Understanding the Mechanism of 5-ASA in Treating Colonic Inflammation

G&H How was the link between 5-aminosalicylic acid (5-ASA) and peroxisome proliferator-activated receptor-γ (PPARγ) initially established?

PD The relationship between 5-ASA and PPARγ was first demonstrated in an experimental model utilizing genetically engineered heterozygous knockout mice, in whom colitis symptoms were induced and 5-ASA administered. The anti-inflammatory effect of 5-ASA in these mice was compared to that in control mice expressing normal levels of PPARγ in the colon. By evaluating microscopic and histologic lesions and levels of tumor necrosis factor-α present in the colonic mucosa, it was found that optimal concentration of 5-ASA in the colon had the greatest anti-inflammatory effect in control mice. In the PPARγ knockout mice, anti-inflammatory effects were greatly lessened. Thus, we conclude that the antiinflammatory effects of 5-ASA are mediated, at least in part, by PPARγ.

G&H What is the role of gut flora in the activation of PPAR γ expression?

PD In the colon, PPARγ is mainly expressed in epithelial cells. The concentration of PPARγ in the colonic mucosa is 100 times greater than that present in the small bowel. This is true in both humans and rodents. We have evaluated PPARγ levels in germ-free mice, compared to mice with normal flora, and those with humanized flora, and we have shown that in germ-free mice, PPARγ levels in the colon are very low but when bacteria are introduced, levels of PPARγ expression are raised. In mouse models, PPARγ has been similarly stimulated with different fractions of live gut bacteria, killed gut bacteria, and lipopolysacchride (LPS), with the understanding that LPS is a key determinant in inducing PPARγ expression and activation. Thus emerges a balance achieved between activity of bacteria and activity of LPS, wherein the former induces inflammation via the epithelial pathway and the latter enhances PPARγ expression activation.

G&H How does this balance between bacteria-mediated inflammation and LPS-induced PPARγ play a part in inflammatory bowel disease manifestation and treatment?

PD The expression of PPARγ is more closely related to the pathophysiology of ulcerative colitis (UC) than Crohn's disease. In patients with UC, there is an unexplained downregulation of expression of PPARγ relative to control patients, and it seems that the downregulation of PPARγ in the colonic mucosa is indicative of the level of inflammation. 5-ASA is a ligand of PPARγ, as has been demonstrated in animal models of UC as well as in vitro, utilizing colonic epithelial cells. As a ligand, 5-ASA binds to PPARγ, and this binding is functional, leading to activation of PPARγ that has an anti-inflammatory effect.

G&H Are there pharmacokinetic similarities between 5-ASA and diabetes therapies that also target PPARγ (ie, thiazolidinediones)?

PD There are similarities in the chemical structures of 5-ASA and thiazolidinediones (TZDs). The main difference is that with 5-ASA there is an almost exclusively topical effect on PPARγ expressed by the epithelial cells, with some minor residual effect on PPARγ expressed in the lymphocytes or macrophages within the mucosal spaces. The TZDs, in contrast, promote systemic expression of PPARγ. 5-ASA, even when orally administered, is mainly concentrated in the terminal and colonic ileum. Even when 5-ASA crosses the gut barrier it is for the most part not free but N-acetylated 5-ASA. Due to this N-acetylation, 5-ASA is not available to bind with and activate PPARγ throughout the body.

G&H What are the next steps in research of 5-ASA and PPARγ as they relate to UC treatment?

PD We need to better understand why the down-regulation of PPARγ expression and activation occurs in patients with UC. As of yet, the etiology of this abnormality remains unknown. It may be due to individual differences in the make-up of colonic flora or the expression of different polyunsaturated fatty acids, which may also regulate levels of PPARγ. Each of these concepts requires further investigation in order to better understand the pathophysiology of UC. We also need to find new compounds that combine the topical activity of 5-ASA with a stronger affinity to PPARγ. The adverse cardiovascular events that have been associated with TZDs stem from their systemic effects, which activate a number of different PPARγ-mediated pathways. In order to retain the excellent safety profile enjoyed by 5-ASA formulations, new compounds must remain topically active.

G&H What progress has been made toward the development of new, more effective compounds?

PD Because we know that 5-ASA is a ligand of PPARγ, we should, theoretically, be able to develop new chemical compounds, similar to 5-ASA but with a stronger affinity to PPARγ with some ease. At present, we have developed several molecules with similarities to 5-ASA, and we have now selected one compound, GED-0507-34 (Giuliani S.p.A., Milan, Italy), which has demonstrated 100–150-fold more anti-inflammatory activity than 5-ASA. This compound holds great promise in in vitro and in vivo experimental models of colitis, and the specificity of this molecule appears to be very good, avoiding any adverse events. Our current goal is to begin phase I study of this molecule at the beginning of 2009.

Figure 1.

5-ASA binds to cytoplasmic PPARγ, inducing its translocation to the nucleus and conformational change. This re-arrangement promotes the recruitment of the co-activator DRIP and the assembly of the active heterodimer PPAR-RXR, leading to the activation of PPRE.

5-ASA

5-aminosalicylic acid

DRIP

vitamin D receptor-interacting protein

PPAR

peroxisome proliferator-activated receptor

PPRE

PPARγ response element

RXR

retinoid X receptor.