Endocrine Abstracts

Context: Cushing’s disease (CD) results from the release of high levels of adrenocorticotrophic hormone (ACTH) from a pituitary adenoma. Increased ACTH secretion stimulates excess cortisol production, causing weight gain, hypertension, diabetes and depression. The only curative treatment is transsphenoidal surgery, but the rate of recurrence is high and there is a lack of suitable medical therapies. RNA-interference is a mechanism of post-transcriptional gene silencing that can be utilised to knock-down the expression of specific genes using small interfering RNAs (siRNA). ACTH is encoded by the POMC gene, so anti-POMC siRNAs might be an effective treatment for CD through silencing POMC expression and reducing ACTH secretion.

Methodology: The effectiveness of RNA-interference at reducing POMC expression was analysed by transfection of murine AtT-20 cells with three anti-POMC siRNAs followed by measurement of extracellular ACTH concentrations by immunoassay. Control transfections were siRNAs with no target sequence and mismatched anti-POMC siRNAs. Cell viability was assessed by trypan blue staining. Induction of IFN-α and IFN-β expression by siRNAs was measured in ELISAs.

Results: At 24 h post-transfection, anti-POMC siRNAs at 10 or 30 nM reduced secretion of ACTH by 81-89% compared with untreated cells (P<0.001). Control siRNAs had no effect upon ACTH secretion. Suppression of ACTH secretion was maintained for upto four days post-transfection. Viable cell counts remained equivalent whether cells were transfected or untreated. Anti-POMC siRNAs did not induce detectable IFN-α or IFN-β.

Conclusions: The results indicated that anti-POMC siRNAs significantly decreased ACTH secretion by AtT-20 cells. Specificity of the anti-POMC siRNAs was indicated by the ineffectiveness of anti-POMC siRNAs with nucleotide mismatches. ACTH reduction was not caused by adverse effects of the transfection process. Anti-POMC siRNAs did not appear to induce an interferon response. RNA-interference by POMC-specific siRNAs could be a novel medical therapy for CD.