DNA Copy Number Aberrations and Expression of ABC Transporter Genes in Breast Tumour: Correlation with the Effect of Neoadjuvant Chemotherapy and Prognosis of the Disease.

One of the important reasons for the ineffectiveness of chemotherapy in breast cancer (BC) is considered to be the formation of a multidrug resistance phenotype in tumour cells, which is caused by the expression of energy-dependent ABC transporters. The aim of this work was to assess chromosomal aberrations and the level of transcripts of all 49 known ABC transporter genes in breast tumours. MATERIALS AND METHODS: The study included 129 patients with breast cancer. A microarray study of all tumour samples was carried out on microchips. RESULTS: This study established that the presence of a deletion in genes ABCB1, ABCB4, ABCB8, ABCC7, ABCC11, ABCC12, ABCF2, and ABCG4 is associated with an objective response to treatment (p ≤ 0.05). A decrease in the expression of genes was associated with a good response to chemotherapy, whereas an increase in expression caused the progression and stabilization of the tumour. Analysis of metastatic-free survival rates showed that the presence of ABCB1/4 and ABCC1/6 deletions was associated with 100% survival (log-rank test p = 0.01 and p = 0.03). CONCLUSIONS: The study showed that the aberrant state of ABC transporter genes, as well as a decrease in the expression of these genes, is a predictor of the effectiveness of therapeutic treatment and a potential prognostic marker of metastatic survival.

Stem gene expression in breast tumors during chemotherapy: Connection with the main clinical and morphological factors and the disease outcome.

Introduction: In this research, we studied how the expression of 14 stem genes (TERT; OCT3; SMO; MYC; SNAI2; MOB3B; KLF4; BMI1; VIM; FLT3; LAT; SMAD2; LMNB2; KLF1), as well as the TGF-ß1 cytokine gene and its TGFBR1 receptor in breast tumors before and after NAC is associated with clinical and morphological parameters and the disease outcome. Materials and Methods: The study included 82 patients with the morphologically verified diagnosis of T1-4N0-3M0 breast cancer (stages IIA - IIIB). The material was paired biopsy samples of tumor and surgical material for each patient. The stem genes expression was analyzed via qPCR. Results: As a result, we found that increased level of stem genes expression in breast tumors is associated with lymphogenic metastasis, young age, small tumor size, expression of estrogen and progesterone receptors, and the luminal B molecular subtype. NAC stimulates the expression of 7 out of 16 stem genes. Patients who further developed hematogenic metastases have twice as many hyperexpressed stem genes in their tumors before the treatment and after NAC than patients with no hematogenic metastases. The expression level of three genes - OCT3, LAT, and LMNB2 - in a residual tumor allows us to predict metastasis-free survival of patients with breast cancer of various molecular subtypes with a 79% accuracy. Conclusion: Thus, stem genes hyperexpression is associated with tumor progression.

Predictive and Prognostic Significance of mRNA Expression and DNA Copies Aberrations of ERCC1, RRM1, TOP1, TOP2A, TUBB3, TYMS, and GSTP1 Genes in Patients with Breast Cancer.

Increasingly, many researchers are focusing on the sensitivity in breast tumors (BC) to certain chemotherapy drugs and have personalized their research based on the assessment of this sensitivity. One such personalized approach is to assess the chemotherapy's gene expression, as well as aberrations in the number of DNA copies-deletions and amplifications with the ability to have a significant effect on the gene's activity. Thus, the aim of this work was to study the predictive and prognostic significance of the expression and chromosomal aberrations of eight chemosensitivity genes in breast cancer patients. MATERIAL AND METHODS: The study involved 97 patients with luminal B breast cancer IIB-IIIB stages. DNA and RNA were isolated from samples of tumor tissue before and after treatment. Microarray analysis was performed for all samples on high-density microarrays (DNA chips) of Affymetrix (USA) CytoScanTM HD Array and Clariom™ S Assay, human. Detection of expression level of seven chemosensitivity genes-RRM1, ERCC1, TOP1, TOP2a, TUBB3, TYMS, and GSTP1-was performed using PCR real-time (RT-qPCR). RESULTS: The expression of the RRM1 (AC scheme), TOP2α, TYMS, and TUBB3 genes in patients with an objective response to treatment (complete and partial regression) is higher than in patients with stabilization and progression (p < 0.05). According to our results, the presence of a high level of GSTP1 in a tumor biopsy is associated with the low efficiency of the NAC CP scheme (p = 0.05). The presence of RRM1 deletion is associated with complete and partial regression, as for the TOP1 and TUBB3 genes (p < 0.05). Higher rates of metastatic survival are associated with a high level of expression and amplification of the GSTP1 gene (log-rank test p = 0.02 and p = 0.05). CONCLUSION: Thus, a complex assessment of the chemotherapy's gene expression is important not only for understanding the heterogeneity and molecular biology of breast cancer but also to obtain a more accurate disease prognosis.

Five-year survival in luminal breast cancer patients: relation with intratumoral activity of proteasomes.

Background: The purpose of the study was to analyze the relationship between the caspase-like (CL) and chymotrypsin-like (ChTL) activities of proteasomes and the 5-year overall and metastasis-free survival rates in patients with luminal breast cancer. Methods: The study included 117 patients with primary operable invasive breast cancer (T1-2N0-1M0). Tissue samples from breast cancer patients were obtained as a result of the radical mastectomy or breast conserving surgery, which was a first line of therapy. The ChTL and CL proteasomes activities in the tumor tissue and in the surrounding adjacent breast tissues were assessed using the fluorometric method. The coefficients of ChTL (cChTL) and CL (cCL) proteasomes activities were also determined. The coefficients were calculated as the ratio of the corresponding proteasomes activity in the tumor tissue to the surrounding adjacent breast tissues. Within 5 years of follow-up, hematogenous metastases occurred in 14% of patients with luminal A breast cancer, in 31% of patients with luminal B human epidermal growth factor receptor-2 (HER-2) negative and in 23% of patients with luminal B HER-2 positive breast cancers. The study protocol was approved by the Local Ethics Committee of the Cancer Research Institute of Tomsk National Research Medical Center. Written informed consent was obtained from all patients. Results: An increase in the ChTL and CL proteasomes activities was shown in all studied molecular subtypes of breast cancer compared to adjacent tissues. It was found that the cChTL of >35.9 U/mg protein and the cCL of >2.21 in breast cancer patients were associated with the development of distant metastases. In patients with luminal A breast cancer, the 5-year metastasis-free survival rates were associated only with the value of cCL of proteasomes (log-rank test: P=0.008). In patients with luminal B HER-2 negative breast cancer, the 5-year metastasis-free survival rates were associated with the levels of ChTL and cCL proteasomes activities (log-rank test: P=0.02 and P=0.04, respectively). Conclusions: The data obtained on the correlation of 5-year metastasis-free survival rates with the level of proteasomes activities indicate the possibility of their use as additional prognostic criteria for breast cancer.

Amplifications of Stemness Gene Loci-New Markers for the Determination of the Need for Neoadjuvant Chemotherapy for Patients with Breast Cancer. A Prospective Study.

In this prospective study, a new strategy for the prescription of neoadjuvant chemotherapy (NAC) was prospectively tested and depended on the presence of stemness gene amplifications in the tumor before treatment, which in our early studies showed a connection with metastasis. The study included 92 patients with grade IIA-IIIB luminal B breast cancer. Patients underwent a biopsy before treatment, and with the use of a CytoScan HD Array microarray (Affymetrix, Santa Clara, CA, USA), the presence of stemness gene amplifications (3q, 5p, 6p, 7q, 8q, 13q, 9p, 9q, 10p, 10q21.1, 16p, 18chr, 19p) in the tumor was determined. In group 1 (n = 41), in the presence of two or more amplifications, patients were prescribed a personalized NAC regimen. In group 2 (n = 21), if there was no amplification of stemness genes in the tumor, then patients were not prescribed NAC, and treatment began with surgery. Group 3 (n = 30) served as a historical control. The frequency of an objective response to NAC in groups 1 and 3 was 79%. Nonmetastatic survival was found in 100% of patients in group 2, who did not undergo NAC. In patients in group 1, the frequency of metastasis was 10% (4/41). At the same time, in patients in group 3, who received NAC, the rate of metastasis was 47% (14/30). The differences between group 1 and group 3 and between group 2 and group 3 were statistically significant, both by Fisher's criterion and a log-rank test. The appointment of NAC was most feasible in patients with clones with stemness gene amplifications in the primary tumor, while in the absence of amplifications, preoperative chemotherapy led to a sharp decrease in metastasis-free survival. This strategy of NAC prescription allowed us to achieve 93% metastatic survival in patients with breast cancer.

DNA methylation markers panel can improve prediction of response to neoadjuvant chemotherapy in luminal B breast cancer.

Despite the advantages of neoadjuvant chemotherapy (NACT), associated toxicity is a serious complication that renders monitoring of the patients' response to NACT highly important. Thus, prediction of tumor response to treatment is imperative to avoid exposure of potential non-responders to deleterious complications. We have performed genome-wide analysis of DNA methylation by XmaI-RRBS and selected CpG dinucleotides differential methylation of which discriminates luminal B breast cancer samples with different sensitivity to NACT. With this data, we have developed multiplex methylation sensitive restriction enzyme PCR (MSRE-PCR) protocol for determining the methylation status of 10 genes (SLC9A3, C1QL2, DPYS, IRF4, ADCY8, KCNQ2, TERT, SYNDIG1, SKOR2 and GRIK1) that distinguish BC samples with different NACT response. Analysis of these 10 markers by MSRE-PCR in biopsy samples allowed us to reveal three top informative combinations of markers, (1) IRF4 and C1QL2; (2) IRF4, C1QL2, and ADCY8; (3) IRF4, C1QL2, and DPYS, with the areas under ROC curves (AUCs) of 0.75, 0.78 and 0.74, respectively. A classifier based on IRF4 and C1QL2 better meets the diagnostic panel simplicity requirements, as it consists of only two markers. Diagnostic accuracy of the panel of these two markers is 0.75, with the sensitivity of 75% and specificity of 75%.

Neoadjuvant Chemotherapy Induces the Appearance of New Copy Number Aberrations in Breast Tumor and is Associated with Metastasis.

BACKGROUND: In this study, we examined the CNA-genetic landscape (CNA - copy number aberration) of breast cancer prior to and following neoadjuvant chemotherapy (NAC) and correlated changes in the tumor landscape with chemotherapy efficiency as well as metastasis-free survival. OBJECTIVE: Breast cancer patients (n = 30) with luminal B molecular subtypes were treated with anthracycline- based therapy. METHODS: To study CNAs in breast tumors, microarray analysis was performed. RESULTS: Three effects of NAC on tumor CNA landscape were identified: 1 - the number of CNAbearing tumor clones decreased following NAC; 2 - there were no alterations in the number of CNAcontaining clones after NAC; 3 - the treatment with NAC increased the number of CNA-bearing clones (new clones appeared). All NAC-treated patients who had new tumor clones with amplification (20%) had a 100% likelihood of metastasis formation. In these cases, NAC contributed to the emergence of potential metastatic clones. Our study identified the following loci - 5p, 6p, 7q, 8q, 9p, 10p, 10q22.1, 13q, 16p, 18Chr and 19p - that were amplified during the treatment with NAC and maybe the markers of potential metastatic clones. In other patients who showed total or partial elimination of CNA-bearing cell clones, no new amplification clones were observed after NAC, and no evidence of metastases was found with follow-up for 5 years (р = 0.00000). CONCLUSION: Our data suggest that the main therapeutic result from NAC is the elimination of potential metastatic clones present in the tumor before treatment. The results showed the necessity of an intelligent approach to NAC to avoid metastasis stimulation.

Amplification of Stem Genes: New Potential Metastatic Makers in Patients with an Early Form of Breast Cancer.

BACKGROUND: According to our previous studies, the presence of amplifications of stem genes can lead to their ectopic expression and this is associated with an increased activity of tumor stem cells in these patients. This leads to a high aggressiveness of the tumor and the development of metastatic disease. The aim was to evaluate the prognostic significance of the presence of amplifications of stem genes and their expression in patients with early breast cancer (BC). METHODS: The study included 28 patients with T1NxM0 BC. We used surgical specimens, including formalin-fixed paraffin-embedded archive materials, for 8 patients. A microarray analysis was performed on high-density DNA chips from CytoScanHDArray to assess the status of copy number aberration (CNA) of stem genes locus. Gene expression was assessed using RT-qPCR. RESULTS: CNA analysis of the studied tumors of patients without chemotherapy showed that 17/18 patients without metastases did not have two or more amplifications of chromosomal regions. Ten patients had visceral metastases. In 9/10 of these patients in the primary tumor there were two or more amplifications of the stem genes locus. Two or more amplifications of stem genes locus were found in 12 patients with stage I. Hematogenous metastases did not develop in all patients. Comparison of metastasis-free survival rates in groups of patients with 1 or without amplifications and with two or more amplifications showed statistically significant differences (P = 0.01). CONCLUSION: Our studies have shown that the presence of clones with two or more amplifications of stem gene in patients with BC T1NxM0 has a significant prognostic value and determines an unfavorable prognosis for distant metastasis.

Genome-wide methylotyping resolves breast cancer epigenetic heterogeneity and suggests novel therapeutic perspectives.

Aim: To provide a breast cancer (BC) methylotype classification by genome-wide CpG islands bisulfite DNA sequencing. Materials & methods: XmaI-reduced representation bisulfite sequencing DNA methylation sequencing method was used to profile DNA methylation of 110 BC samples and 6 normal breast samples. Intrinsic DNA methylation BC subtypes were elicited by unsupervised hierarchical cluster analysis, and cluster-specific differentially methylated genes were identified. Results & conclusion: Overall, six distinct BC methylotypes were identified. BC cell lines constitute a separate group extremely highly methylated at the CpG islands. In turn, primary BC samples segregate into two major subtypes, highly and moderately methylated. Highly and moderately methylated superclusters, each incorporate three distinct epigenomic BC clusters with specific features, suggesting novel perspectives for personalized therapy.

Heterogeneity of Circulating Tumor Cells in Neoadjuvant Chemotherapy of Breast Cancer.

The biological properties of circulating tumor cells (CTCs), and their dynamics during neoadjuvant chemotherapy are important, both for disease progression prediction and therapeutic target determination, with the aim of preventing disease progression. The aim of our study was to estimate of different CTC subsets in breast cancer during the NACT (neoadjuvant chemotherapy). The prospective study includes 27 patients with invasive breast cancer, T2-4N0-3M0, aged 32 to 60 years. Venous heparinized blood samples, taken before and after biopsy, after each courses of chemotherapy (on days 3-7), and before surgical intervention, served as the material for this study. Different subsets of circulating tumor cells were determined on the basis of the expression of EpCAM, CD45, CD44, CD24, and N-Cadherin using flow cytometry. As the result of this study, it has been observed that significant changes in the quantity of the different subsets of circulating tumor cells in patients' blood were observed after carrying out the 3rd course of NACT. NACT causes significant changes in the quantity of six CTC subsets, with various combinations of stemness and epithelial-mesenchymal transition (EMT) properties.

[Targeting of the AKT/m-TOR Pathway: Biomarkers of Resistance to Cancer Therapy--
AKT/m-TOR Pathway and Resistance to Cancer Therapy].

Resistance to cancer therapy continues to be a major limitation for the successful treatment of cancer. There are many published studies on therapy resistance in breast and prostate cancers; however, there are currently no data on molecular markers associated with resistance. The conflicting data were reported regarding the AKT/m-TOR signaling pathway components as markers predicting resistance. The AKT/m-TOR signaling pathway is involved in the development of many human cancers; its activation is related to cell proliferation, angiogenesis, apoptosis, as well as to therapy resistance. Molecular alterations in the AKT/m-TOR signaling pathway provide a platform to identify universal markers associated with the development of resistance to cancer therapy.

Effect of Targeted Therapy With Pazopanib on Expression Levels of Transcription, Growth Factors and Components of AKT/m-TOR Signaling Pathway in Patients with Renal Cell Carcinoma

Background: The effect of the targeted therapy on cancer molecular markers remains currently unknown. The aim of the study was to investigate the expression and content of transcription, growth factors and components of the AKT/m-TOR signaling pathway in kidney cancer patients before and after targeted therapy with pazopanib. Methods: A total of 157 patients with renal cell carcinoma were enrolled into the study. The level of mRNA expression was investigated by real-time PCR, and the contents of transcription and growth factors, as well as the levels of AKT/m- TOR signaling pathway components were determined by ELISA and Western blotting. Results: Targeted therapy with pazopanib resulted in a 3.1-fold decrease in HIF-2α expression that was accompanied by a reduction in the levels of NF-κB p65 and p50, HIF-1α and CAIX. The levels of GSK-3ß and AKT mRNA were increased; however, the levels of corresponding proteins remained low. The targeted therapy with pazopanib did not influence the level of PTEN phosphatase. A 1.9-fold increase in the level of p70 S6 (S371) was observed after therapy. Conclusion: The efficacy of tyrosine kinase inhibitors is associated with the changes in the angiogenic factors. Molecular characteristics of cancer could determine markers of disease progression as well as potential targets for anticancer therapies

Genetic variability in the regulation of the expression cluster of MDR genes in patients with breast cancer.

PURPOSE: We aimed to investigate the association between the polymorphism and expression patterns of multiple drug resistance genes (MDR) in breast cancer (BC). MATERIALS AND METHODS: The MDR gene expression levels were measured in tumor tissues of 106 breast cancer patients using quantitative real-time PCR. Affymetrix CytoScan™ HD Array chips were used to assess genotypes. Pairwise correlation analysis for ABCB1, ABCC1, ABCC2 and ABCG2 gene expression levels was carried out to reveal co-expression clusters. Associations between SNPs of MDR genes and their preoperative expression levels were assessed using analysis of covariance adjusting for covariates. RESULTS: The SNPs associated with the expression of the ABCB1, ABCC1, ABCC2 and ABCG2 genes before NAC were detected. In addition, 21 SNPs associated with the expression of four ABC-transporter genes and involved in the expression regulation were identified. Validation in an independent sample confirmed the association between the MDR cluster genes and 11 SNPs. CONCLUSIONS: Four MDR genes: ABCB1, ABCC1, ABCC2 and ABCG2 were shown to form the functional expression cluster in breast tumor. Further studies are required to discover precise mechanisms of the cluster regulation, thereby providing new approaches and targets to combat the development of the MDR phenotype during chemotherapy.

Deletions of multidrug resistance gene loci in breast cancer leads to the down-regulation of its expression and predict tumor response to neoadjuvant chemotherapy.

Neoadjuvant chemotherapy (NAC) is intensively used for the treatment of primary breast cancer. In our previous studies, we reported that clinical tumor response to NAC is associated with the change of multidrug resistance (MDR) gene expression in tumors after chemotherapy. In this study we performed a combined analysis of MDR gene locus deletions in tumor DNA, MDR gene expression and clinical response to NAC in 73 BC patients. Copy number variations (CNVs) in biopsy specimens were tested using high-density microarray platform CytoScanTM HD Array (Affymetrix, USA). 75%-100% persons having deletions of MDR gene loci demonstrated the down-regulation of MDR gene expression. Expression of MDR genes was 2-8 times lower in patients with deletion than in patients having no deletion only in post-NAC tumors samples but not in tumor tissue before chemotherapy. All patients with deletions of ABCB1 ABCB 3 ABCC5 gene loci--7q21.1, 6p21.32, 3q27 correspondingly, and most patients having deletions in ABCC1 (16p13.1), ABCC2 (10q24), ABCG1 (21q22.3), ABCG2 (4q22.1), responded favorably to NAC. The analysis of all CNVs, including both amplification and deletion showed that the frequency of 13q14.2 deletion was 85% among patients bearing tumor with the deletion at least in one MDR gene locus versus 9% in patients with no deletions. Differences in the frequency of 13q14.2 deletions between the two groups were statistically significant (p = 2.03 × 10(-11), Fisher test, Bonferroni-adjusted p = 1.73 × 10(-8)). In conclusion, our study for the first time demonstrates that deletion MDR gene loci can be used as predictive marker for tumor response to NAC.

Relationship between the expression of phosphorylated heat shock protein beta-1 with lymph node metastases of breast cancer.

BACKGROUND: Heat shock protein beta-1 (HspB1) is a chaperone of the sHsp (small heat shock protein). The common functions of sHsps are chaperone activity, inhibition of apoptosis, regulation of cell development, and cell differentiation, take part in signal transduction. OBJECTIVE: To study the intracellular localization of phosphorylated features and non-phosphorylated forms of HspB1 in primary breast cancer cells and to evaluate their relationship with regional lymphatic metastasis. MATERIAL AND METHODS: Tumor biopsies of breast tissue were collected from 100 patients with a confirmed diagnosis of invasive carcinoma, nonspecific type, between the ages of 31-80 years. Immunohistochemistry was used to determine the intracellular localization of phosphorylated and non-phosphorylated forms of HspB1. RESULTS: The result of this study showed that biopsies from patients with lymph node metastasis exhibited significantly higher levels of phosphorylated forms of HspB1 in the nucleus and cytoplasm compared with the group without lymph node metastasis. Analysis showed that the expression of phosphorylated forms of the chaperone HspB1 correlates with the amount and percentage of lymph node metastases affected. CONCLUSION: The nuclear expression of phosphorylated and non-phosphorylated forms of the chaperone HspB1 is a marker of tumor cells associated with lymphatic metastasis of breast cancer.

Proteasome functioning in breast cancer: connection with clinical-pathological factors.

Breast cancer is one of four oncology diseases that are most widespread in the world. Moreover, breast cancer is one of leading causes of cancer-related deaths in female population within economically developed regions of the world. So far, detection of new mechanisms of breast cancer development is very important for discovery of novel areas in which therapy approaches may be elaborated. The objective of the present study is to investigate involvement of proteasomes, which cleave up to 90% of cellular proteins and regulate numerous cellular processes, in mechanisms of breast cancer development. Proteasome characteristics in 106 patient breast carcinomas and adjacent tissues, as well as relationships of detected proteasome parameters with clinical-pathological factors, were investigated. Proteasome chymotrypsin-like activity was evaluated by hydrolysis of fluorogenic peptide Suc-LLVY-AMC. The expression of proteasome subunits was studied by Western-blotting and immunohistochemistry. The wide range of chymotrypsin-like activity in tumors was detected. Activity in tumors was higher if compared to adjacent tissues in 76 from 106 patients. Multiple analysis of generalized linear models discovered that in estrogen α-receptor absence, tumor growth was connected with the enhanced expression of proteasome immune subunit LMP2 and proteasome activator PA700 in tumor (at 95% confidence interval). Besides, by this analysis we detected some phenomena in adjacent tissue, which are important for tumor growth and progression of lymph node metastasis in estrogen α-receptor absence. These phenomena are related to the enhanced expression of activator PA700 and immune subunit LMP7. Thus, breast cancer development is connected with functioning of immune proteasome forms and activator PA700 in patients without estrogen α-receptors in tumor cells. These results could indicate a field for search of new therapy approaches for this category of patients, which has the worst prognosis of health recovery.

Intratumoral morphological heterogeneity of breast cancer: neoadjuvant chemotherapy efficiency and multidrug resistance gene expression.

In this study, the influence of intratumoral morphological heterogeneity of breast cancer on neoadjuvant chemotherapy (NAC) efficiency was investigated. In particular, we analysed the association of NAC response and pre- and post-NAC expression of the main multidrug resistance (MDR) genes--ABCB1, ABCC1, ABCC5, ABCG1, and ABCG2, with the presence of different morphological structures in breast tumors. In addition, the expression of MDR genes was investigated in different morphological structures and in their microenvironment by comparing probes obtained using laser microdissection. The results of this study showed that tumors with alveolar structures were more frequently NAC-nonresponsive than cases without this structural type (p = 0.0028, Bonferroni-corrected p = 0.014). The presence of trabecular structures in breast tumors was also associated with chemoresistance (p = 0.0272, Bonferroni-corrected p = 0.136). High expression of MDR genes was not found in alveolar structures (including their microenvironment) and in tumors containing this structural type. In contrast, more active MDR genes and expression of the ABCB1 gene were found only in trabecular structures. Taken together, our data indicate that breast tumors with alveolar structures possess resistance to NAC, which is not related to high expression of MDR genes, whereas chemoresistance of tumors with trabecular structures can depend on the expression level of ABCB1.

Phenotypic drift as a cause for intratumoral morphological heterogeneity of invasive ductal breast carcinoma not otherwise specified.

Invasive ductal carcinoma (IDC) not otherwise specified (NOS), the most common type of breast cancer, demonstrates great intratumoral morphological heterogeneity, which encompasses the presence of different types of morphological structures-tubular, trabecular, solid, and alveolar structures and discrete groups of tumor cells, the origins of which remain unclear at present. In this study of 162 IDC NOS patients, we investigated whether the distribution of different types of morphological structures is related to the basic clinicopathological parameters of IDC NOS. Our results showed that in patients with only one type of tumor structure, the presence of any one of the five types was equally probable; however, cases with two types of structures were more likely to contain trabecular structures than the other four types. The development of intratumoral morphological heterogeneity was not associated with menopausal status, tumor size, histological grade, hematogenic metastasis, or recurrence. However, the number of different types of morphological structures was significantly higher in luminal tumors than in triple-negative tumors. An increase in the frequency of lymph node metastasis correlated with the increased number of different types of structures in breast tumors; however, in contrast to premenopausal patients, this association was explained by the presence of alveolar structures in postmenopausal women. In addition, we showed a significant decrease in the numbers of positive lymph nodes in tumors with high numbers of morphological variants. The frequency of lymph node metastases and the number of positive nodes were generally independent features and formed by different mechanisms. Based on the evidence, the term "phenotypic drift" has been designated as the basis for the development of intratumoral morphological heterogeneity of IDC NOS.

Changing the expression vector of multidrug resistance genes is related to neoadjuvant chemotherapy response.

PURPOSE: We aimed to examine the association between alterations in multidrug resistance (MDR) gene expression, measured before and after neoadjuvant chemotherapy (NAC), and short-term response in a cohort of stage IIA-IIIC breast cancer patients (n = 84). METHODS: All patients were treated with two to four preoperative cycles of FAC (5-fluorouracil-adriamycin-cyclophosphamide), CAX (cyclophosphamide-adriamycin-xeloda) or taxane regimes. The expression levels of key MDR genes (ABCB1, ABCC1, ABCC2, ABCC3, ABCC5, ABCG1, ABCG2, GSTP1, and MVP) were evaluated in both tumor tissues obtained pre-therapy and in specimens removed by final surgery, using TaqMan-based quantitative reverse transcriptase PCR. RESULTS: No significant difference in the average level of MDR gene expression in paired breast tumors before and after NAC was found when analyzed in both responsive and non-responsive patients. There was no correlation between the expression levels of MDR genes in pre-NAC tumors and immediate NAC response. In the group with tumor responses, we found a statistically significant downregulation of expression of ABCB1, ABCC1, ABCC2, ABCC5, ABCG1, ABCG2, GSTP1, and MVP genes following NAC in FAC and CAX-treated patients (67-93% of cases). In contrast, we found that expression of these genes was upregulated after NAC, mostly in non-responsive patients (55-96% of cases). Responsiveness to taxotere was related to reduced levels of ABCB1, ABCC2, ABCG1, ABCG2, and MVP mRNA in tumor samples collected after chemotherapy. CONCLUSION: Our results suggest that reductions in MDR gene expression in post-NAC samples in comparison with pre-NAC are associated with tumor response to FAC and CAX as well as taxotere-based NAC, while patients displaying MDR gene upregulation had resistance to therapy.