Frequency of rare BCR-ABL1 fusion transcripts in chronic myeloid leukemia patients.

INTRODUCTION: The hallmark of chronic myeloid leukemia (CML) is the presence of Philadelphia chromosome, its resultant fusion transcript (BCR-ABL1), and fusion protein (p210). Alternate breakpoints in BCR (m-bcr, µ-bcr, and others) or ABL1 result in the expression of few rare fusion transcripts (e19a2, e1a2, e13a3, e14a3) and fusion proteins (p190, p200, p225) whose exact clinical significance remains to be determined. METHODS: Our study was designed to determine the type and frequency of BCR-ABL1 fusion transcripts in 1260 CML patients and to analyze the prognosis and treatment response in patients harboring rare BCR-ABL1 fusion transcripts. RESULTS: The frequency of various BCR-ABL1 fusion transcripts was as follows: e14a2 (60%), e13a2 (34.3%), e1a2 (1.2%), e1a2 + e13a2 (2.0%), e1a2 + e14a2 (1.8%), e19a2 (0.3%), and e14a3 (0.3%). CML patients with e1a2 transcripts had higher rates of disease progression, resistance, or suboptimal response to imatinib and failed to achieve major molecular response. CONCLUSION: Characterization of the specific fusion transcript in CML patients is important owing to the difference in prognosis and response to therapy in addition to the conventional need for monitoring treatment response. CML patients with e1a2 transcripts have to be closely monitored due to the high incidence of disease progression and treatment resistance/failure.

Antioxidant defense system during endometrial receptivity in the guinea pig: effect of ormeloxifene, a selective estrogen receptor modulator.

The role of the antioxidant defense system during endometrial receptivity, a phenomenon crucial for implantation and decidualization, and the effect of ormeloxifene, a selective estrogen receptor modulator, were investigated in the guinea pig, a laboratory mammalian species with interstitial implantation and a long functional luteal phase during each estrous cycle. A sharp rise in the activity of superoxide dismutase (SOD) in both antimesometrial (AM) and mesometrial segments and peroxidase in the AM segment of the uterus was observed on the day of maximal endometrial receptivity. Pretreatment with ormeloxifene resulted in loss of endometrial responsiveness, as evidenced by inhibition of trauma-induced decidualization and the activity of ornithine decarboxylase, a marker of tissue growth and repair. This was associated with a decrease in SOD and estradiol dehydrogenase activities, with corresponding increases in estrone dehydrogenase activity and stimulation of uterine luminal epithelial cell height and a distension of the uterine and glandular lumen. A decrease in peroxidase activity was observed only in the AM segment of the uterus on the imminent day of maximal endometrial receptivity. No effect on peripheral plasma progesterone concentration or surface ultrastructure was evident. These findings demonstrate that SOD plays an important role, with peroxidase having a supplementary role, in the first line of defense against superoxide anion radicals during the period of maximal endometrial receptivity in the guinea pig. Inhibition of endometrial receptivity and decidualization by ormeloxifene administered during the pre-receptive phase appears to be due to a depressed antioxidant defense system via dysregulation of redox-sensitive signaling, resulting in altered cellular toxicity due to increased superoxide radicals, and might contribute to the contraceptive action of ormeloxifene. This might be related to its estrogen antagonistic activity and/or decreased bioavailability of estradiol at a cellular level due to its increased metabolism to biologically less-active estrone via activation of estradiol-17 beta-hydroxysteroid dehydrogenase and suppression of estrone-17 beta-hydroxysteroid dehydrogenase.

Efficacy of picroliv in combination with miltefosine, an orally effective antileishmanial drug against experimental visceral leishmaniasis.

Visceral leishmaniasis (VL) or kala-azar continues to persist as one of the major public health problems in many tropical countries. However, no effective treatment for radical cure of the disease is yet available. Miltefosine, an alkyl phospholipid compound, is the first orally effective drug, which has shown 98% cure rate of VL patients during phase III clinical trial in India. Since this drug requires long course of treatment and has long half-life, there are fairly good chances of emergence of resistance. Furthermore, this drug has produced severe side-effects in some of the cases. We therefore examined the possibility of minimizing these effects by applying miltefosine in lower doses in combination with picrloviv, an immunomodulator against Leishmania donovani in hamsters (Mesocricetus auratus). The picroliv per se showed no antileishmanaial potential. However, when given with suboptimal dose of miltefosine, it enhanced efficacy of the latter from 45 to 86% on day 7 post treatment and from 32 to 64% on day 28 post treatment. Interestingly, the efficacy of this combination was as good as the curative dose of miltefosine alone. Thus, this combination appears to offer a fruitful strategy for treatment of VL.

Chemotherapy of leishmaniasis part II: synthesis and bioevaluation of substituted arylketene dithioacetals as antileishmanial agents.

Some novel aryl substituted ketene dithioacetals have been synthesized using novel synthetic methods. The compounds were screened against Leishmania donovani in hamsters for their activity profile. Some of the compounds inhibited 50-65% parasite growth at 50 mg kg(-1) x 5 days.

Immunomodulatory activity of analog of muramyl dipeptide and their use as adjunct to chemotherapy of Leishmania donovani in hamster.

In search of a potent immunomodulator to be used as an immunoprophylactic agent and as adjunct to chemotherapy against Leishmania infection, two analogs of muramyl dipeptide, viz. N.Ac-norMur-MeVal-D-isoGln (86/448) and N.AcMur-Acc-D-isoGln (89/729) were evaluated for desired activity. Effect of these peptides on cell mediated and humoral immunity was studied by immunizing the peptide treated mouse with sheep red blood cells (SRBC) and determining HA-titer, plaque forming cells assay and delayed type of hypersensitivity (DTH) response after 4-5 days. Both the peptides stimulated cell mediated immunity (CMI), humoral response as well as macrophage function in terms of super oxide anion (O2-) and nitric oxide (NO) generation. Mitogen induced lymphocyte proliferation and production of IL-2 and INF-gamma increased while that of IL-4 and IL-10 decreased by both the peptides showing a typical Th1 type response. After establishing the immunostimulatory activity, these peptides were evaluated for immunoprophylactic efficacy as well as for use as adjunct to chemotherapy with stibanate (SSG) against Leishmania donovani infection in golden hamster. These peptides were found quite effective in both the modes. In adjunct use the treatment may require lower dose of SSG and thereby reduce the chances of drug toxicity.

Hematological and molecular remission with combination chemotherapy in a patient with PLZF-RARalpha acute promyelocytic leukemia (APML).

Patients with acute promyelocytic leukemia (APML) with the t(11;17) translocation usually respond poorly to all-trans retinoic acid (ATRA) and chemotherapy. We describe a patient with promyelocytic leukemia zinc finger/retinoic acid receptor alpha (PLZF/RARalpha) APML who was treated with combination chemotherapy after poor response to arsenic trioxide. He achieved hematological remission in 4 weeks followed by achievement of molecular remission in 8 weeks. Four cycles of consolidation chemotherapy followed by four cycles of maintenance therapy were given over a period of 9 months. At a follow-up of 32 months after achieving hematological remission, he continues to remain in hematological and molecular remission with normal blood parameters and negative reverse transcriptase polymerase chain reaction (RT-PCR) results. Combination chemotherapy can achieve sustained remission in patients with PLZF/RARalpha APML.

Adjunct effect of immunostimulating hexapeptide analogous to human beta-casein fragment (54-59) to sodium stibogluconate against experimental visceral leishmaniasis.

Visceral leishmaniasis (VL) is a major public health problem in many tropical countries of the world. The available chemotherapeutics require parenteral administration and have other limitations like cost, toxicity, variable efficacy or restricted supplies. There is no effective treatment for immunosuppressed patients with leishmaniasis- HIV co-infection. Hence, new therapies, that are effective when treatment with the currently available drugs fails, must be developed. One of the major strategies for effective and safe treatment of leishmaniasis and other infectious diseases, in the last decade, involves the use of immunomodulators as adjunct to chemotherapy. In this context, we studied the immunomodulatory activity of a hexapeptide Val-Glu-Pro-Ile-Gly-Tyr (CDRI compound 89-215) corresponding to (54-59) fragment of human beta-casein in mice and its efficacy in adjunct chemotherapy with SSG using L. donovani/hamster model. The hexapeptide was found to enhance both humoral and CMI responses. In animal model the hexapeptide per se showed no antileishmanial activity. However, when given alongwith suiboptimal dose of SSG, it enhanced the efficacy of SSG from 24% to 80%. The activity was very close to the efficacy (85%) recorded for curative dose of SSG. Adjunct chemotherapy with immunomodulator in visceral leishmaniasis appears to be a fruitful preposition.

Acanthocheilonema viteae: octopamine and its physiological role.

Octopamine acts as an important neurotransmitter and neuromodulator in arthropods, mollusks, and nematodes. In mammals, however, no definite function for this amine has yet been described. By virtue of this difference in the neurophysiological requirement of the mammalian host and nematodes, octopamine offers good opportunity for exploring this area deeply with a view to identify a unique target for filarial chemotherapy. Results of the present study indicated that Acanthocheilonema viteae, the rodent filarial parasite, utilized tyrosine as a precursor for producing octopamine and some other biogenic amines. Octopamine exhibited specific saturable binding with the membrane prepared from the anterior portion of the filariid. This amine induced concentration dependent increase in the membrane potential which possibly caused tonic paralysis of the filariid. The rate of micro filarial release by the female worms also declined in the presence of this amine. The study thus provided preliminary evidences for the presence of an octopamine neurotransmitter system and also about some of the roles it plays in A. viteae.

Synthesis and antileishmanial profile of some novel terpenyl pyrimidines.

Some novel terpenyl pyrimidine derivatives 2(a-d) and 6(a-b) have been synthesised from alpha/beta-ionone keteneacetals 1 and 5. The terpenyl pyrimidine 2e has been synthesised from beta-ionone 3 in two steps in quantitative yield. The pyrimidine derivatives were screened for in-vivo antilesihmanial activity. The compounds 2d, 2e, 6a and 6b showed promising in-vivo antileishmanial activity.

Macrophages in the development of protective immunity against experimental Brugia malayi infection.

The present report compares the macrophage function in rodent hosts susceptible and resistant to the human lymphatic filariid Brugia malayi. Macrophages from both mastomys (resistant) and gerbil (susceptible) infected intraperitoneally (i.p.) with the infective larvae (L3) of B. malayi were isolated from peritoneal lavage at different time-intervals and formation rate of NO, H2O2, O2-, TNF-alpha, glutathione peroxidase and reductase was assayed. NO release was found to be significantly increased in resistant mastomys as compared to gerbils and the release was markedly suppressed by i.p. administration of the NOS inhibitor aminoguanidine (AG). The AG-treated mastomys also demonstrated significantly greater establishment of larvae which correlated well with suppressed formation of NO. Nitric oxide synergizes with superoxide to form peroxynitrite radical (potent oxidant), which is known to be more toxic per se than NO. Results indicate the possible involvement of peroxynitrite in the rapid killing of larvae in the peritoneal cavity of mastomys. In contrast, the production of H2O2 was found to be enhanced in both species indicating that B. malayi L3 could withstand the toxic effects of H2O2. The higher level of glutathione peroxidase and reductase, as observed in mastomys compared with the gerbil after larval introduction, possibly protects the cell against the injurious effect of H2O2. The TNF-alpha level remained virtually unchanged in both the hosts, suggesting an insignificant role for this cytokine in parasite establishment.

Leishmanicidal activity of phenylene bridged C2 symmetric glycosyl ureides.

A number of phenylene bridged C2 symmetric glycosyl uerides with ester (3a-f), alcohol (4a-c) and acid (5a-d) functionalities were prepared by addition of glycosyl amino esters with phenyl diisocyanates and their further reaction with LiAlH(4) or hydrolysis with LiOH. All the compounds were screened for their in vitro and in vivo antileishmanial activity. Most of the compounds exhibited good activity while two of the compounds 3e and 3f reduced the clinical dose of standard drug SSG.

Poorer NF-kappa B signaling by microfilariae in macrophages from BALB/c mice affects their ability to produce cytotoxic levels of nitric oxide to kill microfilariae.

Upon activation with microfilariae (mf), macrophages from C57Bl/6 mice showed higher nuclear factor-kappa B (NF-kappa B) but lower activating protein 1 DNA-binding activity as compared to BALB/c macrophages. The C57Bl/6 macrophages produced cytotoxic levels of nitric oxide (NO) to kill Setaria cervi mf as compared to BALB/c macrophages. Inhibition of the NF-kappa B signal by pyrrolidine dithiocarbamate (PDTC) blocked NO production and microfilaricidal activity of C57Bl/6 macrophages and inclusion of the exogenous NO generator (SNP) in the PDTC treated C57Bl/6 macrophage cultures induced mf cytotoxicity. These results underscore that the NF-kappa B signal (induced in response to mf) is important for the NO-mediated microfilaricidal activity of macrophages.

7-O-[4-methyl piperazine-1-(2-acetyl)]-2H-1-benzopyran-2-one: a novel antifilarial lead compound.

In preliminary studies we found that benzopyrones (coumarins), which are known to exert many biological activities including anti-inflammatory effect, possess promising macrofilaricidal action as well. In order to explore the possibility of combining such a macrofilaricidal activity with the microfilaricidal potential of the known piperazine pharmacophore, we synthesized a series of compounds and evaluated their antifilarial effect. In the present study, one of these compounds, 7-O-[4-methyl piperazine-1-(2-acetyl)]-2H-1-benzopyran-2-one (2), which has shown promising macrofilaricidal action against rodent filariid Litomosoides carinii in cotton rats, was evaluated against infection with Brugia malayi in Mastomys coucha and jird (Meriones unguiculatus). In the B. malayi-M. coucha system, the compound at a dose of 300 mg/kg, oral (p.o.) x5 days showed 53.6% adulticidal and 46.0% microfilaricidal activity along with 46.3% sterilization effect on the female worms. In addition, the compound interfered with the establishment of infective larvae (L(3))-induced infection to an extent of 50% at the same dose level. At 1 microM concentration it inhibited protease activity of B. malayi to 82%. The compound thus provides a novel lead for further synthesis and development of antifilarial agents with macrofilaricidal, microfilaricidal, female-sterilizing and possible larvicidal efficacy.

Modulation of inflammatory mediators by ibuprofen and curcumin treatment during chronic inflammation in rat.

Inflammation is a protective tissue response occurring in three distinct phases, acute, subacute and a chronic proliferative phase. We undertook the present study to understand the overall immune response of the body during adjuvant induced chronic inflammation in rat and the effect of ibuprofen and curcumin on this response. Inflammatory mediators were estimated on day 21 and day 35 after adjuvant injection. The level of C-reactive protein increased to 200% on day 21 and then reduced to 50% on day 35 compared to control. Curcumin and ibuprofen further reduced the increased levels at both the time intervals. Haptoglobin level decreased to 42% on day 21 but increased to 5 times of control on day 35. Curcumin and ibuprofen reduced the increased levels at day 35. No significant change was observed in Prostaglandin-E2 and Leukotriene-B4 levels and in Lymphocyte proliferation. The level of Tumor Necrosis Factor-alpha increased by three folds on day 21, but came down to 88% on day 35. Ibuprofen treatment decreased the raised level on day 21 and increased the reduced level on day 35. Interleukin-1beta increased to 2 folds on day 21 and 10 folds on day 35 which were significantly brought down by curcumin and ibuprofen. Nitric oxide level was reduced at both the time intervals, which were increased by drug treatment.

Preadult stage parasites and multiple timed exposure to infective larvae are involved in development of limb edema in Brugia malayi-infected Indian leaf monkeys (Presbytis entellus).

The pathogenesis of filarial limb edema is not known. The role of parasitological variables and parasite-mediated phenomena in the development of limb edema was investigated in the Presbytis entellus-Brugia malayi model. Infection was initiated with subcutaneous inoculation of infective third-stage larvae (L(3)), and the animals were reexposed to different doses of L(3) at the prepatent, patent, and diminishing microfilaremia (0 to 5% of peak microfilaremia count) stages of infection. A large L(3) inoculum size and repeated inoculation in the ankle region during the prepatent, patent, and diminishing microfilaremia stages of infection were found to be necessary for reproducible induction of limb edema. The preadult stage of the parasite was found to be the most potent inducer of limb edema, followed by L(5) and L(4). The presence of the proinflammatory cytokines tumor necrosis factor alpha, interleukin-1beta, and interleukin-6 in edema fluid in the leg receiving the parasite challenge indicated that the limb edema development was due to parasite-mediated cytokine responses. The absence of bacterial infection or anti-streptolysin O titer in the edema fluid and blood indicated that bacterial infection is not necessary for the development of limb edema.

Expression of adenosine deaminase and 5'-nucleotidase in artificially induced deciduoma in rat and hamster.

Enzymes adenosine deaminase (ADA) and 5-nucleotidase (5-'NT) are known to play active role in tissue/cell proliferation and differentiation. To validate this the two enzymes were studied in artificially induced deciduoma of rat and hamster. The deciduoma was induced by traumatizing one of the uterine horns of progesterone primed animals. Non traumatized horn served as control. The animals were later maintained on progesterone, given alone (Gr.I) or conjointly with estrogen (Gr.II). The weight of each uterine horn was recorded to determine the formation of deciduoma. There was no marked difference between the weights of traumatized and control horn on day 2 post-traumatization (PT), but a progressive rise was noticed after this day in both species. The ADA activity however differed, day and species wise. While in the rats of Gr.I it was low in the traumatized horn on all the days, in the hamsters it was remarkably high from day 2 to 6 PT. In the rats of Gr.II also the activity though was low in the traumatized horn, but on day 2 and 4 only; on day 6 and 7 PT it increased markedly. In hamster, on the contrary, again the enzyme activity was remarkably high on all the three days. The 5'-NT activity, however, did not show any marked difference between the two horns under Gr.I and II in both species. It was rather high in the control horn of each group. The results suggest: (I) the progesterone alone though produces a significant rise in the uterine weight of traumatized horn in both species, the ADA activity increases only in hamster, (2) under the conjoint treatment also the enzyme activity remains high in hamster; and (3) the activity of enzyme 5'-NT does not alter during the deciduoma formation in both the species.

Syntheses and immunomodulatory activity of 3-O-[2'-hydroxy-3'-N,N-disubstituted aminopropan-1'-yl]-alpha-D-glucofuranoses.

A number of 3-O-[2'-hydroxy-3'-N,N-aminopropan-1'-yl]-alpha-D-glucofuranoses were synthesised by regioselective oxirane ring opening in compound 2 with different secondary amines followed by selective deacetalisation. All the compounds were tested for their immunomodulatory potential in vitro; seven of them expressed significant immunostimulant activity.

Sand pits and bucket handles: A case of bilateral traumatic bucket-handle tears of the medial menisci.

Bucket-handle tears of the medial menisci are a common injury in young adult athletes. The incidence of acute bilateral tears occurring simultaneously is anecdotally very small and its occurrence has not been reported. We present the case of an 18-year-old man who sustained bilateral bucket-handle tears of the medial menisci while long jumping. We propose that the mechanism of injury may be analogous to that in footballers. This case report raises awareness of the occurrence of bilateral bucket-handle tears.

Immunomodulatory activity of hexapeptides related to proline rich peptide from colostrum.

Twelve analogues of an immunomodulatory hexapeptide YVPGFP (I) derived from Proline rich peptide (from colostrum) have been synthesized with modifications at positions 2, 4 and 6. In MLR assay one of the analogues exhibited approx 50% inhibition at 0.1 microg/mL concentration in contrast to prednisolone and I which caused around 70 and 20% suppression respectively, at the same concentration.