Investigations on the effects of cobalt-alkyne complexes on leukemia and lymphoma cells: cytotoxicity and cellular uptake

Cobalt-alkyne complexes represent a new class of antiproliferative drugs with high activity on cell lines derived from human solid tumors. These promising results encouraged us to evaluate also their effects against leukemia and lymphoma cells. For this purpose, we selected three cobalt complexes with (2-propyn-1-yl)acetylsalicylate (Co-ASS), 2-propyn-1-ol (Co-Prop) and diphenylacetylene (Co-Diph) ligands and investigated their growth inhibiting properties on the LAMA-84, K-562, SD-1 leukemia and U-937 lymphoma cell lines. The cobalt complexes showed high effects on LAMA-84 cells (IC(50)=7.7-16.8 microM) after 48 and 72 h of incubation, but were inactive (K-562, U-937) or low active (SD-1) on the other cell lines. The proliferation of SD-1 cells was reduced by Co-Prop (IC(50)=18.6 microM) and Co-Diph (IC(50)=7.5 microM) only after a 72 h exposure. The antiproliferative effects did not correlate with the accumulation of the drugs into the tumor cells. The time dependent uptake during 24 h determined by atomic absorption spectroscopy was comparably the same in sensitive LAMA-84 and insensitive K-562 cells.     

A sugar ester and an iridoid glycoside from Scrophularia ningpoensis

From cytotoxic extracts of the roots of Scrophularia ningpoensis Hemsl. (Scrophulariaceae) a new sugar ester, ningposide D (3-O-acetyl-2-O-p-methoxycinnamoyl-alpha(beta)-L-rhamnopyranose) (1) and a new iridoid glycoside, scrophuloside B4 (6-O-(2'-O-acetyl-3'-O-cinnamoyl-4'-O-p-methoxycinnamoyl-alpha-L-rhamnopyranosyl) catalpol) (2) along with known compounds: oleanonic acid (3), ursolonic acid (4), cinnamic acid (5), 3-hydroxy-4-methoxy benzoic acid (6), 5-(hydroxymethyl)-2-furfural (7) and beta-sitosterol (8) were isolated. The structures of the new compounds were elucidated by spectral data (1, 2D NMR, EI, HRESI-MS and MS/MS). Oleanonic acid (3) and ursolonic acid (4) were found to be cytotoxic against a series of human cancer cell lines with IC50=4.6, 15.5 microM on MCF7; 4.2, 14.5 microM on K562; 14.8, 44.4 microM on Bowes; 24.9, 43.6 microM on T24S; 61.3, 151.5 microM on A549, respectively. Beta-sitosterol (8) inhibited Bowes cells growth at IC50=36.5 microM. Scrophuloside B4 (2) showed activity on K562 and Bowes cells at IC50=44.6, 90.2 microM, respectively.     

Cytotoxicity and cellular accumulation of palladium(II) complexes of tetracyclines

We studied the cytotoxic effect and the uptake of Pd(II) complexes of doxycycline (Dox), [Pd(Dox)Cl2], and tetracycline (Tc), [Pd(Tc)Cl2], in chronic myelogenous leukemia cells. The effect of the compounds on macrophage viability was also investigated. Compound 1 is more effective than compound 2 in inhibiting the growth of K562 cells with the IC(50) values of 14.44 and 34.54 microM, respectively. There is a good correlation between cell-growth inhibition and intracellular metal concentrations, determined by inductively coupled plasma atomic emission spectroscopy (ICP-AES). Incubation of the cells with equitoxic concentrations of both compounds yields approximately the same intracellular Pd concentration. At the IC(50) doses, intracellular concentration is ca. 33 x 10(-16) mol/cell for both compounds 1 and 2. This suggests that more [Pd(Tc)Cl2] is needed to produce a cytotoxic effect, because it enters cells more slowly. Both compounds up to 16 microM did not affect the viability of mouse peritoneal macrophages after a 48-h incubation. After 72 h of incubation, the IC(50) values are 22 for [Pd(Dox)Cl2] and 40 microM for [Pd(Tc)Cl(2)]. Therefore, the cytotoxic effect in cancer cells exhibited by both compounds is higher than their effect in macrophages.     

Cadmium- and chromium-induced oxidative stress, DNA damage, and apoptotic cell death in cultured human chronic myelogenous leukemic K562 cells, promyelocytic leukemic HL-60 cells, and normal human peripheral blood mononuclear cells

Sodium dichromate [Cr(VI)] and cadmium chloride [Cd(II)] are both cytotoxic and mutagenic. This study examined the toxic and apoptotic potentials of these two cations on three cell types in vitro, namely, human chronic myelogenous leukemic (CML) K562 cells, promyelocytic leukemic HL-60 cells, and normal human peripheral blood mononuclear cells. The cells were incubated with 0-100 microM concentrations of the two cations for 0, 24, or 48 hours at 37 degrees C. Both Cr(VI) and Cd(II) induced changes in intracellular oxidized states of cells, which were detected using laser scanning confocal microscopy. Cell cycle modulation and apoptosis of the K562 cells by Cr(VI) and Cd(II) were determined by flow cytometry. Significant decreases in the G2/M phase were observed in the Cr(VI) and Cd(II) treated CML cells compared with untreated cells. At 12.5 microM, Cr(VI) induced greater apoptosis in K562 cells as compared with Cd(II). In the K562 cells, 2.2- and 3.0-fold increases in DNA fragmentation were observed following incubation with 12.5 and 25 microM Cr(VI), respectively, and 1.2- and 1.7-fold increases in DNA fragmentation were observed with Cd(II). Furthermore, approximately 2.7- and 4.9-fold increases in cytochrome c reduction were observed following incubation with 12.5 and 25 microM Cr(VI), respectively, and 1.6- and 3.3-fold increases in cytochrome c reduction were observed with Cd(II), demonstrating enhanced production of superoxide anion. Approximately 3.1 to 6-fold increases in hydroxyl radical production were observed following incubation of the K562 cells with these cations at 12.5 and 25 microM concentrations. These results in K562 cells were compared with promyelocytic leukemic HL-60 cells and normal human peripheral blood mononuclear cells. More pronounced effects were observed on K562 and HL-60 cells, and much lesser effects were observed on normal human peripheral blood mononuclear cells. The results demonstrate that both cations are toxic, producing oxidative tissue damage and apoptosis. Furthermore, more drastic effects were observed on K562 and HL-60 cells as compared with normal human peripheral blood mononuclear cells.     

3-Deazauridine triggers dose-dependent apoptosis in myeloid leukemia cells and enhances retinoic acid-induced granulocytic differentiation of HL-60 cells

Therapeutic nucleoside analogue 3-deazauridine (DU) exerts cytotoxic activity against cancer cells by disruption of DNA synthesis resulting in cell death. The present study evaluates whether DU alone at doses 2.5-15 microM or in combination with all trans retinoic acid (RA) or dibutyryl cAMP (dbcAMP) is effective against myelogenous leukemia. The data of this study indicate that DU induces dose-dependent cell death by apoptosis in myeloid leukemia cell lines HL-60, NB4, HEL and K562 as demonstrated by cell staining or flow cytometry and agarose gel electrophoresis. 24h-treatment with DU produced dose-dependent HL-60 cell growth inhibition and dose-independent S phase arrest that was not reversed upon removal of higher doses of DU (10-15 microM). Exposition to nontoxic dose of DU (2.5 microM) for 24h followed by RA or dbcAMP and 96 h-cotreatment with DU significantly enhanced RA- but not dbcAMP-mediated granulocytic differentiation. Cell maturation was paralleled with an increase in the proportion of cells in G1 or G2+M phase. We conclude that, depending on the dose or the sequence of administration with RA, an inhibitor of DNA replication, DU triggers a process of either differentiation or apoptosis in myeloid leukemia cells.     

Induction of apoptosis in K562/ADM cells by gamma-linolenic acid involves lipid peroxidation and activation of caspase-3

Numerous studies have revealed that gamma-linolenic acid (GLA) possesses effective tumoricidal properties while not inducing damage to normal cells or creating harmful systemic side effects. It can exert anti-tumor efficacy against a variety of cancers including leukemia. However, little is known about the effects of GLA on leukemia resistant to chemotherapy, emerging as a serious clinical problem. The present study tested GLA-induced apoptosis in K562/ADM multidrug-resistant (MDR) leukemic cells and investigated its possible mechanisms. Using cell viability, fluorescent staining of nuclei, flow cytometric Annexin V/PI double staining and lactate dehydrogenase (LDH) release, we found that GLA could inhibit cell growth and induce apoptosis and secondary necrosis. The results showed that incubation with GLA concentrations of 10-60 microg/ml caused a dose- and time-dependent decrease of K562/ADM cell viability, and the IC50 value was 50.5 microg/ml at 24 h and 31.5 microg/ml at 48 h. Flow cytometry using Annexin V/PI double staining assessed apoptosis, necrosis and viability. Typical apoptotic nuclei were shown by staining of K562/ADM cells with DNA-binding fluorochrome Hoechst 33342, characterized by chromatin condensation and nuclear fragmentation. On the other hand, after treated K562/ADM cells with 20 microg/ml GLA for 48 h and with 40 microg/ml GLA for 12 h, the LDH release significantly increased, indicated losses of plasma membrane integrity and presence of necrosis. Further, the inhibition of GLA-induced apoptosis by a pan-caspase inhibitor (z-VAD-fmk) suggested the involvement of caspases. The increase of caspase-3 activity with GLA concentration confirmed its role in the process. The results also showed that the malondialdehyde (MDA) content was also significantly elevated, and antioxidant BHT could block GLA cytotoxity, indicating the cytotoxity induced by GLA may be due to lipid peroxidation.     

Phenolic glyscosides, a new class of human recombinant nucleotide pyrophosphatase phosphodiesterase-1 inhibitors

Cytotoxicity and kinetic studies of phenolic glycosides, benzoyl salireposide (1) and salireposide (2), isolated from Symplocos racemosa, were performed against phosphodiesterase I enzyme from snake venom and human nucleotide pyrophosphatase phosphodiesterase-1. Lineweaver-Burk and Dixon plots and their secondary replots showed that these compounds are pure non-competitive inhibitors of both enzymes. K(i) Values of compounds 1 and 2 were found to be 360 and 1000 microM, respectively, against human nucleotide pyrophosphatase phosphodiesterase, and 525 and 1100 microM, respectively, against snake venom phosphodiesterase. IC(50) values of compounds 1 and 2 are 90 microM +/- 0.04 and 383 microM +/- 0.03, respectively, against human nucleotide pyrophosphatase phosphodiesterase and 171 microM +/- 0.02 and 544 microM +/- 0.021, respectively, against snake venom phosphodiesterase. Both compounds were found to be nontoxic up to concentration of 500 microM/mL as >90% cells were viable after 3 h of incubation. These compounds are potential candidates for the therapy of arthritis.     

Synthesis and potent antileukemic activities of 10-benzyl-9(10H)-acridinones

A novel series of 10-benzyl-9(10H)-acridinones and 1-benzyl-4-piperidones were synthesized and tested for their in vitro antitumor activities against CCRF-CEM cells. Assay-based antiproliferative activity study using CCRF-CEM cell lines revealed that the acridone group and the substitution pattern on the benzene unit had significant effect on cytotoxicity of this series of compounds, among which 10-(3,5-dimethoxy)benzyl-9(10H)-acridinone (3b) was found to be the most active compound with IC(50) at about 0.7 microM. Compound 3b was also found to have antiproliferative activity against two other human leukemic cell lines K562 and HL60 using the MTT assay. The antitumor effect of 3b is believed to be due to the induction of apoptosis, which is further confirmed by PI (Propidium iodide) staining and Annexin V-FITC/PI staining assay using flow cytometry analysis.     

Selenium-containing naphthalimides as anticancer agents: design, synthesis and bioactivity

Selenium analogues (4b-4h, and 4j) of two known sulfur compounds were synthesized and tested their anticancer activities. The selenium compound 4b had comparable activity with its sulfur analogue 4a, while DNA-binding study showed these two compounds had similar interaction with ct-DNA, the K(b) was 8.23 and 2.36, respectively. The primary results showed that most compounds had moderate anticancer activities with IC(50) values between 10(-6) and 10(-5) M. Another selenium analogue 4j showed the highest activity with the IC(50) values around 5.3 μM against K562 and MCF-7 cell lines. More importantly, the organochalcogen compounds exhibited stronger anticancer activities against K562 cell line than the other cell lines tested.     

Arsenic trioxide induces apoptosis in chronic myelogenous leukemia K562 cells: possible involvement of p38 MAP kinase

Arsenic trioxide (As(2)O(3)) was recently demonstrated to be an effective inducer of apoptosis in patients with relapsed acute promyelocytic leukemia (APL) as well as in patients with APL in whom all-trans-retinoic acid and conventional chemotherapy failed. Chronic myelogenous leukemia cells are highly resistant to chemotherapeutic drugs. To determine if As(2)O(3) might be useful for the treatment of chronic myelogenous leukemia, we examined the ability of As(2)O(3) to induce apoptosis in K562 cells. In vitro cytotoxicity of As(2)O(3) was evaluated in K562 cells by a MTT assay; the IC(50) value for As(2)O(3) was determined to be 10 microM. When analyzed by agarose gel electrophoresis, the DNA fragments became evident after incubation of the cells with 20 microM As(2)O(3) for 24 h. We also found morphological changes and chromatin condensation of the cells undergoing apoptosis. Activation of caspase-3 was observed 6 h after treatment with 20 microM As(2)O(3) by a Western blot analysis. Next, we examined the MAP kinase-signaling pathway of As(2)O(3)-induced apoptosis in K562 cells. As(2)O(3) at 10 microM strongly induced the activation of p38 and JNK 1/2, while ERK 1/2 was inhibited. In addition, pretreatment of SB203580, a specific inhibitor of p38, inhibited As(2)O(3) induced apoptotic cell death. These results suggest that As(2)O(3) is able to induce the apoptotic activity in K562 cells, and its apoptotic mechanism may be associated with the activation of p38.     

Anti-leukemic activities of Dictyostelium secondary metabolites: a novel aromatic metabolite, 4-methyl-5-n-pentylbenzene-1,3-diol, isolated from Dictyostelium mucoroides suppresses cell growth in human leukemia K562 and HL-60 cells

It has previously been shown that DIF-1, a differentiation-inducing factor of the cellular slime mold Dictyostelium discoideum, possesses antitumor activities in mammalian tumor cells and that neuronal differentiation of PC12 cells can be induced with furanodictines (FDs), aminosugar analogs found in D. discoideum, or dictyoglucosamines (DGs), N-acetyl glucosamine derivatives (DG-A from D. purpureum and DG-B from D. discoideum). Thus, cellular slime molds are attractive natural resources that may provide valuable lead compounds to be utilized in the field of pharmacology and medicine. In this study, we have isolated a novel aromatic compound, 4-methyl-5-n-pentylbenzene-1,3-diol (MPBD), from fruiting bodies of the cellular slime mold D. mucoroides and assessed the in vitro antiproliferative activities of MPBD, FDs, and DGs in human leukemia K562 and HL-60 cells. MPBD at 20-80 microM dose-dependently suppressed cell growth in both K562 and HL-60 cells. While FDs at 10-80 microM did not affect cell growth, DGs at 10-40 microM dose-dependently suppressed cell growth in the cells. Although we failed to find the roles of FDs and DGs in the original organisms, MPBD at 5-20 microM was found to promote stalk cell formation in D. discoideum. The present results indicate that MPBD, DGs or their derivatives may have therapeutic potential in the treatment of cancer and confirm our expectations regarding cellular slime molds as drug resources.     

Tomoeones A-H, cytotoxic phloroglucinol derivatives from Hypericum ascyron

Phloroglucinol derivatives tomoeones A-H (1-8) and three known compounds were isolated from leaves of Hypericum ascyron. Their structures were established based on spectroscopic analyses. They are all acylphloroglucinol derivatives possessing a spiro skeleton with geminal isoprenyl groups and a monoterpene moiety, and they are stereoisomers to each other at C-4 and C-13. They appear to be a class of phloroglucinol derivatives. Cytotoxicities of the isolated phloroglucinol derivatives against human tumor cell lines, including multidrug-resistant (MDR) cancer cell lines, were evaluated. Tomoeone F (6) demonstrated significant cytotoxicity against KB cells with an IC50 value of 6.2 microM. Compound 6 was also cytotoxic against MDR cancer cell lines (KB-C2 and K562/Adr), which was more potent than doxorubicin.     

Benzoyl nitrogen mustard derivatives of benzoheterocyclic analogues of netropsin: synthesis and biological activity

Synthesis, DNA binding properties and biological activity of a series of bis-benzoheterocycle derivatives 5-11, structurally related to the natural dipyrrole antitumor agent netropsin, and tethered to a benzoyl nitrogen mustard (BAM) as alkylating moiety is reported and structure-activity relationships determined. These compounds 5-11 have been evaluated for sequence selective alkylating properties and cytotoxicity against murine L1210 and human K562 leukaemia cells. Using as target sequence a portion of the long terminal repeat of the type-1 human immunodeficiency virus, we found that these compounds induce similar patterns of DNA fragmentation. In addition, the results obtained indicate that all synthesized compounds retain a good antiproliferative activity in the submicromolar range, and generally are more active against L1210 than K562 cells. With respect to both these cell lines, compounds 6, 7, 10 and 11 showed the greatest potency, ranging from 0.3 to 1 microM, while compounds 8 and 9 exhibit the lowest activity (IC(50)=2-12 microM). Among compounds 5-11, the derivative 11 was found to be the most potent member of this class and it is 5 and 10-fold less active than the bis-pyrrole counterpart 2 against K562 and L1210 cell lines, respectively. For compound 11, the substitution of the C-terminus benzofurane with N-methylindole and indole (to give the compounds 5 and 6, respectively) led to a decrease in cytotoxicity, which is more evident against the K562 cell line. Finally, differences were found among compounds 5-11 in induction of K562 differentiation. Some of them (compounds 7, 8 and 9) are potent inducers of erythroid differentiation of K562 cells, and could be proposed for differentiation anti-cancer therapy.     

Induction of differentiation and apoptosis in three human leukemia cell lines by a new compound from Dendrostellera lessertii

It has previously been shown that Dendrostellera lessertii(Thymelaeaceae)has stronganticancer activity.In this study,the antileukemic activity of another new compound from the same plantextract is reported.Promyelocytic(NB4 and HL-60)and erythroleukemia(K562)cells were cultured in thepresence of various concentrations of the new compound(0.5-3.0 μtg/ml)for 3d.The cell numbers werethen determined by trypan blue exclusion test.The new compound inhibited growth and proliferation ofNB4,HL-60 and K562 with IC_(50) values of 1.5,2.0 and 2.5μg/ml,respectively.We also found that the newcompound inhibited cell proliferation in a dose-and time-dependent manner.At low concentrations and after48h of treatment,approximately 50%-70% of NB4 and HL-60 cells were differentiated to monocyte/macrophage lineage and approximately 30%-40% of the treated K562 cells were differentiated in the mega-karyocytic lineage,as evidenced by morphological changes and nitro blue tetrazolium reduction assays.Results of Hoechst 33258 staining also indicated that the new compound induced NB4 and HL-60 cellapoptosis at their respective IC_(50) values after 72h of treatment.Based on the present data,the new com-pound seems a good candidate for further evaluation as an effective chemotherapeutic agent acting throughinduction of differentiation and apoptosis.     

Ecto-alkaline phosphatase activity identified at physiological pH range on intact P19 and HL-60 cells is induced by retinoic acid

The activity of membrane-bound alkaline phosphatase (ALP) expressed on the external surface of cultured murine P19 teratocarcinoma and human HL-60 myeloblastic leukemia cells was studied at physiological pH using p-nitrophenylphosphate (pNPP) as substrate. The rate of substrate hydrolysis catalyzed by intact viable cells remained constant for eight successive incubations of 30 min and was optimal at micromolar substrate concentrations over the pH range 7.4-8.5. The value of apparent K(m) for pNPP in P19 and HL-60 cells was 120 microM. Hydrolytic activity of the ecto-enzyme at physiological pH decreased by the addition of levamisole, a specific and noncompetitive inhibitor of ALP (K(i) P19 = 57 microM; K(i) HL-60 = 50 microM). Inhibition of hydrolysis was reversed by removal of levamisole within 30 min. Retinoic acid (RA), which promotes the differentiation of P19 and HL-60 cells, induced levamisole-sensitive ecto-phosphohydrolase activity at pH 7.4. After its autophosphorylation by ecto-kinase activity, a 98-kDa membrane protein in P19 cells was found to be sensitive to ecto-ALP, and protein dephosphorylation increased after incubation of cells with RA for 24 h and 48 h. Orthovanadate, an inhibitor of all phosphatase activities, blocked the levamisole-sensitive dephosphorylation of the membrane phosphoproteins, while (R)-(-)-epinephrine reversed the effect by complexation of the inhibitor. The results demonstrate that the levamisole-sensitive phosphohydrolase activity on the cell surface is consistent with ecto-ALP activity degrading both physiological concentrations of exogenously added substrate and endogenous surface phosphoproteins under physiological pH conditions. The dephosphorylating properties of ecto-ALP are induced by RA, suggesting a specific function in differentiating P19 teratocarcinoma and HL-60 myeloblastic leukemia cells.     

Synthesis and in vitro antitumor activity of 4(3H)-quinazolinone derivatives with dithiocarbamate side chains

A series of 4(3H)-quinazolinone derivatives with dithiocarbamate side chains were synthesized and tested for their in vitro antitumor activity against human myelogenous leukemia K562 cells. Among them, (3,4-dihydro-2-methyl-4-oxoquinazolin-6-yl)methyl 4-(4-fluorophenyl)piperazine-1-carbodithioate 8q exhibited significant inhibitory activity against K562 cells with IC(50) value of 0.5 microM.     

3-Hydrogenkwadaphnin targets inosine 5'-monophosphate dehydrogenase and triggers post-G1 arrest apoptosis in human leukemia cell lines

3-Hydrogenkwadaphnin (3-HK) is a recently characterized daphnane-type compound isolated from Dendrostellera lessertii with high anti-tumor activity in animal models. Herein, we report on time- and dose-dependent effects of this compound on growth, differentiation, IMPDH inhibition, cell cycle and apoptosis of a panel of human leukemia cell lines (HL-60, K562 and Molt4). The drug decreased the growth of leukemia cells in less than 24 h of treatment. However, longer exposure times and/or higher concentrations were required to promote cell apoptosis. Cell cycle analysis revealed the accumulation of cells in their G1 phase as early as 12 h after drug exposure but sub-G1 population was recorded after 24 h. Occurrence of apoptosis was constantly accompanied by morphological (staining with DNA-binding dyes) and biochemical (DNA fragments) variations among drug-treated cells. Despite these observations, non-activated normal human PBL were insensitive to the drug action. In addition, treatment of PHA-activated PBL, K562, Molt4 and HL-60 cells with a single dose of the drug for 24 h led to the inhibition of IMPDH activity by almost 37, 38, 44 and 50%, respectively. In contrast, no difference in IMPDH activities were seen between normal PBL and the drug treated PBL cells. Restoration of the depleted GTP concentration by exogenous addition of guanosine (25-50 microM) reversed the drug effects on cell growth, DNA fragmentation and apoptosis. Furthermore, the drug effects were potentiated by exogenous addition of hypoxanthine to the drug-treated cells. Reduction of the drug potency on the non-proliferative (retinoic acid treated) HL-60 cells by almost 40%, compared to the proliferative cells, clearly shows type II IMPDH as one of the main targets of the drug. These results suggest that 3-HK may be a powerful candidate for treatment of leukemia.     

Survival of Flavobacterium psychrophilum in rainbow trout (Oncorhynchus mykiss) serum in vitro

Virulent and non-virulent strains of Flavobacterium psychrophilum of different serotypes were examined for survival and growth in non-immune and immune rainbow trout serum, in vitro. A majority of the examined strains consumed complement of non-immune serum, but the complement cascade was not able to cause an immediate (after 3 h incubation) notable reduction in viability of the inoculated cells. After 24 h incubation a more pronounced reduction in the number of viable bacteria was observed in untreated serum as well as in serum heated at 45 degrees C. In serum heated at 56 degrees C this reduction in cell number was not observed, but an increase in cell number did not occur either. The serum survival of one of the examined strains was different from the others in showing cell multiplication after 24 h incubation in normal as well as heat-treated (45 and 56 degrees C) serum. In immune serum no immediate reduction in viability of inoculated cells, of all tested strains, was observed. The number of viable cells showed a slow decrease or remained almost unchanged for up to 72 h post-inoculation in untreated serum, at 5 degrees C as well as 15 degrees C. In heat-treated serum (45 degrees C) the number of viable cells decreased slowly at 5 degrees C and 15 degrees C for up to 72 h. The results suggest that the examined strains were unaffected by the alternative complement reaction present in fish serum as well as by antibodies against F. psychrophilum. However, some unknown component(s) in the fish sera, or lack of nutrients or essential growth factors, inhibited the growth of most of the examined strains in the tested fish sera.     

19-Nor-1alpha,25-dihydroxyvitamin D2 (paricalcitol) exerts anticancer activity against HL-60 cells in vitro at clinically achievable concentrations

19-Nor-1alpha,25-dihydroxyvitamin D2 (paricalcitol) is an analogue of 1,25(OH)2D3 with reduced calcemic effects that is approved in the United States for the suppression of parathyroid hormone in chronic renal failure. Paricalcitol has anticancer activity in prostate cancer cells. We tested the effects of paricalcitol on the HL-60 leukemia cells, studying cellular differentiation, cell cycle changes, apoptosis and cellular proliferation. Paricalcitol at 10(-8)M concentration induced the maturation of HL-60 cells in a time-dependent manner, as shown by increased expression of CD11b differentiation surface antigen. The ability of HL-60 cells to reduce nitroblue tetrazolium (NBT) was markedly increased after exposure to paricalcitol at 10(-8)M for 72 h. Paricalcitol inhibited colony formation of HL-60 cells in a soft agar semisolid media after 10-day incubation (estimated IC50 of 5 x 10(-9) M. Exposure to 10(-8)M paricalcitol for 72 h increased the number of cells in G0/G1 phase, and decreased the number of cells in S phase, and significantly increased the number of HL-60 cells undergoing apoptosis. The concentration required to achieve inhibition of growth of HL-60 cells is comparable to clinically achievable levels. These findings support the clinical evaluation of paricalcitol as an antileukemia agent.