A Stapled Peptide Inhibitor Targeting the Binding Interface of N6-Adenosine-Methyltransferase Subunits METTL3 and METTL14 for Cancer Therapy.

METTL3, a primary methyltransferase catalyzing the RNA N6-methyladenosine (m6A) modification, has been identified as an oncogene in several cancer types and thus nominated as a potentially effective target for therapeutic inhibition. However, current options using this strategy are limited. In this study, we targeted protein-protein interactions at the METTL3-METTL14 binding interface to inhibit complex formation and subsequent catalysis of the RNA m6A modification. Among candidate peptides, RM3 exhibited the highest anti-cancer potency, inhibiting METTL3 activity while also facilitating its proteasomal degradation. We then designed a stapled peptide inhibitor (RSM3) with enhanced peptide stability and formation of the α-helical secondary structure required for METTL3 interaction. Functional and transcriptomic analysis in vivo indicated that RSM3 induced upregulation of programmed cell death-related genes while inhibiting cancer-promoting signals. Furthermore, tumor growth was significantly suppressed while apoptosis was enhanced upon RSM3 treatment, accompanied by increased METTL3 degradation, and reduced global RNA methylation levels in two in vivo tumor models. This peptide inhibitor thus exploits a mechanism distinct from other small-molecule competitive inhibitors to inhibit oncogenic METTL3 activity. Our findings collectively highlight the potential of targeting METTL3 in cancer therapies through peptide-based inhibition of complex formation and proteolytic degradation.

Protective effect of higher free thyroxine levels within the reference range on biliary tract cancer risk: a multivariable mendelian randomization and mediation analysis.

Background: Hepatobiliary cancer (HBC), including hepatocellular carcinoma (HCC) and biliary tract cancer (BTC), is currently one of the malignant tumors that mainly cause human death. Many HBCs are diagnosed in the late stage, which increases the disease burden, indicating that effective prevention strategies and identification of risk factors are urgent. Many studies have reported the role of thyroid hormones on HBC. Our research aims to assess the causal effects and investigate the mediation effects between thyroid function and HBC. Methods: Utilizing the Mendelian randomization (MR) approach, the study employs single nucleotide polymorphisms (SNPs) as instrumental variables (IVs) to explore causal links between thyroid function [free thyroxine (FT4), thyroid stimulating hormone (TSH), hyperthyroidism and hypothyroidism] and HBC. Data were sourced from the ThyroidOmic consortium and FinnGen consortium. The analysis included univariable and multivariable MR analysis, followed by mediation analysis. Results: The study found a significant causal association between high FT4 levels and the reduced risk of BTC, but not HCC. However, TSH, hyperthyroidism and hypothyroidism had no causal associations with the risk of HBC. Notably, we also demonstrated that only higher FT4 levels with the reference range (FT4-RR) could reduce the risk of BTC because this protective effect no longer existed under the conditions of hyperthyroidism or hypothyroidism. Finally, we found that the protective effect of FT4-RR on BTC was mediated partially by decreasing the risk of metabolic syndrome (MetS) and reducing the waist circumference (WC). Conclusion: The findings suggest that higher FT4-RR may have a protective effect against BTC, which is partially mediated by decreased risk of MetS and a reduction in WC. This study highlights the potential role of FT4 in the pathogenesis of BTC and underscores that MetS and WC may play mediation effects as two mediators in this process.

Prediction of malignant esophageal fistula in esophageal cancer using a radiomics-clinical nomogram.

BACKGROUND: Malignant esophageal fistula (MEF), which occurs in 5% to 15% of esophageal cancer (EC) patients, has a poor prognosis. Accurate identification of esophageal cancer patients at high risk of MEF is challenging. The goal of this study was to build and validate a model to predict the occurrence of esophageal fistula in EC patients. METHODS: This study retrospectively enrolled 122 esophageal cancer patients treated by chemotherapy or chemoradiotherapy (53 with fistula, 69 without), and all patients were randomly assigned to a training (n = 86) and a validation (n = 36) cohort. Radiomic features were extracted from pre-treatment CTs, clinically predictors were identified by logistic regression analysis. Lasso regression model was used for feature selection, and radiomics signature building. Multivariable logistic regression analysis was used to develop the clinical nomogram, radiomics-clinical nomogram and radiomics prediction model. The models were validated and compared by discrimination, calibration, reclassification, and clinical benefit. RESULTS: The radiomic signature consisting of ten selected features, was significantly associated with esophageal fistula (P = 0.001). Radiomics-clinical nomogram was created by two predictors including radiomics signature and stenosis, which was identified by logistic regression analysis. The model showed good discrimination with an AUC = 0.782 (95% CI 0.684-0.8796) in the training set and 0.867 (95% CI 0.7461-0.987) in the validation set, with an AIC = 101.1, and good calibration. When compared to the clinical prediction model, the radiomics-clinical nomogram improved NRI by 0.236 (95% CI 0.153, 0.614) and IDI by 0.125 (95% CI 0.040, 0.210), P = 0.004. CONCLUSION: We developed and validated the first radiomics-clinical nomogram for malignant esophageal fistula, which could assist clinicians in identifying patients at high risk of MEF.

TMEM9 promotes lung adenocarcinoma progression via activating the MEK/ERK/STAT3 pathway to induce VEGF expression.

Abnormal Transmembrane protein 9 (TMEM9) expression has been identified in various human tumors. However, the prognostic potential and mechanistic role of TMEM9 in lung adenocarcinoma (LUAD) remain unclear. Here, we first found a significant upregulation of TMEM9 in LUAD tissues, and TMEM9 expression was positively correlated with microvessel density (MVD), T stage, and clinical stage. Survival analysis demonstrated TMEM9 was an independent indicator of poor prognosis in LUAD patients. In addition, downregulation of TMEM9 suppressed tumor growth and metastasis in vitro and in vivo models, and reduced HUVEC proliferation, migration, and tube formation in a cancer cell/HUVEC coculture model. Furthermore, TMEM9 upregulated VEGF expression, and VEGF-neutralizing antibodies reversed HUVEC angiogenesis and cancer cell migration ability caused by overexpression of TMEM9. In contrast, recombinant VEGF (rVEGF) abolished the inhibitory effect of TMEM9-knockdown LUAD cells on HUVEC angiogenesis and tumor cell migration. Moreover, we showed that TMEM9 upregulated VEGF expression by activating the mitogen-activated protein kinase/extracellular signal-regulated kinase/STAT3 (MEK/ERK/STAT3) pathway. Together, our study provides mechanistic insights into the role of TMEM9 in LUAD and highlights the potential of targeting the TMEM9/MEK/ERK/STAT3/VEGF pathway as a novel therapy for preventing LUAD progression.

Fucoidan enhances the anti-tumor effect of anti-PD-1 immunotherapy by regulating gut microbiota.

Currently, the clinical efficacy of anti-PD-1/PD-L1 monotherapy strategies against breast cancer is limited, and low response rates need to be improved. Gut microbiota plays a crucial role in the sensitization process of immunotherapy. As a natural dietary supplement, fucoidan has been reported to have immunomodulatory effects, while some studies have found that oral fucoidan may act as a potential prebiotic to modulate the gut microbiota. Therefore, this study investigated whether fucoidan could enhance the effects of anti-PD-1 monoclonal antibody antitumor immunotherapy by modulating gut microbiota and its metabolites. We found that the anti-tumor effect of the combination treatment was significantly enhanced, while fucoidan significantly improved the composition of the gut microbiota by increasing the number of potentially beneficial bacteria, such as Bifidobacterium, Faecalibaculum and Lactobacillus. Interference with the gut microbiota by antibiotics revealed impaired antitumor efficacy, confirming the necessity of gut microbiota in the antitumor effects of fucoidan in vivo. Metabolomics further revealed that fucoidan may have reversed the metabolic disturbances induced by the breast cancer model through tryptophan metabolism and glycerophospholipid metabolism pathways, with the most significant increase in the content of short-chain fatty acids, especially acetic and butyric acids. These modulations improved the function of effector T cells and suppressed Treg cell production. Thus, our findings suggest that fucoidan combined with the anti-PD-1 monoclonal antibody may be a novel strategy to sensitize breast cancer patients to anti-PD-1 monoclonal antibody immunotherapy. Meanwhile, the gut microbiota might serve as a new biomarker to predict the anti-PD-1 monoclonal antibody response to breast cancer immunotherapy.

Design of donor-acceptor conjugated polymers based on diketopyrrolopyrrole for NIR-II multifunctional imaging.

Diketopyrrolopyrrole (DPP) is an excellent photosensitizer and photothermal agent with the advantages of good planarity, strong electron affinity, high electron mobility, easy purification, easy structural modification and high molar absorption coefficient. It is regarded as one of the ideal choices for the design and synthesis of efficient organic photovoltaic materials. Therefore, two kinds of donor-acceptor (D-A) conjugated polymers were designed and synthesized with DPP as the acceptor, and their optical properties and applications in the near-infrared region were studied. The quantum yield (QY) of PBDT-DPP is 0.46%, and the highest temperature reached within 10 minutes after irradiation with a 660 nm laser is 60 °C. Another polymer, EDOT-DPP, has a QY of 0.48%, and its semiconductor polymer nanoparticle aqueous solution can reach 60 °C within 12 minutes under laser irradiation, achieving photothermal treatment of nude mice tumors. Both polymer NPs have good biocompatibility and promising applications in bioimaging and photothermal therapy.

Integrative analysis of LAG3 immune signature and identification of a LAG3-related genes prognostic signature in kidney renal clear cell carcinoma.

Immune checkpoint blockade (ICB) therapy has resulted in improved overall survival in kidney renal clear cell carcinoma (KIRC), but most treated patients fail to show durable clinical responses. Lymphocyte activation gene-3 (LAG3) is a novel inhibitory immune checkpoint, but its expression pattern, prognostic value, and immunological role in KIRC remain unknown. In this study, we utilized TCGA_KIRC RNA-sequencing data to analyze the relationship between LAG3 expression and clinical features. The single-cell sequencing data and tissue immunofluorescence are employed to investigate the subcellular localization of LAG3 in KIRC. Kaplan-Meier plotter, TIMER, and TISIDB were used to assess the association between LAG3 expression and prognosis, as well as its correlation with immune-related components. We constructed the LAG3 interaction network by using STRING, GeneMANIA, BioGRID, and HitPredict databases. We found that LAG3 is upregulated and correlates with poor prognostic phenotype in KIRC. LAG3 is predominantly expressed on exhausted CD8+ T cells and shows strong co-expression with PDCD1 in KIRC. Moreover, our findings indicated that LAG3 not only inhibits T cell activation but also potentially regulates cell adhesion in KIRC. In conclusion, our study implies that LAG3 can serve as a potential prognostic biomarker for KIRC. Furthermore, blocking both LAG3 and PDCD1 may alleviate resistance to anti-PDCD1 therapy, providing novel insights for immunotherapy decision-making in KIRC patients.

mDia formins form hetero-oligomers and cooperatively maintain murine hematopoiesis.

mDia formin proteins regulate the dynamics and organization of the cytoskeleton through their linear actin nucleation and polymerization activities. We previously showed that mDia1 deficiency leads to aberrant innate immune activation and induces myelodysplasia in a mouse model, and mDia2 regulates enucleation and cytokinesis of erythroblasts and the engraftment of hematopoietic stem and progenitor cells (HSPCs). However, whether and how mDia formins interplay and regulate hematopoiesis under physiological and stress conditions remains unknown. Here, we found that both mDia1 and mDia2 are required for HSPC regeneration under stress, such as serial plating, aging, and reconstitution after myeloid ablation. We showed that mDia1 and mDia2 form hetero-oligomers through the interactions between mDia1 GBD-DID and mDia2 DAD domains. Double knockout of mDia1 and mDia2 in hematopoietic cells synergistically impaired the filamentous actin network and serum response factor-involved transcriptional signaling, which led to declined HSPCs, severe anemia, and significant mortality in neonates and newborn mice. Our data demonstrate the potential roles of mDia hetero-oligomerization and their non-rodent functions in the regulation of HSPCs activity and orchestration of hematopoiesis.

Tributyrin alleviates gut microbiota dysbiosis to repair intestinal damage in antibiotic-treated mice.

Tributyrin (TB) is a butyric acid precursor and has a key role in anti-inflammatory and intestinal barrier repair effects by slowly releasing butyric acid. However, its roles in gut microbiota disorder caused by antibiotics remain unclear. Herein, we established an intestinal microbiota disorder model using ceftriaxone sodium via gavage to investigate the effects of different TB doses for restoring gut microbiota and intestinal injury. First, we divided C57BL/6 male mice into two groups: control (NC, n = 8) and experimental (ABx, n = 24) groups, receiving gavage with 0.2 mL normal saline and 400 mg/mL ceftriaxone sodium solution for 7 d (twice a day and the intermediate interval was 6 h), respectively. Then, mice in the ABx group were randomly split into three groups: model (M, 0.2 mL normal saline), low TB group (TL, 0.3 g/kg BW), and high TB group (TH, 3 g/kg BW) for 11 d. We found that TB supplementation alleviated antibiotics-induced weight loss, diarrhea, and intestinal tissue damage. The 16S rRNA sequence analysis showed that TB intervention increased the α diversity of intestinal flora, increased potential short-chain fatty acids (SCFAs)-producing bacteria (such as Muribaculaceae and Bifidobacterium), and inhibited the relative abundance of potentially pathogenic bacteria (such as Bacteroidetes and Enterococcus) compared to the M group. TB supplementation reversed the reduction in SCFAs production in antibiotic-treated mice. Additionally, TB downregulated the levels of serum LPS and zonulin, TNF-α, IL-6, IL-1ß and NLRP3 inflammasome-related factors in intestinal tissue and upregulated tight junction proteins (such as ZO-1 and Occludin) and MUC2. Overall, the adjustment ability of low-dose TB to the above indexes was stronger than high-dose TB. In conclusion, TB can restore the dysbiosis of gut microbiota, increase SCFAs, suppress inflammation, and ameliorate antibiotic-induced intestinal damage, indicating that TB might be a potential gut microbiota modulator.

The Combination of Inositol Hexaphosphate and Inositol Inhibits Metastasis of Colorectal Cancer Cells by Upregulating Claudin 7.

Inositol hexaphosphate (IP6), a widely found natural bioactive substance in grains, effectively inhibits the progression of colorectal cancer (CRC) when used in combination with inositol (INS). We previously showed that supplementation of IP6 and INS upregulated the claudin 7 gene in orthotropic CRC xenografts in mice. The aim of this study was to elucidate the role of claudin 7 in the inhibition of CRC metastasis by IP6 and INS, and explore the underlying mechanisms. We found that IP6, INS and their combination inhibited the epithelial-mesenchymal transition (EMT) of colon cancer cell lines (SW480 and SW620), as indicated by upregulation of claudin 7 and E-cadherin, and downregulation of N-cadherin. The effect of IP6 and INS was stronger compared to either agent alone (combination index < 1). Furthermore, the silencing of the claudin 7 gene diminished the anti-metastatic effects of IP6 and INS on SW480 and SW620 cells. Consistent with in vitro findings, the combination of IP6 and INS suppressed CRC xenograft growth in a mouse model, which was neutralized by claudin 7. Taken together, the combination of IP6 and INS can inhibit CRC metastasis by blocking EMT of tumor cells through upregulation of claudin 7.

The role of FoxM1 in immune cells.

Forkhead box M1 (FoxM1), a proliferation specific transcriptional modulator, plays a principal role in many physiological and pathological processes. FoxM1-mediated oncogenic processes have been well addressed. However, functions of FoxM1 in immune cells are less summarized. The literatures about the expression of FoxM1 and its regulation on immune cells were searched on PubMed and Google Scholar. In this review, we provide an overview on the roles of FoxM1 in regulating functions of immune cells, including T cells, B cells, monocytes, macrophages, and dendritic cells, and discuss their contributions to diseases.

Clonal hematopoiesis and bone marrow inflammation.

Clonal hematopoiesis (CH) occurs in hematopoietic stem cells with increased risks of progressing to hematologic malignancies. CH mutations are predominantly found in aged populations and correlate with an increased incidence of cardiovascular and other diseases. Increased lines of evidence demonstrate that CH mutations are closely related to the inflammatory bone marrow microenvironment. In this review, we summarize the recent advances in this topic starting from the discovery of CH and its mutations. We focus on the most commonly mutated and well-studied genes in CH and their contributions to the innate immune responses and inflammatory signaling, especially in the hematopoietic cells of bone marrow. We also aimed to discuss the interrelationship between inflammatory bone marrow microenvironment and CH mutations. Finally, we provide our perspectives on the challenges in the field and possible future directions to help understand the pathophysiology of CH.

Anticancer activities of natural antimicrobial peptides from animals.

Cancer is the most common cause of human death worldwide, posing a serious threat to human health and having a negative impact on the economy. In the past few decades, significant progress has been made in anticancer therapies, but traditional anticancer therapies, including radiation therapy, surgery, chemotherapy, molecular targeted therapy, immunotherapy and antibody-drug conjugates (ADCs), have serious side effects, low specificity, and the emergence of drug resistance. Therefore, there is an urgent need to develop new treatment methods to improve efficacy and reduce side effects. Antimicrobial peptides (AMPs) exist in the innate immune system of various organisms. As the most promising alternatives to traditional drugs for treating cancers, some AMPs also have been proven to possess anticancer activities, which are defined as anticancer peptides (ACPs). These peptides have the advantages of being able to specifically target cancer cells and have less toxicity to normal tissues. More and more studies have found that marine and terrestrial animals contain a large amount of ACPs. In this article, we introduced the animal derived AMPs with anti-cancer activity, and summarized the types of tumor cells inhibited by ACPs, the mechanisms by which they exert anti-tumor effects and clinical applications of ACPs.

Epiplakin1 promotes the progression of esophageal squamous cell carcinoma by activating the PI3K-AKT signaling pathway.

BACKGROUND: Epiplakin1 (EPPK1) has been associated with disease progression and unfavorable prognosis of many cancers, but its functional involvement in esophageal squamous cell carcinoma (ESCC) remains to be uncovered. METHODS: The Quantitative Real-time PCR (qPCR) assay was employed to determine the expression of EPPK1 in ESCC tissues and cells. CCK-8 assay, colony forming assay, wound healing assay, and transwell invasion assay were utilized to evaluate the effects of EPPK1 on cell proliferation, migration, and invasion capacity in ESCC cells using small interfering ribonucleic acids. Flow cytometry was performed to estimate the cell apoptotic rate caused by silencing of EPPK1. The proteins related to epithelial-to-mesenchymal transition (EMT), apoptosis, and activation of the phosphatidylinositol 3-kinase/serine threonine protein kinase 1 (PI3K/AKT) signaling pathway were measured by western blot. RESULTS: The expression of EPPK1 was dramatically increased in ESCC tissues and cells compared to that in relative controls. Additionally, silencing of EPPK1 suppressed ESCC cell growth, colony formation, migration, invasion, and EMT, while promoting ESCC cell apoptosis. Furthermore, EPPK1 induced ESCC cell progression via mediating the PI3K/AKT signaling pathway. CONCLUSION: EPPK1 promotes ESCC progression by modulating the PI3K/AKT signaling pathway and could serve as a potential target for ESCC treatment.

An update on the roles of circular RNAs in spinal cord injury.

Spinal cord injury (SCI) is a disabling condition for which therapeutic options are limited. Increasing number of microarray and next-generation sequencing studies have demonstrated that SCI coincides with altered expression of circular RNAs (circRNAs) in the damaged tissue. Emerging functional evidence further pinpointed specific differentially expressed circRNAs (e.g., circ-HIPK3, cicRNA.7079, circRNA_01477, circRNA-2960, and circ_0001723) for their effects on cellular processes relevant to SCI repair and regeneration, including neuronal apoptosis, astrocyte activation, and neuroinflammation, via sponging SCI-related microRNAs. Although circRNAs and their target microRNAs appear to be good candidates for therapeutic exploitation in SCI, further investigation into the efficient delivery of these regulatory molecules in a cell-type specific manner is a pre-requisite for translating these basic discoveries into clinical benefits.

Combined immunotherapy and targeted treatment for primary alveolar soft part sarcoma of the lung: case report and literature review.

Primary acinar soft part sarcoma of the lung (ASPS) is a rare malignancy with unique cellular structure and clinical and genetic characteristics. Most patients do not exhibit clear clinical symptoms, with only a few developing respiratory symptoms. The typical histological characteristics are acinoid or organ-like structures. Immunofluorescence in situ hybridization suggests a rearrangement of the transcription factor E3 gene. Patients respond poorly to chemotherapy and are, thus, primarily treated with surgery and targeted therapy. We report herein a unique case of primary alveolar soft part sarcoma of the lung. The patient was a 24-year-old man with metastases to multiple organs, such as the brain, lungs, pancreas, and liver. The craniocerebral lesions attained partial remission after whole-brain radiotherapy and targeted combined immunotherapy, and other distant metastases completely disappeared after targeted combined immunotherapy (anlotinib and camrelizumab), indicating significant treatment efficacy. Anlotinib is an oral multi-target tyrosine kinase inhibitor (TKI) that exerts its anti-tumor effects by acting on various kinases. Camrelizumab is a humanized immunoglobulin G4 monoclonal antibody that can target PD-1 to block the interaction between PD-L1 and programmed death ligand 2, ultimately causing an anti-tumor effect. This is the first report of successful use of anlotinib combined with camrelizumab in the treatment of advanced primary ASPS. The treatment benefit provides preliminary evidence that targeted therapy, combined with immunotherapy, may be a safe and effective approach to treat primary pulmonary ASPS patients, thus warranting further investigation.

Real-World Data on Osimertinib in Chinese Patients with Pretreated, EGFR T790M Mutation Positive, Advanced Non-Small Cell Lung Cancer: A Retrospective Study.

PURPOSE: As a third-generation EGFR TKI has been taken orally, Osimertinib effectively inhibits mutant EGFR, including T790M EGFR resistance mutations. Here, we examined real-world efficacy and tolerability of Osimertinib among Chinese patients with advanced EGFR T790M-mutant NSCLC. PATIENTS AND METHODS: A total of 106 advanced NSCLC patients who were taking Osimertinib following disease progression after EGFR-TKIs or other treatments were retrospectively recruited in this study. The PFS and OS after Osimertinib treatment were analyzed as the primary endpoints. RESULTS: Osimertinib was used as a second line and ≥3rd line treatment in 22.6% and 77.4% of the patients, respectively. DCR and ORR were 93.4% and 57.5%, respectively. Median PFS was 12.4 12 (95% CI, 10.5-13.5) months. The PFS was 11 (8.0, 14.0) and 12 (10.3,13.7) months (p = 0.373), in patients with and without CNS metastasis, respectively. PFS in 2nd and ≥3rd line treatment was 11 (9.0, 13.0) and 12.4 12 (8.9, 15.1) months (p = 0.799), respectively. In patients with EGFR exon 19 deletion and exon 21 L858 mutation, the median PFS was 11 (9.2, 12.8) and 12 (9.2, 14.8) months, respectively (p = 0.833). Median PFS in the monotherapy group and combined anti-angiogenesis group was 11 (9.9,12.1) and 14 (11.2,16.8) months, respectively. Median OS after Osimertinib initiation was 27 (19.6, 34.4) months: 15 (6.9, 23.1) and 27 (22, 32) months in patients with and without CNS metastasis (p=0.027), 27 (20.3,33.7) months and (undefined) as second line or ≥3rd line of treatment (p = 0.421), respectively. In patients with exon 19 deletion, the median OS was not reached, and in patients with exon 21 L858 mutations, the median OS was 23 (19.1,29.9) months (p=0.027). Median OS in the monotherapy group was 27 (21.7,32.3) months, and in combined anti-angiogenesis group was not reached (p=0.68). CONCLUSION: Osimertinib can effectively treat advanced NSCLC with T790M mutations independently of previous treatment lines.

Association between levels of tumor-infiltrating lymphocytes in different subtypes of primary breast tumors and prognostic outcomes: a meta-analysis.

BACKGROUND: To investigate the impact of the elevation of tumor-infiltrating lymphocytes (TILs) in different molecular subtypes of primary breast cancer, i.e. each 10% increment of TILs and high-level TILs (TILs≥50%) in tumor, on overall survival (OS) and pathological complete response (pCR) and to compare the presentation of high-level TILs across these molecular subtypes. METHODS: Citation retrieval was performed in the PubMed, Cochrane Library, Embase and Web of Science databases. All statistical calculations were performed by the software of StataSE version 12.0. RESULTS: Twenty-two eligible clinical trials including 15,676 unique patients were included for meta-analysis. Each 10% increment of TILs significantly improved OS in human epidermal growth factor receptor 2 (HER2)-overexpression (pooled Hazard ratio (HR), 0.92; 95% CI, 0.89-0.95) and triple-negative (TN) (pooled HR, 0.90; 95% CI, 0.89-0.92) breast tumors but not in luminal tumor subtype (pooled HR, 1.06; 95% CI, 0.99-1.13). It was also associated with an increased pCR rate in breast cancers (pooled Odds ratio (OR), 1.27; 95% CI, 1.19-13.5). High-level TILs were significantly related with a higher pCR rate (pooled OR, 2.73; 95% CI, 2.40-3.01) than low-level TILs. The HER2-amplified (pooled OR, 3.14; 95% CI, 1.95-5.06) and TN (pooled OR, 4.09; 95% CI, 2.71-6.19) phenotypes of breast cancers expressed significantly more high-level TILs than the luminal tumor subtype, although the presentation of those between the former two subsets was not significantly different (pooled OR, 1.30; 95%CI, 0.83-2.04). CONCLUSIONS: The elevation of TILs in breast tumors predicts favorable prognostic outcomes, particularly in the HER2-overexpression and TN subtypes.