Reversal of pancreatitis-induced pain by an orally available, small molecule interleukin-6 receptor antagonist

Abstract

Pancreatic pain resulting from chronic inflammation of the pancreas is often intractable and clinically difficult to manage with available analgesics reflecting the need for more effective therapies. Mechanisms underlying pancreatitis pain are not well understood. Here, the possibility that interleukin-6 (IL-6) may promote pancreatitis pain was investigated with TB-2-081 (3-O-formyl-20R,21-epoxyresibufogenin, EBRF), a small molecule IL-6 receptor antagonist that was semi-synthetically derived from natural sources. The potential activity and mechanism of TB-2-081 was investigated following induction of persistent pancreatitis using dibutyltin dichloride (DBTC) in rats. TB-2-081 displaces binding of IL-6 to the human recombinant soluble IL-6 receptor with apparent high affinity and inhibits IL-6 mediated cell growth. Systemic or oral, but not intrathecal, administration of TB-2-081 reversed DBTC-induced abdominal hypersensitivity in a dose- and time-dependent manner. IL-6 levels were significantly upregulated in the dorsal root ganglia (DRG) of rats with pancreatitis on day 6 after DBTC injection. IL-6 enhanced capsaicin-evoked release of calcitonin gene related peptide from cultured DRG neurons was blocked by TB-2-081. Our data demonstrate that TB-2-081 acts as a systemically available and orally active small molecule IL-6 receptor antagonist. TB-2-081 effectively reduces pancreatitis-induced pain through peripheral mechanisms that are likely due to (a) increased expression of IL-6 in the DRG and (b) IL-6-mediated sensitization of nociceptive neurons. The activity of TB-2-081 implicates an important role for IL-6 in sustaining pancreatitis pain. Strategies targeting IL-6 actions through small molecule antagonists may offer novel approaches to improve therapy of chronic pancreatitis and other chronic pain states.

Preventive analgesia

“…Furthermore, we should expand our analgesic armamentarium from conventional NSAIDs, opioids and other nonopioid analgesics, including regional analgesic techniques to include interventions into more central pain mechanisms, including modification of the microglia response. In this context, the future may be bright and calling for studies of several drugs such as blocking nerve growth factor [29], modulating microglia activation by propentofylline [30] or minocycline [30,31], use of transient receptor potential cation channel, subfamily V, member 1 (TRPV1) antagonists [32] or various cytokine antagonists with a focus on interleukin-1b (IL1b) [33], IL-6 antagonists [34], or cannabinoid receptor antagonists [35]. In this context, we should not be naive and repeat previous history of short-term analgesic interventions, as optimal preventive treatment most probably will require a prolonged intervention until the peripheral inflammatory response and afferent input has resolved.…”

Chikusetsusaponin IVa Butyl Ester (CS-IVa-Be), a Novel IL6R Antagonist, Inhibits IL6/STAT3 Signaling Pathway and Induces Cancer Cell Apoptosis

“…IL6 receptor binding was performed following the method described previously (23). Briefly, 96-well plates (Nunc) were coated with 500 ng/mL of human recombinant soluble IL6 receptor (sIL6R), washed, and blocked.…”

Establishment of a novel cell-based assay for screening small molecule antagonists of human interleukin-6 receptor

“…The positive compounds were re-screened in a secondary screening to test and verify the results. A previous model described by Vardanyan et al [21] was used to examine the functions of the active compounds obtained by the preliminary screening and thus to assess the reliability of the novel assay (several negative compounds in the preliminary screening were included as a control). The procedure is as follows: the plates were coated with soluble IL-6R overnight at 4 °C and pre-blocked with 1% BSA at RT for 1 h. After three washes, the wells were incubated with human IL-6 plus one of eight concentrations of the compounds at RT for 2 h. After washing to remove any unbound IL-6, the wells were incubated with an anti-human IL-6 antibody (primary antibody) at 37 °C for 1 h, and washed and incubated with a secondary antibody (HRP conjugate) at 37 °C for 30 min.…”

“…Some methods derived from soluble receptor-ligand binding assays are also widely used in screening IL-6R antagonists [20,21] . However, each of the assays seemingly has pros and cons.…”

“…For example, the radioactive ligand-receptor binding test is considered to be an effective, sensitive, and classical analysis method, but safer methodologies than radiolabeled assays allow the use of radioactivity to be avoided [22] . A safe method based on the receptor-ligand binding assay mechanism is the enzyme-linked immunosorbent assay (ELISA); this is a quick, simple and extensive approach [21,23] . However, the ELISA still has room for improvement.…”