PROJECT SUMMARY Of the two hallmark proteinopathies, b-amyloid and tau, that define Alzheimer’s disease (AD), studies consistently show that women exhibit higher levels of tau than men. This finding is well-characterized in older women, even those who are considered clinically normal, but the biological mechanism driving this sex difference remains elusive. Sex hormonal changes, either due to menopause or hormone therapy, may be a contributing factor, although effects are equivocal. Sex hormones are not the only sex biological component that could play a role. Sex chromosomes form the genetic foundation by which women are biologically differentiated from men. The X chromosome, in particular, has largely been ignored in the AD field, due to complexities involving measurement and analysis. Women possess two X chromosomes, which has profound implications for sex-specific associations with neurodegenerative disease. To avoid ‘overdosing’ women with X-linked genes coming from two X chromosomes, one X is randomly silenced in each cell throughout the body. This random X inactivation, however, is incomplete, with some genes escaping this inactivation. In human women, approximately 30% of X-linked genes consistently escape inactivation across all tissue types. Of those inactive X escaped genes that have been reported in the literature (n=60), many are involved in the immune system. Due to this phenomenon, women tend to exhibit more robust immune systems and are more likely to suffer from autoimmune diseases. Of relevance to AD, neuroinflammation, exacerbated by robust immune responses, is argued to be a key early driver of pathology in vulnerable brain regions. These key pieces of evidence give rise to an innovative question that has never been tackled: could inactive X escaped genes be the key to explaining female vulnerability to AD pathology? And could this mechanism occur via an inflammatory pathway? As such, in this innovative proposal I put forward a conceptually novel approach to addressing sex differences in AD pathology by focusing on the ‘eXcluded’ chromosome. In collaboration with expert colleagues in the field of AD genetics, I will employ the latest techniques to examine the genomic, transcriptomic, and epigenetic impact of inactive-X escaped genes as a driver of sex differences in in vivo neuroimaging markers of b-amyloid and tau. This study will be unparalleled in its scope, as I will leverage my skills in neuroimaging, data harmonization and statistics to combine three large, deeply-phenotyped cohorts that have b-amyloid and tau positron emission tomography neuroimaging and samples for whole genome and transcriptome sequencing (n=900) and pro-inflammatory markers in blood: the Harvard Aging Brain Study (HABS), the Anti-Amyloid Treatment in Asymptomatic Alzheimer's (A4) study and the Longitudinal Evaluation of Amyloid Risk and Neurodegeneration (LEARN) study. I will also validate my findings in the Religious Orders Study, the Memory and Aging Project (ROS/MAP) and the Alzheimer’s disease Neuroimaging Initiative (ADNI). Elucidating sex-specific mechanisms for AD risk has far-reaching consequences for identifying drug targets, but also better predicting drug response outcomes in men and women in clinical trials.

PROJECT NARRATIVE Women are more likely to exhibit higher levels of Alzheimer’s disease pathology than men, even at the earliest stages of disease, however, no convincing biological mechanism has been proposed to explain this sex- specific vulnerability to disease. I propose a novel conceptualization of the problem through the examination of the X chromosome; specifically, X-linked genes that escape inactivation on the second X chromosome in women. These genes are often involved with immune responses pathways – a known mechanism for greater amyloid and tau pathological burden - and which could have major implications for female susceptibility to Alzheimer’s disease.