Tumor protein p53 (TP53) is the most frequently mutated gene in cancer 1,2. In patients with myelodysplastic syndromes (MDS), TP53 mutations are associated with high-risk disease 3,4 , rapid transformation to acute myeloid leukemia (AML) 5 , resistance to conventional therapies 6-8 and dismal outcomes 9. Consistent with the tumor-suppressive role of TP53, patients harbor both mono-and biallelic mutations 10. However, the biological and clinical implications of TP53 allelic state have not been fully investigated in MDS or any other cancer type. We analyzed 3,324 patients with MDS for TP53 mutations and allelic imbalances and delineated two subsets of patients with distinct phenotypes and outcomes. One-third of TP53-mutated patients had monoallelic mutations whereas two-thirds had multiple hits (multi-hit) consistent with biallelic targeting. Established associations with complex karyotype, few co-occurring mutations, high-risk presentation and poor outcomes were specific to multi-hit patients only. TP53 multi-hit state predicted risk of death and leukemic transformation independently of the Revised International Prognostic Scoring System (IPSS-R) 11. Surprisingly, monoallelic patients did not differ from TP53 wild-type patients in outcomes and response to therapy. This study shows that consideration of TP53 allelic state is critical for diagnostic and prognostic precision in MDS as well as in future correlative studies of treatment response. In collaboration with the International Working Group for Prognosis in MDS (Supplementary Table 1), we assembled a cohort of 3,324 peridiagnostic and treatment-naive patients with MDS or closely related myeloid neoplasms (Extended Data Fig. 1 and Supplementary Fig. 1). Genetic profiling included conventional G-banding analyses (CBA) and tumor-only, capture-based, next-generation sequencing (NGS) of a panel of genes recurrently mutated in MDS, as well as genome-wide copy number probes. Allele-specific copy number profiles were generated from NGS data using the CNACS algorithm 7 (see Methods and Code availability). An additional 1,120 samples derived from the Japanese MDS consortium (Extended Data Fig. 2) were used as a validation cohort. To study the effect of TP53 allelic state on genome stability, clinical presentation, outcome and response to therapy, we performed a detailed characterization of alterations at the TP53 locus. First, we assessed genome-wide allelic imbalances in the cohort of 3,324 patients, to include arm-level or focal (~3 Mb) ploidy alterations and regions of copy-neutral loss of heterozygosity (cnLOH) (Extended Data Fig. 3, Supplementary Figs. 2-4 and Methods).

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To perform a range of ventilatory and nonventilatory behaviors, the diaphragm muscle (DIAm) must be able to generate sufficient forces throughout the lifespan. We hypothesized that sarcopenia impacts DIAm force generation and thus limits performance of expulsive, higher force, nonventilatory behaviors. Male and female mice (n = 79) at 6 and 24 mo of age (100 vs. 70-75% survival, respectively) were used to examine transdiaphragmatic pressure (Pdi) generation across motor behaviors in vivo and in vitro DIAm specific force. We found a significant effect of age on maximum Pdi (20-41% decline during tracheal occlusion and bilateral phrenic nerve stimulation), maximum DIAm specific force (30% decline), and DIAm fatigue resistance (15% increase). There were no differences between sexes in these age effects on DIAm performance. These results support our hypothesis that sarcopenia primarily impacts higher force, nonventilatory motor behaviors of the DIAm. Such functional limitations may have negative implications in the ability of the DIAm to generate forces needed for airway clearance in old age and thereby contribute to age-related respiratory complications.

Introduction Sarcopenia likely comprises muscle fiber denervation and re-innervation, resulting in clustering of muscle fibers of the same type (classified by myosin heavy chain isoform composition). Development of methodology to quantitatively evaluate clustering of muscle fibers according to fiber type is necessary. Methods Fiber type specific immunofluorescence histology was used to quantify fiber clustering in murine diaphragm muscle (n=15) at 6 and 24 months of age. Results With age, fiber type clustering is evidenced by fiber type specific changes in distances between fibers, specifically a 14% decrease to the closest fiber for type I and 24% increase for type IIx and/or IIb fibers (P<0.001). Additionally, a 34% increase to the 3 closest type IIx and/or IIb fibers was found (P<0.001). Discussion This novel method of analyzing fiber type clustering may be useful in examining pathophysiological conditions of motor unit loss in neuromuscular disorders, myopathies, dystrophies, injuries, or amyotrophic lateral sclerosis.

Recent genomic research efforts in multiple myeloma have revealed clinically relevant molecular subgroups beyond conventional cytogenetic classifications. Implementing these advances in clinical trial design and in routine patient care requires a new generation of molecular diagnostic tools. Here, we present a custom capture next-generation sequencing (NGS) panel designed to identify rearrangements involving the IGH locus, arm level, and focal copy number aberrations, as well as frequently mutated genes in multiple myeloma in a single assay. We sequenced 154 patients with plasma cell disorders and performed a head-to-head comparison with the results from conventional clinical assays, i.e., fluorescent in situ hybridization (FISH) and single-nucleotide polymorphism (SNP) microarray. Our custom capture NGS panel had high sensitivity (>99%) and specificity (>99%) for detection of IGH translocations and relevant chromosomal gains and losses in multiple myeloma. In addition, the assay was able to capture novel genomic markers associated with poor outcome such as bi-allelic events involving TP53. In summary, we show that a multiple myeloma designed custom capture NGS panel can detect IGH translocations and CNAs with very high concordance in relation to FISH and SNP microarrays and importantly captures the most relevant and recurrent somatic mutations in multiple myeloma rendering this approach highly suitable for clinical application in the modern era.