Table 2.

In vivo studies of NOB for colon cancer chemoprevention.

Animal Models	Treatment/Dosage	Mechanisms	Detailed Results	References
Colitis-associated colon carcinogenesis model -AOM (12 mg/kg i.p.)/1% DSS in drinking water treated male CD-1 mice (5-week-old)	AIN93G diet containing 0.05% wt NOB (20 weeks)	Cell cycle arrest	Protein expression in colonic mucosa by Western blot - Reduced levels of CDK-2, CDK-4, CDK-6, cyclin D and cyclin E - Increased levels of p21, p27 and p53	[76]
		Anti-inflammatory effects	Immunohistochemical analysis - Reduced expression of iNOS reduced by 35% when compared to the positive control Protein expression in colonic mucosa by Western blot - Increased level of HO-1 - Increased level of NQO1 - Induced translocation of level of Nrf2 transcription factor (Nuclear fraction < Cytoplasmic fraction)
Colitis-associated colon carcinogenesis model -AOM/DSS treated AOM (12 mg/kg i.p.)/1% DSS in drinking water treated male CD-1 mice (5 week old)	AIN93G diet containing 0.05% wt NOB (20 weeks)	Inhibit AOM/DSS-induced colon carcinogenesis	- Prevented shortening of colon length, reduced the increased colon weight/length ratio - Reduced tumor incidence by 40% and tumor multiplicity by 71% - Maintained histological characteristic of normal mucosa	[54]
		Anti-proliferative effect	- Reduced PCNA-positive colonocytes by 69% in mucosal crypts
		Apoptosis-inducing effect	- Increased cleaved caspase-3 positive cells by 2.3-fold in colonic tumor
		Anti-inflammatory effects	- Reduced levels of proinflammatory cytokines - ELISA showed reduction of TNF-α by 51%, IL-1ß by 92% and IL-6 by 69% compared - qRT-PCR analysis showed reduction of TNF-α by 65%, IL-1ß by 69% and IL-6 by 45%
Colon carcinogenesis model -AOM (15 mg/kg i.p.) treated male db/db mice	Diet containing 100 ppm NOB (0.1% wt) (10 weeks)	Inhibit AOM induced colon carcinogenesis	- Reduced frequency of preneoplastic lesions (colonic aberrant crypt foci (ACF) and β-catenin-accumulated crypts (BCAC)) - Reduced incidence of ACF by 68-91% and BCAC by 64–71% - Reduced PCNA-labeling index in ACF by 21% and BCAC by 19%	[83]
Colon carcinogenesis model -AOM (10 mg/kg i.p.)/1% DSS in drinking water treated male CD-1 mice	Diet containing 100 ppm NOB (0.1% wt) (for 17 weeks)	Inhibit AOM/DSS-induced colon carcinogenesis	- Suppressed incidence of neoplasms (adenoma and adenocarcinoma), lowered multiplicity of tumor	[84]
		Inhibit leptin-induced colon carcinogenesis
			- Suppressed serum levels of leptin by 75–84%
Colon carcinogenesis model -AOM (20 mg/kg s.c.) treated male F344 rats	Diet containing NOB (0.01% wt and 0.05% wt) (34 weeks)	Inhibit AOM induced colon carcinogenesis	- Reduced incidence and multiplicity of colonic adenocarcinoma	[74]
		Anti-proliferative effect
			- Increased apoptosis index of adenocarcinoma
		Anti-inflammatory effect
			- Reduced level of PGE2 in colonic adenocarcinoma and surrounding mucosa
Colon carcinogenesis model -AOM (20 mg/kg s.c.) treated male F344 rats	Diet containing NOB (0.01% wt and 0.05% wt) (5 weeks)	Inhibit AOM-induced colon carcinogenesis	- Reduced the frequency of colonic aberrant crypt foci formation - Reduced number of ACF in proximal, middle and distal colon	[41]
		Anti-proliferative effect
			- Reduced MIB-5 labeling index of ACF but not of normal colonic crypts
		Anti-inflammatory effect
			- Reduced level of PGE2 in colonic mucosa
Colon carcinogenesis model -PhIP hydrochloride (100 mg/kg i.g.) treated F344 male rats (twice/week for 10 weeks)	Diet containing NOB (0.05% wt.) (50 weeks)	Inhibit PhIP-induced ACF in transverse colon	- Reduced the total colonic ACF indices in transverse colon	[75]
Colorectal cancer xenograft mouse model -COLO205 cells s.c.	NOB 100 mg/kg i.p. daily for 3 weeks 5-DMN 50 mg/kg and 100 mg/kg i.p. daily for 3 weeks	Anti-tumor effect	- NOB reduced tumor size and weight but not significant as compared to control - 5-DMN reduced tumor size and weight significantly as compared to control	[49]
		Autophagy induction	- 5-DMN increased LC3 expression
		Anti-inflammatory effect
			- 5-DMN increased p53 expression - 5-DMN reduced COX-2 expression
		Anti-angiogenesis
			- 5-DMN reduced VEGF expression

AOM—azoxymethane; DSS—dextran sulfate sodium; i.p.—intraperitoneal injection; s.c.—subcutaneous injection; i.g.—intragastric administration; PCNA—proliferating cell nuclear antigen; ACF—aberrant crypt foci; BCAC—β-catenin-accumulated crypts; PhIP—1-Methyl-6-phenyl-1H-imidazo[4,5-b]pyridin-2-amine; LC3—microtubule-associated protein light chain 3; VEGF—vascular endothelial growth factor.