Pro-inflammatory cytokine lowering anti-inflammatory drugs

National Institute On Aging

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Abstract

Tumor necrosis factor-alpha (TNF) is one of the primary pro-inflammatory cytokines synthesized and released by microglial cells with physiological and pathological roles. On release, TNF may initiate a self-propagating cycle of unchecked inflammation (Jung et al., Front Cell Dev Biol 7:313, 2019). Pharmacologic intervention to interrupt this cycle may be of benefit in neuroinflammation-mediated diseases. In 1993, Moreira et al., (J Exp Med 177:1675-80, 1993) reported that the drug thalidomide (THAL) lowered TNF protein levels post-transcriptionally by accelerating degradation of its mRNA. Unfortunately, THAL is not a particularly potent TNF lowering agent, and induces serious teratogenic adverse effects to embryos in utero, sedation and peripheral neuropathy at clinical doses. Nevertheless, knowledge of THALs TNF lowering activity gave rise to studies to differentiate these actions, understand THALs TNF structure/activity relationship and develop more potent analogs. Generation of analogs with enhanced anti-TNF activity and reduced teratogenic and neurotoxic effects may provide a viable treatment strategy for CNS neuroinflammatory and other inflammatory disorders. Our chemistry modifications to the backbone of THAL and newer analogs (i.e., pomalidomide (POM)) are generating a library of novel agents (issued US patents: 7,973,057 and 8,927,725, and U.S. Patent Application 62/235,105). Our focus is to identify well-tolerated drug-like compounds with more potent anti-TNF activity from our novel library and develop these as experimental drugs to define the role of neuroinflammation in and to treat Alzheimer's disease (AD) and associated disorders. Problem and Aims: AD is a complex disorder that manifests as progressive dementia with few other symptoms. With a long meandering course, AD is associated with deposits of amyloid-B protein (AB) some 20 years prior to dementia development. It also induces intracellular microtubule-associated protein Tau (MAPT) accumulation as neurofibrillary tangles (NFTs) that correlate more closely with dementia (Baranello et al., Curr Alzheimer Res 12:32-46, 2015). NFTs arise some 10 years after AB, and brain atrophy follows after 5 further years. Brain resilience and redundancy protects affected subjects from dementia for 5 further years after atrophy detection. Experimental drugs/strategies that reduce generation of AB oligomers and aggregates, as well as of p-tau have largely formed the treatment approaches thus far developed to combat AD development. Albeit these target are considered initiators of the cascade of events that become self-propagating and then drive AD progression, their toxicity may not be the direct cause of neurodegeneration. This premise may account for the failure of anti-amyloid and anti-tau therapies in clinical AD trials when administered late in the disease course (Becker et al., Nature Rev Drug Discov 13:156, 2014). Presence of soluble and insoluble AB and MAPT can induce microglia activation (McGeer & McGeer, Acta Neuropathol 126:479-97, 2013), and direct evidence of neuroinflammation in AD brain has been shown by in vivo PET imaging (Schuitemaker et al., Neurobiol Aging 34:12836, 2013). Notably, levels of pro-inflammatory cytokines are elevated in plasma and CSF in AD patients, for TNF by as much as 25-fold (Tarkowski et al., J Clin Immunol 19:223-30, 1999). In MCI subjects that progress to develop AD, a rise in CSF TNF levels correlates with disease progression (Tarkowski et al., J Neurol Neurosurg Psychiatry 74:1200-5, 2003). Paralleling this, elevated expression of TNF is reported in entorhinal cortex of 3xTg-AD mice prior to appearance of amyloid and tau pathology, and this increase associates with the onset of cognitive deficits in these mice and later neuronal loss (Janelsins et al. J Neuroinflamm 2:23, 2005). We hypothesize that failure of protein homeostasis leads to accumulation of proteins (e.g., AB, APOE and MAPT) that induce microglial activation and a proinflammatory M1-like response to instigate their removal. Continuing generation of protein (AB, APOE and MAPT) leads to maintenance of a chronic M1 response, lack of transition to an anti-inflammatory M2 response (particularly in aging brain that is already vulnerable to inflammation) with ensuing neuronal impairment observed in animal models and in preclinical AD that eventually leads to cell death. Proinflammatory cytokines, like TNF, induce vascular changes to allow lymphocyte infiltration that may underpin reported cerebral vasculature leakiness of AD patients and related Tg mouse models. Moreover, TNF induces AB production in cellular and animal AD models, further increasing its accumulation and accelerating the entire cascade. Our focus is to understand the time course of development of neuropathology accumulation of inflammatory cytokines and behavioral deficits in mouse models that may reflect the disease pathology in humans. Our aim is to use these models, together with classical evaluations of pharmcokinetics/dynamics and toxicity evaluations, to aid select out from our agents that potently lower TNF - compounds that can be moved to the clinic to mitigate the neuroinflammatory element in AD and associated disorders. Our studies involve: (i) Synthetic chemistry on the backbones of THAL and POM to generate more potent anti-inflammatory agents that are better tolerated (Luo et al. Bioorg Med Chem 26:1547-59, 2018). (ii) Cellular screening for anti-inflammatory actions (Tweedie et al. J Neuroinflamm 9:106, 2012) (iii) Zebrafish and chicken embryo screening for anti-inflammatory, anti-angiogenesis and toxicology screening (Collaborators: Drs. Vargesson, Figg) (Mahony et al. PNAS 110:12703-8, 2013; Beedie et al. Oncotarget 7:33237, 2016). (iv) Pharmacokinetic/dynamic/toxicological evaluations in acute rodent studies (Lecca et al., Elife 9:e54726, 2020). (v) Efficacy evaluations in both acute and chronic rodent models involving inflammation, cognitive impairment and/or AD and related disorders (Russo et al. J Neurochem. 122:11871-92, 2012; Tweedie et al. J Neuroinflamm 9:106, 2012; Belarbi et al. J Neuroinflamm 9:23, 2012; Starke et al. J Neuroinflammation 11:77, 2014; Baratz et al. J Neuroinflamm 12:45, 2015; Wang et al. J Neuroinflamm 13:168, 2016; Batsaikhan et al. Int J Mol Sci 20(3) pii: E502; Tsai et al., Cell Transplant 28:439-57, 2019). (vi) Evaluation of select agents across cellular models of SARS-CoV-2 to assess mitigation of the cytokine storm associated with the disorder, and blocking of viral cell entry (Collaborators: Mannan Baig, Shuibing Chen, Jasper Chan, Dong Seok Kim) In synopsis: Our focus is to use our novel compounds as agents to understand the time-dependent role of neuroinflammation in AD progression in animal models and, concurrently, to select out the most potent with drug-like features as a new treatment intervention for AD and related disorders, creating a preclinical package both on our best agent as well as on the comparator clinically approved cancer drug, POM (as a back up), to support clinical translation. Notably, the drug target in these proposed studies - elevated levels of TNF - has relevance to AD (with the potential to impact acute and chronic neurological endpoints) and to related CNS and systemic disorders driven by inflammation (for independent review see: Tobinick E; Curr Alzheimer Res. 9:99-109, 2012; Ignatowski T et al., CNS Drugs. 28:679-97, 2014; Clark & Vissel, J Neuroinflamm ;13:236, 2016). We additionally evaluate our TNF lowering agents in other neurological disorders in which an inflammatory state is evident, and for which effective treatments are lacking (such as TBI, stroke and Bilirubin-induced Neurological Dysfunction and COVID-19).