Cell intrinsic and extrinsic factors synergize in mice with haploinsufficiency for Tp53, and two human del(5q) genes, Egr1 and Apc.

Therapy-related myeloid neoplasms (t-MN) are a late complication of the successful use of cytotoxic therapy for patients with cancer. A heterozygous deletions of the long arm of chromosome 5 [del(5q)], observed in 40% of patients, is associated with prior exposure to alkylating agents, and a high frequency of TP53 loss or mutation. In previous studies, we demonstrated that haploinsufficiency of 2 del(5q) genes, Egr1, and Apc, individually play a role in the pathogenesis of hematologic disease in mice. We now show that loss of one copy of Egr1 or Tp53 in an Apc haploinsufficient background (Apc (del/+)) accelerated the development of a macrocytic anemia with monocytosis, early features of t-MN. The development of anemia was significantly accelerated by treatment of mice with the alkylating agent, N-ethyl-N-nitrosourea (ENU), regardless of the levels of expression of Egr1 and Tp53. Transplantation of either wild type; Egr1(+/-); Tp53(+/-); Apc(del/+); or Egr1(+/-), Apc(del/+) bone marrow cells into lethally irradiated Apc(del/+) recipients resulted in rapid development of anemia that was further accelerated by administration of ENU to recipients, demonstrating that the Apc(del/+)-induced anemia was cell extrinsic and potentiated by ENU mutagenesis. These data emphasize the synergistic role of cell intrinsic and cell extrinsic (microenvironment) factors in the pathogenesis of t-MN, and raise awareness of the deleterious effects of cytotoxic therapy on the stromal microenvironment.

Nonclinical Safety Profile of BMS-986001, a Nucleoside Transcriptase Inhibitor for Combination Retroviral Therapy.

Nucleoside reverse transcriptase inhibitors (NRTIs)/nucleotide reverse transcriptase inhibitors are key components of combination antiretroviral therapy for HIV infection. First-generation NRTIs are associated with mitochondrial toxicity in patients, mainly due to inhibition of human DNA polymerase γ (hDNA polγ) that manifests as adverse events such as lipodystrophy, lactic acidosis, myopathy, cardiomyopathy, or nephropathy in patients. In chronic nonclinical studies in rodents and nonrodents, eukaryotic (host) mitochondrial toxicity manifests as some drug-specific toxicities similar to human toxicity. BMS-986001, a novel thymidine analog with minimal hDNA polγ inhibition, has demonstrated antiretroviral activity in early clinical studies. The primary toxicity of BMS-986001 in rats and monkeys is bone marrow dyserythropoiesis with associated decreases in red blood cell mass. Additionally, at high doses, severe platelet reductions accompanied by cutaneous petechiae began during weeks 8 and 11 in 3 of 60 monkeys in chronic toxicity studies. In a 6-month study, platelet reductions required euthanasia of the 2 affected monkeys (300 mg/kg/d) at week 14, but with dose reduction (200 mg/kg/d) remaining monkeys had no platelet changes. One affected monkey (200 mg/kg/d) in a 9-month study completed dosing and its platelet counts recovered during a 1-month recovery. Formation of platelet-bound immunoglobulin in the presence of BMS-986001, together with rapid and complete platelet recovery in the absence of BMS-986001, suggested that platelet decreases in monkeys may be immune mediated. No findings indicative of mitochondrial toxicity were observed in rats or monkeys given BMS-986001, suggesting an improved safety profile compared to marketed NRTI or tenofovir disoproxil fumarate.

An ethanolic extract of black cohosh causes hematological changes but not estrogenic effects in female rodents.

Black cohosh rhizome (Actaea racemosa) is used as a remedy for pain and gynecological ailments; modern preparations are commonly sold as ethanolic extracts available as dietary supplements. Black cohosh was nominated to the National Toxicology Program (NTP) for toxicity testing due to its widespread use and lack of safety data. Several commercially available black cohosh extracts (BCE) were characterized by the NTP, and one with chemical composition closest to formulations available to consumers was used for all studies. Female B6C3F1/N mice and Wistar Han rats were given 0, 15 (rats only), 62.5 (mice only), 125, 250, 500, or 1000 mg/kg/day BCE by gavage for 90 days starting at weaning. BCE induced dose-dependent hematological changes consistent with a non-regenerative macrocytic anemia and increased frequencies of peripheral micronucleated red blood cells (RBC) in both species. Effects were more severe in mice, which had decreased RBC counts in all treatment groups and increased micronucleated RBC at doses above 125 mg/kg. Dose-dependent thymus and liver toxicity was observed in rats but not mice. No biologically significant effects were observed in other organs. Puberty was delayed 2.9 days at the highest treatment dose in rats; a similar magnitude delay in mice occurred in the 125 and 250 mg/kg groups but not at the higher doses. An additional uterotrophic assay conducted in mice exposed for 3 days to 0.001, 0.01, 0.1, 1, 10, 100 and 500 mg/kg found no estrogenic or anti-estrogenic activity. These are the first studies to observe adverse effects of BCE in rodents.

Hematological effects of valproate in psychiatric patients: what are the risk factors?

BACKGROUND: Sodium valproate is an anticonvulsant that is also one of the common treatments used for bipolar disorder. The present study was conducted in psychiatric patients with the aim of examining the effects of valproate on hematological parameters and to explore any association with sex and age. METHODS: A list of all psychiatric patients who underwent valproate level estimation in the years 2004-2008 in Newcastle upon Tyne was drawn from the biochemistry database of the local hospital. The names and date of births of these patients were used to draw corresponding hematological data, including hemoglobin, white blood cell count, mean corpuscular volume (MCV), and platelet count, conducted on the same day or within a week of the valproate level measurement. RESULTS: : The data from 126 patients were analyzed. The prevalence of thrombocytopenia (platelet count, <150,000/microL) was found to be approximately 5%. In female subjects, a significant negative correlation was found between serum valproate level and platelet count; also, a positive correlation between valproate level and MCV was found. Neither correlation was found in male subjects. The risk of a low platelet count was found to be significantly increased at serum valproate level above 80 microg/mL in female subjects. The regression analysis in female patients showed a trend toward fall in platelet count and an increase in MCV with increasing age. CONCLUSIONS: In psychiatric patients on valproate therapy, close monitoring of full blood count is required in women particularly at valproate serum level above 80 microg/mL. This may be particularly important in older patients.

Lamivudine-associated macrocytosis in HIV-infected patients.

Macrocytosis has been increasingly observed in HIV-infected population and is an early indicator of bone marrow toxicity. A retrospective study was conducted among these patients in a HIV clinic to assess risk factors of macrocytosis. We found that lamivudine was strongly associated with macrocytosis (OR = 24.6 [2.9-3223.0], P = 0.001). This association may be due to lamivudine's effect on erythrocyte synthesis.

Macrocytosis secondary to hydroxyurea therapy.

A 10-year-old, male neutered Shetland Sheepdog was presented to the University of Florida for evaluation of a well-granulated mast cell tumor. Hydroxyurea therapy was instituted and serial CBCs showed persistent mild anemia and macrocytosis without a corresponding increase in polychromasia. The dog's MCV increased progressively, reaching its highest value of 100.0 fL after 6 months of treatment, and a diagnosis of macrocytosis associated with hydroxyurea therapy was made. The dog's increase in MCV was prominent, and rapidly decreased after the drug was discontinued, consistent with previous observations in human and canine subjects treated with hydroxyurea. Hydroxyurea is a cytotoxic chemotherapeutic agent used in a variety of conditions in human and veterinary medicine, and megaloblastic changes associated with its use have been described in multiple species. This report shows that hydroxyurea treatment is a differential diagnosis for prominent macrocytosis in dogs in the absence of other signs of erythrocyte regeneration.

Life-threatening maternal and fetal macrocytic anemia from antiretroviral therapy.

BACKGROUND: Antiretroviral therapy is recommended for human immunodeficiency virus (HIV)-infected patients during pregnancy to reduce the vertical transmission to the newborn. Complications from this therapy are uncommon. CASE: A 38-year-old HIV-positive pregnant woman was treated with lamivudine and zidovudine. At 28 weeks of gestation, her hemoglobin had fallen to 4.6 g/dL with an mean corpuscular volume (MCV) of 126 microm. At 36 weeks the fetal biophysical profile was abnormal. A pale hydropic infant was delivered via emergency cesarean, with a hemoglobin of 2.1 gm and MCV of 131 microm. The newborn hemoglobin normalized after withdrawal of the neonatal retroviral therapy. CONCLUSION: Maternal-fetal macrocytic anemia may complicate antiretroviral therapy.

Lifetime treatment of mice with azidothymidine (AZT) produces myelodysplasia.

AZT has induced a macrocytic anemia in AIDS patients on long term AZT therapy. It is generally assumed that DNA elongation is stopped by the insertion of AZT into the chain in place of thymidine thus preventing the phosphate hydroxyl linkages and therefore suppresses hemopoietic progenitor cell proliferation in an early stage of differentiation. CBA/Ca male mice started on AZT 0.75 mg/ml H2O at 84 days of age and kept on it for 687 days when dosage reduced to 0.5 mg/ml H2O for a group, another group removed from AZT to see recovery, and third group remained on 0.75 mg. At 687 days mice that had been on 0.75 mg had average platelet counts of 2.5 x 10(6). Histological examination on 9 of 10 mice with such thrombocytopenia showed changes compatible with myelodysplastic syndrome (MDS). A variety of histological patterns was observed. There were two cases of hypocellular myelodysplasia, two cases of hypersegmented myelodysplastic granulocytosis, two cases of hypercellular marrow with abnormal megakaryocytes with bizarre nuclei, one case of megakaryocytic myelosis associated with a hyperplastic marrow, dysmyelopoiesis and a hypocellular marrow and two cases of myelodysplasia with dyserythropoiesis, hemosiderosis and a hypocellular marrow. Above mentioned AZT incorporation may have induced an ineffective hemopoiesis in the primitive hemopoietic progenitor cells, which is known to be seen commonly in the myelodysplastic syndrome.

Macrocytosis associated with divalproex treatment.

Divalproex (DVP) is increasingly prescribed to children and adolescents in psychiatric practices. Among the hematologic adverse effects of DVP, decreased platelet count is well described in the medical literature. However, to date, few studies describe the occurrence of macrocytosis as an adverse effect of divalproex. We report two cases of pediatric patients who developed macrocytosis and decreasing platelet counts secondary to DVP treatment. Because macrocytosis remained an asymptomatic nonprogressive condition for our patients, we support the recommendation for closer surveillance of the patients' complete blood counts for development of anemia in addition to thrombocytopenia.

Hematologic and trace element alterations following chronic maternal ingestion of propylthiourea.

Rats with diets containing 0.2% propylthiouracil (PTU) throughout gestation had progeny with persistent cyanosis and high neonatal mortality. Histological and histochemical studies failed to reveal lung abnormalities in these pups. Studies of the blood of PTU-fed dams demonstrated that hemoglobin, packed cell volume, and numbers of erythrocytes were significantly reduced. In their 21-day fetal young, erythrocytopenia was accompanied by an elevated mean corpuscular volume and a reduced mean corpuscular hemoglobin concentration. Imprints of marrow from dams and of liver and spleens of the young showed normoblastic erythropoiesis. A granulocytic leucocytosis was present in the blood of the PTU-fed dams, whereas their progeny had a granulocytopenic leucopenia. Tissue concentrations of copper, zinc, manganese, magnesium and iron were determined. The most striking changes observed were the significant elevations of copper in the dams' brain, liver and kidneys. No changes in the concentration of any of the trace minerals were found in the livers of the pups. Food restriction to the dam failed to significantly alter maternal or fetal hematologic or trace element concentrations as compared with controls. It is evident that PTU, when fed to pregnant rats, has demonstrable effects on erythropoiesis, granulocytopoiesis, and maternal trace element distribution. It is not presently known whether these phenomena are interrelated.

Anemia and genotoxicity induced by sub-chronic intragastric treatment of rats with titanium dioxide nanoparticles.

Titanium dioxide nanoparticles (TiO2 NPs) are widely used for their whiteness and opacity. We investigated the hematological effects and genotoxicity of anatase TiO2 NPs following sub-chronic oral gavage treatment. TiO2-NPs were characterized by X-ray diffractometry (XRD), X-ray photoelectron spectroscopy (XPS), and transmission electron microscopy (TEM). Wistar rats were treated with anatase TiO2 NPs by intragastric administration for 60 days. Hematological analysis showed a significant decrease in RBC and HCT and a significant increase in MCV, PLT, MPV and WBC at higher doses. Furthermore, abnormally shaped red cells, sometimes containing micronuclei, and hyper-segmented neutrophil nuclei were observed with TiO2 NPs treatment. The micronucleus test revealed damage to chromosomes in rat bone marrow at 100 and 200mg/kg bw; the comet assay showed significant DNA damage at the same doses.

Myeloneuropathy and macrocytosis associated with nitrous oxide abuse.

After four months of excessive nitrous oxide (N2O) exposure, a dentist had myeloneuropathy with spastic paraparesis, Lhermitte's sign, sensory shading, loss of position and vibration sense, ataxia, and impotence. Macrocytosis, hypersegmented neutrophils, and a reduced vitamin B12 level were associated with normal findings on gastric analysis and Schilling test. Complete hematologic and neurologic recovery followed N2O avoidance and vitamin therapy for six months.

Oral treatment with trimethoprim-sulfamethoxazole and zidovudine suppresses murine accessory cell-dependent immune responses.

Trimethoprim-sulfamethoxazole (TMP-SMX), commonly used for prophylaxis of Pneumocystis carinii pneumonia (PCP) in AIDS patients, often produces a high incidence of treatment-limiting reactions. We investigated the effect of oral administration of TMP-SMX alone or in combination with the antiretroviral drug zidovudine (ZDV) on hematopoiesis and cellular immunity in BALB/c mice. Daily treatment for 28 days with TMP-SMX (160:800 mg/kg) had no effect on hematopoiesis or the ex vivo proliferative response of splenic T lymphocytes to allogeneic tumor cells (EL-4) or to concanavalin A (ConA), or that of splenic B cells to lipopolysaccharide (LPS). ZDV at 240 mg/kg/day was not immunosuppressive but caused a mild macrocytic anemia. Combined treatment produced severe pancytopenia, a significant drop in splenic cellularity, and a 61% decrease in the percentage of splenic macrophages. The percentage of splenic CD3+ lymphocytes increased 150% in the TMP-SMX + ZDV group, but the ratios of T-cell subsets and the frequency of B cells remained unchanged. Combined drug treatment did not impair the proliferative response of B cells to LPS or that of T cells to EL-4 cells. In concert with the reduction in the percentage of macrophages, the proliferative response of T lymphocytes to ConA decreased significantly. Optimal ConA-induced T-cell proliferation requires the participation of accessory cells (AC) (e.g., macrophages); EL-4 cells are able to function as AC. These data indicate that ZDV synergizes with TMP-SMX, causing severe hematotoxicity and suppressing AC-dependent immune function, and suggest that this therapeutic regimen may contribute to the immune deterioration in AIDS patients.

Methionine in the treatment of nitrous-oxide-induced neuropathy and myeloneuropathy.

Two cases of severe myeloneuropathy and macrocytic anemia associated with a low serum level of vitamin B12 after prolonged exposure to nitrous oxide are reported. In both cases, the neurological manifestations worsened initially despite B12 supplementation, although in one case the use of methionine seemed to arrest the progression of the disease and accelerate recovery. This offers further support for the biochemical hypothesis of methionine synthetase inhibition by nitrous oxide and reproduces in man previously reported animal studies with methionine. Methionine may be an important first-line therapy in the initial treatment of neuropathy and myeloneuropathy induced by nitrous oxide, and has a hypothetical role in the treatment of subacute combined degeneration of the cord.

Anemia of azaribine in the treatment of psoriasis.

Azaribine is an effective agent in the treatment of psoriasis. In this investigation the extent of clinical dermatologic remission appeared to correlate with the degree of metabolic block induced by 6-azauridylic acid, as quantitated by the urinary excretion of orotic acid and orotidine, and the development of anemia. Following azaribine therapy there was a coordinate rise of the specific activities of erythrocyte orotate phosphoribosyltransferase and orotidine-5'-monophosphate decarboxylase. There was no correlation between the pretreatment activity of these enzymes and the clinical response to azaribine. The anemia of azaribine therapy was mile and of a megaloblastic type. Uridine effectively corrected the azaribine-induced anemia, but led to exacerbation of the patients' psoriasis. Following uridine therapy there was a reduction in the urinary excretion of orotic acid and orotidine, presumable reflecting end-product inhibition or repression of the first steps of a repeated pyrimidine biosynthesis.