Combination of automated high throughput platforms, flow cytometry, and hierarchical clustering to detect cell state.

BACKGROUND: This study examined whether hierarchical clustering could be used to detect cell states induced by treatment combinations that were generated through automation and high-throughput (HT) technology. Data-mining techniques were used to analyze the large experimental data sets to determine whether nonlinear, non-obvious responses could be extracted from the data. METHODS: Unary, binary, and ternary combinations of pharmacological factors (examples of stimuli) were used to induce differentiation of HL-60 cells using a HT automated approach. Cell profiles were analyzed by incorporating hierarchical clustering methods on data collected by flow cytometry. Data-mining techniques were used to explore the combinatorial space for nonlinear, unexpected events. Additional small-scale, follow-up experiments were performed on cellular profiles of interest. RESULTS: Multiple, distinct cellular profiles were detected using hierarchical clustering of expressed cell-surface antigens. Data-mining of this large, complex data set retrieved cases of both factor dominance and cooperativity, as well as atypical cellular profiles. Follow-up experiments found that treatment combinations producing "atypical cell types" made those cells more susceptible to apoptosis. CONCLUSIONS Hierarchical clustering and other data-mining techniques were applied to analyze large data sets from HT flow cytometry. From each sample, the data set was filtered and used to define discrete, usable states that were then related back to their original formulations. Analysis of resultant cell populations induced by a multitude of treatments identified unexpected phenotypes and nonlinear response profiles.

The prevalence of germline BRCA1 and BRCA2 mutations in young women with breast cancer undergoing breast-conservation therapy.

BACKGROUND: Germline mutations of BRCA1 and BRCA2 increase the risk for breast cancer. Mutation carriers selecting breast-conservation therapy (BCT) for treatment of operable breast cancer experience a higher rate of new primary breast cancers. We sought to determine the frequency of BRCA1/BRCA2 mutations in women who underwent BCT. Genetic testing results were compared with the prior probability of mutations in either gene. METHODS: Eighty-nine patients age 39 or younger entered the study. Genetic testing was performed for BRCA1 and BRCA2 and the BRCAPRO model determined the probability of carrying a mutation. RESULTS: Eight mutations were discovered (prevalence, 9.0%). Twenty (22%) uncharacterized sequence variants were found. The prior probability of carrying a mutation was 14%. Mutation carriers had a higher prior probability (.49) compared with women with uncharacterized variants (.09) or with normal genes (.11). CONCLUSIONS: BRCA1 and BRCA2 mutations are common (9%) among unselected young breast cancer patients undergoing BCT.

Respiratory maneuvers decrease irradiated cardiac volume in patients with left-sided breast cancer.

Late cardiac morbidity and mortality among left breast cancer survivors treated with radiation therapy is related to cardiac volume included in the radiation portals. To determine if respiratory maneuvers can help decrease cardiac volume included in the radiation portals for left-sided breast cancer, 17 women with breast cancer, who had undergone left breast radiation therapy, underwent cardiac magnetic resonance imaging (MRI). Cardiac volume within the radiation portals was assessed from a transverse stack of eight, 10-mm thick, contiguous slices, covering the entire heart and obtained during breathholding at (1) endtidal volume (ETid) and (2) deep inspiration. Fourteen subjects (93% of those who completed the study) had inclusion of at least a portion of their heart within the radiation portals at ETid (median: 25.9 cm3, range 4.2-119.1 cm3). In all subjects, inspiratory breathholding decreased irradiated cardiac volume [median change: -18.1 cm3 (-49%), p < or = 0.001 vs. ETid]. In 21% of patients, the entire heart could be displaced outside the radiation field with deep inspiration. Age was not correlated with change or percentage change in cardiac volume with respiratory maneuvers. We conclude that in breast cancer patients, deep inspiratory maneuvers significantly decrease irradiated cardiac volume included in the left breast radiation field. Such an approach during delivery of radiation therapy allows preservation of radiation dosage to the breast, while reducing cardiac involvement and possible associated cardiovascular toxicity.