Contributions of lignification, tissue arrangement patterns, and cross-sectional area to whole-stem mechanical properties in Arabidopsis thaliana.

Plant cells withstand mechanical stress originating from turgor pressure by robustly maintaining the mechanical properties of the cell wall. This applies at the organ scale as well; many plant stems act as pressurized cylinders, where the epidermis is under tension and inner tissues are under compression. The clavata3 de-etiolated3 (clv3-8 det3-1) double mutant of Arabidopsis thaliana displays cracks in its stems because of a conflict between the mechanical properties of the weak epidermis and over-proliferation of inner stem tissues. In this work, we conducted three-point bending tests on various Arabidopsis thaliana mutants, including those displaying the stem cracking phenotype, to examine the differences in their mechanical properties. The clv3-8 det3-1 double mutant exhibited reduced stem stiffness, consistent with reduced differentiation due to the clv3-8 mutation. Yet, in clv3-8, stem cross-sectional area was increased associating with the increase in vascular bundle number, and stem cross-sections displayed various shapes. To uncouple the contribution of geometry and cell-wall differentiation to the mechanical properties of the whole stems, we tested the contribution of lignified fibers to stem stiffness. In order to suppress lignin deposition in stems genetically, we generated multiple higher-order mutants by crossing clv3-8 and/or det3-1 with nst1-1 nst3-1, in which lignin deposition is suppressed. Stem stiffness was reduced markedly in all nst1-1 nst3-1 mutant backgrounds. Overall, our results suggest that stem stiffness relies on the presence of differentiated, lignified, fiber tissue as well as on the alignment or spatial distribution of vascular bundles within the stem organ.

Intratumoral Tumor Infiltrating Lymphocytes (TILs) are Associated With Cell Proliferation and Better Survival But Not Always With Chemotherapy Response in Breast Cancer.

OBJECTIVE: To investigate the clinical relevance of intratumoral tumor infiltrating lymphocytes (TILs) in breast cancer as measured by computational deconvolution of bulk tumor transcriptomes. SUMMARY BACKGROUND DATA: Commonly assessed TILs, located in tumor stroma without direct contact with cancer cells (stromal TILs), correlate with breast cancer treatment response and survival. The clinical relevance of intratumoral TILs has been less studied partly due to their rarity; however, they may have nonnegligible effects given their direct contact with cancer cells. METHODS: In all, 5870 breast cancer patients from TCGA, METABRIC, GSE96058, GSE25066, GSE163882, GSE123845, and GSE20271 cohorts were analyzed and validated. RESULTS: The intratumoral TIL score was established by the sum of all types of lymphocytes using the xCell algorithm. This score was the highest in triple-negative breast cancer (TNBC) and the lowest in the ER-positive/HER2-negative subtype. It correlated with cytolytic activity and infiltrations of dendritic cells, macrophages, and monocytes, and uniformly enriched immune-related gene sets regardless of subtype. Intratumoral TIL-high tumors correlated with higher mutation rates and significant cell proliferation on biological, pathological, and molecular analyses only in the ER-positive/HER2-negative subtype. It was significantly associated with pathological complete response after anthracycline- and taxane-based neoadjuvant chemotherapy in about half of the cohorts, regardless of the subtype. Intratumoral TIL-high tumors correlated with better overall survival in HER2-positive and TNBC subtypes consistently in 3 cohorts. CONCLUSIONS: Intratumoral TILs estimated by transcriptome computation were associated with increased immune response and cell proliferation in ER-positive/HER2-negative and better survival in HER2-positive and TNBC subtypes, but not always with pathological complete response after neoadjuvant chemotherapy.

Contribution of vasculature to stem integrity in Arabidopsis thaliana.

In plants, coordinated growth is important for organ mechanical integrity because cells remain contiguous through their walls. So far, defects in inflorescence stem integrity in Arabidopsis thaliana have mainly been related to epidermal defects. Although these observations suggest a growth-limiting function at the stem cortex, deeper layers of the stem could also contribute to stem integrity. The nac secondary cell wall thickening promoting factor1 (nst1) nst3 double-mutant background is characterized by weaker vascular bundles without cracks. By screening for the cracking phenotype in this background, we identified a regulator of stem cracking, the transcription factor INDETERMINATE DOMAIN9 (IDD9). Stem cracking was not caused by vascular bundle breakage in plants that expressed a dominant repressor version of IDD9. Instead, cracking emerged from increased cell expansion in non-lignified interfascicular fiber cells that stretched the epidermis. This phenotype could be enhanced through CLAVATA3-dependent cell proliferation. Collectively, our results demonstrate that stem integrity relies on three additive mechanical components: the epidermis, which resists inner cell growth; cell proliferation in inner tissues; and growth heterogeneity associated with vascular bundle distribution in deep tissues.

Germinating seedlings and mature shoots of Cuscuta campestris respond differently to light stimuli during parasitism but not during circumnutation.

Seedlings of the parasitic plant genus Cuscuta (dodder) locate hosts by circumnutation, coil around the host near soil level and form a haustorium, establishing a primary parasitism beneath the canopy. Mature shoots elongating from the parasitic region parasitize other hosts on the upper surfaces of their canopy. Although parasitism by dodder is stimulated by blue and far-red light, and inhibited by red light, the responses to light signals during the developmental stages are not comprehensively understood. Therefore, we compared the effects of different types of light on both circumnutation and parasitism by germinating seedlings and mature shoots of Cuscuta campestris. Seedlings established parasitism under blue and far-red light, but not under red light, as has been reported repeatedly. By contrast, mature shoots exhibited coiling around the host and haustoria formation even under a red light as well as under blue and far-red light. These findings indicate that C. campestris modified its response to red light during the transition from young seedlings to mature shoots, facilitating parasitism. Light quality did not affect the circumnutation of either seedlings or mature shoots, indicating that circumnutation and the coiling movement that leads to parasitism were regulated by different environmental signals.

Delayed Diagnosis and Prognostic Impact of Breast Cancer During the COVID-19 Pandemic.

BACKGROUND: Breast screening services were suspended for several months owing to the coronavirus disease 2019 (COVID-19) pandemic. We estimated the potential impact on breast cancer mortality using long-term global observations. However, the magnitude of the impact may vary across countries; therefore, we conducted an analysis and modeling study of this impact in Japan. PATIENTS AND METHODS: We compared the clinicopathological features of breast cancers between the nonpandemicgroup (April 1, 2019 to October 31, 2019) and the pandemic group (April 1, 2020 to October 31, 2020). We also compared the estimated 10-year survival rates between the two groups based on the weighted average of the 10-year survival rate by clinical stage and site (2004-2007). RESULTS: Results...Pandemic-related disruption decreased the number of breast cancer cases from296 to 249 during both 7-month periods. The percentage of patients with stage IIB or higher disease was significantly higher in the pandemic group than in the non-pandemic group (22.0% vs. 31.3%, P = 0.0133). The percentage of cases with a Ki-67 labeling index higher than 20% tended to be higher in the pandemic group than in the non-pandemic group (62.2% vs. 54.4%). The estimated 10-year survival rate was lower in the pandemic group than in the non-pandemic group (83.9% vs. 87.9%, 95% confidence interval of the difference: 0.87-8.8, P > 0.05). CONCLUSION: We found more aggressive and advanced disease afterthe suspension of breast cancer screening services owing to the COVID-19 pandemic. This may have affected the long-term clinical outcomes of patients with breast cancer.

Elevated bile acid metabolism and microbiome are associated with suppressed cell proliferation and better survival in breast cancer.

Bile acids are metabolized by the gut microbiome and are involved in fat absorption. Contrary to their carcinogenic role in gastrointestinal cancers, bile acids have been reported to inhibit cancer cell proliferation in breast cancer. The microbiome of breast cancer tissues may also influence cancer proliferation. We hypothesized that bile acid metabolism reflects its accumulation and is associated with certain microbiomes, breast cancer biology, and patient survival. Transcriptomic and clinicopathological information of a total of 6050 patients in three large open primary breast cancer cohorts (GSE96058, METABRIC, TCGA) and 16S rRNA gene sequence microbiome data of breast cancer tissues in TCGA were analyzed by high and low bile acid metabolism scores calculated by gene set variation analysis (GSVA). Breast cancers with high bile acid metabolism had a significantly improved survival across all three cohorts. Metabolic pathways related to the production and regulation of bile acids were consistently enriched in high bile acid metabolism groups across all cohorts. On the other hand, the low bile acid metabolism group was associated with higher Ki67 expression and Nottingham histological grade, as well as enrichment of cell proliferation-related gene sets. Intratumoral heterogeneity, homologous recombination deficiency, mutational load, activation of cancer immunity, and infiltration of anticancer immune cells were also higher in this group. Gammaretrovirus, Hymenobacter, Anaerococcus, and Collimonas were significantly more abundant in the high bile acid metabolism group compared to Lactobacillus, Ruegeria, and Marichromatium in the low metabolism group. Surprisingly, almost all Hallmark cell proliferation-associated gene sets were highly enriched in all three microorganisms that were abundant in the low bile acid metabolism group. In conclusion, microorganisms abundant in the breast tumor microenvironment with low bile acid metabolism are associated with aggressive cancer biology, including increased cell proliferation and poor survival.

Development of a novel BRCAness score that predicts response to PARP inhibitors.

BACKGROUND: BRCAness is a characteristic feature of homologous recombination deficiency (HRD) mimicking BRCA gene mutation in breast cancer. We hypothesized that a measure to quantify BRCAness that causes synthetic lethality in BRCA mutated tumors will identify responders to PARP inhibitors. METHODS: A total of 6753 breast cancer patients from 3 large independent cohorts were analyzed. A score was generated by transcriptomic profiling using gene set variation analysis algorithm on 34 BRCA1-mutation related genes selected by high AUC levels in ROC curve between BRCA1 mutation and wildtype breast cancer. RESULTS: The score was significantly associated with BRCA1 mutation, high mutation load and intratumoral heterogeneity as expected, as well as with high HRD, DNA repair and MKi67 expression regardless of BRCA mutations. High BRCAness tumors enriched not only DNA repair, but also all five Hallmark cell proliferation-related gene sets. High BRCAness tumors were significantly associated with higher cytolytic activity and with higher anti-cancerous immune cell infiltration. Not only did the breast cancer cell lines with BRCA-mutation show high score, but even the other cells in human breast cancer tumor microenvironment were contributing to the score. The BRCAness score was the highest in triple-negative breast cancer consistently in all 3 cohorts. BRCAness was associated with response to chemotherapy and correlated strongly with response to PARP inhibitor in both triple-negative and ER-positive/HER2-negative breast cancer. CONCLUSIONS: We established a novel BRCAness score using BRCA-mutation-related gene expressions and found that it associates with DNA repair and predicts response to PARP inhibitors regardless of BRCA mutation.

ß-Tubulin Isoforms Related to Docetaxel Sensitivity in 2D and 3D Cultured TNBC Cell Lines.

BACKGROUND/AIM: We previously reported that the half maximal inhibitory concentration (IC50) values of cisplatin and epirubicin correlated in 13 triple-negative breast cancer (TNBC) cell lines between two-dimensional (2D) and three-dimensional (3D) culture methods. However, the IC50 values of docetaxel (DTX) did not correlate between the two culture methods. We hypothesized that this non-correlation is partly associated with differences in expression of the ß-tubulin isoform, the target molecule of DTX and in morphology depending on the culture method. MATERIALS AND METHODS: We investigated the expression levels of ß-tubulin isoforms by real-time polymerase chain reaction and morphology of spheroid formation in the 13 TNBC cell lines cultured using the 2D and 3D culture methods. RESULTS: Tubulin ß class I (TUBB) expression levels were negatively correlated with the IC50 value of DTX in the 2D culture method (R=-0.360), whereas tubulin ß class IIa (TUBB2a) expression levels were positively correlated in the 3D culture method (R=0.398). There was no significant difference in the expression levels of ß-tubulin isoforms between the 2D and 3D culture methods. The spheroids were classified morphologically into three types: round, mass, and grape-like. However, no clear association was found between DTX sensitivity and morphology. CONCLUSION: The non-correlation of the IC50 values of DTX between the 2D and 3D culture methods does not appear to be due to the changes in ß-tubulin isoforms. Morphology in the 3D culture method may play some role in drug sensitivity.

Tissue-targeted inorganic pyrophosphate hydrolysis in a fugu5 mutant reveals that excess inorganic pyrophosphate triggers developmental defects in a cell-autonomous manner.

Excess PPi triggers developmental defects in a cell-autonomous manner. The level of inorganic pyrophosphate (PPi) must be tightly regulated in all kingdoms for the proper execution of cellular functions. In plants, the vacuolar proton pyrophosphatase (H+-PPase) has a pivotal role in PPi homeostasis. We previously demonstrated that the excess cytosolic PPi in the H+-PPase loss-of-function fugu5 mutant inhibits gluconeogenesis from seed storage lipids, arrests cell division in cotyledonary palisade tissue, and triggers a compensated cell enlargement (CCE). Moreover, PPi alters pavement cell (PC) shape, stomatal patterning, and functioning, supporting specific yet broad inhibitory effects of PPi on leaf morphogenesis. Whereas these developmental defects were totally rescued by the expression of the yeast soluble pyrophosphatase IPP1, sucrose supply alone canceled CCE in the palisade tissue but not the epidermal developmental defects. Hence, we postulated that the latter are likely triggered by excess PPi rather than a sucrose deficit. To formally test this hypothesis, we adopted a spatiotemporal approach by constructing and analyzing fugu5-1 PDF1 pro ::IPP1, fugu5-1 CLV1 pro ::IPP1, and fugu5-1 ICL pro ::IPP1, whereby PPi was removed specifically from the epidermis, palisade tissue cells, or during the 4 days following seed imbibition, respectively. It is important to note that whereas PC defects in fugu5-1 PDF1 pro ::IPP1 were completely recovered, those in fugu5-1 CLV1 pro ::IPP1 were not. In addition, phenotypic analyses of fugu5-1 ICL pro ::IPP1 lines demonstrated that the immediate removal of PPi after seed imbibition markedly improved overall plant growth, abolished CCE, but only partially restored the epidermal developmental defects. Next, the impact of spatial and temporal removal of PPi was investigated by capillary electrophoresis time-of-flight mass spectrometry (CE-TOF MS). Our analysis revealed that the metabolic profiles are differentially affected among all the above transgenic lines, and consistent with an axial role of central metabolism of gluconeogenesis in CCE. Taken together, this study provides a conceptual framework to unveil metabolic fluctuations within leaf tissues with high spatio-temporal resolution. Finally, our findings suggest that excess PPi exerts its inhibitory effect in planta in the early stages of seedling establishment in a tissue- and cell-autonomous manner.

Left sided breast cancer is associated with aggressive biology and worse outcomes than right sided breast cancer.

Breast cancer is more common on the left side than the right side. We aim to evaluate differences in clinicopathological and genomic characteristics based on laterality. We analyzed survival outcomes and clinical characteristics of 881,320 patients recorded by the Surveillance, Epidemiology, and End Results (SEER) program. The Cancer Genome Atlas (TCGA) was used to explore genomic and clinical features from 1,062 patients. Gene expression data was used to quantitate cytolytic activity and hallmark gene-sets were used for gene set enrichment analysis. An institutional retrospective review was conducted on 155 patients treated with neoadjuvant chemotherapy (NACT). Patient characteristics were summarized by pathological complete response (pCR). Left sided tumors were found to be more prevalent than right sided tumors. No major clinicopathological differences were noted by laterality. Left sided breast cancer demonstrated poorer outcomes versus right sided tumors (HR 1.05, 95% CI 1.01-1.08; p = 0.011). Cell proliferation gene sets, including E2F Targets, G2M Checkpoint, Mitotic spindle, and MYC Targets, were enriched on the left side compared to the right. Left sided tumors had lower pCR rates versus right sided tumors (15.4% versus 29.9%, p = 0.036). Our findings suggest that left sided breast cancer is associated with aggressive biology and worse outcomes compared to right sided breast cancer.

Intratumoral PDGFB gene predominantly expressed in endothelial cells is associated with angiogenesis and lymphangiogenesis, but not with metastasis in breast cancer.

PURPOSE: Platelet-derived growth factor B (PDGFB) is known to play essential roles in angiogenesis and lymphangiogenesis during development, and tumor growth and vessel stabilization in experimental models. However, whether these findings could be translated to breast cancer patients remains unclear. We hypothesized that PDGFB gene expression is associated with angiogenesis, cell proliferation, and clinical outcomes in breast cancer patients. METHODS: A total of 7635 primary breast cancer patients with full transcriptome and clinical data available from 13 independent cohorts were analyzed using in silico approach. The median value was used to divide each cohort into high and low PDGFB expression groups. RESULTS: High PDGFB gene expression was associated with increased expression of angiogenesis-related genes, higher amount of vascular cell infiltrations, and with enrichment of angiogenesis gene set, lymphangiogenesis-related gene expressions, lymphangiogenesis-related cell infiltrations, and enrichmentof lymphangiogenesis gene set in GSE96058 and validated by TCGA cohorts; however, not with lymphatic metastasis. PDGFB expression was neither associated with cell proliferation as assessed by Ki67 expression nor with Nottingham histological grade, or response to neoadjuvant chemotherapy. We found that PDGFB was most extensively expressed by endothelial and perivascular-like cells in the tumor microenvironment, and minimally by cancer cells consistently in two single-cell sequence cohorts. High PDGFB expression enriched TGFß, epithelial-mesenchymal transition, hypoxia, and cancer stem cell-associated pathways. However, no association with distant metastasis was observed. Disease-specific and disease-free survival were worse in the high PDGFB expression group consistently in TCGA and METABRIC cohorts. CONCLUSION: PDGFB is predominantly expressed in endothelial cells and is associated with angiogenesis and lymphangiogenesis, but not with cellular proliferation or metastasis in breast cancer.

NR2F1, a Tumor Dormancy Marker, Is Expressed Predominantly in Cancer-Associated Fibroblasts and Is Associated with Suppressed Breast Cancer Cell Proliferation.

BACKGROUND: Tumor dormancy is a crucial mechanism responsible for the late recurrence of breast cancer. Thus, we investigated the clinical relevance of the expression of NR2F1, a known dormancy biomarker. METHODS: A total of 6758 transcriptomes of bulk tumors from multiple breast cancer patient cohorts and two single-cell sequence cohorts were analyzed. RESULTS: Breast cancer (BC) with high NR2F1 expression enriched TGFß signaling, multiple metastases, and stem cell-related pathways. Cell proliferation-related gene sets were suppressed, and MKi67 expression was lower in high NR2F1 BC. In tumors with high Nottingham grade, NR2F1 expression was found to be lower. There was no consistent relationship between NR2F1 expression and metastasis or survival. Cancer mutation rates, immune responses, and immune cell infiltrations were lower in high NR2F1 tumors, whereas the infiltration of stromal cells including cancer-associated fibroblasts (CAFs) was higher. NR2F1 was predominantly expressed in CAFs, particularly inflammatory CAFs, rather than in cancer cells, consistently in the two single-cell sequence cohorts. CONCLUSIONS: NR2F1 expression in breast cancer is associated with tumor dormancy traits, and it is predominantly expressed in CAFs in the tumor microenvironment.

Intratumoral lymphatic endothelial cell infiltration reflecting lymphangiogenesis is counterbalanced by immune responses and better cancer biology in the breast cancer tumor microenvironment.

Lymphangiogenesis, the generation of new lymphatic vessels from existing ones, results from the dynamic interactions of lymphatic endothelial cells and the tumor microenvironment (TME). It is well known that lymphangiogenesis occurs during the initial stage of metastasis in various types of malignant tumors. However, it is currently not used as a biomarker partially because gold standard method to quantify it is labor and cost intensive. We hypothesized that the quantity of intratumoral lymphatic endothelial cells (iLECs) in the TME is an indicator of lymphangiogenesis and a predictor of metastatic potential and overall survival in breast cancer. We analyzed a total of 4145 breast cancer patients from the Cancer Genome Atlas (TCGA) and GSE96058 by quantifying their iLECs using the xCell algorithm, and correlated these scores with patient survival, tumor grade, and cancer stage. We also assessed various pro- and anti-cancer gene sets for each tumor to characterize tumor behavior and aggressiveness. As we expected, high-iLEC breast cancer demonstrated enriched lymphoangiogenesis and angiogenesis gene sets and was associated with increased expressions of related genes. Also enriched were inflammatory response and immune response-related gene sets; IL2/STAT5 pathway, IL6/JAK/STAT3 pathway, TNFα pathway, allograft rejection, and complement as well as cancer stemness related gene sets like Notch signaling, Hedgehog signaling, epithelial mesenchymal transition, and Wnt beta-catenin signaling. Tumors with high-iLEC showed higher proportions of stromal cells and fewer anti-cancer immune cells. On the other hand, iLEC score did not correlate with patient survival or lymph node metastasis. Surprisingly, breast cancers with fewer iLECs demonstrated enriched E2F Targets, G2M Checkpoint, MYC Targets v1, and MTORC1 signaling which are cancer cell proliferation-related gene sets and exhibited an abundance of pro-cancer immune cells. The amount of iLEC correlated inversely with Ki67 expression and histological grade, which is in agreement that low-iLEC breast cancer was associated with enhanced cancer cell proliferation. In conclusion, while iLECs can be used as a surrogate for lymphangiogenesis in breast cancer, low-iLEC tumors also exhibit features which correspond to aggressive tumor biology, which may explain why the amount of iLECs was not associated with patient survival in our cohorts.

APOBEC3F expression in triple-negative breast cancer is associated with tumor microenvironment infiltration and activation of cancer immunity and improved survival.

The apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) causes a point mutation from cytidine to uracil in DNA and/or RNA. The role of APOBEC3A and APOBEC3B in breast cancer has been well described, whereas that of APOBEC3F remains unknown. To investigate the clinical relevance of APOBEC3F expression, we analyzed a total of 3000 breast cancer cases from multiple independent large patient cohorts including METABRIC, TCGA, GSE75688, and GSE114725. High expression of APOBEC3F was associated with improved disease-specific and overall survival in triple negative breast cancer (TNBC). APOBEC3F is not usually a reflection of cancer cell biology in TNBC or luminal breast cancer, except for homologous recombination deficiency in TNBC. In the TNBC homologous recombination deficiency group, APOBEC3F expression was not consistently associated with intratumor heterogeneity, mutation rates, or neoantigens. APOBEC3F expression did not correlate with response to any of the drugs tested in breast cancer cell lines in vitro. However, high APOBEC3F expression was associated with enrichment of several immune-related gene sets and immune activity. High APOBEC3F expression also accompanied higher infiltration of anti-cancer immune cell infiltration in TNBC. However, in luminal breast cancer, high APOBEC3F tumor significantly enriched not only immune-related gene sets, but also cell proliferation-, metastasis-, and apoptosis-related gene sets. Analysis of single-cell transcriptomes showed APOBEC3F exclusively expressed in immune cells and significantly associated with cytolytic activity of the immune cells, immune response, and immune cell proliferation. Expression of immune checkpoint genes was uniformly elevated in APOBEC3F-high tumors. We conclude that APOBEC3F is exclusively expressed in immune cells and this expression is associated with enhanced anti-cancer immune response as well as improved survival in TNBC.

High RAD51 gene expression is associated with aggressive biology and with poor survival in breast cancer.

PURPOSE: Although the DNA repair mechanism is important in preventing carcinogenesis, its activation in established cancer cells may support their proliferation and aggravate cancer progression. RAD51 cooperates with BRCA2 and is essential in the homologous recombination of DNA repair. To this end, we hypothesized that RAD51 gene expression is associated with cancer cell proliferation and poor prognosis of breast cancer (BC) patients. METHODS: A total of 8515 primary BC patients with transcriptome and clinical data from 17 independent cohorts were analyzed. The median value was used to divide each cohort into high and low RAD51 expression groups. RESULTS: High RAD51 expression enriched the DNA repair gene set and was correlated with DNA repair-related genes. Nottingham histological grade, Ki67 expression and cell proliferation-related gene sets (E2F Targets, G2M Checkpoint and Myc Targets) were all significantly associated with the high RAD51 BC group. RAD51 expression was positively correlated with Homologous Recombination Deficiency, as well as both mutational burden and neoantigens that accompanied a higher infiltration of immune cells. Primary BC with lymph node metastases was associated with high expression of RAD51 in two cohorts. There was no strong correlation between RAD51 expression and drug sensitivity in cell lines, and RAD51 expression was lower after the neoadjuvant chemotherapy compared to before the treatment. High RAD51 BC was associated with poor prognosis consistently in three independent cohorts. CONCLUSION: RAD51 gene expression is associated with aggressive cancer biology, cancer cell proliferation, and poor survival in breast cancer.

Beneficial effects of transdermal administration of tamoxifen on capsular contracture after breast implantation in murine models.

BACKGROUND AND AIM: Capsular contracture is the most common complication with smooth-type silicone implants. We investigated the preventive effect of an active metabolite of tamoxifen, 4-hydroxytamoxifen (4-OH TAM), on capsular contracture. METHODS: A silicone sheet was implanted into the back of 28 female ICR mice. Mixtures of gel with 0.2% 4-OH TAM and 0.1% 4-OH TAM were administered transdermally once a day for 4 weeks. Saline was administered to the control. After killing the mice, capsular thickness was measured in H&E-stained specimens. Estrogen receptor (ER), α-smooth muscle actin (α-SMA), and transforming growth factor-ß (TGF-ß) expressions were immunohistochemically investigated in the capsules. RESULTS: The capsule was thinner in the 0.2% 4-OH TAM gel group than in the control group (control, 0.1% 4-OH TAM gel, 0.2% 4-OH TAM gel: 52.8 ± 3.4 µm, 54.2 ± 6.8 µm, 46.4 ± 3.3 µm, respectively). ER was found in most fibroblasts of all samples. α-SMA expression in the capsule was significantly lower in the 4-OH TAM gel groups than in the control group (control = 70.0 ± 3.4%, 0.1% 4-OH TAM = 57.0 ± 3.4%, 0.2% 4-OH TAM = 49.4 ± 4.9%). TGF-ß expression was significantly reduced by the 4-OH TAM gel injections dose-dependently (control = 67.3 ± 2.2%, 0.1% 4-OH TAM = 52.4 ± 3.1%, 0.2% 4-OH TAM = 45.1 ± 2.4%). CONCLUSIONS: The transdermal administration of 0.1% and 0.2% 4-OH TAM gels inhibited capsule development. The inhibition of TGF-ß expression is a mechanism by which 4-OH TAM suppresses fibroblast growth, preventing capsular formation.

Different members of the APOBEC3 family of DNA mutators have opposing associations with the landscape of breast cancer.

APOBEC enzymes are strong mutagenic factors. In breast cancer, expression of APOBEC3B is increased and associated with mutation load and poor outcome. Other APOBEC3s can also mutate DNA but their clinical significance in breast cancer and its underpinnings have not been comprehensively studied. In our examination of 1,091 breast carcinoma cases, high expression of APOBEC3A or APOBEC3B genes was associated with greater tumor burden of mutations and other genomic aberrations. Expression of none of the five APOBEC3C-H genes had any correlation with these features, including T[C-T/G]W mutations, but their high expression levels indicated a robust anti-cancer immune response within tumors, with elevated CD8+ T cell abundance, T cell receptor diversity, and immune cytolytic activity. Concordantly, survival analyses of this and two other cohorts with > 3,000 patients each showed favorable prognostic benefit of high APOBEC3C-H expression for both cancer progression and mortality. A detrimental prognostic value was observed for APOBEC3A and APOBEC3B. Single-cell data revealed cancer epithelial and stromal immune cells as major sources of APOBEC3B and APOBEC3C-H expression in tumors, respectively. These observations on opposing associations with breast cancer of different APOBEC3s highlight the contrasting roles of these enzymes, promoting cancer through mutagenesis while antagonizing it through immune response.

Increased intratumor heterogeneity, angiogenesis and epithelial to mesenchymal transition pathways in metaplastic breast cancer.

Metaplastic breast cancer (MBC) constitutes a rare but unique histologic entity with poor prognosis. We hypothesized that MBC possesses unique genetic profile and tumor immune microenvironment. MBC cases were identified from a total of 10827 breast cancer entries in the Cancer Genome Atlas Data Set (TCGA) and the AACR-GENIE (Genomics Evidence Neoplasia Information Exchange) cohorts. Tumor infiltrated immune cells were estimated by xCell. Baseline clinical characteristics were compared, and gene set enrichment analysis (GSEA) was performed. MBC comprised 0.66% of the cohorts (1.2% of TCGA and 0.6% of GENIE). MBC cases were predominantly triple-negative (TNBC) (8 (61.5%) vs 151 (14.4%), P<0.001), and high Nottingham histological grade (8 (61.5%) vs 222 (21.1%), P=0.02) compared to non-MBC in the TCGA cohort. Increased infiltration of M1 macrophages (P=0.012), dendritic cells (P<0.001) and eosinophils (P=0.036) was noted in the MBC cohort however there was no difference in cytolytic activity (P=0.806), CD4 memory (P=0.297) or CD8 T-cells (P=0.864). Tumor mutation burden was lower in the MBC compared to the non-MBC, median: 0.4 vs 1.6/Mb in the TCGA-TNBC cohort (P=0.67) and 3.0 vs 4.0/Mb (P=0.1) in the GENIE-cohort. MBC had increased intratumor heterogeneity (P<0.001), macrophage regulation (P=0.008) and TGF-beta response (P<0.001). Disease-specific survival was decreased in MBC (P=0.018). Angiogenesis and epithelial-to-mesenchymal transition pathways were enriched in triple-negative MBC by GSEA (P=0.004 and P<0.001, respectively). Our results suggest that high intratumor heterogeneity, enriched angiogenesis and EMT pathway expression represent possible mechanisms leading to worse disease-specific survival found in metaplastic breast cancer.

An auxin signaling network translates low-sugar-state input into compensated cell enlargement in the fugu5 cotyledon.

In plants, the effective mobilization of seed nutrient reserves is crucial during germination and for seedling establishment. The Arabidopsis H+-PPase-loss-of-function fugu5 mutants exhibit a reduced number of cells in the cotyledons. This leads to enhanced post-mitotic cell expansion, also known as compensated cell enlargement (CCE). While decreased cell numbers have been ascribed to reduced gluconeogenesis from triacylglycerol, the molecular mechanisms underlying CCE remain ill-known. Given the role of indole 3-butyric acid (IBA) in cotyledon development, and because CCE in fugu5 is specifically and completely cancelled by ech2, which shows defective IBA-to-indoleacetic acid (IAA) conversion, IBA has emerged as a potential regulator of CCE. Here, to further illuminate the regulatory role of IBA in CCE, we used a series of high-order mutants that harbored a specific defect in IBA-to-IAA conversion, IBA efflux, IAA signaling, or vacuolar type H+-ATPase (V-ATPase) activity and analyzed the genetic interaction with fugu5-1. We found that while CCE in fugu5 was promoted by IBA, defects in IBA-to-IAA conversion, IAA response, or the V-ATPase activity alone cancelled CCE. Consistently, endogenous IAA in fugu5 reached a level 2.2-fold higher than the WT in 1-week-old seedlings. Finally, the above findings were validated in icl-2, mls-2, pck1-2 and ibr10 mutants, in which CCE was triggered by low sugar contents. This provides a scenario in which following seed germination, the low-sugar-state triggers IAA synthesis, leading to CCE through the activation of the V-ATPase. These findings illustrate how fine-tuning cell and organ size regulation depend on interplays between metabolism and IAA levels in plants.

A case of hereditary metachronous bilateral triple-negative breast cancer that was highly sensitive to carboplatin.

A 52-year-old woman with a strong family history of breast cancer was diagnosed as having triple-negative breast cancer (TNBC) in her right breast. Neoadjuvant chemotherapy (NAC; four cycles of epirubicin/cyclophosphamide/5-fluorouracil) was performed, followed by breast-conserving surgery and axillary lymph node dissection. Histopathological analysis of the surgical specimens demonstrated a few focal tumor cells remaining in the stroma, but not a pathological complete response (pCR). Weekly paclitaxel was subsequently added to the treatment regimen. A total of 17 months after the adjuvant treatments, TNBC recurred in her left breast with massive lymph node metastasis. Because of the early recurrence after standard treatment, NAC was administered together with carboplatin and paclitaxel. Histopathological analysis of the partially resected breast and axillary lymph nodes demonstrated a pCR. No recurrent disease was found 2 years after the second TNBC treatment. This case underlines the importance of platinum-based chemotherapy and prophylactic mastectomy for patients with BRCA dysfunction.