Importance: Obesity, especially visceral obesity, is an established risk factor associated with all-cause mortality. However, the inadequacy of conventional anthropometric measures in assessing fat distribution necessitates a more comprehensive indicator, body roundness index (BRI), to decipher its population-based characteristics and potential association with mortality risk.Objective: To evaluate the temporal trends of BRI among US noninstitutionalized civilian residents and explore its association with all-cause mortality.Design, Setting, and Participants: For this cohort study, information on a nationally representative cohort of 32 995 US adults (age ≥20 years) was extracted from the National Health and Nutrition Examination Survey (NHANES) from 1999 to 2018 and NHANES Linked Mortality File, with mortality ascertained through December 31, 2019. Data were analyzed between April 1 and September 30, 2023.Exposures: Biennial weighted percentage changes in BRI were calculated. Restricted cubic spline curve was used to determine optimal cutoff points for BRI.Main Outcome and Measures: The survival outcome was all-cause mortality. Mortality data were obtained from the Centers for Disease Control and Prevention website and linked to the NHANES database using the unique subject identifier. Weibull regression model was adopted to quantify the association between BRI and all-cause mortality.Results: Among 32 995 US adults, the mean (SD) age was 46.74 (16.92) years, and 16 529 (50.10%) were women. Mean BRI increased gradually from 4.80 (95% CI, 4.62-4.97) to 5.62 (95% CI, 5.37-5.86) from 1999 through 2018, with a biennial change of 0.95% (95% CI, 0.80%-1.09%; P < .001), and this increasing trend was more obvious among women, elderly individuals, and individuals who identified as Mexican American. After a median (IQR) follow-up of 9.98 (5.33-14.33) years, 3452 deaths (10.46% of participants) from all causes occurred. There was a U-shaped association between BRI and all-cause mortality, with the risk increased by 25% (hazard ratio, 1.25; 95% CI, 1.05-1.47) for adults with BRI less than 3.4 and by 49% (hazard ratio, 1.49; 95% CI, 1.31-1.70) for those with BRI of 6.9 or greater compared with the middle quintile of BRI of 4.5 to 5.5 after full adjustment.Conclusions and Relevance: This national cohort study found an increasing trend of BRI during nearly 20-year period among US adults, and importantly, a U-shaped association between BRI and all-cause mortality. These findings provide evidence for proposing BRI as a noninvasive screening tool for mortality risk estimation, an innovative concept that could be incorporated into public health practice pending consistent validation in other independent cohorts.

Background: Aspartate β-hydroxylase (ASPH) is silent in normal adult tissues only to re-emerge during oncogenesis where its function is required for generation and maintenance of malignant phenotypes. Exosomes enable prooncogenic secretome delivering and trafficking for long-distance cell-to-cell communication. This study aims to explore molecular mechanisms underlying how ASPH network regulates designated exosomes to program development and progression of breast cancer. Methods: Stable cell lines overexpressing or knocking-out of ASPH were established using lentivirus transfection or CRISPR-CAS9 systems. Western blot, MTT, immunofluorescence, luciferase reporter, co-immunoprecipitation, 2D/3-D invasion, tube formation, mammosphere formation, immunohistochemistry and newly developed in vitro metastasis were applied. Results: Through physical interactions with Notch receptors, ligands (JAGs) and regulators (ADAM10/17), ASPH activates Notch cascade to provide raw materials (especially MMPs/ADAMs) for synthesis/release of pro-metastatic exosomes. Exosomes orchestrate EMT, 2-D/3-D invasion, stemness, angiogenesis, and premetastatic niche formation. Small molecule inhibitors (SMIs) of ASPH’s β-hydroxylase specifically/efficiently abrogated in vitro metastasis, which mimics basement membrane invasion at primary site, intravasation/extravasation (transendothelial migration), and colonization/outgrowth at distant sites. Multiple organ-metastases in orthotopic and tail vein injection murine models were substantially blocked by a specific SMI. ASPH is silenced in normal adult breast, upregulated from in situ malignancies to highly expressed in invasive/advanced ductal carcinoma. Moderate-high expression of ASPH confers more aggressive molecular subtypes (TNBC or Her2 amplified), early recurrence/progression and devastating outcome (reduced overall/disease-free survival) of breast cancer. Expression profiling of Notch signaling components positively correlates with ASPH expression in breast cancer patients, confirming that ASPH-Notch axis acts functionally in breast tumorigenesis. Conclusions: ASPH-Notch axis guides particularly selective exosomes to potentiate multifaceted metastasis. ASPH’s pro-oncogenic/pro-metastatic properties are essential for breast cancer development/progression, revealing a potential target for therapy.

Background: Interactions between tumor and microenvironment determine individual response to immunotherapy. Triple negative breast cancer (TNBC) and hepatocellular carcinoma (HCC) have exhibited suboptimal responses to immune checkpoint inhibitors (ICIs). Aspartate β-hydroxylase (ASPH), an oncofetal protein and tumor associated antigen (TAA), is a potential target for immunotherapy. Methods: Subcutaneous HCC and orthotopic TNBC murine models were established in immunocompetent BALB/c mice with injection of BNL-T3 and 4 T1 cells, respectively. Immunohistochemistry, immunofluorescence, H&E, flow cytometry, ELISA and in vitro cytotoxicity assays were performed. Results: The ASPH-MYC signaling cascade upregulates PD-L1 expression on breast and liver tumor cells. A bio-nanoparticle based λ phage vaccine targeting ASPH was administrated to mice harboring syngeneic HCC or TNBC tumors, either alone or in combination with PD-1 blockade. In control, autocrine chemokine ligand 13 (CXCL13)-C-X-C chemokine receptor type 5 (CXCR5) axis promoted tumor development and progression in HCC and TNBC. Interactions between PD-L1⁺ cancer cells and PD-1⁺ T cells resulted in T cell exhaustion and apoptosis, causing immune evasion of cancer cells. In contrast, combination therapy (Vaccine+PD-1 inhibitor) significantly suppressed primary hepatic or mammary tumor growth (with distant pulmonary metastases in TNBC). Adaptive immune responses were attributed to expansion of activated CD4⁺ T helper type 1 (Th1)/CD8⁺ cytotoxic T cells (CTLs) that displayed enhanced effector functions, and maturation of plasma cells that secreted high titers of ASPH-specific antibody. Combination therapy significantly reduced tumor infiltration of immunosuppressive CD4⁺/CD25⁺/FOXP3⁺ Tregs. When the PD-1/PD-L1 signal was inhibited, CXCL13 produced by ASPH⁺ cancer cells recruited CXCR5⁺/CD8⁺ T lymphocytes to tertiary lymphoid structures (TLSs), comprising effector and memory CTLs, T follicular helper cells, B cell germinal center, and follicular dendritic cells. TLSs facilitate activation and maturation of DCs and actively recruit immune subsets to tumor microenvironment. These CTLs secreted CXCL13 to recruit more CXCR5⁺ immune cells and to lyse CXCR5⁺ cancer cells. Upon combination treatment, formation of TLSs predicts sensitivity to ICI blockade. Combination therapy substantially prolonged overall survival of mice with HCC or TNBC. Conclusions: Synergistic antitumor efficacy attributable to a λ phage vaccine specifically targeting ASPH, an ideal TAA, combined with ICIs, inhibits tumor growth and progression of TNBC and HCC.

Background: Signaling pathways critical for embryonic development re-emerge in adult pancreas during tumorigenesis. Aspartate β-hydroxylase (ASPH) drives embryonic cell motility/invasion in pancreatic development/differentiation. We explored if dysregulated ASPH is critically involved in pancreatic cancer pathogenesis.Methods: To demonstrate if/how ASPH mediates malignant phenotypes, proliferation, migration, 2-D/3-D invasion, pancreatosphere formation, immunofluorescence, Western blot, co-immunoprecipitation, invadopodia formation/maturation/function, qRT-PCR, immunohistochemistry (IHC), and self-developed in vitro metastasis assays were performed. Patient-derived xenograft (PDX) models of human pancreatic ductal adenocarcinoma (PDAC) were established to illustrate in vivo antitumor effects of the third-generation small molecule inhibitor specifically against ASPH’s β-hydroxylase activity. Prognostic values of ASPH network components were evaluated with Kaplan-Meier plots, log-rank tests, and Cox proportional hazards regression models.Results: ASPH renders pancreatic cancer cells more aggressive phenotypes characterized by epithelial–mesenchymal transition (EMT), 2-D/3-D invasion, invadopodia formation/function as demonstrated by extracellular matrix (ECM) degradation, stemness (cancer stem cell marker upregulation and pancreatosphere formation), transendothelial migration (mimicking intravasation/extravasation), and sphere formation (mimicking metastatic colonization/outgrowth at distant sites). Mechanistically, ASPH activates SRC cascade through direct physical interaction with ADAM12/ADAM15 independent of FAK. The ASPH-SRC axis enables invadopodia construction and initiates MMP-mediated ECM degradation/remodeling as executors for invasiveness. Pharmacologic inhibition of invadopodia attenuates in vitro metastasis. ASPH fosters primary tumor development and pulmonary metastasis in PDX models of PDAC, which is blocked by a leading compound specifically against ASPH enzymatic activity. ASPH is silenced in normal pancreas, progressively upregulated from pre-malignant lesions to invasive/advanced stages of PDAC. Expression profiling of ASPH-SRC network components independently/jointly predicts clinical outcome of PDAC patients. Compared to a negative-low level, a moderate-very high level of ASPH, ADAM12, activated SRC, and MMPs correlated with curtailed overall survival (OS) of pancreatic cancer patients (log-rank test,ps < 0.001). The more unfavorable molecules patients carry, the more deleterious prognosis is destinated. Patients with 0–2 (n = 4), 3–5 (n = 8), 6–8 (n = 24), and 9–12 (n = 73) unfavorable expression scores of the 5 molecules had median survival time of 55.4, 15.9, 9.7, and 5.0 months, respectively (p < 0.001).Conclusion: Targeting the ASPH-SRC axis, which is essential for propagating multi-step PDAC metastasis, may specifically/substantially retard development/progression and thus improve prognosis of PDAC.

MicroRNA-21 inhibits mitochondria-mediated apoptosis in keloid MicroRNA-21 inhibits mitochondria-mediated apoptosis in keloid