Identification of Genes Transcriptionally Responsive to the Loss of MLL Fusions in MLL-Rearranged Acute Lymphoblastic Leukemia

Introduction

MLL-rearranged acute lymphoblastic leukemia (ALL) in infants (<1 year) is characterized by high relapse rates and a dismal prognosis. To facilitate the discovery of novel therapeutic targets, we here searched for genes directly influenced by the repression of various MLL fusions.

Methods

For this, we performed gene expression profiling after siRNA-mediated repression of MLL-AF4, MLL-ENL, and AF4-MLL in MLL-rearranged ALL cell line models. The obtained results were compared with various already established gene signatures including those consisting of known MLL-AF4 target genes, or those associated with primary MLL-rearranged infant ALL samples.

Results

Genes that were down-regulated in response to the repression of MLL-AF4 and MLL-ENL appeared characteristically expressed in primary MLL-rearranged infant ALL samples, and often represented known MLL-AF4 targets genes. Genes that were up-regulated in response to the repression of MLL-AF4 and MLL-ENL often represented genes typically silenced by promoter hypermethylation in MLL-rearranged infant ALL. Genes that were affected in response to the repression of AF4-MLL showed significant enrichment in gene expression profiles associated with AF4-MLL expressing t(4;11)+ infant ALL patient samples.

Conclusion

We conclude that the here identified genes readily responsive to the loss of MLL fusion expression potentially represent attractive therapeutic targets and may provide additional insights in MLL-rearranged acute leukemias.     

Structural basis for the recruitment of ERCC1-XPF to nucleotide excision repair complexes by XPA

The nucleotide excision repair (NER) pathway corrects DNA damage caused by sunlight, environmental mutagens and certain antitumor agents. This multistep DNA repair reaction operates by the sequential assembly of protein factors at sites of DNA damage. The efficient recognition of DNA damage and its repair are orchestrated by specific protein-protein and protein-DNA interactions within NER complexes. We have investigated an essential protein-protein interaction of the NER pathway, the binding of the XPA protein to the ERCC1 subunit of the repair endonuclease ERCC1-XPF. The structure of ERCC1 in complex with an XPA peptide shows that only a small region of XPA interacts with ERCC1 to form a stable complex exhibiting submicromolar binding affinity. However, this XPA peptide is a potent inhibitor of NER activity in a cell-free assay, blocking the excision of a cisplatin adduct from DNA. The structure of the peptide inhibitor bound to its target site reveals a binding interface that is amenable to the development of small molecule peptidomimetics that could be used to modulate NER repair activities in vivo.     

HPV testing for cervical cancer screening in Croatia

Opportunistic screening based on the Pap smear has been undertaken in Croatia since 1953. However, cervical cancer remains an important health problem in Croatia when compared to European countries with organised screening programmes. In Croatia, in addition to screening based on well established cytology, Human papillomavirus (HPV) testing is widely used as secondary test as a triage to borderline cytology and as a follow-up after treatment of severe cervical lesions. Many different approaches for HPV testing arose in Croatia over the last decade depending on the needs of each medical institution involved. Presently, there is an urgent need for better networking between the laboratories, the implementation of quality assessment and the adaptation of a uniform system of referring to and reporting of HPV testing. In conclusion, the best possible organisation for HPV testing would be essential for implementation of HPV testing as primary screening test in Croatia, an thus ultimately and hopefully, the more successful cervical cancer control.     

Atherosclerotic lesions of supra-aortic arteries in diabetic patients

The aim of this prospective study was to determine the prevalence and localization of stenotic atherosclerotic lesions of supra-aortic arteries in diabetic patients according to age and sex. Angiograms obtained by digital subtraction angiography were analyzed in 150 diabetic patients (study group) and 150 non-diabetic patients (control group) with symptoms of cerebral ischemia. Diabetic patients were found to have a significantly higher prevalence of stenotic atherosclerotic lesions of the internal carotid artery. Lesions of the large supra-aortic arteries were significantly more common in the left than in the right side of the neck (p < 0.001), but the difference between the diabetic and the non-diabetic group did not reach statistical significance. Hemodynamic conditions were found to be more important than diabetes for the occurrence of atherosclerotic lesions in these arteries. Changes in the proximal segment of the left common carotid artery were the most common finding in diabetic patients, hence attention should be paid to this localization on control examinations.     

Intercostal catheter analgesia is more efficient vs. intercostal nerve blockade for post-thoracotomy pain relief

A pain after thoracotomy may result in a postoperative hypoventilation and lead to atelectases and pneumonia. This study was aimed to compare two analgesic regimens after posterolateral thoracotomy. 80 patients (40-70 years) undergoing thoracotomy were randomized to intercostal catheter analgesia (group A, n = 40) and intercostal nerve block (group B, n = 40). Patients in group A were given 20 mL of 0.5% bupivacaine injections twice a day by intercostal catheter. Intercostal nerve blockade was performed using 5 mL of 0.5% bupivacaine. Intercostal nerve in thoracotomy wound, nerves below and above thoracotomy wound was also injected. PaO2, PaCO2, FVC, FEV1 and visual analog pain scale (VAS) were obtained preoperatively, 24, 48 and 72 hours after operation. Postoperative complications were recorded at the patient discharge. Differences between groups were calculated using Mann-Whitney, KW test and chi square test. The arterial blood gas analyses did not show statistically significant change in any group and time according to the baseline values. FVC and FEV1 decreased significantly in both groups at first postoperative day according to baseline measurements. Patients in B group had significantly higher FEV1 values in the third postoperative day (73.05 +/- 11.25 in A vs. 83.50 +/- 9.17 in B group, p < 0.05). Intercostal catheter analgesia resulted in significantly lower postoperative VAS scores and reduced opioid requirement as compared to intercostal nerve blockade. No differences in the postoperative complications were observed between groups.     

Genome-wide local ancestry approach identifies genes and variants associated with chemotherapeutic susceptibility in African Americans

Chemotherapeutic agents are used in the treatment of many cancers, yet variable resistance and toxicities among individuals limit successful outcomes. Several studies have indicated outcome differences associated with ancestry among patients with various cancer types. Using both traditional SNP-based and newly developed gene-based genome-wide approaches, we investigated the genetics of chemotherapeutic susceptibility in lymphoblastoid cell lines derived from 83 African Americans, a population for which there is a disparity in the number of genome-wide studies performed. To account for population structure in this admixed population, we incorporated local ancestry information into our association model. We tested over 2 million SNPs and identified 325, 176, 240, and 190 SNPs that were suggestively associated with cytarabine-, 5'-deoxyfluorouridine (5'-DFUR)-, carboplatin-, and cisplatin-induced cytotoxicity, respectively (p≤10(-4)). Importantly, some of these variants are found only in populations of African descent. We also show that cisplatin-susceptibility SNPs are enriched for carboplatin-susceptibility SNPs. Using a gene-based genome-wide association approach, we identified 26, 11, 20, and 41 suggestive candidate genes for association with cytarabine-, 5'-DFUR-, carboplatin-, and cisplatin-induced cytotoxicity, respectively (p≤10(-3)). Fourteen of these genes showed evidence of association with their respective chemotherapeutic phenotypes in the Yoruba from Ibadan, Nigeria (p<0.05), including TP53I11, COPS5 and GAS8, which are known to be involved in tumorigenesis. Although our results require further study, we have identified variants and genes associated with chemotherapeutic susceptibility in African Americans by using an approach that incorporates local ancestry information.     

Atmospheric pollen season in Zagreb (Croatia) and its relationship with temperature and precipitation

The number of individuals allergic to plant pollen has recently been on a constant increase, especially in large cities and industrial areas. Therefore, monitoring of airborne pollen types and concentrations during the pollen season is of the utmost medical importance. The research reported in this paper aims to determine the beginning, course and end of the pollen season for the plants in the City of Zagreb, to identify allergenic plants, and to assess the variation in airborne pollen concentration as a function of temperature and precipitation changes for the year 2002. A volumetric Hirst sampler was used for airborne pollen sampling. Qualitative and quantitative pollen analysis was performed under a light microscope (magnification ×400). In the Zagreb area, 12 groups of highly allergenic plants (alder, hazel, cypress, birch, ash, hornbeam, grasses, elder, nettles, sweet chestnut, artemisia and ambrosia) were identified. Birch pollen predominated in spring, the highest concentrations being recorded in February and March. Grass pollen prevailed in May and June, and pollen of herbaceous plants of the genus Urtica (nettle) and of ambrosia in July, August and September. Air temperature was mostly higher or considerably higher than the annual average in those months, which resulted in a many days with high and very high airborne pollen concentrations. The exception was April, when these concentrations were lower because of high levels of precipitation. This also held for the first half of August and the second half of September. Pollen-sensitive individuals were at high risk from February till October because of the high airborne pollen concentrations, which only showed a transient decrease when the temperature fell or there was precipitation.     

Molecular variants of human papilloma viruses type 16 and 6 in women with different cytological results detected by RFLP analysis

HPV infections are common and the presence of the same high-risk type in cervical specimens can be due to reinfection or persistence. Persistent infection is the most important predictor for development of cervical carcinoma. The aim of this study was to validate PCR-RFLP with two sets of primers: MY09/MY11 that amplify a fragment of L1 and P1/P2 that amplify a fragment of E1 ORF. PCR product of MY09/MY11 was digested with a set of 6 restriction enzymes (RE) and PCR product of P1/P2 with a set of 12 RE. Cervical samples from 110 women patients of the University Gynecologic Clinic CHC Zagreb were analyzed. There were 98 (89.1%) PCR positive samples detected with P1/P2 primers, and 94 (85.5%) PCR positive samples detected with MY09/MY11 primers. Seven HPV types were detected with P1/P2-RFLP technique and 17 with MY09/MY11-RFLP PCR positive samples amplified with both primer pairs agreed with each other in 82 samples; 16 samples were only positive with P1/P2 and 12 samples were only positive by MY09/MY11. HPV 16 was detected in 39 samples with MY09/11-RFLP, out of these two variants (two different patterns) were found with P1/P2 using Dde I, Hae III and Eco I. HPV 6 was detected in 9 samples with MY09/11-RFLP, out of these two variants were found with P1/P2 using HinfI. Combining these two PCR-RFLP methods subtypes of HPV 16 and HPV 6 were detected.     

Novel amidino substituted 2-phenylbenzothiazoles: Synthesis,antitumor evaluation in vitro and acute toxicity testing in vivo

The efficient synthesis of new bis-substituted nitro-amidino, amino-amidino (10a, 10b–13a, 13b) and previously prepared diamidino 2-phenyl-benzothiazoles (9a, 9b) is described. The compounds 11a and 11b were prepared by recently developed methodology of the key precursors in zwitterionic form 8a and 8b with 4-nitrobenzoylchloride in a very good yield (70%). All compounds except diamidino-substituted 2-phenylbenzothiazole 9a show exceptionally prominent tumor cell-growth inhibitory activity and cytotoxicity, whereby the special selectivity of amino-amidine 2-phenylbenzothiazole 12a towards MCF-7 and H 460 cells makes this compound a prospective lead compound that should be further evaluated in animal models. All in vivo tested compounds (12a, 12b, 13a and 13b) are absorbed from mice gastrointestinal system. LD₅₀ are between 67.33 and 696.2 mg/kg body weight (OECD/EPA toxicity categories 2–3).     

Early odontoblastic layer response to cavity preparation and acid etching in rats

The aim of this study was to establish the early odontoblastic layer response and quantitatively to estimate the number of odontoblasts after cavity preparation with and without acid etching. Half of 56 cavities prepared on rats' first upper molars were acid etched. Qualitative and morphometric analyses were made on histological and ultrathin sections 5 min, 6 h, 24 h and 72 h post-operatively. Under the etched cavity, a greater disarrangement of odontoblasts was found, modifications in nuclear shape and condensed chromatin 5 min. post-operatively. An additional reduction of odontoblast number was detected and an increase of aspirated cell number 5 min, 6 h and 24 h post-operatively, pronounced hyperaemia 6, 24 and 72 hours post-operatively and increased odontoblast number 72 hours post-operatively, compared to unetched cavities. In conclusion, injury to the odontoblastic layer was greater, but numerical renewal of the odontoblastic layer began earlier in etched cavities compared to unetched cavities.