Results
This method of analysis enabled the detection of lipid markers in fluid cytology by direct analysis. Samples were classified into cancerous and non-cancerous groups following histopathological assessment. Subsequent statistical analysis showed statistically significant clustering of lipid species in fat, benign and cancerous lymph node samples. The structure of the most relevant biomarker lipids was determined based on exact mass using Lipid Maps as a reference library. Phosphatidic acids (PA: m/z 782.58) and Phosphatidylglycerols (PG: m/z 782.54) were shown to be of greater abundance in the cancerous cytology compared to benign cytology.
Conclusions & Discussion
This proof-of-principle work shows that this novel analytical platform can be utilised to provide rapid, highly accurate identification of cancerous cytology to reduce diagnostic uncertainty. The clinical translation of this cytological identification technique to alternative solid and cystic tumours, such as thyroid and pancreatic cancers where cellular pathological identification is of variable specificity, would be profound.
References & Acknowledgements:
(1)-Cancer Research UK Statistical Information Team 2010. UK Oesophageal Cancer Statistics. [online] Available at: http://www.cancerresearchuk.org/health-professional/cancer-statistics/statistics-by-cancer-type/oesophageal-cancer/mortality
(2)-Cancer Research UK Statistical Information Team 2010. UK Stomach Cancer Mortality Statistics. [online] Available at: http://www.cancerresearchuk.org/health-professional/cancer-statistics/statistics-by-cancer-type/stomach-cancer/mortality
(3)-The NHS Information Centre, 2010. National Oesophago-gastric Cancer Audit 2010. [online] Available at: http://www.hscic.gov.uk/catalogue/PUB02758/clin-audi-supp-prog-oeso-gast-2010-rep1.pdf
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