<p>Molecular profiles of mesenchymal stem cells (MSCs) are needed to standardize the composition and effectiveness of MSC therapeutics. This study employs RNA sequencing to identify genes to be used in concert with CD264 as a molecular profile of aging MSCs at a clinically relevant culture passage. CD264⁻ and CD264⁺ populations were isolated by fluorescence-activated cell sorting from passage 4 MSC cultures. CD264⁺ MSCs exhibited an aging phenotype relative to their CD264⁻ counterpart. Donor-matched CD264^−/+ mRNA samples from 5 donors were subjected to pair-ended, next-generation sequencing. An independent set of 5 donor MSCs was used to validate differential expression of select genes with quantitative reverse transcription PCR. Pairwise differential expression analysis identified 2,322 downregulated genes and 2,695 upregulated genes in CD264⁺ MSCs relative to donor-matched CD264⁻ MSCs with a Benjamini–Hochberg adjusted <i>p</i>-value (BH <i>p</i>_<i>adj</i>) &lt; 0.1. Nearly 25% of these genes were unique to CD264^−/+ MSCs and not differentially expressed at a significance level of BH <i>p</i>_<i>adj</i> &lt; 0.1 in previous RNA sequencing studies of early- vs. late-passage MSCs. Least Absolute Shrinkage and Selection Operator regression identified microtubule-associated protein 1A (<i>MAP1A</i>) and pituitary tumor-transforming gene 1 (<i>PTTG1</i>) as predictive genes of CD264⁺ MSCs. Combined <i>MAP1A</i> and <i>PTTG1</i> expression correctly classified CD264 status of MSC samples with an accuracy of 100%. Differential expression and predictive ability of <i>MAP1A</i> and <i>PTTG1</i> compared favorably with that of existing senescence markers expressed in early passage CD264^−/+ MSCs. This study provides the first linkage of <i>MAP1A</i> to CD264, aging and senescence. Our findings have application as quality metrics to standardize the composition of MSC therapies and as molecular targets to slow/reverse cellular aging.</p>