Genes of cell-cell interactions, chemotherapy detoxification and apoptosis are induced during chemotherapy of acute myeloid leukemia

<p>Abstract</p> <p>Background</p> <p>The molecular changes <it>in vivo </it>in acute myeloid leukemia cells early after start of conventional genotoxic chemotherapy are incompletely understood, and it is not known if early molecular modulations reflect clinical response.</p> <p>Methods</p> <p>The gene expression was examined by whole genome 44 k oligo microarrays and 12 k cDNA microarrays in peripheral blood leukocytes collected from seven leukemia patients before treatment, 2–4 h and 18–24 h after start of chemotherapy and validated by real-time quantitative PCR. Statistically significantly upregulated genes were classified using gene ontology (GO) terms. Parallel samples were examined by flow cytometry for apoptosis by annexin V-binding and the expression of selected proteins were confirmed by immunoblotting.</p> <p>Results</p> <p>Significant differential modulation of 151 genes were found at 4 h after start of induction therapy with cytarabine and anthracycline, including significant overexpression of 31 genes associated with p53 regulation. Within 4 h of chemotherapy the BCL2/BAX and BCL2/PUMA ratio were attenuated in proapoptotic direction. FLT3 mutations indicated that non-responders (5/7 patients, 8 versus 49 months survival) are characterized by a unique gene response profile before and at 4 h. At 18–24 h after chemotherapy, the gene expression of p53 target genes was attenuated, while genes involved in chemoresistance, cytarabine detoxification, chemokine networks and T cell receptor were prominent. No signs of apoptosis were observed in the collected cells, suggesting the treated patients as a physiological source of pre-apoptotic cells.</p> <p>Conclusion</p> <p>Pre-apoptotic gene expression can be monitored within hours after start of chemotherapy in patients with acute myeloid leukemia, and may be useful in future determination of therapy responders. The low number of patients and the heterogeneity of acute myeloid leukemia limited the identification of gene expression predictive of therapy response. Therapy-induced gene expression reflects the complex biological processes involved in clinical cancer cell eradication and should be explored for future enhancement of therapy.</p

Gene graph of the <i>M.tuberculosis</i> H37Rv <i>1754c-1765c, Mb1785-1787</i>, <i>MT1808</i> and <i>MT1812-1813</i> regions.

<p>The graph is based on region z-scores, with each line representing one isolate analyzed by CGH. Region z-scores of more than ±4.42 (absolute value) occurring in one or more samples, corresponding to a standard normal P-value of 0.00001 (1e-5) indicate deletions (above the 0-line) or extra regions (below 0-line). Red lines represent isolates of the Beijing lineage, green lines; ST42 lineage and blue lines; non-Beijing isolates other than ST42.</p

PCR results from the 22 regions chosen for verification in 60 clinical <i>M. tuberculosis</i> isolates.

<p>The dendogram based on PCR results from 16 deletions and 6 extra regions chosen for verification on 60 clinical <i>M. tuberculosis</i> isolates (28 Beijing, 13 ST42 and 19 other non-Beijing) was produced by unweighted pair group method (UPGMA) clustering using a Pearson correlation based distance measure.</p

Regions chosen for PCR verification based on the CGH array analysis.

<p>The dendogram based on CGH array analysis of 22 <i>M. tuberculosis</i> isolates from Myanmar (11 Beijing, 4 ST42, 2 ST89 and 5 previously unreported [UR] shared-types [ST]) was produced by weighted pair group method (WPGMA) clustering using a Pearson correlation based distance measure defined on regional z-score values, corresponding to P-values between 1e-3 and 1e-16. The green color indicates extra regions that are present in the clinical isolates, but not in the reference strain <i>M. tuberculosis</i> H37Rv, whereas the red color depicts deletions in the clinical isolates compared to the reference strain <i>M. tuberculosis</i> H37Rv.</p

Genetic diversity among 22 <i>Mycobacterium tuberculosis</i> isolates tested by CGH.

<p>The dendogram was produced by weighted pair group method (WPGMA) clustering using a Pearson correlation based distance measure defined on regional z-score values. The red color indicates deletions in the clinical isolates compared to the reference strain <i>M. tuberculosis</i> H37Rv, whereas the green color shows extra regions in the clinical isolates not present in the reference strain <i>M. tuberculosis</i> H37Rv.</p

Primers used for the PCR verification of deleted and extra regions as observed by CGH analysis.

*<p>:<a href="http://xrl.us/6cg3" target="_blank">http://xrl.us/6cg3</a></p><p>The annealing temperature for each primer set and the expected PCR product size based on the published sequences of <i>M. tuberculosis</i> H37Rv, <i>M. bovis</i> AF22197 or <i>M. tuberculosis</i> CDC1551 are also provided.</p