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Results of enzyme-linked immunosorbent assay of the soluble forms of PD-1/PD-L immune checkpoint receptor and ligand (sPD-1 and sPD-L1) in pretreatment blood serum of 88 breast cancer patients at various disease stages aged 30-83 years are presented. The control group included 55 practically healthy women aged 19-82 years. Serum sPD-1 and sPD-L1 levels in breast cancer patients highly significantly (p<0.0001) differ from control and these changes are opposite: soluble receptor level is more than 6-fold decreased, while soluble ligand concentration - 5.5 fold increased. Both markers separately, as well as their ratio demonstrate very high sensitivity (94-100%) and specificity (95-100%) in relation to healthy control. No statistically significant associations of sPD-1 and sPD-L1 levels with clinical stage, individual TNM system criteria, tumor histological structure, grade, receptor status, and molecular type were established. In particular, no significant peculiarities of the markers' levels in triple negative breast cancer successfully treated with anti-PD-1/PD-L1 preparations were revealed. Long-term follow-up and dynamic studies of sPD-1 and sPD-L1serum levels in the course of treatment are required for evaluation of their independent from clinical and morphological factors prognostic significance and the possibility of application as low invasive tests for prediction and monitoring of corresponding targeted therapy efficiency., Competing Interests: The authors declare no conflict of interest.

It is known that microRNAs (miRNAs) are able to dynamically regulate gene expression. At the same time, methylation can reduce expression of miRNA encoding genes and, therefore, reduce their inhibitory effects on mRNAs of target genes, including those of oncogenes, that promoting the development of tumors of different localization. The role of miRNA hypermethylation in the pathogenesis of ovarian cancer is not completely understood; so we conducted a search for new hypermethylated and potentially suppressor miRNA genes in ovarian tumors. Four new miRNA genes (MIR-107, MIR-130b, MIR-203a, MIR-1258) commonly hypermethylated (28-52%) in tumor tissues vs 4-7% in paired histologically normal tissues, p < 0.01, were identified in a representative set of 54 ovarian cancer samples using methylation-specific PCR. It was shown that hypermethylation of MIR-130b, MIR-203a, and MIR-1258 genes is significantly (p < 0.05) associated with metastasis of ovarian cancer. These results suggest the involvement of four miRNAs (miR-107, miR-130b, miR-203a, and miR-1258) and hypermethylation of their encoding genes in the pathogenesis of ovarian cancer.

In patients with endometrial cancer (N=94), endometrial polyps (N=28), endometrial hyperplasia (N=25), and healthy women (N=77), the serum contents of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 were measured by ELISA. Both carcinoma and benign neoplasms were accompanied by significant elevation of MMP-7 and TIMP-2 in blood serum. The greatest elevation (in comparison with the control) was observed for MMP-7, although serum concentration of this marker was practically identical in patients with carcinoma and benign tumors. In contrast, the levels of MMP-2 and TIMP-1 were lower in cancer patients in comparison with the control; in these patients, the levels of MMP-9 and TIMP-1 were also lower than the corresponding levels in patients with polyps and endometrial hyperplasia. There were no significant correlations between the levels of examined markers with tumor metastasizing, its histological structure, and differentiation degree of endometrial cancer. No differences were observed between examined serological markers in patients with polyps and endometrial hyperplasia of various severities. The examined MMPs and TIMPs cannot be advanced as potential diagnostic markers of endometrial cancer, but they can be used to monitor and prognosticate the disease and to assess effectiveness of the targeted therapy.

MicroRNA and methylation are important epigenetic mechanisms in the pathogenesis of cancer. The role of a group of microRNA hypermethylated genes in the pathogenesis of ovarian cancer was studied and their diagnostic and prognostic potential was evaluated. Studies on a representative sample of 54 ovarian cancer specimens with the use of methyl-specific PCR resulted in detection of five microRNA genes (MIR-9-1, MIR-9-3, MIR-107, MIR-1258, and MIR-130b) methylated in the majority of tumor specimens in comparison with paired specimens of histologically intact tissue (37-57% vs. 4-9%, p<0.01). Methylation of three genes (MIR-9-1, MIR-9-3, and MIR-130b) was significantly (p≤0.05) associated with the parameters of ovarian cancer progress (clinical stage, differentiation degree, tumor size, and presence of metastases). These findings attest to oncosuppressive role of the studied microRNA genes (MIR-9-1, MIR-9-3, MIR-107, MIR-1258, and MIR-130b) in the pathogenesis and progress of ovarian cancer and indicated their prognostic potential.

Methylation of promoter CpG islands and microRNA (miRNA) interactions with mRNAs of target genes are epigenetic mechanisms that play a crucial role in deregulation of gene expression and signaling pathways in tumors. Altered expression of six chromosome 3p genes (RARB(2), SEMA3B, RHOA, GPX1, NKIRAS1, and CHL1) and two miRNA genes (MIR-129-2 and MIR-9-1) was observed in primary clear cell renal cell carcinomas (ccRCCs, 31-48 samples) by RT-PCR and qPCR. Significant downregulation (p < 0.05, Fisher's exact test) was observed for SEMA3B, NKIRAS1, and CHL1; and differential expression, for the other chromosome 3p and miRNA genes. Methylation-specific PCR with primers to RARB(2), SEMA3B, MIR-129-2, and MIR-9-1 showed that their methylation frequency was significantly (p < 0.05, Fisher's exact test) elevated in the ccRCC samples. Significant correlations between promoter methylation and expression were confirmed for SEMA3B and observed for the first time for RARB(2), GPX1, and MIR-129-2 in ccRCC (Spearman's correlation coefficient rs ranging 0.31-0.60, p < 0.05). The MIR-129-2 and RARB(2) methylation frequencies significantly correlated with ccRCC progression. MIR-129-2 methylation correlated with upregulation of RARB(2), RHOA, NKIRAS1, and CHL1 (rs ranging 0.35-0.53, p < 0.05). The findings implicate methylation in regulating RARB(2), SEMA3B, GPX1, and MIR-129-2 and indicate that miR-129-2 and methylation of its gene affect RARB(2), RHOA, NKIRAS1, and CHL1 expression.

Medical histories of 101 urothelial bladder cancer patients were compared with the results of morphological analysis and biomolecular detection of human papilloma viruses (HPV) in the tumor specimens. DNA of HPV16 (the major type of virus responsible for appearance of cervical carcinoma) was detected in 38 specimens, while mRNA of E6 and E7 oncogenes and E7 oncoprotein of HPV16 were observed in 13 specimens. HPV-positive bladder cancer was characterized by higher degree of cell anaplasia than HPV-negative cancer; in the primary bladder tumor, HPV was detected more often than in recurrent bladder cancer. These data attest to involvement of HPV16 in the genesis of bladder cancer. No correlations of HPV status of bladder tumor with patient's sex, age, and invasion into the muscle layer were revealed.

IGF-1, IGF-2, and IGFBP-1,2,3 were assayed in blood serum of patients with malignant ovarian tumors (n=44), borderline ovarian tumors (n=11), and benign ovarian tumors (n=12) as well as in healthy women (n=33). In blood serum of patients with malignant ovarian tumors, the level of IGF-1 was lower and IGFBP-1 was higher than in other groups. In patients with malignant and borderline ovarian tumors, the level of IGFBP-2 was higher than in healthy women and in patients with benign ovarian tumors. There was no correlation between most examined parameters and the clinical and morphological peculiarities of ovarian tumors. The study revealed IGF/IGFBP imbalance in patients with malignant ovarian tumor and showed that IGFBP-2 proved to be a potential diagnostic serological marker w with 90% sensitivity and 90% specificity.

Immunohistochemical method was used to assay for Snail family regulatory proteins of epithelial-mesenchymal transition, their NF-κB coactivator, and the components of VEGF signaling pathway (VEGF and its receptors VEGFR1 and VEGFR2) in 157 specimens of breast tumors. Most tumors did not express SNAI1, while 65% tumors demonstrated mid- or high-level SNAI2 expression. There were significant correlations between the expression of SNAI1, SNAI2, and their NF-κB co-activator. Correlation was also detected between expression of Snail and VEGFR1 protein families in the tumors. In addition, the study revealed tumoral co-expression of SNAI2 and VEGFR2. The data attest to coordinated activation of regulatory proteins of epithelial-mesenchymal transition and the major components of VEGF signaling pathway in breast tumors.

Background: The Arp2/3 complex is required for cell migration and invasion. The Arp2/3 complex and its activators, such as the WAVE complex, are deregulated in diverse cancers. Here we investigate the expression of Arpin, the Arp2/3 inhibitory protein that antagonises the WAVE complex., Methods: We used qRT-PCR and reverse phase protein arrays in a patient cohort with known clinical parameters and outcome, immunofluorescence in breast biopsy cryosections and breast cancer cell lines., Results: Arpin was downregulated at the mRNA and protein levels in mammary carcinoma cells. Arpin mRNA downregulation was associated with poor metastasis-free survival (MFS) on univariate analysis (P=0.022). High expression of the NCKAP1 gene that encodes a WAVE complex subunit was also associated with poor MFS on univariate analysis (P=0.0037) and was mutually exclusive with Arpin low. Arpin low or NCKAP1 high was an independent prognosis factor on multivariate analysis (P=0.0012) and was strongly associated with poor MFS (P=0.000064)., Conclusions: Loss of the Arp2/3 inhibitory protein Arpin produces a similar poor outcome in breast cancer as high expression of the NCKAP1 subunit of the Arp2/3 activatory WAVE complex.

A battery of tests for detection human papillomavirus DNA, mRNA corresponding to viral oncogenes, and viral oncoprotein E7 in cancer bladder urothelium was piloted in 35 samples of bladder cancer. DNA of human papillomavirus type 16 (causes cervical cancer) was found in 16 (46%) samples; E6/E7 oncogene transcript and E7 oncoprotein of human papillomavirus type 16 were detected in 10 and 7 human papillomavirus DNA-positive samples, respectively. These findings attest to association of bladder cancer with human papillomavirus in Russia.

Cell migration requires the generation of branched actin networks that power the protrusion of the plasma membrane in lamellipodia. The actin-related proteins 2 and 3 (Arp2/3) complex is the molecular machine that nucleates these branched actin networks. This machine is activated at the leading edge of migrating cells by Wiskott-Aldrich syndrome protein (WASP)-family verprolin-homologous protein (WAVE, also known as SCAR). The WAVE complex is itself directly activated by the small GTPase Rac, which induces lamellipodia. However, how cells regulate the directionality of migration is poorly understood. Here we identify a new protein, Arpin, that inhibits the Arp2/3 complex in vitro, and show that Rac signalling recruits and activates Arpin at the lamellipodial tip, like WAVE. Consistently, after depletion of the inhibitory Arpin, lamellipodia protrude faster and cells migrate faster. A major role of this inhibitory circuit, however, is to control directional persistence of migration. Indeed, Arpin depletion in both mammalian cells and Dictyostelium discoideum amoeba resulted in straighter trajectories, whereas Arpin microinjection in fish keratocytes, one of the most persistent systems of cell migration, induced these cells to turn. The coexistence of the Rac-Arpin-Arp2/3 inhibitory circuit with the Rac-WAVE-Arp2/3 activatory circuit can account for this conserved role of Arpin in steering cell migration.

MicroRNA regulates gene expression, is involved in many cellular processes, and plays an important role in the development of cancer. The regulation of the expression of miRNA genes can be achieved by methylating their CpG islands, which is shown in different types of tumors. The methylation of miRNA genes in clear cell renal cell carcinoma (CCRCC) has mainly been studied for the miR-9 and miR-34 families. The methylation of six miRNA genes (miR-124a-2, -124a-3, -9-1, -9-3, -34b/c, -129-2) was investigated with the use of representative set of CCRCC samples (46 cases). Methylation of three genes miR-124a-2, -124a-3, and -129-2 was studied in kidney tumors for the first time. Methylation analysis was performed using methyl specific PCR. It is shown that the frequency of methylation of six genes (miR-124a-2, -124a-3, -9-1, -9-3, -34b/c and -129-2) was significantly higher in tumor samples than in samples of histologically normal tissue (P < 3 x 10(-5) by Fisher's exact test). These results suggest the properties of tumor suppressors for the six miRNA genes indicated in CCRCC. We also found correlations between the methylation frequency of some miRNA genes and signs of the progression of CCRCC (tumor size, clinical stage, loss of differentiation, and metastasis).

Malignant cell transformation requires changes in the ability of cells to migrate. The disruption of actin cytoskeleton and intercellular adhesions is an important component of the acquisition of invasive properties in epithelial malignancies. The invasive ability of carcinoma cells is associated with reduced expression of adhesion junction molecules and increased expression of mesenchymal markers, frequently referred to as epithelial-to-mesenchymal transition (EMT). Standard features of the EMT program in cancer cells include fibroblastic phenotype, downregulation of the epithelial marker E-cadherin, induction of Snail-family transcription factors, as well as expression of mesenchymal proteins. We compared the epithelial and mesenchymal marker profiles of nonmalignant HaCaT keratinocytes to the corresponding profiles of cervical carcinoma cell lines C-33A, SiHa, and CaSki. The characteristics of the EMT appeared to be more developed in SiHa and CaSki cervical cancer cells. Further activation of the EMT program in cancer cells was induced by epidermal growth factor. Decreased epithelial marker E-cadherin in CaSki cells was accompanied by increased mesenchymal markers N-cadherin and vimentin. Downregulated expression of E-cadherin in SiHa and CaSki cells was associated with increased expression of Snail transcription factor. Our goal was to study actin reorganization in the EMT process in cell cultures and in tissue. We found that β-cytoplasmic actin structures are disorganized in the cervical cancer cells. The expression of β-cytoplasmic actin was downregulated.

The study objective was an immunohistochemical evaluation of pAkt expression in 81 CIN and microinvasive cervical cancer tissue samples and 10 samples of relatively "normal" cervical epithelium of HPV-infected women. PAkt expression showed significant up-regulation in CIN2, CIN3 and microinvasive cancer in compare to CIN1 and "normal" epithelium. The rate of pAkt- positive cells increased progressively by cervical neoplasia grade advancement reaching 7 +/- 5% in CIN2, 15 +/- 13% in CIN3 and 17 +/- 15% in microinvasive cancer. The rate of pAkt-positive cases in general was 1,7-fold higher in CIN3 (41%) than in CIN2 (24%). pAkt expression in conjunction with other markers may be used in immunohistochemical studies for individual CIN outcome prognosis and prospectively in immunocytochemical tests for CIN grade diagnostics improvement before using invasive methods. To elaborate multicomponent system of markers with their indexation there is a need for further investigations with greater number of cases.

The correlation of morphological mistakes in neoplasia grade verification from visibility of transformation zone (TZ) and patient age was studied in 503 patients with CIN and microinvasive cervical cancer. The square of ectocervix lesion was defined by LeiseCap software in colposcopic working station Leisegang 3MV. The exclusive significance of TZ in HPV-associated cancerogenesis was confirmed clinically. We've established that the neoplasia stage increases with age while lesion extension and TZ visibility decrease dramatically leading to the subsequent decrease in colposcopy sensitivity and adequacy of guided biopsy. The critical age for underdiagnosis of latent lesions seems to be 35 years. The diagnostic and therapeutic value of conization and the large loop excision of the TZ (LLETZ) as the procedures with the optimal TZ excision in patients with visibly unchanged ectocervix are confirmed in cases when CIN 2-3 and microinvasive cancer are suspected.

Enzyme immunoassay showed that the content of matrix metalloproteinases (MMP) 2 and 7 in tumors was higher than in the adjacent histologically intact tissue in 91 and 76% patients with breast cancer, respectively, while MMP-9 levels in the tumor and intact tissue were virtually the same. Serum concentrations of MMP-2 and MMP-7 did not correlate with their levels in the tumors, were within the normal range, and virtually did not decrease after removal of the primary tumor. Serum levels of MMP-9 in patients were significantly lower than in the control and increased after surgery in 85% patients. No clear-cut relationship between the studied parameters and clinical morphological prognostic factors of breast cancer was detected.

Expression of Ki-67, thymidine phosphorylase (TP) and PTEN were assessed in various grades of cervical intraepithelial neoplasia (CIN) in order to evaluate their potentials of predicting the gravity of possible damage to the epithelium as well as pro- or regression of CIN. Ki-67 and TP levels were shown to correlate directly with CIN grade. It was suggested that a small number of cases of Ki-67 and TP expression absence (15%), exclusively in CIN3 samples, be due to imminent progression to invasive cancer. Both separately and in combination, Ki-67 and TP expression indices should be regarded as having a potential as markers for cervical carcinoma diagnosis, grade and clinical course.

Hyperexpression of p16(INK4a) protein is an early marker of cervical cancer. Hyperexpression of INK4a gene encoding this protein at the level of mRNA and p16(INK4a) was detected in tumor cells of some patients with bladder cancer associated with human papilloma virus-16. However, in contrast to cervical cancer, this phenomenon in urothelial carcinomas does not correlate with expression of human papilloma virus-16 oncogenes E6 and E7.

A female patient with recurrent bladder cancer underwent complex examination. The primary tumor removed in 2004 showed human papillomavirus (HPV) 16 DNA, mRNA corresponding to HPV16 oncogene E7, as well as HPV16 protein E7. The patient is a smoker who has been working at a chemical factory for over 20 years. During tumor recurrence in 2009, there was no DNA of high-risk HPV types in the cancer cells. HPV16 E7protein and cellular p 16(INK4alpha), an indicator of HPV-induced carcinogenesis, were not found. Colposcopy revealed no precancerous changes in the epithelium of the cervix uteri. The cervical epitheliocytes contained no high-risk HPV DNA, E7 and p16(INK4alpha) proteins. It seems expedient to continue in vitro studies of the possible role of HPV in urothelial carcinogenesis on an experimental model.

The most common forms of luminal breast cancer (BC) were compared with basal-like and Her2/neu3+ BC. Their primary classification was based on morphological diagnosis and the expression of estrogen receptor (ER), progesterone receptor (PR), and Her2/neu receptors. Monoclonal antibodies to actins and keratins were used for the study. Basal-like BC cells (ER/PR/Her2/ neu-) were regularly stained with antibodies to basal keratins 5/6 and 17, smooth muscle alpha-actin, and p63. Luminal keratin 8 staining was reduced. The solid regions had beta-actin staining with disappearance in the scirrhous component. beta-actin and basal keratins were also observed in metaplastic BC with ER/PR/Her2/neu3+. Immunomorphology using cytoskeletal markers along with the expression of steroid hormone and Her2/neu receptors may be used in the diagnosis of basal-like forms of BC.

Earlier in some small cell lung (SCLC) and non-small cell lung carcinoma (NSCLC) cell lines, methylation of CpG-island was found in the SEMA3B region, which belongs to the first intron according to the NCBI data base (Build 36). The aim of this work was to study methylation of two SEMA3B CpG-islands: promoter and intronic in clear cell renal cell carcinoma (RCC). Using methyl specific PCR and bisulfite sequencing, it was shown for the first time that promoter CpG-island was methylated in RCC with high frequency 56% (34/61), and intronic CpG-island - with somewhat lower frequency 35% (17/48). Significant reverse correlation was estimated between mRNA level decrease and methylation of promoter CpG-island in RCC for the first time (P < or = 0.05 by Fisher's exact test), no correlation was determined for intronic CpG-island. This result suggested that methylation of promoter CpG-island contributed into inactivation of SEMA3B gene-suppressor in RCC.

The initial levels of soluble Fas antigen (sFas), leptin, and vascular endothelium growth factor (VEGF) were measured in the sera of 100 patients with ovarian cancer and benign tumors and in 60 healthy women aged 28-65 years. Serum levels of sFas and VEGF were elevated in the total group of patients with ovarian tumors, while leptin levels were the same as in healthy women. The studied parameters did not depend on the age of patients and healthy women. The levels of sFas and leptin were virtually the same in benign and malignant ovarian tumors, while VEGF concentration was higher in patients with ovarian cancer. The mean serum levels of sFas, VEGF, and leptin in patients with poorly and moderately differentiated serous ovarian cancer were 2-fold higher than in well-differentiated tumors (p<0.05), while serum concentrations of sFas and leptin increased with the disease stage progress in patients with ovarian cancer (p<0.05). According to the data of unifactorial analysis, the increase in serum levels of sFas and VEGF in ovarian cancer patients correlated with short duration of the relapse-free period. Multifactorial analysis showed that the disease stage (p=0.006), presence of ascites (p=0.03), VEGF concentration (p=0.02), and the sFas/leptin coefficient (p=0.045) are highly significant independent factors for predicting the relapse-free survival of patients with serous ovarian cancer.

The short arm of chromosome 3 (3p) contains several critical regions harboring the set of genes with tumor suppressor activities. The RASSF1A gene (LUCA region, 3p21.31) shows various functions which can be associated with tumorigenesis. Among 3p genes this gene can be most frequently methylated in epithelial tumors of various locations. Here two independent methods (methyl-specific PCR and methyl-sensitive restriction analysis) were used to show significant correlation of methylation level of promoter region of this gene with grade and clinical stage of clear cell renal cell carcinoma (RCC) for the first time. Analysis of 23 polymorphic markers of 3p using the representative set of samples (80 cases RCC), described clinically and histological, permitted to reveal significant correlation between frequency of allelic alterations in some critical regions of 3p (LUCA and AP20) and RCC progression, as distinct from the whole 3p. These data suggest that methylation of promoter region of the RASSF1A gene is associated with RCC progression, and besides, structure-functional alterations in other 3p genes can be also related with RCC progression. In addition, significant correlation between RASSF1A methylation events and allelic losses in the close polymorphic marker was shown here, pointing to the role of "two hit" model for this tumor-suppressor gene inactivation in RCC.

The data of preoperative diagnosis and morphological examination were compared for 144 patients with prostatic carcinoma T1-4N0-XM0 subjected to radical prostatectomy in 1997-2007. In assessment of prostatic capsule invasion, sensitivity of the rectal examination was 21.7%, specificity--89.8%, diagnostic efficacy--68.1%, PPV--50.0%, NPV--70.9%, AUC under ROC curve--0.558 +/- 0.053 (p = 0.348); sensitivity of transrectal ultrasonic investigation--21.7%, specificity--89.8%, diagnostic efficacy--68.8%, PPV--52.6%, NPV--71.2%, AUC under ROC curve--0.563 +/- 0.053 (p = 0.211). Factors of a poor prognosis of prostatic capsule invasion were PSA > 10 ng/ml (p = 0.028) and Gleason score > 7 (p = 0.052). Combined use of these two parameters raises quality of preoperative assessment of category T [sensitivity--80.0%, specificity--55.1%, diagnostic efficacy--56.3%, PPV--80.4%, NPV--44.9%, AUC under ROC curve--0.624 +/- 0.049 (p = 0.017)]. Sensitivity of clinical assessment of N category was 11.1% in 100% specificity, 94.4% diagnostic efficacy, 100% PPV, 94.4% NPV, 0.556 +/- 0.107 (p = 0.577) AUC under ROC curve. A single significant prognostic factor of pN+ category was PSA > 10 ng/ml (p = 0.014). Sensitivity of histological examination of biopsy material in relation to true Gleason's parameter (< 7 or > 7) was 59.4%, specificity 89.3%, diagnostic efficacy 82.6%, PPV 61.3%, NPV 88.5%, AUC under ROC curve 0.743 +/- 0.056 (p < 0.0001). Thus, combined use of a baseline PSA concentration with a borderline value > 10 ng/ml and biopsy Gleason score > 7 raises quality of preoperative evaluation of extraprostatic tumor extension and condition of regional lymph nodes.

Oncoprotein E7 HPV16 was detected by immunohistochemical staining with specific polyclonal antiserum [Fiedler et al., 2004] in 7 out of the 24 (29.2%) studied bladder cancer specimens. The result is in good agreement with the hypothesis that HPVs take part in the carcinogenesis of the urothelium. However, some of the observations made seem rather hard to be interpreted at present. The latter include the detection of E7 HPV16 in a small number of cancer cells in a few bladder cancer specimens being examined; the presence of this protein in the cytoplasm, rather in the cancer cell nuclei, and its detection in some morphologically normal bladder urothelial specimens from non-cancer patients. Thus, the hypothesis that HPVs are implicated in the carcinogenesis of the bladder urothelium deserves further verification.

The distribution of beta- and gamma-cytoplasmic actins was compared in the normal cells and dysplastic malignant breast epithelial cells. In the normal luminal epithelium, beta- and gamma-cytoplasmic actins were located in different cell compartments: gamma-actin was more expressed in the apical parts of epithelial cells while beta-actin was in their basolateral domain. Polarized distribution of actinic isoforms was partially preserved in the papillomas and fibroadenomas; a more pronounced coexpression of isoforms was detected in the dysplastic proliferates. In ductal and lobular in situ carcinoma cells, gamma-actin filamentous structures were absent while the gamma-cytoplasmic actin network throughout the cytoplasm was increased. It is generally accepted that the enhanced motility of cancer cells as to the nonmalignant situation is crucial in the process of cancer invasion. The authors' findings suggest that specific monoclonal antibodies to beta- and gamma-cytoplasmic actins may be used as supplementary markers that can differentiate benign and malignant breast neoplasms.

Allelic imbalances (AI) of polymorphic markers at the short arm of chromosome 3 (3p) were mapped using DNA samples of renal cell carcinoma (RCC, 80 cases), breast carcinoma (BC, 95 cases), and epithelial ovarian cancer (EOC, 50 cases) at the same dense panel of markers (up to 24 loci). Six regions with the increased AI frequency (versus the average values determined for all the analyzed 3p markers) at RCC, BC or EOC were found in 3p chromosome. Four 3p regions presumably contain suppressor genes of tumor growth (TSG) observed in the epithelial tumors of various types. Region between D3S2409 and D3S3667 markers in the 3q21.31 region was identified in this study for the first time. The AI peak in D3S2409-D3S3667 region was statistically significant (P < 0.001, according to Fisher) when representative sample of 95 BC patients was analyzed. The data on increased frequency of polymorphic marker allele amplification suggest that the D3S2409-D3S3667 region contains both putative TSG and protooncogenes.

Expression of the STAT1 gene belonging to the group of interferon-regulated genes was analyzed in cervical tumors and cell lines harboring the genome of human papilloma viruses (HPV) of so-called high risk group. Expression of this gene in invasive carcinomas was maintained on a definite level that was not significantly distinct from that in adjacent normal (control) tissue. Tumors from different patients differ from each other by expression level of the STAT1 gene. These variations can be attributed to the heterogeneity of tumor cell population and different ratio between normal and tumor cells, as well as to putative persistence of intra-individual variability of STAT1 expression in normal cell population. It was demonstrated that viral genome status (episomal or integrative) did not influence STAT1 gene transcription. In conclusion, these data demonstrate that the STAT1 gene is expressed in an individual and specific manner both in HPV-positive cervical tumors and cell lines harboring transforming genes of these viruses.

RHOA protein, a member of small GTPases family, is implicated in cell morphogenesis, adhesion, and in cell cycle regulation. RHOA gene (3p21.31) exhibits cell transformation activity, and therefore gene is considered as a potential oncogene. The aim of this study was to investigate RHOA transcription and copy number changes in three epithelial tumors (breast, renal cell and epithelial ovarian carcinomas, 45 tumor/normal pairs altogether). EII, HhaI, AciI n Bsh1236I). Hypomethylation of the RHOA promoter region in tumor DNA was observed two times more frequently than increased methylation. Moreover, all (15) cancer cases with hypomethylation of the RHOA gene showed a 2-10 fold increased expression of RHOA. It was concluded that gene copy multiplication and demethylation of the RHOA promoter region can contribute to transcription activation of this gene in epithelial tumors.

An immunomorphological study of 30 breast carcinomas was conducted. In accordance with the histological characteristics they had been previously diagnosed as lobular infiltrative carcinomas (LIC). We have studied E-cadherin cell-cell adhesion molecules and also epithelial markers and basement membranes. We have analyzed the cryostate and paraffin sections of in situ structures, invasion components and metastases. E-cadherin staining was negative, positive or mixed. The E-cadherin negative group consisted of 17 carcinomas of classic LIC accompanied with dyshesive invasion. The E-cadherin positive group consisted of 6 carcinomas. According to our results this diagnosis was changed to ductal infiltrative carcinoma. The E-cadherin mixed group consisted of 7 special type carcinomas with double lobular-ductal differentiation, which had in situ structures of negative or positive cells. In these cases in situ structures consisting of both E-cadherin negative and positive parts were also observed. It is shown that E-cadherin expression can be used in doubtful cases in order to differentiate between the lobular and ductal forms of breast carcinoma.

Lobular carcinoma ranks second among most frequent tumors of the breast. Metastases to the lymph nodes and distant organs are one of the most important prognostic factors. 60% regional lymph node metastases were morphologically diagnosed during surgical removal of the tumor. Distant metastases rate was 1% at the beginning of the treatment and reached 10% within 5-year follow-up. Cases of secondary uterus and epiploon lobular carcinoma are rare and tumor location in the breast should be proved.

Hyperexpression of epidermal growth factor receptor (EGFR) is often identified as unfavorable prognosis for different epithelial cancers. The study was concerned with an attempt of establishing a relationship between EGFR expression, on the one hand, and patient's clinico-morphological status, prognosis and efficacy of chemotherapy for stage III-IV serous ovarian carcinoma, on the other. EGFR hyperexpression predominated in advanced aggressive tumors and involved a significantly shorter period preceding tumor progression. Similarly, overall survival median in patients with EGFR hyperexpression (21+/-4 months) appeared lower than without it (42+/-8 months). In serous ovarian carcinoma stage III-IV, EGFR hyperexpression should be considered sufficient for prognosis of chemotherapy efficacy, pre-progression time and survival.

Methylation of the promoter CpG-islands of the candidate tumor suppressor gene RASSF1A (3p21.31) was studied in primary tumors of kidney, breast and ovary (172 cases). Methylation-specific PCR (MSP) and methyl-sensitive restriction endonuclease digestion followed by PCR (MSRA) were applied. Statistically significant correlation (P << 10(-6)) was shown for the results of the MSP and MSRA, and the data of bisulfite sequencing reported earlier. The frequency of RASSF1A methylation according to MSP and MSRA was 86% (25/29) and 94% (50/53) in renal cell carcinoma (RCC) and 64% (18/28) and 78% (32/41)--in breast carcinoma (BC) samples, and 59% (17/29) and 73% (33/45) in ovarian epithelial tumors (OET), respectively. The use of several methyl-sensitive restriction enzymes (HpaII, HhaI, Bsh12361, AciI) enhanced the sensitivity of MSRA and allowed to analyze methylation status of 18 CpG-pairs in the RASSF1A CpG-island. Density of methylation of the RASSF1A CpG-island was 72% (644/900) in RCC, 63% (361/576) in BC, and 58% (346/594) in OET samples (18 CpG-pairs multiplied to the number of samples shown methylation were assumed as 100%). The RASSF1A gene methylation was also observed in samples of morphologically normal tissues adjacent to corresponding tumors (11-35%), but it was not detected in blood DNAs of healthy donors (0/15). The RASSF1A methylation frequency did not show significant correlation to tumor stage, grade and metastases (P = 0.3-1.0). The RASSF1A gene methylation was observed more frequently than other investigated aberrations--hemi- and homozygous deletions inside or around this gene. These observations are consistent with the hypothesis that the RASSF1A gene methylation is an early event in the carcinogenesis and one of the dominant ways of its inactivation.

We carried out a retrospective immunohistochemical study of Ki-67, PCNA, Bcl-2, BAX, BclX, and VEGF expression in tumors of two groups of breast cancer patients with favorable and unfavorable course of the disease. Considerably enhanced VEGF expression was detected in tumors of patients with early relapses of breast cancer. High VEGF expression was paralleled by high level of Ki-67 and PCNA expression in tumors. It can be hypothesized that expression of VEGF, Ki-67, and PCNA in primary tumor can be used for predicting the course of breast cancer or detecting the patients at a high risk of early relapses.

Data are described on a retrospective immunohistochemical study of the Ki-67, PCNA, Bcl-2, BAX, BclX and VEGF expression in tumors of two groups of patients with breast cancer (BC) and with the favorable and unfavorable clinical courses. A reliably high level of VEGF expression was detected in tumors of patients with an early BC relapse. Apart from VEGF, higher levels of the Ki-67 and PCNA expression were registered in tumors of the discussed patient's category. It can be suggested that the expression parameters of VEGF, Ki-67 and PCNA in the primary tumor can be made use of in evaluating the BD prognostications or describing the features of high-risk groups with a high probability of early disease relapses.

Expression of the markers was studied immunocytochemically on cytological preparations of breast tumor, ploidy was assessed by flow cytofluorimetry. The material was divided into 2 groups: diploid--41.4% cases and aneuploid--58.6%. Hyperexpression of Her-2/neu in the first group was observed in 54.5%, in the 2nd group in 48.6% cases. Expression of Ki-67 in 63.4% and in 68.9%, respectively. Tumours with high proliferative activity were numerous in aneuploid tissues, and in diploid tumours moderate activity prevailed (p = 0.006). Direct correlation between the markers was observed, high expression of Ki-67 more frequently was associated with positive expression of Her-2/neu. Thus, aneuploid tumours with high proliferative activity and hyperexpression of Her-2/neu are more aggressive tumours of a large size.

The tumor of the mandible in this case was first qualified as ameloblastoma. Although in the last WHO classification clear-cell odontogenic tumour is considered to be a benign tumour, repeated recurrences and distant metastases described in the literature prove its malignant character.

Distribution of cell-cell adhesion molecules of E-cadherin (EC) and beta-catenin (BC) infiltrating breast carcinoma (BC) and markers of epithelium, basal membrane has been studied by immunofluorescence in 50 infiltrating BC tissue samples including 17 ductal carcinomas, 23 lobular carcinomas and 10 combined carcinomas. Two types of EC-BC structure alterations were found: homogeneity disorder accompanied by appearance of local thickening and gaps or complete lack of these structures. Alterations of type 1 were found in all the analyzed samples of ductal and combined carcinomas and partially in lobular carcinomas, in in situ structures and in invasion components as well. Alterations of type 2 were found in 16 cases of lobular carcinomas of 23 examined. Our data suggest the existence of two groups of lobular BC: carcinomas with partial depletion of EC-BC structures and carcinomas which lost these structures. Further investigations are needed to evaluate clinical importance of the difference found.

The study of molecular-biological markers for prediction of recurrence-free survival in breast cancer stage I-IIA demonstrates that high expression of thymidilate synthetase and high maximal density of microvessels are prognostically effective and that the prognosis is influenced by expression of both Bcl-2 and Bax. Low recurrence-free survival was observed in Bcl-2-/Bax patients (31%, median 44 months) while such survival was high in Bcl-2+/Bax patients (86%, median was not reached). The findings can be used for prognostication of a breast cancer clinical course.

The study of different epithelial antigen expression has been performed in 183 breast cancers. In 73 patients regional lymph nodes were studied as well. Panepithelial antigen Egp-34 (Mab HEA-125) was expressed in 100% of primary tumors and metastases. Antigen MUC-1 (Mab ICO-25) was identified in 93% of breast cancers. Monomorphic type of expression in tumor cells was typical for Egp-34, MUC-1 being in certain cases expressed as a proportion of cells. Additional immunohistochemical study of regional lymph nodes with Mab to Egp34 and MUC-1 provides a 10% increase in the rate of breast cancer metastases detection compared to histological examination alone. The carcinoembryonic antigen (CEA) was useful in identification of metastases only in CEA-positive breast cancers.

The detection of the biological parameters of the tumor before the treatment beginning becomes of more importance. The present study aimed to carry out the comparative analysis of the molecular markers expression (P53, Ki-67, Her-2/neu, Bcl-2, Bax, ER, FasL and CD95) at the cytologic and the correspondent histologic samples. The 18 tissue samples of the breast cancer were investigated. The immunocytochemical and the immunohistochemical methods of the molecular markers determination were used. Our study showed the correlation between two methods and the possibility of the use of the immunocytochemical staining as routine method of the molecular markers expression determination.

An immunomorphological study of 52 samples of normal thyroid tissue, 10 adenomas, 42 well differentiated carcinomas and 2 metastases has been performed using markers of the epithelium, extracellular matrix and angiogenesis. Basal keratin N 17 of the compound epithelium occurred only in malignant tumors. The highest number of the keratin N 17-positive cells was registered in sclerosing A-cell cancer, medullary and mixed C-cell cancer which is very aggressive. These cells locate in the foci of proliferation of tumor structures and in the regions of infiltrative growth. Multiple defects of basal membranes and prevalence of capillaries marked by antibodies to CD-31 over those identified by the antibodies to factor VIII were found in the areas of strong expression of keratin 17. Thus, the above markers detect foci of aggressive growth even in well differentiated thyroid carcinomas. This fact may be of prognostic value in oncomorphological practice.

The outcomes of treatment were studied in 190 patients with breast cancer who had undergone radical mastectomy with telethoracoscopic parasternal lymphodissection. An association of this parameter with the sizes of a primary tumor, with the extent of damage to the axillary lymph collector, with the histological pattern of a tumor, and with the site of a primary focus was analyzed. Radical removal of the parasternal lymphatic chain was proved by scintimammography in the pre- and postoperative periods. The high diagnostic value of telethoracoscopic parasternal lymphodissection was shown as a miniinvasive axilliary of radical mastectomy for breast cancer, which makes it possible to establish a stage of the disease reliably and to plan chemoradiation treatment adequately.

Allelic deletions along the short arm of human chromosome 3 were mapped in 57 pairs of DNA samples from tumor and normal tissue of renal carcinoma patients in order to locate potential tumor suppressor genes. Twenty highly polymorphic microsatellite markers were used for deletion mapping. Allelic deletions were found in most of the samples (91%). Extended terminal deletions (56%) prevailed over shorter internal and multiple deletions and dominated (65%) in the most aggressive histopathological kidney cancer subtype, clear-cell carcinoma. Frequency analysis of loss of heterozygosity allowed detection of the human chromosome 3 regions most essential for renal carcinomas: the region adjacent to the gene VHL (3p26-p25), the region of homozygous deletions AP20 (3p22-p21.33), and a new region between markers D3S2420 and D3S2409 (3p21.31, 2.2 Mbp).

The investigation involved 30 patients with locally advanced breast cancer (T3-4N1-2M0). Combination therapy comprised two courses: carboplatin 300 mg/m2, i.v., dropwise, on day 1; doxorubicin 30 mg/m2, i.v., bolus-flow, on days 1 and 8; 5-fluorouracil 350 mg/m2, i.v., bolus-flow, on days 1 and 8, and irradiation of the breast and regional metastasis area (single target dose--2 Gy, total target dose--40 Gy). Overall clinical response was 96.7% (29/30), mammography-wise--83.3% (25/30). All patients were found operable and radical mastectomy was performed in 25. Therapeutic effect stage III-IV was histologically confirmed in 40% (25/30), stage I-II--60% (15/25). Median overall and recurrence-free survival was not reached within 36 months in 24/30, relapse-free survival was been reported in 16/24 (66.6%), tumor progression--8/24 (33.4%). Three-year; host-mastectomy recurrence-free survival--68.8 +/- 16.0%.

Epidermal growth factor receptors (EGFR) were identified in the membrane fraction of tumor in 21 out of 58 (36%) endometrial cancer patients. EGFR ligands--EGF-like peptides--were found in tumor cytosols in 36% of patients. However, EGFR and EGF-like peptides were observed in 7% of tumors only; there were no receptors or ligands in 39% of patients. Both EGFR and steroid receptors were expressed in 20 tumor patients. The frequency of EGFR expression in endometrial carcinomas was practically identical to that of normal endometrium at the proliferative stage of menstrual cycle and half that of hyperplastic endometrium. Postmenopausal patients tended to show a decrease in hormonal sensitivity of carcinoma and EGFR expression frequency rate and a lower degree of endometrial carcinoma cell differentiation. Also, a significant rise in the EGFR expression frequency and mean concentration was found in conjunction with a deeper invasion of tumor into the myometrium.

Phospatidylinositol 3-kinase (PI 3-kinase) expression was analysed by Western blotting with monoclonal antibodies to the p85 subunit in a series of tumour and adjacent mammary gland samples collected at surgery from 33 breast cancer patients. Seventy-nine percent of the investigated pairs of the samples were characterised by an increased level of PI 3-kinase in the tumour in comparison with the adjacent mammary gland. PI 3-kinase activation was not associated with tumour steroid receptor status, histologic grade and other clinico-morphological characteristics. Furthermore, immunoblotting of epidermal growth factor receptor (EGFR) in the tumours with increased PI 3-kinase and corresponding adjacent tissues revealed no association between EGFR and PI 3-kinase activation. Thus, increased PI 3-kinase expression appears to be a widespread feature of breast cancer not associated with the main biological markers of its prognosis and hormone sensitivity.

A rare case of aggressive ovarian tumor in a 38-year-old female is described. Cytological diagnosis of the intraoperative smears was difficult because of the presence of the epithelial and non-epithelial components. The latter consisted of two types: spindle cells and roundish undifferentiated cells with multinuclear forms. Histologically, it was a malignant mesodermal (Mullerian) mixed tumor. Round mesenchymal cells expressed desmin and vimentin while spindle cells expressed myoglobin. Non-epithelial component was verified as leiomyosarcoma and rhabdomyosarcoma. Cells of the epithelial component expressed cytokeratins and epithelial membrane antigen.