3 June 2022 - - Data from US-based RNAi therapeutics company Alnylam Pharmaceuticals, Inc's (NASDAQ: ALNY) preclinical research on delivery of lipophilic short interfering RNA (siRNA) conjugates to extra-hepatic tissues, including the central nervous system, were published online in Nature biotechnology, the company said.

The published data provide early evidence of a potential role for 2'-O-hexadecyl -conjugated siRNAs in treating diseases of the CNS, eye, and lung.

The framework for designing siRNAs for extrahepatic applications leverages Alnylam's decades long investment in the siRNA delivery platform.

The full manuscript titled "Expanding RNAi therapeutics to extrahepatic tissues with lipophilic conjugates" will appear in the June issue of Nature biotechnology.

The published data show that conjugation of C16 to metabolically stable siRNAs enables robust and long-lasting gene silencing in the CNS, eye and lung in rodents and non-human primates with broad cell type specificity and a favorable nonclinical safety profile.

The manuscript also provides evidence demonstrating C16-siRNA-mediated gene silencing translates into efficacy in an in vivo mouse model of neurodegenerative disease.

Based on the published results, a single intrathecal administration of C16 conjugated siRNA targeting SOD1 in rodents resulted in a widespread uptake of the conjugate in neurons, astrocytes, and microglial cells and a dose dependent SOD1 knockdown with greater than 75 % target knockdown at the highest dose of 0.9 mg.

These results were recapitulated in NHPs treated with C16-conjugated siRNA targeting amyloid beta precursor protein, with up to 70 and 80% reductions in the spinal cord and brain, respectively, at three months post a single IT dose of 60 mg.

These reductions were sustained beyond three months based on biomarker measurements in the cerebrospinal fluid.

The APP-siRNAs were well-tolerated in NHPs and there were no test item-related microscopic findings in the examined brain, spinal cord, and dorsal root ganglia sections.

The published data also provide a pre-clinical proof-of-concept, whereby administration of a C16-siRNA targeting both intracellular and extracellular APP in a mouse model of Alzheimer's disease produced a sufficiently potent and durable knockdown in the CNS to alter both the physiological deficits in the diseased mice, such as Aβ deposition and inflammation, as well as, normalizing behavioral deficits as measured by an open field test.

Reduction of APP expression was correlated with reduced expression of Iba1, a marker normally upregulated in human disease, and increased levels of glutamate, a metabolite associated with improved cognition.

The safety and efficacy of ALN-APP, an investigational RNAi therapeutic conjugated to C16 and targeting APP, is currently being evaluated in a Phase 1 clinical study for the treatment of early onset Alzheimer's disease.

C16 conjugation also enables ocular delivery in NHP via intravitreal administration with greater than 95 % target knockdown noted in the retinal pigmented epithelium at 100 μg per eye of conjugated siRNA targeting transthyretin.

Similarly, intranasal administration of C16 lipophile conjugated siRNA targeting mouse SOD1 resulted in broad uptake of the conjugate to mouse lung tissue, including bronchioles and alveoli, and 57% SOD1 mRNA knockdown sustained for two months post a 10 mg/kg dose.

ALN-APP is an investigational, intrathecally administered RNAi therapeutic targeting amyloid precursor protein in development in collaboration with Regeneron Pharmaceuticals for the treatment of Alzheimer's disease and cerebral amyloid angiopathy.

Genetic mutations that increase production of APP or alter its cleavage cause early-onset AD, early-onset CAA, or both.

ALN-APP is designed to decrease APP mRNA in the central nervous system, to decrease synthesis of APP protein and all downstream intracellular and extracellular APP-derived cleavage products, including amyloid beta.

Reducing APP protein production is expected to reduce the secretion of Aβ peptides that aggregate into extracellular amyloid deposits and reduce the intraneuronal APP cleavage products that trigger the formation of neurofibrillary tangles and cause neuronal dysfunction in Alzheimer's disease.

ALN-APP is the first program utilizing Alnylam's C16 conjugate technology, which enables enhanced delivery to cells in the CNS. The safety and efficacy of ALN-APP have not been evaluated by the FDA, EMA, or any other health authority.