Natural and chemotherapy-induced clonal evolution of tumors

Evolution and natural selection of tumoral clones in the process of transformation and the following carcinogenesis can be called natural clonal evolution. Its main driving factors are internal: genetic instability initiated by driver mutations and microenvironment, which enables selective pressure while forming the environment for cell transformation and their survival. We present our overview of contemporary research dealing with mechanisms of carcinogenesis in different localizations from precancerous pathologies to metastasis and relapse. It shows that natural clonal evolution establishes intratumoral heterogeneity and enables tumor progression. Tumors of monoclonal origin are of low-level intratumoral heterogeneity in the initial stages, and this increases with the size of the tumor. Tumors of polyclonal origin are of extremely high-level intratumoral heterogeneity in the initial stages and become more homogeneous when larger due to clonal expansion. In cases of chemotherapy-induced clonal evolution of a tumor, chemotherapy becomes the leading factor in treatment. The latest research shows that the impact of chemotherapy can radically increase the speed of clonal evolution and lead to new malignant and resistant clones that cause tumor metastasis. Another option of chemotherapy-induced clonal evolution is formation of a new dominant clone from a clone that was minor in the initial tumor and obtained free space due to elimination of sensitive clones by chemotherapy. As a result, in ~20% of cases, chemotherapy can stimulate metastasis and relapse of tumors due to clonal evolution. The conclusion of the overview formulates approaches to tumor treatment based on clonal evolution: in particular, precision therapy, prediction of metastasis stimulation in patients treated with chemotherapy, methods of genetic evaluation of chemotherapy efficiency and clonal-oriented treatment, and approaches to manipulating the clonal evolution of tumors are presented

Copy number aberrations landscape of a breast tumor, connection with the efficiency of neoadjuvant chemotherapy

The research involved 80 patients diagnosed with breast cancer (BC). Each patient had their tumor biopsy material sampled before their treatment. We studied the tumor tissue using the CytoScan HD Array (Affymetrix, USA) microarray to evaluate the CNA landscape. We studied the frequency of segmental and numerical CNA occurrence, their association with the efficiency of neoadjuvant chemotherapy (NAC). We found that the biggest number of amplifications (with frequency over 60%) were found on in the following locuses; 1q32.1 1q32.3, 1q42.13, 1q42.2, 1q43. The biggest frequency of deletions (more than in 58% of the patients) was found in these locuses: 16q21, 16q23.2, 16q23.3, 17p12, 17p13.1. However, we found the locuses with full absence of segmental chromosome anomalies. We observed trisomy most frequently in the 7, 8, 12, and 17 chromosomes, and monosomy in the 3, 4, 9, 11, 18, and X-chromosomes. We demonstrated the connection between the high frequency of cytobands with CNA in the patients’ tumors and the efficiency of NAC. We also identified the cytobands, whose CNA are linked to the response to NAC

Microarray research of allelic imbalance in breast cancers during neoadjuvant chemotherapy

For single nucleotide polymorphism – SNP (SNP – Single Nucleotide Polymorphism) is characterized by phenomenon of allelic imbalance (AI). The phenomenon of allelic imbalanced (AI) is typical of many genes in different malignancies. Allelic imbalanced may result allelic deletions (loss of one copy of the locus) or amplification of one allele, resulting in only a single allelic variant of SNP is determined in the tumor in the PCR. Phenomenon of AI in tumors of breast cancer (BC) is considered in one or more genes. The phenomenon of AI during chemotherapy (CT) and especially in the longer format of the gene had not previously been studied. Thus, the aim of this work was to study microarray imbalanced allele in mammary tumors during neoadjuvant chemotherapy (NCT). Materials and methods: The study included 26 breast cancer patients with stage IIA – IIIC. The patients in the neoadjuvant mode received 2–4 courses of chemotherapy regimens FAC or CAX. DNA from 26 paired samples before treatment and operational samples were isolated by dialing QIAampDNA miniKit (Qiagen, Germany). Microarray analysis was performed on DNA chips of high density company Affymetrix (USA) CytoScanTM HD Array, which contains more than 750 thousand SNP. Microarray analysis was performed on SNP genotypes DNA tumor tissue before and after treatment for each patient and recorded as change occurring allelic imbalanced tumor tissue genotype (AA > AB > AB AA BB > AB > AB BB) during therapy. Results: The frequency of the AI in breast tumor during NCT was highly variable (within 0.9–66.5%) of the studied SNP (6850 – 497,979 SNP). For each patient, frequency shift genotype (homozygous in heterozygous genotype, and vice versa) was calculated as a percentage of all the shifts. Changes in the wild or mutant heterozygous genotypes (AA or BB > AB) were combined into one group; the second group was the sum of the change in the heterozygous genotype homozygous wild genotype or the mutant (AB > AA or BB). We have found that the direction of the AI was significantly associated effect of NCT. In the group of patients with partial regression, the direction of AI change from homozygous to heterozygous genotype often occurs (AA or BB > AB) (9/14, 64%), whereas patients with no response to the NCT (with stabilization or progression) have the opposite effect. All these patients (12/12 cases) have the direction of the change of AI from heterozygous genotype to homozygous (AA or AB > BB) (p = 0.00071). AI during chemotherapy at the level of the marked tendency (Log-rank test, p = 0.062) is associated with 5-year metastasis-free survival. Low metastasis-free survival rate is observed in patients with AI in the direction of the change from the heterozygous to homozygous genotype, while 100% survival is noted in patients with change from homozygous to heterozygous genotypes, and this imbalanced t allele is a favorable prognostic factor. Conclusion: Allelic imbalanced in breast tumor during NCT phenomenon is massive and may affect up to 67% of SNP. AI may occur in the direction of change from homozygous to heterozygous genotype, and it is associated with a good response to treatment and 100% metastasis-free survival. Apparently, this can be explained by the fact that the change from homozygous genotypes to heterozygous occur due to partial destruction of tumor cells by chemotherapy, resulting in the increase of stromal elements. In contrast, AI in the direction change from heterozygous to homozygous genotype during NCT is associated with no response to chemotherapy due to metastasis and occurring of new mutant clones in the tumor

HPV-negative cervical cancer: a distinct type of the uterine cervix with poor prognosis

The real-time PCR method was used to study cervical scrapings from 116 patients with stag

Disease-free and overall survival of patients diagnosed with HPV-associated or HPV-negative cervical cancer

The real-time PCR method is used to study scrapings of cervical epithelium and outer portion of the cervix in 116 patients aged 24–79 years with stage I–IV primary cervical cancer. The comprehensive survey included colposcopy, cytological and histological analysis, detection and genotyping of high-risk human papillomavirus. In 84 patients (72.4%) the presence of human papillomavirus (HPV) of high carcinogenic risk (HCR) is found, in 32 patients (27.6%) the presence of the virus has not been inspected in the tumor. A significant decrease in the survival rate as well as the prevalence of the worst prognosis for patients with HPV-negative cervical cancer are shown

Предиктивная и прогностическая значимость явления потери гетерозиготности в генах ABC-транспортеров в опухоли молочной железы

ABC-transporter family genes have been well studied and their involvement in the development of drug resistance has been assessed. The presence of aberrant conditions in these genes can affect the treatment and prognosis of the disease. Loss of heterozygosity (LOH) is one of these conditions; it is a common event in cancer development. Therefore, the aim of this study was to investigate the relationship between LOH in ABC transporter genes in breast cancer and response to chemotherapy and disease prognosis. Material and Methods. A total of 130 breast cancer patients were included in the study. Microarray analysis was performed on Affymetrix CytoScan™ HD Array high-density DNA chips to assess LOH status. Chromosome Analysis Suite 4.1 software (Affymetrix, USA) was used to process microarray results. Results. Forty-nine ABC transporter genes were evaluated for LOH. The frequency of LOH ranged from 6.9 % to 90 %. An association analysis identified two genes: ABCG5 and ABCG8, in which the presence of LOH was associated with a lack of objective response to neoadjuvant chemotherapy. The presence of LOH in the ABCA5, ABCA6, ABCA8, ABCA9, ABCA10 and ABCC3 genes was associated with high rates of metastasis-free survival (log-rank test, p<0.04). conclusion. The presence of loss of heterozygosity in the ABC transporter genes was found to have no significant effect on the response to chemotherapy. However, a high prognostic potential of ABCA family genes was found

Genome-wide association study of loss of heterozygosity and metastasis-free survival in breast cancer patients

One of the factors providing the diversity and heterogeneity of malignant tumors, particularly breast cancer, are genetic variations, due to gene polymorphism, and, especially, the phenomenon of loss of heterozygosity (LOH). It has been shown that LOH in some genes could be a good prognostic marker. Aim: To perform genome-wide study on LOH in association with metastasis free survival in breast cancer. Materials and Methods: The study involved 68 patients with breast cancer. LOH status was detected by microarray analysis, using a high density DNA-chip CytoScanTM HD Array (Affymetrix, USA). The Chromosome Analysis Suite 3.1 (Affymetrix, USA) software was used for result processing. Results: 13,815 genes were examined, in order to detect LOH. The frequency of LOH varied from 0% to 63%. The association analysis identified four genes: EDA2R, PGK1, TAF9B and CYSLTR1 that demonstrated the presence of LOH associated with metastasis-free survival (log-rank test, p < 0.03). Conclusions: The presence of LOH in EDA2R, TAF9B, and CYSLTR1 genes is associated with metastasis-free survival in breast cancer patients, indicating their potential value as prognostic markers

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

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), TOP2a, 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