Study on the mechanism of heterogeneous tumor-associated macrophages in three subtypes of breast cancer through the integration of single-cell RNA sequencing and in vitro experiments.

BACKGROUND: Tumor-associated macrophages (TAM) exert a significant influence on the progression and heterogeneity of various subtypes of breast cancer (BRCA). However, the roles of heterogeneous TAM within BRCA subtypes remain unclear. Therefore, this study sought to elucidate the role of TAM across the following three BRCA subtypes: triple-negative breast cancer, luminal, and HER2. MATERIALS AND METHODS: This investigation aimed to delineate the variations in marker genes, drug sensitivity, and cellular communication among TAM across the three BRCA subtypes. We identified specific ligand-receptor (L-R) pairs and downstream mechanisms regulated by VEGFA-VEGFR1, SPP1-CD44, and SPP1-ITGB1 L-R pairs. Experimental verification of these pairs was conducted by co-culturing macrophages with three subtypes of BRCA cells. RESULTS: Our findings reveal the heterogeneity of macrophages within the three BRCA subtypes, evidenced by variations in marker gene expression, composition, and functional characteristics. Notably, heterogeneous TAM were found to promote invasive migration and epithelial-mesenchymal transition (EMT) in MDA-MB-231, MCF-7, and SKBR3 cells, activating NF-κB pathway via P38 MAPK, TGF-ß1, and AKT, respectively, through distinct VEGFA-VEGFR1, SPP1-CD44, and SPP1-ITGB1 L-R pairs. Inhibition of these specific L-R pairs effectively reversed EMT, migration, and invasion of each cancer cells. Furthermore, we observed a correlation between ligand gene expression and TAM sensitivity to anticancer drugs, suggesting a potential strategy for optimizing personalized treatment guidance. CONCLUSION: Our study highlights the capacity of heterogeneous TAM to modulate biological functions via distinct pathways mediated by specific L-R pairs within diverse BRCA subtypes. This study might provide insights into precision immunotherapy of different subtypes of BRCA.

Pooled analysis of NeoCARH and NeoCART trials: patient-reported outcomes in patients with early-stage breast cancer receiving platinum-based or anthracycline-based neoadjuvant chemotherapy.

PURPOSE: Anthracycline-based or platinum-based neoadjuvant chemotherapy belongs to the standard treatment for early-stage breast cancer (EBC) that is either triple-negative or human epidermal growth factor receptor 2 positive (HER2 +). Currently, there is a paucity of data comparing their impact on health-related quality of life (HRQoL). METHODS: Triple-negative or HER2 + EBC from our two prospective randomized controlled trials, neoCARH and neoCART, were divided into two groups based on the neoadjuvant chemotherapy regimens they received: anthracycline-based or platinum-based group. HRQoL was the exploratory endpoint in these two trials, which was assessed using the European Organization for Research and Treatment of Cancer Quality of Life-Core30 and Breast23 questionnaires. The primary variable of interest was the C30 summary score (C30-SumSc). Assessments were carried out at baseline, after neoadjuvant chemotherapy, and 1 year and 2 years after diagnosis. RESULTS: The mean questionnaires' compliance rate was 95.0%. After neoadjuvant chemotherapy, 210 patients had evaluable HRQoL data, the mean least square change from baseline for the platinum-based group was - 15.997 (95% confidence interval (CI): - 17.877 to - 14.117), and it was - 20.156 (95% CI: - 22.053 to - 18.258) for the anthracycline-based group (difference: 4.159, 95% CI: 1.462 to 6.855, P = 0.003, minimal important difference = 3). For the majority of the domains of interest assessed by the C30 and BR23 questionnaires, the platinum-based group demonstrated superior outcomes in comparison to the anthracycline-based group. CONCLUSION: Patients receiving platinum-based or anthracycline-based regimens both experienced worsened HRQoL after neoadjuvant chemotherapy; however, the former provided relatively better HRQoL compared with the latter. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: NCT03140553. Registered 4 May 2017 (neoCARH). NCT03154749. Registered 16 May 2017 (neoCART).

YY1 mediated DCUN1D5 transcriptional activation promotes triple-negative breast cancer progression by targeting FN1/PI3K/AKT pathway.

Triple-negative breast cancer (TNBC) is more aggressive and has a higher metastasis rate compared with other subtypes of breast cancer. Due to the lack of drug-targetable receptors, chemotherapy is now the only available systemic treatment for TNBC. However, some patients might still develop drug resistance and have poor prognosis. Therefore, novel molecular biomarkers and new treatment targets are urgently needed for patients with TNBC. To provide molecular insights into TNBC progression, we investigated the function and the underlying mechanism of Defective in cullin neddylation 1 domain containing 5 (DCUN1D5) in the regulation of TNBC. By TCGA dataset and surgical specimens with immunohistochemical (IHC) staining method, DCUN1D5 was identified to be significantly upregulated in TNBC tumor tissues and negatively associated with prognosis. A series of in vitro and in vivo experiments were performed to confirm the oncogenic role of DCUN1D5 in TNBC. Overexpression of FN1 or PI3K/AKT activator IGF-1 could restore the proliferative and invasive ability induced by DCUN1D5 knockdown and DCUN1D5 could act as a novel transcriptional target of transcription factor Yin Yang 1 (YY1). In conclusion, YY1-enhanced DCUN1D5 expression could promote TNBC progression by FN1/PI3K/AKT pathway and DCUN1D5 might be a potential prognostic biomarker and therapeutic target for TNBC treatment.

Genome-wide in vivo CRISPR screen identifies TGFß3 as actionable biomarker of palbociclib resistance in triple negative breast cancer.

Triple negative breast cancer (TNBC) remains exceptionally challenging to treat. While CDK4/6 inhibitors have revolutionized HR + breast cancer therapy, there is limited understanding of their efficacy in TNBC and meaningful predictors of response and resistance to these drugs remain scarce. We conducted an in vivo genome-wide CRISPR screen using palbociclib as a selection pressure in TNBC. Hits were prioritized using microarray data from a large panel of breast cancer cell lines to identify top palbociclib sensitizers. Our study defines TGFß3 as an actionable determinant of palbociclib sensitivity that potentiates its anti-tumor effects. Mechanistically, we show that chronic palbociclib exposure depletes p21 levels, contributing to acquired resistance, and that TGFß3 treatment can overcome this. This study defines TGFß3 as an actionable biomarker that can be used to improve patient stratification for palbociclib treatment and exploits the synergistic interaction between CDK4/6 and TGFß3 to propose a new combinatorial treatment for TNBC.

Manubriectomy made easy.

Bone metastasis is the most common form of distant metastasis encountered within the breast cancer population. Surgical resection of bone metastases is a curative treatment option in patients who present with an isolated solitary lesion and no other associated disease. This decision is typically made following a multidisciplinary discussion. Patients can also be put forward for surgical excision of bone metastases following inadequate response to chemotherapy or radiotherapy.  With tumours located in the manubrium of the sternum, surgery serves not only to resect the bone metastasis but to provide suitable chest wall reconstruction. The goal of this approach is to maintain the structural and bony stability of the chest wall as well as that of associated structures, e.g. rib insertion or articulation of the shoulder girdle. A widely utilized approach involves excising the area of metastasis within the manubrium followed by implanting a bone cement prosthesis. Titanium plates are used to fix the bone prosthesis to the sternal body inferiorly and to the remainder of the manubrium superiorly.  We present a step-by-step video tutorial for performing a lower hemi-manubriectomy in a patient with triple-negative breast cancer. Our goal is to describe the fundamental principles and surgical techniques used to perform this procedure followed by the postoperative outcomes.

Exploring the regulatory role of FBXL19-AS1 in triple-negative breast cancer through the miR-378a-3p/OTUB2 axis.

The regulatory potential of long noncoding RNA (lncRNA) FBXL19-AS1 has been highlighted in various cancers, but its effect on triple-negative breast cancer (TNBC) remains unclear. Here, we aimed to elucidate the role of FBXL19-AS1 in TNBC and its underlying mechanism. RT-qPCR was employed to detect the expressions of FBXL19-AS1 and miR-378a-3p in tissues and cells. Immunohistochemical staining and western blot were utilized to detect the expression levels of proteins. Cell activities were detected using flow cytometry, CCK-8, and transwell assay. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were deployed to investigate interactions of different molecules. Protein-protein interaction (PPI) network, gene ontology (GO), and Kyoto encyclopedia of genes and genomes (KEGG) pathways were used to analyze the downstream pathway. In vivo xenograft model was conducted to detect the effect of FBXL19-AS1 on tumor growth. FBXL19-AS1 was overexpressed in TNBC tissues and cell lines compared with counterparts. FBXL19-AS1 knockdown suppressed TNBC cell activities, whereas its overexpression exhibited the opposite effect. Mechanistically, FBXL19-AS1 was found to interact with miR-378a-3p. Further analysis revealed that miR-378a-3p exerted tumor-suppressive effects in TNBC cells. Additionally, miR-378a-3p targeted and downregulated the expression of ubiquitin aldehyde binding 2 (OTUB2), a deubiquitinase associated with TNBC progression. In vivo experiments substantiated the inhibitory effects of FBXL19-AS1 knockdown on TNBC tumorigenesis, and a miR-378a-3p inhibitor partially rescued these effects. The downstream pathway of the miR-378a-3p/OTUB2 axis was explored, revealing connections with proteins involved in modifying other proteins, removing ubiquitin molecules, and influencing signaling pathways, including the Hippo signaling pathway. Western blot analysis confirmed changes in YAP and TAZ expression levels, indicating a potential regulatory network. In summary, FBXL19-AS1 promotes exacerbation in TNBC by suppressing miR-378a-3p, leading to increased OTUB2 expression. The downstream mechanism may be related to the Hippo signaling pathway. These findings propose potential therapeutic targets for TNBC treatment.

In vitro study on anti-proliferative and anti-cancer activity of picrosides in triple-negative breast cancer.

Picrorhiza kurroa, an "Indian gentian," a known Himalayan medicinal herb with rich source of phytochemicals like picrosides I, II, and other glycosides, has been traditionally used for the treatment of liver and respiratory ailments. Picrosides anti-proliferative, anti-oxidant, anti-inflammatory and other pharmacological properties were evaluated in treating triple-negative breast cancer (TNBC). Picroside I and II were procured from Sigma-Aldrich and were analyzed for anti-cancer activity in triple-negative breast cancer (MDA-MB-231) cells. Cell viability was analyzed using MTT and trypan blue assays. Apoptosis was analyzed through DNA fragmentation and Annexin V/PI flow cytometric analysis. Wound healing and cell survival assays were employed to determine the inhibition of invasion capacity and anti-proliferative activity of picrosides in MDA-MB-231 cells. Measurement of intracellular ROS was studied through mitochondrial membrane potential assessment using DiOC6 staining for anti-oxidant activity of picrosides in MDA-MB-231 cells. Both Picroside I and II have shown decreased cell viability of MDA-MB-231 cells with increasing concentrations. IC50 values of 95.3 µM and 130.8 µM have been obtained for Picroside I and II in MDA-MB-231 cells. Early apoptotic phase have shown an increase of 20% (p < 0.05) with increasing concentrations (0, 50, 75, and 100 µM) of Picroside I and 15% (p < 0.05) increase with Picroside II. Decrease in mitochondrial membrane potential of 2-2.5-fold (p < 0.05) was observed which indicated decreased reactive oxygen species (ROS) generation with increasing concentrations of Picroside I and II. An increasing percentage of 70-80% (p < 0.05) cell population was arrested in G0/G1 phase of cell cycle after Picroside I and II treatment in cancer cells. Our results suggest that Picroside I and II possess significant anti-proliferative and anti-cancer activity which is mediated by inhibition of cell growth, decreased mitochondrial membrane potential, DNA damage, apoptosis, and cell cycle arrest. Therefore, Picroside I and II can be developed as a potential anti-cancer drug of future and further mechanistic studies are underway to identify the mechanism of anti-cancer potential.

Enhancing therapeutic efficacy: sustained delivery of 5-fluorouracil (5-FU) via thiolated chitosan nanoparticles targeting CD44 in triple-negative breast cancer.

Our current study reports the successful synthesis of thiolated chitosan-based nanoparticles for targeted drug delivery of 5-Fluorouracil. This process was achieved through the ionic gelation technique, aiming to improve the efficacy of the chemotherapeutic moiety by modifying the surface of the nanoparticles (NPs) with a ligand. We coated these NPs with hyaluronic acid (HA) to actively target the CD44 receptor, which is frequently overexpressed in various solid malignancies, including breast cancer. XRD, FTIR, SEM, and TEM were used for the physicochemical analysis of the NPs. These 5-Fluorouracil (5-FU) loaded NPs were evaluated on MDA-MB-231 (a triple-negative breast cell line) and MCF-10A (normal epithelial breast cells) to determine their in vitro efficacy. The developed 5-FU-loaded NPs exhibited a particle size within a favorable range (< 300 nm). The positive zeta potential of these nanoparticles facilitated their uptake by negatively charged cancer cells. Moreover, they demonstrated robust stability and achieved high encapsulation efficiency. These nanoparticles exhibited significant cytotoxicity compared to the crude drug (p < 0.05) and displayed a promising release pattern consistent with the basic diffusion model. These traits improve the pharmacokinetic profile, efficacy, and ability to precisely target these nanoparticles, offering a potentially successful anticancer treatment for breast cancer. However, additional in vivo assessments of these formulations are obligatory to confirm these findings.

Comprehensive characterization of stemness-related lncRNAs in triple-negative breast cancer identified a novel prognostic signature related to treatment outcomes, immune landscape analysis and therapeutic guidance: a silico analysis with in vivo experiments.

BACKGROUND: Cancer stem cells (CSCs) and long non-coding RNAs (lncRNAs) are known to play a crucial role in the growth, migration, recurrence, and drug resistance of tumor cells, particularly in triple-negative breast cancer (TNBC). This study aims to investigate stemness-related lncRNAs (SRlncRNAs) as potential prognostic indicators for TNBC patients. METHODS: Utilizing RNA sequencing data and corresponding clinical information from the TCGA database, and employing Weighted Gene Co-expression Network Analysis (WGCNA) on TNBC mRNAsi sourced from an online database, stemness-related genes (SRGs) and SRlncRNAs were identified. A prognostic model was developed using univariate Cox and LASSO-Cox analysis based on SRlncRNAs. The performance of the model was evaluated using Kaplan-Meier analysis, ROC curves, and ROC-AUC. Additionally, the study delved into the underlying signaling pathways and immune status associated with the divergent prognoses of TNBC patients. RESULTS: The research identified a signature of six SRlncRNAs (AC245100.6, LINC02511, AC092431.1, FRGCA, EMSLR, and MIR193BHG) for TNBC. Risk scores derived from this signature were found to correlate with the abundance of plasma cells. Furthermore, the nominated chemotherapy drugs for TNBC exhibited considerable variability between different risk score groups. RT-qPCR validation confirmed abnormal expression patterns of these SRlncRNAs in TNBC stem cells, affirming the potential of the SRlncRNAs signature as a prognostic biomarker. CONCLUSION: The identified signature not only demonstrates predictive power in terms of patient outcomes but also provides insights into the underlying biology, signaling pathways, and immune status associated with TNBC prognosis. The findings suggest the possibility of guiding personalized treatments, including immune checkpoint gene therapy and chemotherapy strategies, based on the risk scores derived from the SRlncRNA signature. Overall, this research contributes valuable knowledge towards advancing precision medicine in the context of TNBC.

Regulation of cellular and molecular markers of epithelial-mesenchymal transition by Brazilin in breast cancer cells.

Breast cancer is the most common invasive neoplasm and the leading cause of cancer death in women worldwide. The main cause of mortality in cancer patients is invasion and metastasis, where the epithelial-mesenchymal transition (EMT) is a crucial player in these processes. Pharmacological therapy has plants as its primary source, including isoflavonoids. Brazilin is an isoflavonoid isolated from Haematoxilum brasiletto that has shown antiproliferative activity in several cancer cell lines. In this study, we evaluated the effect of Brazilin on canonical markers of EMT such as E-cadherin, vimentin, Twist, and matrix metalloproteases (MMPs). By Western blot, we evaluated E-cadherin, vimentin, and Twist expression and the subcellular localization by immunofluorescence. Using gelatin zymography, we determined the levels of secretion of MMPs. We used Transwell chambers coated with matrigel to determine the in vitro invasion of breast cancer cells treated with Brazilin. Interestingly, our results show that Brazilin increases 50% in E-cadherin expression and decreases 50% in vimentin and Twist expression, MMPs, and cell invasion in triple-negative breast cancer (TNBC) MDA-MB-231 and to a lesser extend in MCF7 ER+ breast cancer cells. Together, these findings position Brazilin as a new molecule with great potential for use as complementary or alternative treatment in breast cancer therapy in the future.

Ternary heterostructure-driven photoinduced electron-hole separation enhanced oxidative stress for triple-negative breast cancer therapy.

Zinc oxide nanoparticles (ZnO NPs) stand as among the most significant metal oxide nanoparticles in trigger the formation of reactive oxygen species (ROS) and induce apoptosis. Nevertheless, the utilization of ZnO NPs has been limited by the shallowness of short-wavelength light and the constrained production of ROS. To overcome these limitations, a strategy involves achieving a red shift towards the near-infrared (NIR) light spectrum, promoting the separation and restraining the recombination of electron-hole (e--h+) pairs. Herein, the hybrid plasmonic system Au@ZnO (AZ) with graphene quantum dots (GQDs) doping (AZG) nano heterostructures is rationally designed for optimal NIR-driven cancer treatment. Significantly, a multifold increase in ROS generation can be achieved through the following creative initiatives: (i) plasmonic Au nanorods expands the photocatalytic capabilities of AZG into the NIR domain, offering a foundation for NIR-induced ROS generation for clinical utilization; (ii) elaborate design of mesoporous core-shell AZ structures facilitates the redistribution of electron-hole pairs; (iii) the incorporation GQDs in mesoporous structure could efficiently restrain the recombination of the e--h+ pairs; (iv) Modification of hyaluronic acid (HA) can enhance CD44 receptor mediated targeted triple-negative breast cancer (TNBC). In addition, the introduced Au NRs present as catalysts for enhancing photothermal therapy (PTT), effectively inducing apoptosis in tumor cells. The resulting HA-modified AZG (AZGH) exhibits efficient hot electron injection and e--h+ separation, affording unparalleled convenience for ROS production and enabling NIR-induced PDT for the cancer treanment. As a result, our well-designed mesoporous core-shell AZGH hybrid as photosensitizers can exhibit excellent PDT efficacy.

Neoadjuvant radiotherapy in ER+, HER2+, and triple-negative -specific breast cancer based humanized tumor mice enhances anti-PD-L1 treatment efficacy.

Pre-operative radiation therapy is not currently integrated into the treatment protocols for breast cancer. However, transforming immunological "cold" breast cancers by neoadjuvant irradiation into their "hot" variants is supposed to elicit an endogenous tumor immune defense and, thus, enhance immunotherapy efficiency. We investigated cellular and immunological effects of sub-lethal, neoadjuvant irradiation of ER pos., HER2 pos., and triple-negative breast cancer subtypes in-vitro and in-vivo in humanized tumor mice (HTM). This mouse model is characterized by a human-like immune system and therefore facilitates detailed analysis of the mechanisms and efficiency of neoadjuvant, irradiation-induced "in-situ vaccination", especially in the context of concurrently applied checkpoint therapy. Similar to clinical appearances, we observed a gradually increased immunogenicity from the luminal over the HER2-pos. to the triple negative subtype in HTM indicated by an increasing immune cell infiltration into the tumor tissue. Anti-PD-L1 therapy divided the HER2-pos. and triple negative HTM groups into responder and non-responder, while the luminal HTMs were basically irresponsive. Irradiation alone was effective in the HER2-pos. and luminal subtype-specific HTM and was supportive for overcoming irresponsiveness to single anti-PD-L1 treatment. The treatment success correlated with a significantly increased T cell proportion and PD-1 expression in the spleen. In all subtype-specific HTM combination therapy proved most effective in diminishing tumor growth, enhancing the immune response, and converted non-responder into responder during anti-PD-L1 therapy. In HTM, neoadjuvant irradiation reinforced anti-PD-L1 checkpoint treatment of breast cancer in a subtype -specific manner. According to the "bench to bedside" principle, this study offers a vital foundation for clinical translating the use of neoadjuvant irradiation in the context of checkpoint therapy.

Identification of shared gene signatures in major depressive disorder and triple-negative breast cancer.

BACKGROUND: Patients with major depressive disorder (MDD) have an increased risk of breast cancer (BC), implying that these two diseases share similar pathological mechanisms. This study aimed to identify the key pathogenic genes that lead to the occurrence of both triple-negative breast cancer (TNBC) and MDD. METHODS: Public datasets GSE65194 and GSE98793 were analyzed to identify differentially expressed genes (DEGs) shared by both datasets. A protein-protein interaction (PPI) network was constructed using STRING and Cytoscape to identify key PPI genes using cytoHubba. Hub DEGs were obtained from the intersection of hub genes from a PPI network with genes in the disease associated modules of the Weighed Gene Co-expression Network Analysis (WGCNA). Independent datasets (TCGA and GSE76826) and RT-qPCR validated hub gene expression. RESULTS: A total of 113 overlapping DEGs were identified between TNBC and MDD. The PPI network was constructed, and 35 hub DEGs were identified. Through WGCNA, the blue, brown, and turquoise modules were recognized as highly correlated with TNBC, while the brown, turquoise, and yellow modules were similarly correlated with MDD. Notably, G3BP1, MAF, NCEH1, and TMEM45A emerged as hub DEGs as they appeared both in modules and PPI hub DEGs. Within the GSE65194 and GSE98793 datasets, G3BP1 and MAF exhibited a significant downregulation in TNBC and MDD groups compared to the control, whereas NCEH1 and TMEM45A demonstrated a significant upregulation. These findings were further substantiated by TCGA and GSE76826, as well as through RT-qPCR validation. CONCLUSIONS: This study identified G3BP1, MAF, NCEH1 and TMEM45A as key pathological genes in both TNBC and MDD.

CircTRIM1 encodes TRIM1-269aa to promote chemoresistance and metastasis of TNBC via enhancing CaM-dependent MARCKS translocation and PI3K/AKT/mTOR activation.

Peptides and proteins encoded by noncanonical open reading frames (ORFs) of circRNAs have recently been recognized to play important roles in disease progression, but the biological functions and mechanisms of these peptides and proteins are largely unknown. Here, we identified a potential coding circular RNA, circTRIM1, that was upregulated in doxorubicin-resistant TNBC cells by intersecting transcriptome and translatome RNA-seq data, and its expression was correlated with clinicopathological characteristics and poor prognosis in patients with TNBC. CircTRIM1 possesses a functional IRES element along with an 810 nt ORF that can be translated into a novel endogenously expressed protein termed TRIM1-269aa. Functionally, we demonstrated that TRIM1-269aa, which is involved in the biological functions of circTRIM1, promoted chemoresistance and metastasis in TNBC cells both in vitro and in vivo. In addition, we found that TRIM1-269aa can be packaged into exosomes and transmitted between TNBC cells. Mechanistically, TRIM1-269aa enhanced the interaction between MARCKS and calmodulin, thus promoting the calmodulin-dependent translocation of MARCKS, which further initiated the activation of the PI3K/AKT/mTOR pathway. Overall, circTRIM1, which encodes TRIM1-269aa, promoted TNBC chemoresistance and metastasis by enhancing MARCKS translocation and PI3K/AKT/mTOR activation. Our investigation has yielded novel insights into the roles of protein-coding circRNAs and supported circTRIM1/TRIM1-269aa as a novel promising prognostic and therapeutic target for patients with TNBC.

Detection of circulating tumor cells by means of machine learning using Smart-Seq2 sequencing.

Circulating tumor cells (CTCs) are tumor cells that separate from the solid tumor and enter the bloodstream, which can cause metastasis. Detection and enumeration of CTCs show promising potential as a predictor for prognosis in cancer patients. Furthermore, single-cells sequencing is a technique that provides genetic information from individual cells and allows to classify them precisely and reliably. Sequencing data typically comprises thousands of gene expression reads per cell, which artificial intelligence algorithms can accurately analyze. This work presents machine-learning-based classifiers that differentiate CTCs from peripheral blood mononuclear cells (PBMCs) based on single cell RNA sequencing data. We developed four tree-based models and we trained and tested them on a dataset consisting of Smart-Seq2 sequenced data from primary tumor sections of breast cancer patients and PBMCs and on a public dataset with manually annotated CTC expression profiles from 34 metastatic breast patients, including triple-negative breast cancer. Our best models achieved about 95% balanced accuracy on the CTC test set on per cell basis, correctly detecting 133 out of 138 CTCs and CTC-PBMC clusters. Considering the non-invasive character of the liquid biopsy examination and our accurate results, we can conclude that our work has potential application value.

Posttranslational protein modifications as gatekeepers of cancer immunogenicity.

Triple-negative breast cancer (TNBC) presents a formidable challenge in oncology due to its aggressive phenotype and the immunosuppressive nature of its tumor microenvironment (TME). In this issue of the JCI, Zhu, Banerjee, and colleagues investigated the potential of targeting the OTU domain-containing protein 4 (OTUD4)/CD73 axis to mitigate immunosuppression in TNBC. They identified elevated CD73 expression as a hallmark of immunosuppression in TNBC. Notably, the CD73 expression was regulated by OTUD4-mediated posttranslational modifications. Using ST80, a pharmacologic inhibitor of OTUD4, the authors demonstrated the restoration of cytotoxic T cell function and enhanced efficacy of anti-PD-L1 therapy in preclinical models. These findings underscore the therapeutic potential of targeting the OTUD4/CD73 axis in TNBC.

Fusogenic vesicular stomatitis virus combined with natural killer T cell immunotherapy controls metastatic breast cancer.

BACKGROUND: Metastatic breast cancer is a leading cause of cancer death in woman. Current treatment options are often associated with adverse side effects and poor outcomes, demonstrating the need for effective new treatments. Immunotherapies can provide durable outcomes in many cancers; however, limited success has been achieved in metastatic triple negative breast cancer. We tested whether combining different immunotherapies can target metastatic triple negative breast cancer in pre-clinical models. METHODS: Using primary and metastatic 4T1 triple negative mammary carcinoma models, we examined the therapeutic effects of oncolytic vesicular stomatitis virus (VSVΔM51) engineered to express reovirus-derived fusion associated small transmembrane proteins p14 (VSV-p14) or p15 (VSV-p15). These viruses were delivered alone or in combination with natural killer T (NKT) cell activation therapy mediated by adoptive transfer of α-galactosylceramide-loaded dendritic cells. RESULTS: Treatment of primary 4T1 tumors with VSV-p14 or VSV-p15 alone increased immunogenic tumor cell death, attenuated tumor growth, and enhanced immune cell infiltration and activation compared to control oncolytic virus (VSV-GFP) treatments and untreated mice. When combined with NKT cell activation therapy, oncolytic VSV-p14 and VSV-p15 reduced metastatic lung burden to undetectable levels in all mice and generated immune memory as evidenced by enhanced in vitro recall responses (tumor killing and cytokine production) and impaired tumor growth upon rechallenge. CONCLUSION: Combining NKT cell immunotherapy with enhanced oncolytic virotherapy increased anti-tumor immune targeting of lung metastasis and presents a promising treatment strategy for metastatic breast cancer.

Lactylproteome analysis indicates histone H4K12 lactylation as a novel biomarker in triple-negative breast cancer.

Objective: The established link between posttranslational modifications of histone and non-histone lysine (K) residues in cell metabolism, and their role in cancer progression, is well-documented. However, the lactylation expression signature in triple-negative breast cancer (TNBC) remains underexplored. Methods: We conducted a comprehensive lactylproteome profiling of eight pairs of TNBC samples and their matched adjacent tissues. This was achieved through 4-Dimensional label-free quantitative proteomics combined with lactylation analysis (4D-LFQP-LA). The expression of identified lactylated proteins in TNBC was detected using immunoblotting and immunohistochemistry (IHC) with specific primary antibodies, and their clinicopathological and prognostic significance was evaluated. Results: Our analysis identified 58 lactylation sites on 48 proteins, delineating the protein lactylation alteration signature in TNBC. Bioinformatic and functional analyses indicated that these lactylated proteins play crucial roles in regulating key biological processes in TNBC. Notably, lactylation of lysine at position 12 (H4K12lac) in the histone H4 domain was found to be upregulated in TNBC. Further investigations showed a high prevalence of H4K12lac upregulation in TNBC, with positive rates of 93.19% (137/147) and 92.93% (92/99) in TNBC tissue chip and validation cohorts, respectively. H4K12lac expression correlated positively with Ki-67 and inversely with overall survival (OS) in TNBC (HR [hazard ratio] =2.813, 95%CI [credibility interval]: 1.242-6.371, P=0.0164), suggesting its potential as an independent prognostic marker (HR=3.477, 95%CI: 1.324-9.130, P=0.011). Conclusions: Lactylation is a significant post-translational modification in TNBC proteins. H4K12lac emerges as a promising biomarker for TNBC, offering insights into the lactylation profiles of TNBC proteins and linking histone modifications to clinical implications in TNBC.

Identification of restrictive molecules involved in oncolytic virotherapy using genome-wide CRISPR screening.

Oncolytic viruses (OVs) offer a novel approach to treat solid tumors; however, their efficacy is frequently suboptimal due to various limiting factors. To address this challenge, we engineered an OV containing targets for neuron-specific microRNA-124 and Granulocyte-macrophage colony-stimulating factor (GM-CSF), significantly enhancing its neuronal safety while minimally compromising its replication capacity. Moreover, we identified PARP1 as an HSV-1 replication restriction factor using genome-wide CRISPR screening. In models of glioblastoma (GBM) and triple-negative breast cancer (TNBC), we showed that the combination of OV and a PARP inhibitor (PARPi) exhibited superior efficacy compared to either monotherapy. Additionally, single-cell RNA sequencing (scRNA-seq) revealed that this combination therapy sensitized TNBC to immune checkpoint blockade, and the incorporation of an immune checkpoint inhibitor (ICI) further increased the survival rate of tumor-bearing mice. The combination of PARPi and ICI synergistically enhanced the ability of OV to establish durable tumor-specific immune responses. Our study effectively overcomes the inherent limitations of OV therapy, providing valuable insights for the clinical treatment of TNBC, GBM, and other malignancies.

Exogenous Metabolic Modulators Improve Response to Carboplatin in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) lacks targeted therapies, leaving cytotoxic chemotherapy as the current standard treatment. However, chemotherapy resistance remains a major clinical challenge. Increased insulin-like growth factor 1 signaling can potently blunt chemotherapy response, and lysosomal processes including the nutrient scavenging pathway autophagy can enable cancer cells to evade chemotherapy-mediated cell death. Thus, we tested whether inhibition of insulin receptor/insulin-like growth factor 1 receptor with the drug BMS-754807 and/or lysosomal disruption with hydroxychloroquine (HCQ) could sensitize TNBC cells to the chemotherapy drug carboplatin. Using in vitro studies in multiple TNBC cell lines, in concert with in vivo studies employing a murine syngeneic orthotopic transplant model of TNBC, we show that BMS-754807 and HCQ each sensitized TNBC cells and tumors to carboplatin and reveal that exogenous metabolic modulators may work synergistically with carboplatin as indicated by Bliss analysis. Additionally, we demonstrate the lack of overt in vivo toxicity with our combination regimens and, therefore, propose that metabolic targeting of TNBC may be a safe and effective strategy to increase sensitivity to chemotherapy. Thus, we conclude that the use of exogenous metabolic modulators, such as BMS-754807 or HCQ, in combination with chemotherapy warrants additional study as a strategy to improve therapeutic responses in women with TNBC.