Clinically relevant ex vivo fatty acid profiles from a lipid model for colorectal adenocarcinoma
The challenge we face today is identification of the mechanisms through which dietary factors perturb fundamental fatty acid (FA) pathways in cancer cells. In principle, insight into a lipid model for the cancer entity under assessment is required. This prompted and investigation into FA role-players that drive cellular over-production and apoptotic pathways under circumstances where environmental factors impede essential fatty acid metabolism (EFAM) that contributes to colorectal cancer (CRC). Ultimately, better understanding of FA metabolic pathways followed under colorectal pathological conditions can contribute to improved therapies by which growth and recurrence of this disease may be obviated. PURPOSE OF STUDY: The primary goal of this study was to construct clinically relevant ex vivo FA profiles from a lipid model for colorectal adenocarcinoma, not previously reported in the literature. Aims were to identify prominent FA role-players and to debate their involvement in different signaling pathways. The final purpose of this study was to present a rationale for adjuvant FA therapies to improve the management of CRC. MATERIAL AND METHODS: The study group consisted of CRC biopsies (TNM staging, mostly T2/T3) and normal colorectal mucosal biopsies, (n8 of each group). Biopsy selection was hampered by difficult circumstances, since most patients received treatment prior to surgery. All the lipid analyses were personally conducted at the Biolipid Division of the Department Biochemistry at the University of the Free State (UFS) as follows: lipids were extracted; total lipids (TLs) were fractionated into neutral lipids (NLs) and phospholipids (PLs); phosholipid subclasses, i.e. phosphatidylcholine, (PC), phosphatidylserine (PS), phosphatidylethanolamine (PE) and phosphatidylinositol (PI), were separated; and fatty acid methyl esters of all the classes and subclasses were analyzed. Lipid analyses were done according to well established and internationally accepted gaschromatography (GLC) and thin layer chromatography (TLC) techniques. Statistical analyses were performed by a biostatistician attached to the UFS, and p-values and 95% confident intervals (95% CI) for median of differences, as well as specific ratios were calculated. MAIN RESULTS AND DISCUSSION: The ex vivo phospholipid FA profile for CRC cells revealed linoleic acid (LA), arachidonic acid (AA), palmitic acid (PA) and oleic acid (OA) as main role-players involved in CRC. An extensive literature study revealed that: prolonged inflammatory responses and oxidative events, marked by the up-regulation of cyclooxygenases (COXs) and inducible nitric oxide synthase (iNOS), contribute to colorectal carcinogenesis; and prolonged cytokine mediated responses, orchestrated by nuclear factor-kappa Beta (NF-κβ) with consequent immunodeficiency, are prevalent in CRC patients. Past research revealed without doubt that high dietary AA intake (meat) and COX-2 metabolism play a pivotal role during colorectal carcinogenesis. CRC is a stress-related cancer and it is plausible that high iron intake and lipids contribute to chronic inflammation and radical-mediated cell damage in this age-related disease. Current research directions in CRC concern LOX metabolism and peroxisome proliferator activator receptors (PPARs) are receiving much attention, especially PPARδ/β that was an enigma and elicited controversial debate in adenomas (polyps) and adenocarcinoma. Taken together at this point in time, the impact of environmental factors on essential fatty acid metabolism (EFAM) that impede LA conversion to AA via delta-6 and -5 desaturase (Δ6d and Δ5d) pathways with up-regulation of the fatty acid synthase (FAS) and Δ9d pathways, and the down-regulation of LA oxidative metabolism via the 15 LOX-1 pathway with up-regulation of the COX-2 and 15- LOX-2 pathways of AA, is characteristic of colorectal carcinogenesis. Seen in context, the lipid findings of this study integrated with evidence-based information in the literature encompasses the following: excessive dietary linoleic acid (LA) intake that can contribute to cell proliferation, radical-mediated cell damage, and the production of conjugated linoleic acid (CLA) in the gut; high dietary AA intake and COX-2 over-expression that play a pivotal role during carcinogenesis and immune responses with consequent exhaustion of the AA source; enhanced FAS and Δ9d activities that contribute to, respectively, enhanced palmitic acid (PA) and oleic acid (OA), is prevalent in CRC cells. The high PA content in CRC cells apparently activates PPARδ/β and thereby suppresses apoptosis, a crucial factor in the pathobiology of CRC. The link between OA and CRC still needs proper clarification. It is conceivable that the high saturated fatty acid (SFA) content observed in CRC cells, exogenous intake with red meat and endogenous production by enhanced FAS activity, contribute to colorectal carcinogenesis and immunodeficiency in CRC patients, since it can stimulate COX-2 expression (and PGE2 activity) and downregulate the Th1 immune pathway that make CRC a Th2 dominant disease. Taken in consideration that lipid rafts rich in SFAs down-regulate the Th1 cytokine subset, particularly interleukin-2 (IL-2) that stimulates lymphocyte production, it is plausible that management of the CRC patient is hampered by immunodeficiency. All the altered cell signaling pathways in CRC debated in this study served as sound foundation for clinical intervention with adjuvant FA therapeutic strategies to improve CRC management. CONCLUSION: The identification of prominent FA role-players in CRC cells that serve as ligands for specific PPAR family members contributed to the assessment of FA driven proliferation and apoptotic pathways characteristic of CRC. It is conceivable that enhanced PA (and LA) contributes to PPARδ/β over-expression that can suppress PPARγ (and PPARα) activity and then tilts the scale in favor of apoptotic resistance and survival of CRC cells. From an epidemiological viewpoint, excessive iron and SFAs seem to be important co-factors in the multifactorial etiology of CRC. The biohydroxygenation of LA to different CLA products in the gut and to what extent harmful CLA products may pose a danger need to be fully explored, especially since the uptake of beneficial products is apparently limited. Among current therapeutic options in the field of lipids, concurrent therapy with a COX-2 inhibitor (Celecoxib) and docosahexaenoic acid (DHA) to improve membrane fluidity and the impact of drug therapy entered clinical trials and although the outcome is still awaited, the impact of DHA on lymphocyte production may be a concern. Based on a mountain of evidence presented in this study, it is suffice to say that there is a rationale for additional adjuvant FA therapies that can be included in the existing therapeutic regime for CRC management. Firstly, CLA and EPA therapy for prevention of polyp recurrence and improvement of immunocompetence in CRC patients and secondly, GLA and EPA therapy in the management of patients with a history of polyp growths is proposed.