Hanmei Xu

Since its elucidation, the tricarboxylic acid (TCA) cycle has been regarded as a fully defined enzymatic pathway. Here, we report the discovery of a previously unrecognized micropeptide component of this central metabolic system. Through analyzing ribosome profiling data from the mouse sORFs.org database, we discovered APPM (ATP production–promoting micropeptide), a 29–amino acid peptide encoded by a non-canonical small open reading frame within the mouse Ctsb gene. APPM localizes to the mitochondrial matrix and inner membrane, where it directly binds to the E1 subunit (OGDH) of the α-ketoglutarate dehydrogenase complex (OGDHc). This interaction modulates OGDHc enzymatic activity, thereby regulating NADH and ATP production. We identified tyrosine-13 as the critical residue mediating the APPM-OGDH interaction and functional activation. Exogenous APPM administration increased ATP levels in vivo and, in a mouse model of heart failure, restored cardiac function, outperforming the reference drug trimetazidine. Collectively, our findings identify APPM as the first micropeptide regulator of the OGDH complex, expanding the known repertoire of the TCA cycle and presenting a potential therapeutic strategy for bioenergetic disorders.

Triple-negative breast cancer (TNBC) is the most malignant breast cancer subtype. The 5-year overall survival rate of TNBC patients is 77%, but once cancer metastasis occurs, this rate drops to 12%. To date, the discovery of biomarkers associated with TNBC metastasis remains a major challenge. In this study, we identified a novel translated product, XLH-36 micropeptide, encoded by lncRNA (long non-coding RNA) C5orf66-AS1, which acts as an “oncogenic driver” in TNBC. Through conservation analyses across 101 species, we found that XLH-36 is highly conserved in humans and primates. Analysis of RNA-seq data from 1295 breast cancer patients, including 165 TNBC patients from different cohorts, revealed significantly elevated XLH-36 expression levels in breast cancer and TNBC. Specifically, TNBC patients with low C5orf66-AS1 (encoding XLH-36) expression demonstrated a 20% higher overall survival rate compared to those with high C5orf66-AS1 expression over a 50-month follow-up. XLH-36 knockout inhibited the growth and metastasis of TNBC xenografts in mice. Furthermore, we found that XLH-36 directly binds Gemin4, causing it to remain in the cytoplasm and preventing it from carrying out its role in promoting S100A4 mRNA splicing in the nucleus. This leads to a compensatory increase in ICAM1 levels, ultimately resulting in the promotion of epithelial-to-mesenchymal transition (EMT) in TNBC cells and an increase in tumor metastasis. In summary, our findings highlight the crucial role of XLH-36 in TNBC metastasis, which could be exploited in the development of therapeutic and diagnostic strategies for TNBC patients.

Background: Micropeptides, encoded by non-coding RNAs, play a pivotal role in various cellular functions. While several micropeptides have already been linked to HCC, their roles remain incompletely understood. Our study identifies MP60, a conserved micropeptide strongly associated with HCC progression. Methods and Results: By analyzing The Cancer Genome Atlas (TCGA) dataset, we assessed the coding potential of long non-coding RNAs (lncRNAs) with significant expression changes in HCC. Our findings reveal that ENST0000614292, a transcript of LINC01138, exhibited the highest coding potential, encoding a putative 60-amino-acid micropeptide, which we have named MP60 and confirmed the expression of MP60 in HCC tissues, with a nuclear localization. MP60 directly interacts with RNA-binding motif protein 10 (RBM10) and downregulates its expression. Additionally, MP60 modulates EMT. Functional analyses demonstrated that MP60 promotes cellular proliferation and migration, while reducing cellular adhesion, translated by enhanced tumorigenesis in vivo. Notably, MP60 expression is markedly increased in HCC tissues and is associated with a poorer prognosis. Conclusions: These findings identify MP60 as a potential biomarker and therapeutic target in HCC, linking its oncogenic effects to EMT modulation and tumor progression.

Background Chimeric antigen receptor (CAR) T cells have demonstrated remarkable breakthroughs in treating hematologic malignancies, yet their efficacy in solid tumors is limited by the immunosuppressive microenvironment. Sympathetic nerves significantly contribute to this immunosuppressive milieu in solid tumors. However, the impact of tumor sympathetic denervation on enhancing CAR T-cell antitumor efficacy remains unclear. Methods We screened for sympathetic gene sets in various types of cancers and investigated the association of sympathetic nerves with immunosuppression in renal clear cell carcinoma. Using antibodies to block the nerve growth factor (NGF) pathway, we explored sympathetic nerve distribution in tumor tissues and tumor progression. Additionally, we engineered CAR T cells to secrete NGF single chain fragment variable (scFv) to achieve tumor immunosympathectomy and assessed their antitumor efficacy. Bulk RNA sequencing and single-cell RNA sequencing analyses were conducted to evaluate changes in immune cell phenotypes within the tumor microenvironment. Results Blocking the NGF pathway with antibodies effectively reduced sympathetic nerve distribution in tumor tissues and delayed tumor progression. CAR T cells engineered to secrete NGF scFv achieved a similar tumor immunosympathectomy and exhibited enhanced tumor suppression. RNA sequencing analyses revealed that this augmented effect was primarily due to the inhibition of the terminal exhaustion phenotype in tumor-infiltrating CD8 T cells and the prevention of macrophage polarization from M1 to M2. This approach maintained a stronger antitumor immune state at the tumor site. Additionally, splenic T cells also exhibited a more potent immune effector phenotype following the infusion of NGF scFv-secreting CAR T cells. Conclusions Our results suggest that immunosympathectomy is a novel approach to weaken tumor microenvironment immunosuppression and synergistically enhance CAR T-cell efficacy against solid tumors.

High efficacy, selectivity and cellular targeting of therapeutic agents has been an active area of investigation for decades. Currently, most clinically approved therapeutics are small molecules or protein/antibody biologics. Targeted action of small molecule drugs remains a challenge in medicine. In addition, many diseases are considered ‘undruggable’ using standard biomacromolecules. Many of these challenges however, can be addressed using nucleic therapeutics. Nucleic acid drugs (NADs) are a new generation of gene-editing modalities characterized by their high efficiency and rapid development, which have become an active research topic in new drug development field. However, many factors, including their low stability, short half-life, high immunogenicity, tissue targeting, cellular uptake, and endosomal escape, hamper the delivery and clinical application of NADs. Scientists have used chemical modification techniques to improve the physicochemical properties of NADs. In contrast, modified NADs typically require carriers to enter target cells and reach specific intracellular locations. Multiple delivery approaches have been developed to effectively improve intracellular delivery and the in vivo bioavailability of NADs. Several NADs have entered the clinical trial recently, and some have been approved for therapeutic use in different fields. This review summarizes NADs development and evolution and introduces NADs classifications and general delivery strategies, highlighting their success in clinical applications. Additionally, this review discusses the limitations and potential future applications of NADs as gene therapy candidates.

Background: Liver fibrosis is a pathological response to liver damage induced by multiple etiologies including NASH and CCl₄, which may further lead to cirrhosis and hepatocellular carcinoma (HCC). Despite the increasing understanding of liver fibrosis and HCC, clinical prognosis and targeted therapy remain challenging.Methods: This study integrated single-cell sequencing analysis, bulk sequencing analysis, and mouse models to identify highly expressed genes, cell subsets, and signaling pathways associated with liver fibrosis and HCC. Clinical prediction models and prognostic genes were established and verified through machine learning, survival analysis, as well as the utilization of clinical data and tissue samples from HCC patients. The expression heterogeneity of the core prognostic gene, along with its correlation with the tumor microenvironment and prognostic outcomes, was analyzed through single-cell analysis and immune infiltration analysis. In addition, the cAMP database and molecular docking techniques were employed to screen potential small molecule drugs for the treatment of liver fibrosis and HCC.Result: We identified 40 pathogenic genes, 15 critical cell subsets (especially Macrophages), and regulatory signaling pathways related to cell adhesion and the actin cytoskeleton that promote the development of liver fibrosis and HCC. In addition, 7 specific prognostic genes (CCR7, COL3A1, FMNL2, HP, PFN1, SPP1 and TENM4) were identified and evaluated, and expression heterogeneity of core gene SPP1 and its positive correlation with immune infiltration and prognostic development were interpreted. Moreover, 6 potential small molecule drugs for the treatment of liver fibrosis and HCC were provided.Conclusion: The comprehensive investigation, based on a bioinformatics and mouse model strategy, may identify pathogenic genes, cell subsets, regulatory mechanisms, prognostic genes, and potential small molecule drugs, thereby providing valuable insights into the clinical prognosis and targeted treatment of liver fibrosis and HCC.

Triple negative breast cancer (TNBC) is a subtype of breast cancer with the highest degree of malignancy and the worst prognosis. The application of immunotherapy for TNBC is limited. This study was to verify the potential application of chimeric antigen receptor-T cells (CAR-T cells) targeting CD24 named as 24BBz in treatment of TNBC. 24BBz was constructed by lentivirus infection and then was co-culture with breast cancer cell lines to evaluate the activation, proliferation and cytotoxicity of engineered T cells. The anti-tumor activity of 24BBz was verified in the subcutaneous xenograft model of nude mice. We found that CD24 gene was significantly up-regulated in breast cancer (BRCA), especially in TNBC. 24BBz showed antigen-specific activation and dose-dependent cytotoxicity against CD24-positive BRCA tumor cells in vitro. Furthermore, 24BBz showed significant anti-tumor effect in CD24-positive TNBC xenografts and T cells infiltration in tumor tissues, while some T cells exhibited exhaustion. No pathological damage of major organs was found during the treatment. This study proved that CD24-specific CAR-T cells have potent anti-tumor activity and potential application value in treatment of TNBC.

Peptide–drug conjugates (PDCs) are an emerging targeted therapeutic drug following on from the relative success of antibody–drug conjugates (ADCs). In this class, peptides are used to target payload molecules at the disease sites, thereby reducing toxicity and improving the physicochemical properties of the payload. A PDC is composed of three parts: peptide, linker and toxin molecule, and in this structure, the selection of the target in addition to the affinity and stability of the peptide are the keys to the success of PDCs. Since the development of ADCs, drugs have undergone several updates – can PDCs leverage the experience and lessons learned from the development of ADCs over the years to achieve new success? This review presents a systematic introduction of each component of PDCs, as well as the characteristics of PDCs under investigation, with the prospect of PDC development to deepen understanding of their mechanism of action.