Papers

• Glucocorticoids (GC) are synthesised from cholesterol through steroidogenesis. Predominantly in the adrenal cortex, although local GC production also has been reported in the lungs, intestine and skin.

• Glucocorticoids are lipophilic and diffuse easily through the cell membranes. Once in the cell cytosol, GCs bind to glucocorticoid receptors (GR) and translocate into the nucleus as GC–GR complexes via binding to proteins known as importins. In the nucleus, the GR interacts with DNA and proteins to alter gene expression.

  

• Following oral administration of prednisone, prednisolone concentrations in the plasma of healthy dogs are approximately sixfold higher than the concentrations of prednisone
• Ciclosporin A (CsA) is derived from the soil fungus Beauveria nivea
• Ciclosporin A is classified as a calcineurin inhibitor; its primary immunosuppressive effect is the inhibition of T-lymphocyte function
• Activated calcineurin dephosphorylates nuclear factor of activated T cells (NFAT), thereby allowing its translocation into the nucleus, which sub- sequently upregulates transcription of genes important for innate and adaptive immunity such as interleukin (IL)-2, IL-4, tumour necrosis factor (TNF)-α and TNF- γ. Interleukin-2 in particular, is a potent T-lymphocyte stimulator: it induces proliferation and differentiation of naïve CD8+ T cells into effector T cells by promoting secretion of granzyme B and perforins; it stimulates the proinflammatory activity of T helper (Th)-dependent B cells, antigen-presenting cells, mast cells, basophils and eosinophils; and it influences the differentiation of CD4+ T cells into Th1 or Th2, and subsequently affects the proliferation and differentiation of natural killer (NK) cells and B cells
• Ciclosporin A acts by binding to intracellular cyclophilin, which creates a complex that has a high affinity for calcineurin. The binding of the ciclosporin–cyclophilin complex with calcineurin inhibits its function of dephosphorylating NFAT and thus prevents the translocation of NFAT into the nucleus. Without NFAT in the nucleus, the aforementioned inflammatory cytokines are not transcribed and full T-lymphocyte activation is impaired.
• Ciclosporin is primarily metabolised in the liver by the cytochrome P450 3A (CPY3A) family of metabolising enzymes. Several drugs that also are metabolised by this enzyme will affect the blood concentration of CsA if given concurrently. Of these drugs, azole anti- fungals (e.g. ketoconazole and fluconazole) are often used along with CsA (especially in large dogs) because they decrease the metabolism of CsA, and therefore increase blood and skin CsA concentrations. This enables a CsA dose reduction of 50%–70%,45 which reduces the overall cost of therapy.
• The most common adverse effect reported with CsA is gastrointestinal (GI) upset, with vomiting (28%) and diarrhoea (14%) the most frequently reported clinical signs in dogs Gastrointestinal upset is usually transient and resolves with dose reduction. Anecdotally, storing CsA capsules (Atopica) in the freezer for 30–60min before oral administration reduces the incidence of vomiting, and the author has found this beneficial in preventing vomiting in some dogs. When stored in a –20°C freezer for one month, the stability and absorption of CsA in dogs is not impacted.
• Other adverse effects include gingival hyperplasia, cutaneous papillomatosis and op- portunistic infections. Opportunistic bacterial (Nocardia spp., Burkholderia cepacian complex)81–83 and fungal (Alternaria spp., Curvularia spp. and Aspergillus spp.)

Similar to dogs, the most common adverse effects of oral CsA in cats are diarrhoea and vomiting. In cats that were previously infected with feline herpesvirus-1 (FHV-1), administration of oral CsA at the dose of 7mg/ kg/day for 42days could result in reactivation of FHV-1 yet the clinical signs were mild and self-limiting. Acute fatal toxoplasmosis secondary to CsA therapy has been reported in cats yet is still considered rare.

Azothioprine It is a pro-drug of 6-mercaptopurine (6-MP) that exerts its immunosuppressive effects by interfering with nucleotide synthesis. Because AZA is a cytotoxic drug, it should be used with caution with other alkylating agents that interfere with DNA synthesis (e.g. cyclophosphamide) as it may lead to profound myelosuppression. There is some evidence that concurrent use of GC and AZA may increase the risk of acute pancreatitis.

CHLORAMBUCIL Chlorambucil is an anticancer drug from the nitrogen mustard group. Chlorambucil is generally well-tolerated in cats and dogs, although GI effects such as inappetence, vomiting and diarrhoea may be seen. However, cytotoxic myelosuppression can occur 7–14days after initiation of treatment. Other less common adverse effects include reversible myoclonus and Fanconi syndrome, both of which were reported only in cats.

MYCOPHENOLATE MOFETIL Mycophenolate mofetil (MMF) is a pro-drug that is converted to mycophenolic acid (MPA), where it exerts its pharmacological activity. Interestingly, MPA was first isolated from Penicillium stonoliferum in 1913 where it was discovered to possess antibiotic, antiviral and anti- inflammatory properties.

Mycophenolate mofetil exerts its immunosuppressive effect by inhibiting the formation of guanine nucleotides.

OCLACITINIB Oclacitinib is a Janus kinase (JAK) inhibitor Janus kinases are intracellular, nonreceptor tyrosine kinases. They reside in the cytoplasm of cells and are attached to the intracellular/proximal portion of Type I and Type II cytokine receptors. There are four family members of the JAKs—JAK1, JAK2, JAK3 and TYK2— and they play an important role in the transduction of cytokine-mediated signals through the JAK–signal transducers and activators of transcription (JAK–STAT) pathway. Upon binding of a ligand (e.g. cytokines and growth factors) to the receptor, the JAKs are activated and phosphorylate the intracellular domain of the receptor, thereby creating docking sites for the STAT molecules, which are signalling proteins. Once docked, the STAT molecules are further phosphorylated by the JAKs and then released back into the cytoplasm, where they form dimers with other phosphorylated STAT molecules. The dimerised STAT molecules translocate into the nucleus and bind with the DNA to transcript genes that regulate immunity, inflammation and haematopoiesis. Cytokine receptors can be grouped based on which JAKs are associated with the receptor complex.

While cytokines involved in allergy, inflammation and pruritus bind receptor complexes that utilise JAK1, the binding of other cytokines or growth factors involved in haematopoiesis (e.g. erythropoietin) and innate immunity (e.g. IL-12) activate receptors which are associated with the pairing of JAK2/JAK2 or JAK2/TYK2.156 Higher extra-label doses of oclacitinib are associated with immunosuppressive activity. Lymphocyte- enriched cells treated with 10μM of oclacitinib (equivalent to 3–4mg/kg twice daily) resulted in reduced secretion of IL-2, IL-15, IL-18 and IFN-γ, which inhibits the proliferation of T cells. Higher doses of oclacitinib also have been shown to induce apoptosis of canine CD4+ and CD8+ T cells in vitro. In a more recent study, oclacitinib prevented the generation (rather than inducing depletion) of regulatory T cells and the production of IL-10, which are important to maintain immune tolerance. Taken together, higher doses ofoclacitinib or doses used concurrently with another immunosuppressive drug can have immunosuppressive effects on the immune system.

Pharmacokinetics and pharmacodynamics Adverse effects Owing to the MoA, higher doses of oclacitinib could lead to impaired T-cell proliferation and increased risk of infection. Indeed, during the initial safety study, oclacitinib induced cutaneous papillomas, demodicosis and bacterial pneumonia in several 6- and 12-month-old dogs (package insert; Apoquel, Zoetis). In a retrospective study of 53 dogs with AD treated with prolonged twice-daily administration of oclacitinib (0.4–0.6mg/ kg twice daily), pyoderma, gastrointestinal signs and otitis externa were the most common adverse effect reported; only three dogs developed mild neutropaenia. In an feline immunodeficiency virus (FIV)-positive cat, treatment with oclacitinib (1mg/kg twice daily) for feline atopic skin syndrome resulted in fatal disseminated toxoplasmosis; the clinical signs developed fivemonths after initiation of this therapy. In cats, doses of 1–2 mg/kg every 12 h for 28 days were shown to be safe and well-tolerated, although the higher doses did result in gastrointestinal signs in two of 10 cats. Long-term studies with a large number of cats have not been performed and therefore, the long-term safety of oclacitinib in cats is as yet unknown. At present, oclacitinib is not licensed for use in cats.

BRUTON'S TYROSINE KINASE INHIBITOR Bruton's tyrosine kinase (BTK) is an important signalling protein that serves as a link between the B-cell receptor (BCR) and B-cell proliferation and survival. Adverse effects reported from the study involving dogs with PF treated with PRN473 included immune-mediated polyarthritis (n = 1; Week 12), mast cell tumour (n = 1; Week 4), peripheral lymphad- enopathy (n = 2) and pancreatitis (n = 1). Of the four dogs that received PRN1008 for canine PF, only one dog had pyometra, and increased ALT and aspartate aminotransferase. In tolerability studies using an unrelated BTKi (G278), hepatotoxicity was reported in dogs.

• It has been observed that different pruritogenic compounds sensitise the TRPV1 channel in neurons through different intracellular signalling cascades and transmit the signal to the brain where the scratch reflex is produced.
• For example, some authors report that the pruritic action of interleukin (IL)-31 produced by T helper 2 (Th2) cells occurs through the IL-31RA colocalised in TRPV1-positive neurons on the dorsal root ganglion.
• Their experiments in vitro and in vivo in a murine model demonstrate that both TRPV1 and TRPA are critical for IL-31 to evoke the sensation of itch through a neuro-immunological circuit where T cells and sensory neurons participate.
• Among the TRPV1 antagonists is the hydroxymethoxy- iodobenzyl glycoamide pelargonate (HMBG) molecule.

• Doxycycline and ketoconazole -> 15 days later erythema and crusts over dorsal muzzle (sparsely haired area) and bilaterally on dorsal aspect of pinnal folds -> doxycycline withdrawn and crusts resolved
• Phototoxicity is a well-known adverse effect of doxycycline use in humans and is mediated by excited-state singlet oxygen and free radical damage to mitochondria -> signal transduction pathways that lead to the production of proinflammatory cytokines and arachidonic acid metabolites also are activated, resulting in an inflammatory response that has the clinical appearance of an exaggerated sunburn reaction.