Characterisation of the effect of novel anti-cancer components of Pseudechis australis and Crotalus vegrandis snake venom on acute lymphoblastic leukaemia cells

James Boncan, Karen McCloskey, Ken Mills, Sara Dobbin, Diego Cobice, Stephen McClean

Research output: Contribution to journalMeeting abstractpeer-review

Abstract

Introduction/Background & aims:
Acute lymphoblastic leukaemia(ALL) is a haematological malignancy affecting lymphoid lineage cells accounting for approximately 230 UK deaths annually. Standard of care-therapies for ALL are poorly tolerated and have poor clinical responses. Snake venom represents a natural source of bioactive chemicals with numerous in vitro studies identifying several venom components with anti-cancer activity [1]. Here, we aimed to identify active components within crude snake venom and characterise their mechanisms in ALL cells.

Method/Summary of work:
Reh and NALM6, B cell precursor ALL cell lines were used. Crude snake venom from Pseudechis australis(PA) and Crotalus vegrandis (CV) was fractionated using gel-filtration chromatography with active fractions subject to MALDI-ToF mass spectrometry [2]. Fractions were numbered sequentially. Cell viability and cytotoxicity was assessed using Cell Titer-Glo luminescence and CellTox-Green assays, respectively. Lactate dehydrogenase (LDH) release was measured with LDH-Glo cytotoxicity assay kits and luminescent caspase 3/7 assays were used to study apoptosis. In functional assays, at least N= 3 were performed; occasionally only N= 2 was achievable due to limited snake venom fraction volume. Data sets were compared with ANOVA with p< 0.05 considered as significant. 

Results/Discussion:
In viability assays using Reh cells, PA venom fractions, PA60-63 and PA68-71 reduced viability after 24, 48, and 72 h(N=3,p< 0.05). MALDI-ToF analysis of these fractions revealed dis-tinct peaks of 13 kDa, indicative of the phospholipase A2 family of enzymes. Reh cells treated with a commercially bought phospholipase A2isolated from Crotalus adamanteus snake venom also displayed reduced cell viability at 24, 48, and 72 h (IC5063.5μg/ml, 58.7μg/ml, and 57.9μg/ml, respectively, N=3,p< 0.05). In NALM6 cells, CV venom fractions, CV110-115, CV125, CV131-137, and CV139-149reduced cell viability (N=3;p< 0.05). MALDI signals were observed for CV110–115, CV125, and CV131–133 venom fractions, at 9 kDa and 11 kDa, slightly below the phospholipase A2range (13 kDa). Reh cell cytotoxicity was increased by active PA fractions (N=3,p< 0.05). CV112, CV114, CV132, and CV134 evoked cytotoxicity and loss of NALM6 membrane integrity (N= 2). LDH release was not evoked in PA venom-treated Reh cells (N=3,p< 0.05). Finally, neither PA-treated Reh nor CV-treated NALM6 exhibited increased caspase 3/7 activity (N= 2).

Conclusion(s):
PA and CV-derived venom fractions reduced ALL cell viability. MALDI-ToF analysis identified phospholipase A2 family as an anticancer component of interest within PA snake venom. Functional assays highlight two potential mechanisms for reduced viability in PA-treated Reh and CV-treated NALM6; both appear to be apoptosis- and necrosis-independent,  indicating  suppression of ALL cell proliferation.

Original languageEnglish
Pages (from-to)468-469
Number of pages2
JournalBritish Journal of Pharmacology
Volume178
Issue number2
DOIs
Publication statusPublished - 08 Dec 2020
EventPharmacology 2020 - Online (Virtually)
Duration: 14 Dec 202018 Dec 2020
https://live.bethereglobal.com/s/pharmacology-2020

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