BACKGROUND: The direct link between inflammatory bowel diseases and colorectal cancer is well documented. Previous studies have reported that some lactic acid bacterial strains could inhibit colon cancer progression however; the exact molecules involved have not yet been identified. So, in the current study, we illustrated the tumor suppressive effects of the newly identified Lactobacillus acidophilus DSMZ 20079 cell-free pentasaccharide against colon cancer cells. The chemical structure of the purified pentasaccharide was investigated by MALDI-TOF mass spectrum, 1D and 2D Nuclear Magnetic Resonance (NMR). The anticancer potentiality of the purified pentasaccharide against both Human colon cancer (CaCo-2) and Human breast cancer (MCF7) cell lines with its safety usage pattern were evaluated using cytotoxicity, annexin V quantification and BrdU incorporation assays. Also, the immunomodulatory effects of the identified compound were quantified on both LPS-induced PBMC cell model and cancer cells with monitoring the immunophenotyping of T and dendritic cell surface marker. At molecular level, the alteration in gene expression of both inflammatory and apoptotic pathways were quantified upon pentasaccharide-cellular treatment by RTqPCR. RESULTS: The obtained data of the spectroscopic analysis, confirmed the structure of the newly extracted pentasaccharide; (LA-EPS-20079) to be: α-D-Glc(1→2)][α-L-Fuc(1→4)]α-D-GlcA(1→2)α-D-GlcA(1→2)α-D-GlcA. This pentasaccharide, recorded safe dose on normal mammalian cells ranged from 2 to 5 mg/ml with cancer cells selectivity index, ranged of 1.96-51.3. Upon CaCo-2 cell treatment with the non-toxic dose of LA-EPS-20079, the inhibition percentage in CaCo-2 cellular viability, reached 80.65 with an increase in the ratio of the apoptotic cells in sub-G0/G1 cell cycle phase. Also, this pentasaccharide showed potentialities to up-regulate the expression of IKbα, P53 and TGF genes. CONCLUSION: The anticancer potentialities of LA-EPS-20079 oligosaccharides against human colon cancer represented through its regulatory effects on both apoptotic and NF-kappa B inflammatory pathways.

Oligosaccharides, exopolysaccharide, LAB, Colon cancer, NFkB pathway, T cells immunophenotyping

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
2,2,2	aDGlcp	96.00	72.15	74.28	71.29	72.85	62.22
2,2,4	aLFucp	100.28	68.99	69.96	72.52	68.05	16.24
2,2	aDGlcpA	95.39	76.03	70.36	77.96	71.58	174.12
2	aDGlcpA	96.	75.84	71.53	72.16	72.62	176.15
	aDGlcpA	92.18	76.03	71.99	72.85	72.15	176.19


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
2,2,2	aDGlcp	4.68	3.53	3.73	3.43	3.99	3.73-3.83
2,2,4	aLFucp	4.55	3.78	3.80	3.78	4.19	1.27
2,2	aDGlcpA	5.08	3.73	4.12	3.73	4.18	-
2	aDGlcpA	5.14	3.67	3.91	3.56	4.18	-
	aDGlcpA	5.21	3.63	3.80	3.55	4.18	-


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
2,2,2	aDGlcp	96.00/4.68	72.15/3.53	74.28/3.73	71.29/3.43	72.85/3.99	62.22/3.73-3.83
2,2,4	aLFucp	100.28/4.55	68.99/3.78	69.96/3.80	72.52/3.78	68.05/4.19	16.24/1.27
2,2	aDGlcpA	95.39/5.08	76.03/3.73	70.36/4.12	77.96/3.73	71.58/4.18	
2	aDGlcpA	96./5.14	75.84/3.67	71.53/3.91	72.16/3.56	72.62/4.18	
	aDGlcpA	92.18/5.21	76.03/3.63	71.99/3.80	72.85/3.55	72.15/4.18	

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

C[C@@H]1O[C@@H](O[C@@H]2[C@@H](C(=O)O)O[C@H](O[C@H]3[C@@H](O[C@H]4[C@@H](O)O[C@H](C(=O)O)[C@@H](O)[C@@H]4O)O[C@H](C(=O)O)[C@@H](O)[C@@H]3O)[C@H](O[C@H]3O[C@H](CO)[C@@H](O)[C@H](O)[C@H]3O)[C@H]2O)[C@@H](O)[C@H](O)[C@@H]1O

854.67 g/mol (C₃₀H₄₆O₂₈)