Epidemiological studies have found that heavy alcohol use is associated with increased risk for Alzheimer’s disease (AD), with frequent drinking earlier in adulthood increasing risk. The increases in neuroinflammation featured in both heavy alcohol use and AD may be partially responsible for this link. However, it is unknown if abstinence mitigates this risk. We hypothesized that binge ethanol during mid adult life would persistently increase AD pathology even after prolonged abstinence. Male and female 3xTg-AD mice (APPSwe, tauP301, Psen1tm1Mpm) which feature progressive amyloid (Aβ) and tau pathology, received chronic binge ethanol (5g/kg/day, 5-days-on/2-days-off, i.g.) or water during adulthood (from 5.5 to 9 months of age), followed by abstinence and assessment at 14 months of age. The effects of ethanol on protective AD genes (e.g., APOE and TREM2) as well as proinflammatory genes were measured by PCR. Levels of pathologic tau and Aβ were measured by immunohistochemistry and western blot. Ethanol caused persistent reductions in protective AD genes: APOE (25% reduction, *p < 0.05), TREM2 (28%, *p < 0.05), LPL (40%, **p < 0.01), and CTSD (24%, *p < 0.05) and promoted a proinflammatory gene signature in female, but not male cortex. Concurrently, ethanol increased total and hyperphosphorylated tau (AT8) in piriform cortex and hippocampus of females, but not males. Levels of AT8 were negatively correlated with APOE (R = –0.67, *p < 0.05) and TREM2 (R = –0.78, **p < 0.005) suggesting protective roles in pathogenesis. No differences were found in levels of main regulators of tau phosphorylation state (GSK3β, PKA, PP2A), suggesting ethanol disrupted clearance of tau. Therefore, we measured the effect of ethanol on lysosomes, which degrade tau, and lysosomal localization of tau using co-immunofluorescence. In females, ethanol caused a persistent reduction in mature LAMP1 lysosomes in CA1 of hippocampus (35%, *p < 0.05), along with a 60% increase in total tau (*p < 0.05). Thus, chronic binge ethanol during mid adult life causes a persistent enhancement of tau pathology in cortical and hippocampal brain regions of females. Persistent AD pathology was associated with an increased proinflammatory signature and a reduction of mature lysosomes. This implicates binge ethanol exposure with increased risk of AD pathologic progression in females.

We show the therapeutic potential of myeloid-directed immunomodulatory treatment for medulloblastoma. Patients with medulloblastoma, the most common malignant pediatric brain tumor, need new treatments, as standard therapy produces disabling neurotoxicities and fails 20% of patients. About 30% of medulloblastomas show hyperactivation of the SHH (Sonic Hedgehog) signaling pathway, and these tumors have large myeloid populations in the tumor microenvironment (TME). We analyzed whether activating toll-like receptors on these myeloid cells would slow tumor growth. Our prior single-cell RNA sequencing studies in both patient samples and mouse models showed that 10% of the cells in SHH medulloblastoma are myeloid cells and that these cells uniquely express TLR7 and TLR8. We treated mice genetically engineered to develop SHH medulloblastomas with the TLR7/8 agonist resiquimod, administered systemically either as free drug or in polyoxazoline nanoparticles (POx-resiquimod), and compared POx-resiquimod+radiotherapy to radiotherapy alone. We found that POx-resiquimod extended the survival time of mice with medulloblastoma, while free drug failed to show benefit. PK studies showed that POx-resiquimod increased tumor drug exposure, consistent with increased efficacy. Mechanistically, POx-resiquimod increases tumor myeloid populations and decreased the fraction of myeloid cells that expressed IGF1. POx-resiquimod plus radiation therapy, moreover, was superior to either POx-resiquimod alone, or radiotherapy alone. Together our data show that the TLR7/8 agonist resiquimod, delivered in nanoparticle formulation, produces a significant anti-tumor effect in SHH medulloblastoma, and increased the efficacy of radiotherapy. As radiotherapy is the mainstay of current medulloblastoma treatment, we propose that TLR7/8-agonist therapy such as resiquimod may be added to current regimens to reduce the radiation dose needed for efficacy, and to increase the fraction of successfully treated patients.

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