Whole-exome sequencing reveals an association of rs112065068 in TGOLN2 gene with distant metastasis of non-small cell lung cancer.

Early prediction and prevention of recurring illness is critical for improving the survival rates of patients with non-small cell lung cancer (NSCLC). Previously, we demonstrated that the presence of premalignant epithelial changes in the small bronchi distant to the primary tumor is associated with NSCLC progression: isolated basal cell hyperplasia (iBCH) indicates a high risk of distant metastasis, BCH combined with squamous metaplasia (BCHSM) - a high risk of locoregional recurrence. Here, we aimed to identify germline single nucleotide variants (SNVs) and insertions and deletions (InDels) associated with distant metastasis and locoregional recurrence in cases with iBCH and BCHSM using whole-exome sequencing of 172 NSCLC patients. The rs112065068 of the TGOLN2 gene was identified only in iBCH patients and was associated with a high risk of distant metastasis (P < .001) and worse metastasis-free survival (HR = 4.19 (95 %CI 1.97-8.93); P < .001). This variant was validated in a group of 109 NSCLC patients using real-time PCR and Sanger sequencing analyses. To our knowledge, this study is the first to identify a germline variant associated with NSCLC distant metastasis.

Integrin-associated transcriptional characteristics of circulating tumor cells in breast cancer patients.

Background: Integrins enable cell communication with the basal membrane and extracellular matrix, activating signaling pathways and facilitating intracellular changes. Integrins in circulating tumor cells (CTCs) play a significant role in apoptosis evasion and anchor-independent survival. However, the link between CTCs expressing different integrin subunits, their transcriptional profile and, therefore, their functional activity with respect to metastatic potential remains unclear. Methods: Single-cell RNA sequencing of CD45-negative cell fraction of breast cancer patients was performed. All CTCs were divided into nine groups according to their integrin profile. Results: СTCs without the gene expression of integrins or with the expression of non-complementary α and ß subunits that cannot form heterodimers prevailed. Only about 15% of CTCs expressed integrin subunits which can form heterodimers. The transcriptional profile of CTCs appeared to be associated with the spectrum of expressed integrins. The lowest potential activity was observed in CTCs without integrin expression, while the highest frequency of expression of tumor progression-related genes, namely genes of stemness, epithelial-mesenchymal transition (EMT), invasion, proinflammatory chemokines and cytokines as well as laminin subunits, were observed in CTCs co-expressing ITGA6 and ITGB4. Validation on the protein level revealed that the median of integrin ß4+ CTCs was higher in patients with more aggressive molecular subtypes as well as in metastatic breast cancer patients. One can expect that CTCs with ITGA6 and ITGB4 expression will have pronounced metastatic potencies manifesting in expression of EMT and stemness-related genes, as well as potential ability to produce chemokine/proinflammatory cytokines and laminins.

Spatial Heterogeneity of Integrins and Their Ligands in Primary Breast Tumors.

BACKGROUND: The diversity of cell-cell interactions in different regions of a tumor reflects the functional heterogeneity of cancer, which poses challenges in early diagnosis, selection of treatment strategies, and prognosis of breast cancer. Cancer cells interact with each other to form different morphological structures in the tumor and stromal host cells via integrins. The objective of this study was to characterize the morphological and spatial heterogeneity of primary breast tumors in the context of expression profiles of integrins and their ligands. METHODS: We studied spatial transcriptomics using the 10X Visium approach and the Niche Interactions and Communication Heterogeneity in Extracellular Signaling (NICHES) algorithm to map ligand-receptor signaling pathways and visualize the heterogeneity of signaling archetypes in tumor clusters. RESULTS: Cluster analysis of the expression profiles of tumor spots from the samples indicated pronounced inter-tumoral heterogeneity. Integrin-ligand functional clusters were associated with intratumoral heterogeneity, which was manifested by the presence of several morphological loci as observed in histological tumor samples. Inter-tumoral heterogeneity was manifested by a different number of functional clusters, ranging from 2 to 9 for each tumor sample. The main characteristic of these clusters was the significant predominance of non-complementary integrin subunits. Of the 42 functional integrin-ligand pairs in 21 clusters of five samples, 41 pairs occurred only once. The exception was the laminin subunit alpha-5 (LAMA5)-integrin beta 4 (ITGB4) pair, which was detected in two clusters of different samples. CONCLUSIONS: The spatial heterogeneity of integrin-ligand expression clusters in breast cancer contributes significantly to the functional heterogeneity of the tumor, which sets the stage for many scenarios of parenchymatous-stromal relationships, some of which may be effective in the emergence of metastasizing tumor seed cells. The intra- and inter-tumoral spatio-functional heterogeneity of the tumor tissue that we discovered may largely explain why it is difficult to achieve success in most patients with breast cancer using any therapeutic strategy targeting one molecule of the vast array, regardless of the importance of its pathogenetic significance.

The Association of Integrins ß3, ß4, and αVß5 on Exosomes, CTCs and Tumor Cells with Localization of Distant Metastasis in Breast Cancer Patients.

Integrins are cell adhesion receptors, which play a role in breast cancer invasion, angiogenesis, and metastasis. Moreover, it has been shown that exosomal integrins provide organotropic metastasis in a mouse model. In our study, we aimed to investigate the expression of integrins ß3, ß4, and αVß5 on exosomes and tumor cells (circulating tumor cells and primary tumor) and their association with the localization of distant metastasis. We confirmed the association of exosomal integrin ß4 with lung metastasis in breast cancer patients. However, we were unable to evaluate the role of integrin ß3 in brain metastasis due to the rarity of this localization. We established no association of exosomal integrin αVß5 with liver metastasis in our cohort of breast cancer patients. The further evaluation of ß3, ß4, and αVß5 integrin expression on CTCs revealed an association of integrin ß4 and αVß5 with liver, but not the lung metastases. Integrin ß4 in the primary tumor was associated with liver metastasis. Furthermore, an in-depth analysis of phenotypic characteristics of ß4+ tumor cells revealed a significantly increased proportion of E-cadherin+ and CD44+CD24- cells in patients with liver metastases compared to patients with lung or no distant metastases.

Spatial Profile of Tumor Microenvironment in PD-L1-Negative and PD-L1-Positive Triple-Negative Breast Cancer.

The problem of finding more precise stratification criteria for identifying the cohort of patients who would obtain the maximum benefit from immunotherapy is acute in modern times. In our study were enrolled 18 triple-negative breast cancer patients. The Ventana SP142 test was used for PD-L1 detection. Spatial transcriptomic analysis by 10x Genomics was used to compare PD-L1-positive and PD-L1-negative tumors. The seven-color multiplex immunofluorescence (by Akoya) was used for the detection of the type of cells that carried the PD1 receptor and the PD-L1 ligand. Using pathway analysis, we showed that PD-L1-positive tumors demonstrate signatures of a cell response to cytokines, among others, and PD-L1-negative tumors demonstrate signatures of antigen presentation. PD-L1-positive and PD-L1-negative tumors have different tumor microenvironment (TME) compositions according to CIBERSORT analysis. Multiplex immunohistochemistry (IHC) confirmed the prevalence of PD1-negative M2 macrophages and PD1-negative T lymphocytes in PD-L1-positive tumors. PD-L1-positive tumors are not characterized by direct contact between cells carrying the PD1 receptor and the PD-L1 ligand. So, the absence of specific immune reactions against the tumor, predominance of pro-tumor microenvironment, and rare contact between PDL1 and PD1-positive cells may be the potential reasons for the lack of an immune checkpoint inhibitor (ICI) effect in triple-negative breast cancer patients.

Heterogeneity of Circulating Epithelial Cells in Breast Cancer at Single-Cell Resolution: Identifying Tumor and Hybrid Cells.

Circulating tumor cells and hybrid cells formed by the fusion of tumor cells with normal cells are leading players in metastasis and have prognostic relevance. This study applies single-cell RNA sequencing to profile CD45-negative and CD45-positive circulating epithelial cells (CECs) in nonmetastatic breast cancer patients. CECs are represented by transcriptionally-distinct populations that include both aneuploid and diploid cells. CD45- CECs are predominantly aneuploid, but one population contained more diploid than aneuploid cells. CD45+ CECs mostly diploid: only two populations have aneuploid cells. Diploid CD45+ CECs annotated as different immune cells, surprisingly harbored many copy number aberrations, and positively correlated to tumor grade. It is noteworthy that cancer-associated signaling pathways areabundant only in one aneuploid CD45- CEC population, which may represent an aggressive subset of circulating tumor cells. Thus, CD45- and CD45+ CECs are highly heterogeneous in breast cancer patients and include aneuploid cells, which are most likely circulating tumor and hybrid cells, respectively, and diploid cells. DNA ploidy analysis can be an effective instrument for identifying tumor and hybrid cells among CECs. Further follow-up study is needed to determine which subsets of circulating tumor and hybrid cells contribute to breast cancer metastasis.

LIMCH1 as a New Potential Metastasis Predictor in Breast Cancer.

BACKGROUND AND OBJECTIVE: High LIMCH1 expression in lung and renal cancer is determined as a favorable prognostic factor. However, prognostic value of LIMCH1 expression in breast cancer has not been studied yet. Therefore, this study was performed to determine the prognostic value of LIMCH1 expression in breast cancer patients. METHODS: This retrospective study included 89 patients with invasive breast carcinoma of no special type. These patients referred to  Cancer Research Institute of Tomsk National Research Medical Center from 2007 to 2018. LIMCH1 protein expression in tumor cells was detected by immunohistochemical analysis in this study. Statistical analysis was done to investigate the possible relationship between LIMCH1 protein expression and clinicopathological parameters, risk of metastasis, distant metastasis free survival, and overall survival. RESULTS: IHC analysis of breast cancer tissue samples revealed that LIMHC1 protein expression was found in 29.2% (26/89) of the cases. Lymph node and distant metastases were more frequent in patients with LIMCH1 protein expression. LIMCH1 protein expression increased the risk of distant metastasis based on our findings. LIMCH1 protein affected metastatic-free survival regardless of the T, as well as other clinical and pathological parameters (p=0.0146, HR=3.2058 (1.26; 8.17)). Moreover, LIMCH1 protein expression was associated with worse overall survival (p=0.0071, HR=2.73 (1.28; 5.85)) in our breast cancer patients. CONCLUSION: LIMCH1 protein expression was associate with metastases development, providing prognostic stratification. In breast cancer, LIMCH1 protein expression was found as an unfavorable prognostic factor of distant metastasis-free survival based on our findings.

The Novel Association of Early Apoptotic Circulating Tumor Cells with Treatment Outcomes in Breast Cancer Patients.

Stemness and epithelial-mesenchymal plasticity are widely studied in the circulating tumor cells of breast cancer patients because the roles of both processes in tumor progression are well established. An important property that should be taken into account is the ability of CTCs to disseminate, particularly the viability and apoptotic states of circulating tumor cells (CTCs). Recent data demonstrate that apoptosis reversal promotes the formation of stem-like tumor cells with pronounced potential for dissemination. Our study focused on the association between different apoptotic states of CTCs with short- and long-term treatment outcomes. We evaluated the association of viable CTCs, CTCs with early features of apoptosis, and end-stage apoptosis/necrosis CTCs with clinicopathological parameters of breast cancer patients. We found that the proportion of circulating tumor cells with features of early apoptosis is a perspective prognosticator of metastasis-free survival, which also correlates with the neoadjuvant chemotherapy response in breast cancer patients. Moreover, we establish that apoptotic CTCs are associated with the poor response to neoadjuvant chemotherapy, and metastasis-free survival expressed at least two stemness markers, CD44 and CD133.

B Lymphocytes Are a Predictive Marker of Eribulin Response and Overall Survival in Locally Advanced or Metastatic Breast Cancer: A Multicenter, Two-Cohort, Non-Randomized, Open-Label, Retrospective Study.

Triple-negative breast cancer has no specific treatment and unfavorable prognosis. Eribulin is one of the drugs widely used in this cohort of patients. In addition to its antimitotic effect, eribulin has an immunomodulant effect on the tumor microenvironment. In this study, we discover immunological markers, such as tumor-infiltrating lymphocytes, CD8+, CD4+, FoxP3+, CD20+ lymphocytes, and their PD1 positivity or negativity, with the ability to predict benefits from eribulin within locally advanced or metastatic triple-negative breast cancer. The primary objective was to explore the association of composition of immune cells in the microenvironment with response to eribulin. The key secondary objective was overall survival. Seven-color multiplex immunofluorescence was used to phenotype lymphocytes in the primary tumor. It has been shown that the PD1-negative-to-PD1-positive B cells ratio in primary tumors more than 3 is an independent predictor of the short-term effectiveness of eribulin [OR (95%CI) 14.09 (1.29-153.35), p=0.0029] and worse overall survival [HR (95%CI) 11.25 (1.37-70.25), p=0.0009] in patients with locally advanced or metastatic triple-negative breast cancer.

Premalignant Changes in the Bronchial Epithelium Are Prognostic Factors of Distant Metastasis in Non-Small Cell Lung Cancer Patients.

BACKGROUND: The study assessed the possibility of dividing patients into groups based on the assessment of morphological changes in the epithelium of small-caliber bronchi located near the primary tumor in order to predict high and low risks of distant metastasis of non-small cell lung cancer. METHODS: In 171 patients with non-small cell lung cancer (T1-4N0-3M0) in small-caliber bronchi taken at a distance of 3-5 cm from the tumor, various variants of morphological changes in the bronchial epithelium (basal cell hyperplasia (BCH), squamous cell metaplasia (SM), and dysplasia (D)) were assessed. Long-term results of treatment, namely, distant metastasis, were assessed after 2 and 5 years. RESULTS: During the follow-up period, distant metastases were found in 35.1% (60/171) of patients. Most often, they were observed in patients of the high-risk group: BCH+SM-D- (51.6%, 40/95) and BCH-SM+D+ (54.4%, 6/11). Less often, distant metastases were observed in low-risk group patients: BCH+SM+D- (6.7%, 3/45) and BCH-SM-D- (10.0%, 2/20). Tumor size, grade, and stage were significant predictors of metastasis only in the high-risk group. The 5-year metastasis-free survival was better in the low-risk group of distant metastases. CONCLUSIONS: Isolated BCH or dysplasia in small bronchi distant from foci of tumor is associated with a high-risk distant metastasis and less 5-year metastasis-free survival.

Parameters of Tumor Microenvironment Determine Effectiveness of Anti-PD-1/PD-L1 Therapy.

Undoubtedly, one of the most promising approaches to the treatment of cancer is creation of the pathogenetically based therapeutic drugs. Researchers from all over the world are trying to answer the question on how to select a target that would be effective and, in general, they are quite successful at that. The Nobel Prize-winning discovery of mechanisms for regulating activity of the immune system cells through checkpoint molecules, as well as discovery of the ability of tumor cells to use these mechanisms to suppress immune responses was an impetus for the development of modern immunotherapy, and now such inhibitors of the immune checkpoints as PD-1/PD-L1 are included in the routine chemotherapy. Use of such drugs can prolong the patient's life, but, unfortunately, not cure the disease. This is partially due to heterogeneity of tumor cells and microenvironment, but the main reasons may be in the complex relationships between the tumor and microenvironment, which, at times, are so plastic that they can change, adjusting to newly emerging conditions. Main characteristic of the tumor microenvironment is the type of the ongoing immune-inflammatory response (IIR), and since inhibitors of the immune checkpoints act on the cells involved in IIR, it is obvious that the outcomes of cancer therapy, including outcomes of hyperprogressive disease, can be associated with this parameter. The presented review reveals the essence of interactions between the tumor and its microenvironment during therapy with PD-L1 inhibitors.

Heterogeneous Manifestations of Epithelial-Mesenchymal Plasticity of Circulating Tumor Cells in Breast Cancer Patients.

To date, there is indisputable evidence of significant CTC heterogeneity in carcinomas, in particular breast cancer. The heterogeneity of CTCs is manifested in the key characteristics of tumor cells related to metastatic progression - stemness and epithelial-mesenchymal (EMT) plasticity. It is still not clear what markers can characterize the phenomenon of EMT plasticity in the range from epithelial to mesenchymal phenotypes. In this article we examine the manifestations of EMT plasticity in the CTCs in breast cancer. The prospective study included 39 patients with invasive carcinoma of no special type. CTC phenotypes were determined by flow cytometry before any type of treatment. EMT features of CTC were assessed using antibodies against CD45, CD326 (EpCam), CD325 (N-cadherin), CK7, Snail, and Vimentin. Circulating tumor cells in breast cancer are characterized by pronounced heterogeneity of EMT manifestations. The results of the study indicate that the majority of heterogeneous CTC phenotypes (22 out of 24 detectable) exhibit epithelial-mesenchymal plasticity. The variability of EMT manifestations does not prevent intravasation. Co-expression of EpCAM and CK7, regardless of the variant of co-expression of Snail, N-cadherin, and Vimentin, are associated with a low number of CTCs. Intrapersonal heterogeneity is manifested by the detection of several CTC phenotypes in each patient. Interpersonal heterogeneity is manifested by various combinations of CTC phenotypes in patients (from 1 to 17 phenotypes).

Prognosis of Different Types of Non-Small Cell Lung Cancer Progression: Current State and Perspectives.

Despite advances in diagnostics and therapy of non-small cell lung cancer (NSCLC), the problem of prognosis and prevention of tumor progression is still highly important. Even if NSCLC is diagnosed in the early stages, almost a quarter of patients develop relapse; most of them die from recurrent disease. A large number of different markers have been proposed to predict the risk of NSCLC progression; however, none of them are used in clinical practice. It is obvious that this situation is related to the economic and methodological complexity of the proposed markers and/or their insufficient efficiency due to a lack of effective study models and tumor heterogeneity. Another reason may be that potential markers are developed for NSCLC progression in general, which is represented by at least four pathogenetically-distinct processes: synchronous lymph node metastasis, local, regional, and distant recurrence. In this review, we summarize data from published literature on clinicopathological, genetic, and molecular factors associated with different types of NSCLC progression and emphasize challenges and approaches to developing prognostic factors. In conclusion, we highlight the importance of further studies to reveal molecular mechanisms of NSCLC progression and the need for differential analysis of markers of local, regional, and distant recurrences for disease prognosis.

Neoadjuvant chemotherapy combined with intraoperative radiotherapy is effective to prevent recurrence in high-risk non-small cell lung cancer (NSCLC) patients.

BACKGROUND: Basal cell hyperplasia (BCH) and squamous metaplasia (SM) in the small bronchi distant from the tumor is associated with a high risk of non-small cell lung cancer (NSCLC) recurrence. Here, we assessed whether neoadjuvant chemotherapy (NAC), intraoperative radiotherapy (IORT), or adjuvant chemotherapy (AC) is effective to prevent recurrence in NSCLC patients (n=171) with different premalignant lesions in the small bronchi. METHODS: BCH, SM, and dysplasia (D) were identified in the samples of lung tissue distant from the tumor. NSCLC patients were treated by surgery, different combinations of NAC and IORT, and AC. RESULTS: Based on the type of bronchial lesions, NSCLC patients were classified into four groups: BCH+SM-D- (55.6%, 95/171), BCH+SM+D- (26.3%; 45/171), BCH-SM+D+ (6.4%, 11/171), and BCH-SM-D- (11.7%, 20/171). During 5 years, recurrent carcinoma was found in 13.4% (23/171) of patients and represented by metachronous metastases in the thoracic lymph nodes (82.6%, 19/23) and by a relapse in the bronchial stump (17.4%, 4/23). Recurrence was frequent in BCH+SM+D- patients (87.0%, 20/23), rare in BCH+SM-D- and BCH-SM-D- patients (13.0%, 3/23), and absent in BCH-SM+D+ patients (0/23). The 5-year recurrence-free survival was also shorter in BCH+SM+D- patients (HR 27.35; 95% CI: 6.31-118.48; P<0.0001). In the high-risk (BCH+SM+D-) group, recurrence occurred mainly in cases without NAC and IORT (88.2%, 15/17) and was absent (0/15) when these therapies were combined. NAC- and IORT-negative patients also showed poor overall survival (HR 4.35; 95% CI: 1.96-9.66; P<0.0001) and tended to have decreased recurrence-free survival (P=0.075). Importantly, the recurrence rate was not different between AC-treated and AC-naïve BCH+SM+D- patients. CONCLUSIONS: The combination of NAC and IORT is an effective strategy to prevent recurrence in high-risk NSCLC patients.

The Activity of KIF14, Mieap, and EZR in a New Type of the Invasive Component, Torpedo-Like Structures, Predetermines the Metastatic Potential of Breast Cancer.

Intratumor morphological heterogeneity reflects patterns of invasive growth and is an indicator of the metastatic potential of breast cancer. In this study, we used this heterogeneity to identify molecules associated with breast cancer invasion and metastasis. The gene expression microarray data were used to identify genes differentially expressed between solid, trabecular, and other morphological arrangements of tumor cells. Immunohistochemistry was applied to evaluate the association of the selected proteins with metastasis. RNA-sequencing was performed to analyze the molecular makeup of metastatic tumor cells. High frequency of metastases and decreased metastasis-free survival were detected in patients either with positive expression of KIF14 or Mieap or negative expression of EZR at the tips of the torpedo-like structures in breast cancers. KIF14- and Mieap-positive and EZR-negative cells were mainly detected in the torpedo-like structures of the same breast tumors; however, their transcriptomic features differed. KIF14-positive cells showed a significant upregulation of genes involved in ether lipid metabolism. Mieap-positive cells were enriched in genes involved in mitophagy. EZR-negative cells displayed upregulated genes associated with phagocytosis and the chemokine-mediated signaling pathway. In conclusion, the positive expression of KIF14 and Mieap and negative expression of EZR at the tips of the torpedo-like structures are associated with breast cancer metastasis.

Heterogeneity of Stemlike Circulating Tumor Cells in Invasive Breast Cancer.

The presence of stem and epithelial-mesenchymal-transition (EMT) features in circulating tumor cells (CTCs) determines their invasiveness, adaptability to the microenvironment, and resistance to proapoptotic signals and chemotherapy. It also allows them to fulfil the role of metastatic "seeds". We evaluated the heterogeneity of stem CTCs by their CD44, ALDH1, and CD133 expression depending on N-cadherin expression in breast-cancer patients. A total of 38 female patients were selected for this study. CTC phenotypes were determined by flow cytometry before any type of treatment. Multiplex immunofluorescence was used for the evaluation of tumor-cell heterogeneity in primary lesions. In patients who had CD44-CD24- CTCs, a subset of cells with the expression of other stem-cell markers (CD133 and ALDH1) were detected. Expression of CD133 and/or ALDH1 may be associated with expression of N-cadherin: all populations of N-cadherin+ CTCs demonstrate stem features; in the absence of N-cadherin expression, true nonstem (CD44-CD24-CD133-ALDH1-) cells are found. The heterogeneity of stem marker expression in CTCs was observed regardless of N-cadherin expression. In our study, stromal cell-derived factor-1 (SDF-1) receptor expression in CTCs did not depend on stemlike traits, but was instead associated with N-cadherin expression. Subpopulations of tumor cells, detected both in tumors and blood, were identified. Breast cancer was characterized by pronounced interpersonal and intrapersonal heterogeneity of CTCs by the presence and combination of various stem features and N-cadherin expression. To complete the characterization of stemlike features of CTCs, we suggest the simultaneous use of the three stem markers.

Single Tumor Cells With Epithelial-Like Morphology Are Associated With Breast Cancer Metastasis.

Introduction: The identification of tumor cells that can be potential metastatic seeds would reach two key aims-prognosis of metastasis risk and appointment of the optimal adjuvant therapy to prevent metastatic disease. Single tumor cells (STCs) located out of multicellular structures can most likely demonstrate features that are needed to initiate metastasis. Methods: One-hundred-and-thirty-five patients with invasive breast carcinoma of no special type have been enrolled. Molecular subtypes of breast cancer were categorized according to St. Gallen recommendations. Hematoxylin and eosin staining was used to identify STCs with epithelial-like morphology (eSTCs) in breast tumors. Immunofluorescence staining was applied to evaluate stemness and epithelial-mesenchymal transition (EMT) in STCs. The correlation between STCs and recurrence and metastasis-free survival (MFS) was performed using the Kaplan-Meier method and the log-rank test. Results: Distant metastasis was more frequent in eSTC-positive than eSTC-negative patients (28.0% vs. 9.4%, p = 0.007). When tumor types were analyzed separately, distant metastasis tended to be more frequent in eSTC-positive than eSTC-negative patients for HER2-positive cancer [75.0% (3/4) vs. 12.5% (1/8), p = 0.066]. In luminal A [22.7% (5/22) vs. 10.0% (3/30), p = 0.259], luminal B [21.1% (4/19) vs. 6.7% (2/30), p = 0.189], and triple-negative [40.0% (2/5) vs. 11.8% (2/17), p = 0.209] cancers, distance metastasis was not associated with eSTCs. Median MFS was not reached in eSTC-positive and eSTC-negative patients. eSTC-positive patients had a higher risk of breast cancer metastasis [hazard ratio (HR) 3.57, 95% confidence interval (CI): 1.46-8.71; p = 0.001]. When tumor types were analyzed separately, a higher risk of breast cancer metastasis occurred only in HER2-positive patients (HR 8.49, 95% CI: 1.29-55.59; p = 0.016). Immunofluorescence analysis revealed mesenchymal-like STCs (mSTCs) and inter- and intra-tumor heterogeneity in STCs. There were breast tumors with either eSTCs or mSTCs and tumors with both types of STCs. Both eSTCs and mSTCs were represented by cells with different stem and/or EMT phenotypes. Conclusions: STCs with epithelial-like morphology contribute to breast cancer metastasis and represent an attractive model for studying mechanisms of metastatic seeding. The assessment of STCs in histological sections of breast tumors can be a simple and effective method for the prediction of metastasis risk.

Gene Expression Profiling Revealed 2 Types of Bronchial Basal Cell Hyperplasia and Squamous Metaplasia With Different Progression Potentials.

The premalignant process preceding squamous cell lung cancer is not inevitable; it can stop at any of the bronchial lesions: basal cell hyperplasia (BCH), squamous metaplasia (SM), and dysplasia and then progress or regress. At present, the mechanisms underlying the progression of the bronchial lesions remain undefined. Previously, we hypothesized that bronchial lesions that presented individually or combined with each other in the bronchi of lung cancer patients mirror the different "scenarios" of the premalignant process: individual BCH-the stoppage at the stage of hyperplasia, BCH plus SM-the progression of hyperplasia to metaplasia, and SM plus dysplasia-the progression of metaplasia to dysplasia. In this study, we analyzed gene expression profiles of BCH, SM, and dysplasia depending on their cooccurrence in the bronchi of lung cancer patients. The immune response gene expression was found to be a key difference between the individual BCH and BCH combined with SM lesions and a potential mechanism that determines the progression of hyperplasia to metaplasia. Upregulation of the cell cycle and downregulation of the cilium assembly genes mainly distinguished SM that copresented with dysplasia from SM that copresented with BCH and is a probable mechanism of the progression of metaplasia to dysplasia. Dysplasia showed mainly overexpression of the cell division genes and underexpression of the inflammation genes. Thus, this study demonstrates the significant gene expression differences between the premalignant lesions depending on their cooccurrence in the bronchi and sheds light on the mechanisms of the precancerous process preceding squamous cell lung cancer.

P-cadherin-induced decorin secretion is required for collagen fiber alignment and directional collective cell migration.

Directional collective cell migration (DCCM) is crucial for morphogenesis and cancer metastasis. P-cadherin (also known as CDH3), which is a cell-cell adhesion protein expressed in carcinoma and aggressive sarcoma cells and associated with poor prognosis, is a major DCCM regulator. However, it is unclear how P-cadherin-mediated mechanical coupling between migrating cells influences force transmission to the extracellular matrix (ECM). Here, we found that decorin, a small proteoglycan that binds to and organizes collagen fibers, is specifically expressed and secreted upon P-cadherin, but not E- and R-cadherin (also known as CDH1 and CDH4, respectively) expression. Through cell biological and biophysical approaches, we demonstrated that decorin is required for P-cadherin-mediated DCCM and collagen fiber orientation in the migration direction in 2D and 3D matrices. Moreover, P-cadherin, through decorin-mediated collagen fiber reorientation, promotes the activation of ß1 integrin and of the ß-Pix (ARHGEF7)/CDC42 axis, which increases traction forces, allowing DCCM. Our results identify a novel P-cadherin-mediated mechanism to promote DCCM through ECM remodeling and ECM-guided cell migration.

Premalignant lesions of squamous cell carcinoma of the lung: The molecular make-up and factors affecting their progression.

Squamous cell carcinoma (SCC), one of the most common forms of lung cancer, shows accelerated progression and aggressive growth and usually is observed at advanced stages. SCC originates from morphological changes in the bronchial epithelium that occur during chronic inflammation: basal cell hyperplasia, squamous metaplasia, and dysplasia I-III. However, the process is not inevitable; it can be stopped at any stage, remain in the stable state indefinitely and either progress or regress. The reasons and mechanisms of different scenarios of the evolution of premalignant lesions in the respiratory epithelium are not fully understood. In this review, we summarized the literature data (including our own data) regarding genetic, epigenetic, transcriptomic and proteomic profiles of the premalignant lesions and highlighted factors (environmental causes, inflammation, and gene polymorphism) that may govern their progression or regression. In conclusion, we reviewed strategies for lung cancer prevention and proposed new models and research directions for studying premalignant lesions and developing new tools to predict the risk of their malignant transformation.