[Study of testicular cancer gene expression in samples of oral leukoplakia and squamous cell carcinoma of the mouth].

Cancer-testis (CT) antigens are normally expressed mostly in human germ cells, there is also an aberrant expression in some tumor cells. This expression profile makes them potential tumor growth biomarkers and a promising target for tumor immunotherapy. Specificity of CT genes expression in oral malignant and potentially malignant diseases, e.g. oral leukoplakia, is not yet studied. Data on CT genes expression profile in leukoplakia would allow developing new diagnostic methods with potential value for immunotherapy and prophylaxis of leukoplakia malignization. In our study we compared CT genes expression in normal oral mucosa, oral leukoplakia and oral squamous cell carcinoma. We are the first to describe CT genes expression in oral leukoplakia without dysplasia. This findings make impossible differential diagnosis of oral leukoplakia and squamous cell carcinoma on the basis of CT genes expression. The prognostic value of CT genes expression is still unclear, therefore the longitudinal studies are necessary.

[Genetic bases of diversity of the repertoire of immunoglobulins in application to diagnostics of clonality of B-cell lymphoid populations].

Molecular mechanisms underlying the formation of B-cell lymphomas in connection with processes associated with the maturation of B lymphocytes are reviewed. The currently used diagnostic methods do not always distinguish lymphomas from reactive changes of the lymphoid tissue. The principle of the molecular genetic method ofclonality detection in lymphocyte populations, technical problems, and the strategy of its application in clinical diagnostics of lymphomas are described in detail.

[Trisomy of chromosome 8 in Ph-negative cells of the bone marrow in patients with chronic myeloid leukemia treated with inhibitors of BCR-ABL tyrosine kinases].

AIM: To analyse clinical implications of chromosome 8 trisomy in Ph-negative cells of the bone marrow in patients with chronic myeloid leukemia (CML) treated with inhibitors of tyrosinkinases (ITK). MATERIAL AND METHODS: A total of 386 patients with CML (chronic phase--288, acceleration phase--77) received imatinib (400-800 mg/day). Because of resistance and/or intolerance some patients were switched to ITK II (nilotinib, dasatinib, bozutinib). This study included 8 CML patients (7 in a chronic phase, 1 in acceleration phase) treated with BCR-ABL ITK inhibitors of the first (imatinib) and the second line (ITK-II). The standard cytogenetic examination, on demand--investigation of the interphase nuclei with FISH, in some cases morphological, cytochemical and histological examinations of the bone marrow were made. RESULTS: The existence of a Ph-negative clone with trisomy of chromosome 8 had no negative effect on the course of the disease. The patients showed a stable hematological and cytogenetic response and no need in changing treatment policy. In long-term follow-up Ph-negative clone with trisomy of the chromosome 8 persisted without a clear trend to rise in most patients. CONCLUSION: Detection of a Ph-negative clone with chromosome 8 trisomy at early stages suggests parallel existence of Ph-positive and Ph-negative clones. None of the patients had myelodisplasia.

[Prognosis factors in imatinib mesilate therapy in patients with a chronic phase of Ph-positive chronic myeloid leukemia: data from a multicenter non-randomized trial in Russia].

AIM: To reveal prognostically significant factors affecting efficacy of glivek therapy in untreated (duration of the disease < or = 6 months) and pretreated (duration of the disease > 6 months) patients with chronic myeloid leukemia (CML) in a chronic phase. MATERIAL AND METHODS: A total of 338 patients (64 untreated and 274 pretreated) with a chronic-phase CML on glivek therapy entered the trial. RESULTS: Five-year survival on glivek was high (89, 98 and 88% in untreated and pretreated patients, respectively). Incidence of transformation in the acceleration phase and blast crisis was low both in untreated and pretreated patients (1.6 and 11%, respectively) and correlated with the rate of a complete cytogenetic response (CCR). Untreated patients had no factors affecting treatment efficacy negatively, CCR probability was 96%. Blastemia, thrombocytosis and splenomegaly reduced CCR probability significantly in pretreated patients. Slow reduction of the tumor mass, late achievement of a complete hematological response and a cytogenetic response decreased probability of CCR. CONCLUSION: Glivek is a drug of choice for patients with chronic-phase CML. High probability of CCR both in untreated and pretreated patients lowers the risk of the disease transformation into the phase of acceleration/blast crisis and raises overall survival in both groups.

[Cytogenetic response as a marker of efficacy of chronic myeloid leukemia therapy with a BCR-ABL thyrosine kinase inhibitor glivek].

AIM: Clinical practice with the drug glivek (imatinibe mesilate, ST1571) blocking activity of oncoprotein p210 shows that a cytogenetic response can be reached in 50-60% of patients with chronic myeloid leukemia (CML), in a late chronic phase (CP) in resistance to or intolerance of interferon alpha (IF-alpha) and in 24-43% of patients in the acceleration phase (AP). This study aimed at assessment of the rate and stability of a cytogenetic response (CR) and long-term results of survival in CML patients on glivek. MATERIAL AND METHODS: Glivek was given to 195 CML patients (median of the treatment duration was 42 months, 1-156 months, of the patients' age--46 years). 79 patients were in CP, 116--in AP. The doses were 400 mg/day and 116 mg/day, respectively. Karyotype was studied before the treatment and later after each 6 months. RESULTS: A considerable CR was achieved in 57% patients in CP and 44%--in AP. Of them complete CR was obtained in 48 and 35%, respectively. Marked CR is a favourable prognostic factor. Survival of patients with marked CR in CP (97% 0 and AP (89%) was significantly higher than without CR (58 and 47%, respectively, p < 0.05). Marked CR persisted in 95% cases in both phases of CML. In complete CR, a repeated study of karyotype revealed residual number of Ph+ cells both in CP and AP in 86% patients. This demonstrates necessity to take glivek continuously in achievement of a complete CR by karyotypic test. Glivek inhibits the disease progression, lowers annual lethality. 42-month (median of glivek treatment duration) overall survival reached 91 and 59% in CP and AP, respectively. CONCLUSION: CR is an integral index prognosticating CML course. Survival rose significantly in patients with marked CR both in CP and AP of CML. Marked CR is persistent in continuous glivek therapy. The rate of a CR depends much on the disease stage.

[Transcription and mRNA splicing of the human lactoferrin gene controlled by the regulatory region of the bovine alphaS1 casein gene in the mammary gland of transgenic mice and in mouse embryonic stem cells].

The regulatory region of the bovine alphaS1 casein gene was used to obtain two genetic constructs for expression of human lactoferrin in the mammary gland of transgenic animals. Several transfected mouse embryonic stem cell (ESC) lines and primary transgenic mice were generated with these constructs. Recombinant lactoferrin was not detected in milk of transgenic mice by Western blotting. However, a recombinant transcript was found in RNAs isolated from mammary glands of transgenic females during lactation and from transfected ESC lines.

Drug discrimination analysis of NMDA receptor channel blockers as nicotinic receptor antagonists in rats.

RATIONALE: Antagonists acting at the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors inhibit various phenomena associated with exposures to nicotine (e.g., tolerance, sensitization, dependence, and intravenous self-administration). These effects are often discussed in terms of nicotine-induced glutamate release with subsequent glutamate-dependent stimulation of dopamine metabolism and neuronal plasticity in brain areas critically involved in drug-addiction mechanisms. However, it is also well established that certain types of NMDA receptor antagonists (channel blockers) potently bind to nicotinic receptors and may act as nicotinic receptor antagonists. OBJECTIVE: The present study aimed to evaluate the discriminative-stimulus effects of the NMDA receptor channel blockers (+)MK-801, dextromethorphan, and memantine in rats trained to discriminate nicotine from its vehicle. METHODS: Adult male Wistar rats were trained to discriminate 0.6 mg/kg nicotine from saline under a two-lever, fixed-ratio 10 schedule of food reinforcement. During test sessions, injections of (+)MK-801 (0.03--0.3 mg/kg, i.p.), dextromethorphan (30 mg/kg, s.c.), or memantine (1--10 mg/kg, i.p.) were co-administered with s.c. nicotine (0.075--0.6 mg/kg; interaction tests) or saline (generalization tests). Additional interaction and generalization tests were conducted with the selective nicotinic receptor antagonists mecamylamine (0.1--3 mg/kg, s.c.) and MRZ 2/621 (0.3--10 mg/kg, i.p.), and the mGlu5 receptor antagonist MPEP (3--10 mg/kg, i.p.). RESULTS: In generalization tests, none of the compounds produced any appreciable levels of substitution for nicotine. The nicotine discriminative-stimulus control was dose dependently attenuated by mecamylamine (ED(50)=0.67 mg/kg) and MRZ 2/621 (ED(50)=9.7 mg/kg). Both agents produced a marked downward shift in the nicotine dose-response curve. Memantine and MPEP slightly attenuated nicotine discriminative-stimulus effects, while (+)MK-801 and dextromethorphan did not affect the nicotine-appropriate responding. CONCLUSIONS: NMDA receptor channel blockers, such as (+)MK-801, dextromethorphan, and memantine, have minimal interactions with the discriminative-stimulus effects of nicotine.

Effects of NMDA receptor channel blockers, MK-801 and memantine, on locomotor activity and tolerance to delay of reward in Wistar-Kyoto and spontaneously hypertensive rats.

N-Methyl-D-aspartate (NMDA) receptor blockade enhances motor activity and stimulates dopamine metabolism, effects shared with classical psychostimulant drugs. The present study aimed to characterize behavioral effects of two NMDA receptor channel blockers, MK-801 and memantine, in both Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats. In Experiment 1, SHR rats demonstrated higher spontaneous locomotor activity and spent more time in the central area of the open field apparatus compared with WKY rats. Rats of both strains pre-treated with MK-801 (0.01-0.3 mg/kg) or memantine (1-32 mg/kg) demonstrated dose-dependent increases in the total distance traveled and time spent in the central area. Experiment 2 was based on the two-lever discrete-trial delayed reinforcement task in which rats could press one lever to obtain one pellet immediately or another lever for four pellets delivered after a variable delay (0-60 s). Tolerance to delay of reward did not differ between strains. MK-801 (0.03-0.3 mg/kg) and memantine (1-10 mg/kg) produced small, but significant, facilitation of the large-reward lever responding and markedly impaired operant performance at higher dose levels (increased number of missed trials). For both experiments, effects of MK-801 and memantine were more pronounced in WKY compared with SHR rats. Additional studies are needed to address the utility of noncompetitive NMDA receptor blockers in the treatment of attention deficit and hyperactivity disorder.

[Therapeutic efficacy of imatinib mesylate (glivec) in chronic phase of myeloid leukemia]. / Effektivnost' terapii imatiniba mezilatom (glivekom) v khronicheskoi faze mieloleikoza.

AIM: To evaluate efficacy and tolerance of glivek in chronic myeloid leukemia (CML) in patients who failed interferon-alpha (If-a) preparations. MATERIAL AND METHODS: 79 patients in a chronic phase of Ph + CML with hematological and cytogenetic resistance or intolerance of If-a. The response to glivec was assessed by achievement of a complete hematological remission and the cytogenetic effect (the degree of reduction of cell clone Ph+ in bone marrow). Tolerance and safety of the drug was studied by monthly standard clinicohematological tests. RESULTS: Not only a hematological remission (92.4%), but also partial (46.8%) or complete (27.8%) elimination of BCR-ABL +/- cells were achieved after 12 months of the treatment. Glivec was well tolerated. Hematological toxicity primarily as neutropenia and thrombocytopenia were observed in 54.4 and 42% patients, respectively. Neutropenia of the third degree which made impossible to continue the treatment was observed in 29.1% patients; throbocytopenia of the third degree was registered in 16.5% patients. Among most frequent non-hematological side effects there were moderate edema, nausea, leg muscle convulsions, weight gain, arthralgias, skin eruption. All the complications were transient, were managed in all cases with only a short-time discontinuation of glivec therapy. CONCLUSION: High activity of glivec at early stages of CML allows using this drug as a first-line therapy in patients with CML.

[Production of recombinant endostatin in milk from transgenic mice]. / Poluchenie rekombinantnogo éndostatina v moloke transgennykh myshei.

Using the bovine alpha S1-casein gene, a genetic construct with an endostatin-coding fragment of the mouse collagen XVIII cDNA was designed to express endostatin in milk of transgenic animals. Several transgenic mice were obtained. The mice secreted endostatin in milk at 70-300 ng/microliter and transmitted this character to their progeny.

[Functional activity and expression of P-glycoprotein in chronic myeloid leukemia]. / Funktsional'naia aktivnost' i ékspressiia P-glikoproteina pri khronicheskom mieloleikoze.

AIM: To evaluate the prognostic significance of P-glycoprotein (Pgp) in chronic myeloid leukemia (CML). MATERIALS AND METHODS: Functional activity (rhodamine 123 test) and expression of Pgp (binding of UIC2 monoclonal antibodies by cells) were evaluated by flow cytofluorometry. A total of 141 samples of peripheral blood from 121 patients with various stages of CML were examined. RESULTS: The number of patients whose cells express functionally active Pgp increases during the blast crisis (BC) in comparison with the chronic phase (CP). Repeated testing of patients with BC and CP showed that Pgp-expressing cells can disappear from the peripheral blood of patients despite the treatment by Pgp preparations and substrates. However the number of cases with expression and functional activity of Pgp increases in the course of BC. Several patients in whom functionally active Pgp was not detected during diagnosis of BC had longer BC phase than patients with the active protein. CONCLUSION: These data suggest that active Pgp contributes to CML BC (presumably to patient's response to therapy) but this contribution is not decisive.

[Osmotic activity of plasma in plasmapheresis in patients with multiple myeloma]. / Osmoticheskaia aktivnost' plazmy pri plazmafereze u bol'nykh mnozhestvennoi mielomoi.

AIM: To control safety and efficiency of therapeutic plasmapheresis (PA) by osmolality, colloido-osmotic pressure (COP), total protein concentration before and after the procedure in patients with paraproteinemic hemoblastosis. MATERIAL AND METHODS: 20 patients with multiple myeloma have undergone 42 PA procedures conducted by two techniques: continuous flow centrifugation on blood fractioners or intermittent centrifugation of blood in plastic containers. The removed plasma volume averaged 1/3 (group 1) or 2/3 of the plasma volume (group 2). The removed protein reached 62-197 g. Isotonic sodium chloride solution and/or reopolyglucin (20-60 g) replaced the removed plasm. Total protein concentration was measured colorimetrically in biuretic reaction, plasma osmolality--cryoscopically and COP--on Knauer osmometer. RESULTS: PA leads to a short decline in osmolality (97.0-99.1%), of total protein concentration (82.8-78.6%) and of COD (79.2% in replacement with saline and 90.2% in replacement with dextran). During recovery after the procedure plasma osmotic activity and protein concentration return to the baseline. CONCLUSION: In elimination of 1/3 of plasma volume and crystalloid infusion, hemodilution promotes release of abnormal proteins from the tissues into the circulation and thereafter removal them from the organism. In removal of 1/2 and more of plasma volume, COP demans correction made by administration of colloids, e.g. solution of low molecular dextran. There is a potential danger of COD lowering several hours after PA due to different speed of dextran elimination and mobilization of protein reserve.

[Adaptation of reverse transcriptase polymerase chain reaction for clinical diagnosis of expression of MDR1 gene]. / Adaptatsiia metoda obratnotranskriptaznoi tsepnoi reaktsii dlia klinicheskoi diagnostiki ékspressii gena MDR1.

Chemotherapy of malignant tumors is ineffective usually because of tumor cell resistance to it. Two types of resistance are known: cell resistance to a certain drug and multiple drug resistance (MDR). MDR covers a wide spectrum of drugs with different chemical structure and mechanisms of action. The most frequent cause of MDR is hyperexpression in the plasma membrane of P glycoprotein cells, which is coded for by MDR1 gene realizing active release of many cytotoxic substances from cells (Pgp-MDR). Acquisition of MDR phenotype by patient's cells impedes therapy and is often a poor prognostic sign, and therefore testing of material from cancer patients for MDR phenotype is important for selecting tumor therapy. We adapted the reverse transcriptase polymerase chain reaction (RT-PCR) to evaluating the MDR1 gene expression in peripheral blood cells of patients with hemoblastosis, assessed its sensitivity and specificity, and carried out clinical trials with blood samples from patients with MDR. Comparison of the results of RT-PCR with the findings of other methods used for detection of Pg-MDR showed their good correlation in the majority of cases. These results recommend these method for clinical practice in patients with hemoblastosis.

[Molecular cloning of the gene for secreted beta-galactosidase of the filamentous fungus Penicillium canescens]. / Molekuliarnoe klonirovanie gena sekretiruemoi beta-galaktozidazy mitselial'nogo griba Penicillium canescens.

A secreted beta-galactosidase gene from P. canescens was cloned using synthetic oligonucleotide probes and its nucleotide sequence was partially determined. The gene was shown to be present in the genome as a single copy. The deduced primary structure of the signal peptide and the first 25 amino acid residues of the mature protein was established. The structure analysis of the beta-galactosidase gene revealed at least one short intron 64 bp long. The transcription start point was found to be located 89 bp upstream the initiation codon. Some peculiarities of the signal peptide cleavage mechanism are discussed.

[Cloning preprolactin cDNA from Pacific salmon in Escherichia coli]. / Klonirovanie kDNK preprolaktina tikhookeanskogo lososia v Escherichia coli.

Prolactin coding mRNA was shown to be a prevalent part of chum salmon (Oncorhynchus keta) pituitary poly(A)-RNA during the spawning period. Clone lambda gtPrk12 was selected from the pituitary cDNA library by means of hybridization with the prolactin probe, and a nucleotide sequence of the insertion was determined and compared to the prolactin coding sequences from rainbow trout and Pacific chinook salmon, which had been published earlier. The sequences compared exhibited a significant homology. The deduced amino acid sequence of the chum salmon prolactin differed from a sequence determined directly in a single position. The prolactin-coding sequence can be used for constructing the bacterial strain producing prolactin.

[Nucleotide sequences encoding glycinin B4 polypeptide of the cultivated soybean Glycine max and its presumed wild-type ancestor Glycine soja in connection with the origin of cultivated soybean]. / Nukleotidnye posledovatel'nosti, kodiruiushchie polipeptid B4 glitsinina kul'turnoi soi Glycine max i ee predpolagaemogo dikogo predka Glycine soja v sviazi s vyiasneniem proiskhozhdeniia kul'turnoi soi.

Nucleotide sequences of cDNAs encoding soybean glycinin B4 polypeptide were compared in three soybean cultivars and two plant introductions of wild soybean Glycine soja. Only two nucleotide substitutions were found in three cultivars G. max, as compared with G. max and G. soja having nucleotide sequences which contain four nucleotide substitutions. These data serve as additional evidence, at molecular level, indicating the origin of G. max from G. soja. On the other hand, the time period required for four nucleotide substitutions' accumulation, as calculated from parameters of molecular evolution of 11S globulins, is much longer than the term having passed after soybean domestication.

[The parameters of the molecular evolution of the 11S globulins of plants]. / Nekotorye parametry molekuliarnoi évoliutsii 11S-globulinov rastenii.

Nucleotide sequences of cloned cDNA coding for soybean storage protein glycinin and deduced amino acid sequences of basic polypeptides (subfamily II) of glycinin are compared with the amino acid sequences of 11S globulins from other plants. An average number of amino acid substitutions in various evolutionary branches is calculated. A proportional dependence is established between the average number of substitutions per site (the evolutionary distance) and the hypothetical term of divergence of corresponding taxa. The evolution rate of 11S globulins and a term of divergence of the two subfamilies of 11S globulin polypeptides in Fabaceae is estimated.

An attempt to elucidate the origin of cultivated soybean via comparison of nucleotide sequences encoding glycinin B4 polypeptide of cultivated soybean, Glycine max, and its presumed wild progenitor, Glycine soja.

Nucleotide sequences of cDNAs encoding soybean glycinin B4 polypeptide were compared for three soybean cultivars and two introductions of wild soybean, G. soja. For three G. max cultivars, only two nucleotide substitutions were found, while G. max and G. soja nucleotide sequences had four substitutions. These data give added proof that G. max originated from G. soja. On the other hand, the time required for the accumulation of four nucleotide substitutions (calculated from the parameters of 11S globulin molecular evolution) appeared to be longer than the duration of the soybean domestication period.