Biomedical subjects

Anna Lapuk

Publications and source records attributed to Anna Lapuk.

8 recordsLinked to original sources

PubMed2006

Organic cation transporters are determinants of oxaliplatin cytotoxicity.

Although the platinum-based anticancer drugs cisplatin, carboplatin, and oxaliplatin have similar DNA-binding properties, only oxaliplatin is active against colorectal tumors. The mechanisms for this tumor specificity of platinum-based compounds are poorly understood but could be related to differences in uptake. This study shows that the human organic cation transporters (OCT) 1 and 2 (SLC22A1 and SLC22A2) markedly increase oxaliplatin, but not cisplatin or carboplatin, accumulation and cytotoxicity in transfected cells, indicating that oxaliplatin is an excellent substrate of these transporters. The cytotoxicity of oxaliplatin was greater than that of cisplatin in six colon cancer cell lines [mean +/- SE of IC(50) in the six cell lines, 3.9 +/- 1.4 micromol/L (oxaliplatin) versus 11 +/- 2.0 micromol/L (cisplatin)] but was reduced by an OCT inhibitor, cimetidine, to a level similar to, or even lower than that of, cisplatin (29 +/- 11 micromol/L for oxaliplatin versus 19 +/- 4.3 micromol/L for cisplatin). Structure-activity studies indicated that organic functionalities on nonleaving groups coordinated to platinum are critical for selective uptake by OCTs. These results indicate that OCT1 and OCT2 are major determinants of the anticancer activity of oxaliplatin and may contribute to its antitumor specificity. They also strongly suggest that expression of OCTs in tumors should be investigated as markers for selecting specific platinum-based therapies in individual patients. The development of new anticancer drugs, specifically targeted to OCTs, represents a novel strategy for targeted drug therapy. The results of the present structure-activity studies indicate specific tactics for realizing this goal.

Animals↗

PubMed2006

Decoding the fine-scale structure of a breast cancer genome and transcriptome.

A comprehensive understanding of cancer is predicated upon knowledge of the structure of malignant genomes underlying its many variant forms and the molecular mechanisms giving rise to them. It is well established that solid tumor genomes accumulate a large number of genome rearrangements during tumorigenesis. End Sequence Profiling (ESP) maps and clones genome breakpoints associated with all types of genome rearrangements elucidating the structural organization of tumor genomes. Here we extend the ESP methodology in several directions using the breast cancer cell line MCF-7. First, targeted ESP is applied to multiple amplified loci, revealing a complex process of rearrangement and co-amplification in these regions reminiscent of breakage/fusion/bridge cycles. Second, genome breakpoints identified by ESP are confirmed using a combination of DNA sequencing and PCR. Third, in vitro functional studies assign biological function to a rearranged tumor BAC clone, demonstrating that it encodes anti-apoptotic activity. Finally, ESP is extended to the transcriptome identifying four novel fusion transcripts and providing evidence that expression of fusion genes may be common in tumors. These results demonstrate the distinct advantages of ESP including: (1) the ability to detect all types of rearrangements and copy number changes; (2) straightforward integration of ESP data with the annotated genome sequence; (3) immortalization of the genome; (4) ability to generate tumor-specific reagents for in vitro and in vivo functional studies. Given these properties, ESP could play an important role in a tumor genome project.

Breast Neoplasms↗

PubMed2006

A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes.

Recent studies suggest that thousands of genes may contribute to breast cancer pathophysiologies when deregulated by genomic or epigenomic events. Here, we describe a model "system" to appraise the functional contributions of these genes to breast cancer subsets. In general, the recurrent genomic and transcriptional characteristics of 51 breast cancer cell lines mirror those of 145 primary breast tumors, although some significant differences are documented. The cell lines that comprise the system also exhibit the substantial genomic, transcriptional, and biological heterogeneity found in primary tumors. We show, using Trastuzumab (Herceptin) monotherapy as an example, that the system can be used to identify molecular features that predict or indicate response to targeted therapies or other physiological perturbations.

Breast Neoplasms↗

PubMed2006

Genomic and transcriptional aberrations linked to breast cancer pathophysiologies.

This study explores the roles of genome copy number abnormalities (CNAs) in breast cancer pathophysiology by identifying associations between recurrent CNAs, gene expression, and clinical outcome in a set of aggressively treated early-stage breast tumors. It shows that the recurrent CNAs differ between tumor subtypes defined by expression pattern and that stratification of patients according to outcome can be improved by measuring both expression and copy number, especially high-level amplification. Sixty-six genes deregulated by the high-level amplifications are potential therapeutic targets. Nine of these (FGFR1, IKBKB, ERBB2, PROCC, ADAM9, FNTA, ACACA, PNMT, and NR1D1) are considered druggable. Low-level CNAs appear to contribute to cancer progression by altering RNA and cellular metabolism.

Breast Neoplasms↗

PubMed2005

Construction and application of a full-coverage, high-resolution, human chromosome 8q genomic microarray for comparative genomic hybridization.

BACKGROUND: Array-based comparative genomic hybridization (aCGH) enables genome-wide quantitative delineation of genomic imbalances. A high-resolution contig array was developed specifically for chromosome 8q because this chromosome arm is frequently altered in many human cancers. METHODS: A minimal tiling path contig of 702 8q-specific bacterial artificial chromosome (BAC) clones was generated with a novel computational tool (BAC Contig Assembler). BAC clones were amplified by degenerative oligonucleotide primer (DOP) polymerase chain reaction and subsequently printed onto glass slides. For validation of the array DNA samples of gastroesophageal and prostate cancer cell lines, and chronic myeloid leukemia specimens were used, which were previously characterized by multicolor fluorescence in situ hybridization and conventional CGH. RESULTS: Single and double copy gains were confidently demonstrated with the 8q array. Single copy loss and high-level amplifications were accurately detected and confirmed by bicolor fluorescence in situ hybridization experiments. The 8q array was further tested with paraffin-embedded prostate cancer specimens. In these archival specimens, the copy number changes were confirmed. In fresh and archival samples, additional alterations were disclosed. In comparison with conventional CGH, the resolution of the detected changes was much improved, which was demonstrated by an amplicon of 0.7 Mb and a deletion of 0.6 Mb, both spanned by only six BAC clones. CONCLUSIONS: A comprehensive array is presented, which provides a high-resolution method for mapping copy number alterations on chromosome 8q.

Adenocarcinoma↗

PubMed2004

The RAB25 small GTPase determines aggressiveness of ovarian and breast cancers.

High-density array comparative genomic hybridization (CGH) showed amplification of chromosome 1q22 centered on the RAB25 small GTPase, which is implicated in apical vesicle trafficking, in approximately half of ovarian and breast cancers. RAB25 mRNA levels were selectively increased in stage III and IV serous epithelial ovarian cancers compared to other genes within the amplified region, implicating RAB25 as a driving event in the development of the amplicon. Increased DNA copy number or RNA level of RAB25 was associated with markedly decreased disease-free survival or overall survival in ovarian and breast cancers, respectively. Forced expression of RAB25 markedly increased anchorage-dependent and anchorage-independent cell proliferation, prevented apoptosis and anoikis, including that induced by chemotherapy, and increased aggressiveness of cancer cells in vivo. The inhibition of apoptosis was associated with a decrease in expression of the proapoptotic molecules, BAK and BAX, and activation of the antiapoptotic phosphatidylinositol 3 kinase (PI3K) and AKT pathway, providing potential mechanisms for the effects of RAB25 on tumor aggressiveness. Overall, these studies implicate RAB25, and thus the RAB family of small G proteins, in aggressiveness of epithelial cancers.

Animals↗

PubMed2004

Computational BAC clone contig assembly for comprehensive genome analysis.

Comparative genomic hybridization (CGH) has proved to be a powerful tool for the detection of genome copy number changes in human cancers and in other diseases caused by segmental aneusomies. Array versions of CGH allow the definition of these aberrations, with resolution determined by the size and distribution of the array elements. Resolution approaching 100 kb can be achieved by use of arrays comprising bacterial artificial chromosomes (BACs) distributed contiguously across regions of interest. We describe here a computer program that automatically assembles contigs of minimally overlapping BAC clones, using information about BAC end-sequences and the normal genome DNA sequence. We demonstrate the characteristics of contigs assembled and annotated by use of this approach for regions of recurrent abnormality in human ovarian and breast cancers at chromosome bands 3q25-q27 and 8q24 and chromosome arm 20q. We also show illustrative analyses of regions of amplification in these regions in breast and ovarian tumor cell lines by use of array CGH with arrays comprising contiguous BACs.

Breast Neoplasms↗

PubMed2003

End-sequence profiling: sequence-based analysis of aberrant genomes.

Genome rearrangements are important in evolution, cancer, and other diseases. Precise mapping of the rearrangements is essential for identification of the involved genes, and many techniques have been developed for this purpose. We show here that end-sequence profiling (ESP) is particularly well suited to this purpose. ESP is accomplished by constructing a bacterial artificial chromosome (BAC) library from a test genome, measuring BAC end sequences, and mapping end-sequence pairs onto the normal genome sequence. Plots of BAC end-sequences density identify copy number abnormalities at high resolution. BACs spanning structural aberrations have end pairs that map abnormally far apart on the normal genome sequence. These pairs can then be sequenced to determine the involved genes and breakpoint sequences. ESP analysis of the breast cancer cell line MCF-7 demonstrated its utility for analysis of complex genomes. End sequencing of approximately 8,000 clones (0.37-fold haploid genome clonal coverage) produced a comprehensive genome copy number map of the MCF-7 genome at better than 300-kb resolution and identified 381 genome breakpoints, a subset of which was verified by fluorescence in situ hybridization mapping and sequencing.

Animals↗