Exosomal miR-361-3p promotes the viability of breast cancer cells by targeting ETV7 and BATF2 to upregulate the PAI-1/ERK pathway.

BACKGROUND: Malignant progression is the major cause of poor prognosis in breast cancer (BC) patients. Plasma exosomal miRNAs have been reported to be involved in tumor progression, but their roles in BC remain unclear. METHODS: We performed plasma exosomal miRNA sequencing on 45 individuals, including healthy controls and nonmetastatic and metastatic BC patients. We examined the correlation between miRNA expression in tumor tissues and plasma exosomes in BC patients by qRT‒PCR. The effects of exosomal miR-361-3p on BC cells were determined by CellTiter-Glo, migration and wound healing assays. The target genes of miR-361-3p and downstream pathways were explored by dual-luciferase reporter assay, RNA knockdown, rescue experiments, and western blotting. We utilized murine xenograft model to further assess the impact of plasma exosomal miR-361-3p on the malignant progression of BC. RESULTS: We found that the expression level of plasma exosomal miR-361-3p gradually increased with malignant progression in BC patients, and the expression of miR-361-3p in plasma exosomes and BC tissues was positively correlated. Consistently, exosomal miR-361-3p enhanced the migration and proliferation of two BC cell lines, MDA-MB-231 and SK-BR-3. Furthermore, our data showed that miR-361-3p inhibited two novel target genes, ETV7 and BATF2, to activate the PAI-1/ERK pathway, leading to increased BC cell viability. Finally, the consistency of the in vivo experimental results supported that elevated plasma exosomal miR-361-3p promote the malignant progression of BC. CONCLUSIONS: We found for the first time that plasma exosomal miR-361-3p was associated with malignant progression in BC patients. Mechanistically, exosomal miR-361-3p can enhance the migration and proliferation of BC cells by targeting the ETV7 and BATF2/PAI-1/ERK pathways. Our data suggest that plasma exosomal miR-361-3p has the potential to serve as a biomarker for predicting malignant progression in BC patients.

Clinicopathological features and prognosis of patients with HER2-low breast cancer.

BACKGROUND: Low human epidermal growth factor receptor 2 (HER2) expression is an emerging concept in breast cancer that is defined as immunohistochemistry (IHC) 1 + or IHC 2 + and negative in situ hybridization (ISH) but has been poorly investigated. The aim of our study was to determine the frequency of low HER2 expression among HER2-negative breast cancers and compare the clinicopathological features and prognosis of HER2-low patients with those of HER2-zero patients. METHODS: We collected the data of 684 patients with primary HER2-negative breast cancer who underwent surgery between January 2012 and September 2021 from our self-built database. Clinicopathological features, recurrence-free interval (RFI) and breast cancer-specific survival (BCSS) were compared between HER2-low and HER2-zero (IHC 0) patients. RESULTS: Among the 684 patients, 512 (74.9%) patients had low HER2 expression, and 172 (25.1%) patients had zero HER2 expression. The average age was 57.7 ± 12.6 years, 472 (69.0%) patients were aged < 65 years, and 212 (31.0%) patients were aged ≥ 65 years. Compared to HER2-zero tumors, HER2-low tumors had a higher proportion of hormone receptor (HR)-positive cases (89.6% vs. 75.6%, P < 0.001) and a lower rate of histological grade III cases (29.4% vs. 38.8%, P < 0.001). Kaplan-Meier analysis showed that low HER2 expression was associated with prolonged RFI in breast cancer patients, especially in HR + breast cancer patients (P = 0.028) and < 65-year-old breast cancer patients (P = 0.000). Multivariate Cox regression analysis showed that low HER2 expression was a low-risk factor for RFI (HR: 0.531, 95% CI: 0.292-0.967, P = 0.038) but had no influence on BCSS (P = 0.272). CONCLUSIONS: HER2-low tumors had a higher proportion of HR positivity and a lower rate of histological grade III than HER2-zero tumors. Low HER2 expression seemed to be a protective factor for RFI, especially in patients with HR + and those younger than 65 years.

Carbon nanoparticles localized clipped node dissection combined with sentinel lymph node biopsy with indocyanine green and methylene blue after neoadjuvant therapy in node positive breast cancer in China: initial results of a prospective study.

BACKGROUND: This study aimed to evaluate the feasibility of applying carbon nanoparticles (CNs) to visualize clip-marked metastatic nodes in combination with indocyanine green (ICG) and methylene blue (MB) as sentinel lymph node (SLN) tracers for targeted axillary dissection (TAD) after neoadjuvant therapy (NAT). METHODS: This feasibility trial enrolled 40 patients with node-positive breast cancer, and 38 patients completed NAT and surgery were included in the final analysis. Before NAT or surgery, clip-marked nodes were localized with CNs by ultrasonography. After NAT, the clip-marked nodes were removed under the guidance of carbon-tattooing and confirmed by radiography. SLNs were mapped with ICG and MB. Axillary lymph node dissection (ALND) was performed for all patients after TAD. RESULTS: The clip-marked nodes were retrieved in all patients. The SLN identification rate was 100%. 29 of 38 clipped-nodes were SLNs. The false-negative rate was 6.25% (2/32,95% CI:1.09% ~ 22.22%) for TAD nodes and 9.38% (3/32,95% CI 3.24%-24.22%) for SLNs, and 18.75% for clipped-nodes (6/32, 95% CI:7.86% ~ 37.04%). No adverse events were reported during clip placement, CNs localization, or the TAD procedure. CONCLUSIONS: The TAD procedure with CNs to locate and visualize clipped nodes as well as SLN tracing with ICG and MB has good accessibility in China, and this technique could be easily mastered by experienced surgeons. The modified TAD technique has also demonstrated good predictive ability for residual axillary lesions after NAT, and is worth of further evaluation.

Short-term and long-term outcomes of indocyanine green for sentinel lymph node biopsy in early-stage breast cancer.

BACKGROUND: Indocyanine green (ICG) is becoming a frequently used sentinel lymph node (SLN) tracer of breast cancer in China. However, there is still a lack of data on its safety. We reported the clinical outcome of ICG as a tracer of SLN over a median 67-month follow-up period to evaluate its feasibility in clinically node-negative patients with breast cancer. METHODS: A total of 194 consecutive patients underwent sentinel lymph node biopsy (SLNB) with ICG, radioisotopes (RI) and methylene blue (MB), or with ICG and MB. The SLN mapping data by each tracer was recorded, and safety outcomes were analyzed through follow-up. RESULTS: With the triad mapping (N = 44), the identification rate of SLN by ICG was 95.5%, slightly higher than that of MB (86.4%) and comparable with RI (95.5%) and combined methods (95.5%, 100%) (p = 0.068). Analysis of all candidates (N = 194) demonstrated that the identification rate of SLN by ICG or by ICG and MB was 99%, significantly higher than that by MB (92.8%) (p < 0.0001). No tracer-related allergic reaction and permanent skin staining of ICG were observed. Local disease progression was reported in 2 of the 194 patients at the ipsilateral axilla. After remedial axillary lymph node dissection, no disease progression was detected at follow-up. CONCLUSIONS: ICG as an SLN tracer is more accurate than MB and comparable to the combined methods and has good clinical safety. ICG can be considered a useful supplement or suitable alternative to traditional tracers.

Dual Roles of GSNOR1 in Cell Death and Immunity in Tetraploid Nicotiana tabacum.

S-nitrosoglutathione reductase 1 (GSNOR1) is the key enzyme that regulates cellular homeostasis of S-nitrosylation. Although extensively studied in Arabidopsis, the roles of GSNOR1 in tetraploid Nicotiana species have not been investigated previously. To study the function of NtGSNOR1, we knocked out two NtGSNOR1 genes simultaneously in Nicotiana tabacum using clustered regularly interspaced short palindromic repeats (CRISPR)/caspase 9 (Cas9) technology. To our surprise, spontaneous cell death occurred on the leaves of the CRISPR/Cas9 lines but not on those of the wild-type (WT) plants, suggesting that NtGSNOR1 negatively regulates cell death. The natural cell death on the CRISPR/Cas9 lines could be a result from interactions between overaccumulated nitric oxide (NO) and hydrogen peroxide (H2O2). This spontaneous cell death phenotype was not affected by knocking out two Enhanced disease susceptibility 1 genes (NtEDS11a/1b) and thus was independent of the salicylic acid (SA) pathway. Unexpectedly, we found that the NtGSNOR1a/1b knockout plants displayed a significantly (p < 0.001) enhanced resistance to paraquat-induced cell death compared to WT plants, suggesting that NtGSNOR1 functions as a positive regulator of the paraquat-induced cell death. The increased resistance to the paraquat-induced cell death of the NtGSNOR1a/1b knockout plants was correlated with the reduced level of H2O2 accumulation. Interestingly, whereas the N gene-mediated resistance to Tobacco mosaic virus (TMV) was significantly enhanced (p < 0.001), the resistance to Pseudomonas syringae pv. tomato DC3000 was significantly reduced (p < 0.01) in the NtGSNOR1a/1b knockout lines. In summary, our results indicate that NtGSNOR1 functions as both positive and negative regulator of cell death under different conditions and displays distinct effects on resistance against viral and bacterial pathogens.