Compared to other subtypes of breast cancer, triple-negative breast cancers (TNBC) have fewer treatment options and exhibit a worse prognosis. Through integrated transcriptomic, metabolomic, immunohistochemical, spatial, and clinical analyses, we identify the mitochondrial enzyme, α-aminoadipate aminotransferase (AADAT) as a previously unrecognized metabolic immune checkpoint in TNBC. AADAT mRNA and protein were significantly upregulated in human TNBC, and high AADAT expression was associated with reduced intra-tumoral CD8+ T-cell density and inferior survival. Genetic silencing of AADAT in orthotopic murine TNBC models curtailed primary tumor growth and distant metastasis in a CD8+ T-cell-dependent manner, enhanced effector T-cell activation, and sensitized tumors to dual PD-1/CTLA-4 blockade. Mechanistically, unbiased metabolomics showed increased malate levels after AADAT knockdown. Additionally, 4-hydroxyphenylpyruvate, an essential precursor for coenzyme Q10(CoQ10) biosynthesis, decreased following AADAT knockdown, suggesting an impaired mitochondrial electron transport chain. CoQ10 supplementation restored metabolic balance and reversed malate accumulation caused by AADAT knockdown, indicating that AADAT helps maintain CoQ10-supported redox homeostasis, thereby preventing malate buildup and export. Notably, malate addition directly boosted CD8+ T-cell oxidative metabolism, increased the NAD+/NADH ratio and reactive oxygen species, and augmented TNF-α and IFN-γ production. In vivo, malate supplementation in drinking water phenocopied AADAT knockdown, restored the response to paclitaxel plus anti-PD-1 therapy in multiple independent syngeneic TNBC models with de novo or acquired resistance to immunotherapy, reduced tumor burden, and prolonged survival. In patient cohorts, higher spatially clustered intra-tumoral malate is associated with co-localization of functional CD8+ T cells, decreased exhausted T-cell neighborhoods, and superior post-chemotherapy outcomes. These data position AADAT as a central metabolic orchestrator of immune escape in TNBC and nominate oral malate as a readily translatable adjuvant to reverse chemo-immunotherapy resistance in TNBC.

The lack of interoperability among clinical and research data systems poses a significant barrier to cancer researchers interested in evaluating novel mechanistic hypotheses or translating innovative treatment strategies from the laboratory to the clinic. To address this gap in knowledge, we developed an innovative, web-based, data discovery, visualization and analysis tool (nSight™) that allows researchers to quickly and easily query clinical/research data and construct de-identified cancer cohorts. Guiding principles for development of the tool were focused on ease of use, intuitiveness, self-service, and presentation of structured but de-identified data to the end user. nSight™ provides users with information on patient demographics, disease histology, diagnostic procedures and therapeutic interventions, timeline of disease progression/recurrence, along with available molecular profiling/sequencing data and indicators of participation in epidemiologic or lifestyle studies for specific cancer patient cohorts. The platform also allows users to obtain summary statistics based on demographic, histologic and clinical factors as well as perform basic survival analysis using Kaplan-Meier curves between specific patient cohorts. nSight™ is an intuitive, user-friendly tool that enables visualization, integration and analysis of multimodal clinical and research data without placing high technical demands or time constraints on researchers. The platform is designed for research feasibility assessment, cohort development, and retrospective data discovery, which in turn should help investigators identify potential study populations and explore novel hypotheses.

Racial disparities in the clinical outcomes of triple-negative breast cancer (TNBC) have been well-documented, but the underlying biological mechanisms remain poorly understood. To investigate these disparities, we employed a multi-omic approach integrating imaging mass cytometry and spatial transcriptomics to characterize the tumor microenvironment (TME) in self-identified Black American (BA) and White American (WA) TNBC patients. Our analysis revealed that the TME in BA patients is marked by a network of endothelial cells, macrophages, and mesenchymal-like cells, which correlates with reduced patient survival. In contrast, the WA TNBC microenvironment is enriched in T-cells and neutrophils, indicative of T-cell exhaustion and suppressed immune responses. Ligand-receptor and pathway analyses further demonstrated that BA TNBC tumors exhibit a relatively "immune-cold" profile, while WA TNBC tumors display features of an "inflamed" TME, suggesting the evolution of a unique immunosuppressive mechanism. These findings provide insight into racially distinct tumor-promoting and immunosuppressive microenvironments, which may contribute to the observed differences in clinical outcomes among BA and WA TNBC patients.

Myeloid and lymphoid malignancies consist of a heterogenous population of neoplasms, all of which contain distinct genomic alterations used for prognostic, diagnostic, and therapeutic intervention strategies. To clinically evaluate all hematological malignancies using a single test in our laboratory, we have developed a Next-Generation Sequencing assay (NGS) (the PanHeme Assay) that identifies clinically relevant SNV, INDEL (including FLT3-ITDs), CNV, and Fusion mutations known to be important across all myeloid and lymphoid cancers. The PanHeme Assay consists of 358 full coding DNA genes, 125 RNA-based fusions genes, as well as a genome-wide SNP backbone for calling chromosomal copy number changes. This comprehensive assay allows for a single automated laboratory and bioinformatics solution reducing sample-to-report turnaround time to <3 days. Since the integration of this clinical assay, >4,000 unique patients with >6,000 unique tests have been performed leading to >46,000 NGS-based mutations identified. This large reservoir of data includes SNV, INDEL, FLT3-ITD, full chromosome, arm, and gene level gains and losses, as well as clinically relevant fusions, including IGH enhancer driven modifications across all hematological malignancies. This large cohort of data has allowed us to focus on several intriguing questions related to hematological testing, progression, stratification of disease and monitoring. To this end we have focused on interpreting the role mutations associated with Clonal Hematopoiesis of Indeterminate Potential (CHIP) have as a reservoir of potential for cancer progression and relapse. Furthermore, we have investigated the concordance of mutation calling between blood and bone marrow biopsies taken within close proximity within the same patient to better understand the need for invasive BM biopsy procedures and how this can further relate to serial monitoring of disease at future timepoints. Beyond these evaluations we have also focused on the ability of this comprehensive assay, which contains classical mutational profiling as well as molecular cytogenomics capabilities, to show that using CNV and mutation calls derived from the PanHeme assay coupled to hematologic data accurately recapitulate IPSS-M risk stratification and predicts survival in MDS without the need of classical cytogenetics. The final investigation of this clinical data set was to assess the most prevalent mutations identified in AML outside of FLT3-ITDs and NPM1 mutations to help us to develop a comprehensive MRD panel which would cover a larger set of AML patients than current testing modalities. By investigating common mutations within this cohort we can create a small “hotspot” panel to concisely target recurrent mutations in AML patients for serial monitoring, which will be further developed into a clinical MRD assay within our lab. The PanHeme assay has become absolutely necessary for prognostication, diagnosis, and therapy selection within our clinics due to its robust content and extremely fast turn-around time from sample-to-clinical reporting. Beyond this, the assay has also created a large data cohort which can be utilized to address key concepts in the myeloid and lymphoid diseases. This data set has become paramount and has opened up interesting channels around diagnosis, risk stratification, monitoring, and evolution of disease.

The canonical mechanism behind tamoxifen's therapeutic effect on estrogen receptor α/ESR1+ breast cancers is inhibition of ESR1-dependent estrogen signaling. Although ESR1+ tumors expressing wild-type p53 were reported to be more responsive to tamoxifen (Tam) therapy, p53 has not been factored into choice of this therapy and the mechanism underlying the role of p53 in Tam response remains unclear. In a window-of-opportunity trial on patients with newly diagnosed stage I-III ESR1+/HER2/wild-type p53 breast cancer who were randomized to arms with or without Tam prior to surgery, we reveal that the ESR1-p53 interaction in tumors was inhibited by Tam. This resulted in functional reactivation of p53 leading to transcriptional reprogramming that favors tumor-suppressive signaling, as well as downregulation of oncogenic pathways. These findings illustrating the convergence of ESR1 and p53 signaling during Tam therapy enrich mechanistic understanding of the impact of p53 on the response to Tam therapy.

PDF file - 41KB, The combination of Y15 and temozolomide decreases more significantly viability of U251 cells than each agent alone.

PDF file - 175K, Table 1. Concentration of Curaxin-137 (muM) in mouse plasma and tissues (mean +/-SD in muM, n=4) 10 weeks after start of treatment (14 weeks of age) Table 2. Histopathological findings in old animals in each of treatment group (two mice per group)

<p>Melanoma Immune Profile (MIP) correlates with metastasis cohort including patients with local recurrence only.</p>

PDF file - 11K, List of antibodies used in the study

Supplementary Table 4 from Tumor Suppressor Activity of CCAAT/Enhancer Binding Protein α Is Epigenetically Down-regulated in Head and Neck Squamous Cell Carcinoma

Supplementary Table 3 from Tumor Suppressor Activity of CCAAT/Enhancer Binding Protein α Is Epigenetically Down-regulated in Head and Neck Squamous Cell Carcinoma

Supplementary Table 1 from Tumor Suppressor Activity of CCAAT/Enhancer Binding Protein α Is Epigenetically Down-regulated in Head and Neck Squamous Cell Carcinoma

PDF file - 790K, Absence of morphological changes in organs of MMTV-neu mice treated with Curaxin-137 from 4 to 14 weeks of age.

<p>Comparison of patient demographics in validation cohort to original publication discovery and test cohorts.</p>

Supplementary Table 2 from Tumor Suppressor Activity of CCAAT/Enhancer Binding Protein α Is Epigenetically Down-regulated in Head and Neck Squamous Cell Carcinoma

Supplementary Materials and Methods with detailed experimental procedures. Supplemental Table 1 shows the list of antibodies. Supplemental Table 2 shows the list of shRNAs and their sequences. Supplemental Table 3 shows the list of siRNAs and their sources.

Supplementary Figure Legends 1-2 from Tumor Suppressor Activity of CCAAT/Enhancer Binding Protein α Is Epigenetically Down-regulated in Head and Neck Squamous Cell Carcinoma