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To study the precise mechanism of cytotoxic activity of PGD 2 or Δ 12 -PGJ 2 (a biological active metabolite of PGD 2 ), we examined the effect of various compounds on PGD 2 or Δ 12 -PGJ 2 cytottoxic, using a human neuroblastoma cell line (NCG). Cycloheximide (CHM) specifically protected PGD 2 cytotoxicity on NCG cells. When Δ 12 -PGJ 2 was tested, CHM exhibited a similar rescue effect. Puromycin, mitomycin C, and α-amanitin did not affect PGD 2 or Δ 12 -PGJ 2 cytotoxicity. Emetine showed a variable and no consistent rescue effect CHM may have been active at the primary site where PGD 2 or Δ 12 -PGJ 2 exerts its cytotoxicity. This is the first report indicating that CHM reduces the cytotoxicity induced by PGD 2 or Δ 12 -PGJ 2 .

Serum ferritin level was measured serially in three malignant histiocytosis (MH) and five virus-associated hemophagocytic syndrome (VAHS) patients. Serum ferritin levels were greater than 1000 ng/ml at the early phase of disease. When disseminated intravascular coagulation (DIC) developed in patients, serum ferritin levels elevated further to greater than 12,000 ng/ml. MH patients were treated by combination chemotherapy, and VAHS patients were given prednisone. Each MH and VAHS patient died within 1 week from the onset of DIC, without decrease of serum ferritin level. One MH patient with continued high serum ferritin levels for 3 months after DIC also died. The remaining patients with decreased serum ferritin values less than 1000 ng/ml at 3 months after DIC are now alive with normal serum ferritin levels. These results suggest that serum ferritin level in histiocytic proliferative disorders is a useful indicator of disease activity in both neoplastic and reactive conditions rather than only a marker of malignant process.

Eighteen children with refractory acute lymphocytic leukemia (ALL) who had been heavily pretreated, were treated with combination etoposide and cytosine arabinoside (ara-C) chemotherapy. Seventeen of these 18 patients were in their first to third relapses; the remaining patient had never responded to induction therapy. The drug combination of etoposide followed by ara-C was administered as an intravenous (IV) infusion twice a week for two consecutive weeks, a total of four doses. The dosage was 150 mg/m2/dose for each drug. Seven (39%) of the 18 patients attained a complete remission (CR) and three (17%) attained a partial remission (PR). Complete response was obtained in two of eight patients in first marrow relapse, and in five of nine patients in second and third relapse. Five patients achieved a CR after one course of therapy and two achieved a CR after two courses of therapy. Of three patients who had previously received teniposide, two attained a CR with this combination. The duration of these responses was brief with a median of 1 month, ranging from 0.5 to 3 months, with the exception of one case, which has been in remission for 2.5 + months. Although myelosuppression was observed, none of the patients died from infection or bleeding. Allergic reaction with fever and rash was observed in two patients. The efficacy of the etoposide and ara-C combination for refractory childhood ALL is encouraging in several current reinduction regimens.

Effects of 5-bromo-2'-deoxyuridine (BrdU) were studied on two neuroblastoma and two leukemia cell lines, in terms of the relationship between prostaglandin (PG) synthesis and cell growth/differentiation. After treatment with BrdU (5 micrograms/ml), cell growth of the 4 cell lines was inhibited and one neuroblastoma cell line (GOTO) showed flattened morphology with positive S-100 protein, one of the differentiation markers for Schwann or glial cells. In the 4 cell lines, BrdU treatment reduced [1-(14)C]-arachidonic acid incorporation into phosphatidylinositol and phosphatidylethanolamine and was associated with an increase into phosphatidylcholine and triglyceride. BrdU treatment also increased fractions of 6-keto-PGF1 alpha and PGF2 alpha, with a decreased TXB2 fraction. The decreased ratio of TXB2/6-keto-PGF1 alpha or increased 6-keto-PGF1 alpha fraction correlated significantly with cell growth inhibition, suggesting that the changes in the balance of endogenous PGs might be associated with BrdU-induced cell growth inhibition with or without differentiation of neuro-blastoma and leukemia cells in culture.

Tissue specimens obtained at autopsy from seven childhood cases of malignant histiocytosis were studied by immunohistochemistry. Clinically, the majority of the cases showed sustained fever, hepatosplenomegaly, pancytopenia, and DIC. The pretreatment diagnosis was based on their typical clinical manifestations and bone marrow smear findings. Although three patients temporarily responded to exchange transfusion and chemotherapy, all seven patients eventually died of active disease. Postmortem examination revealed the proliferation of atypical histiocytes appearing in variable degrees of maturation in the lymph nodes, liver, spleen, bone marrow, lungs, and central nervous system. Immunohistochemical staining for lysozyme, nonspecific cross-reacting antigen (NCA), alpha 1-antitrypsin (alpha 1 AT), alpha and beta subunits of S100 protein (S100 alpha, beta), and concanavalin A receptors (ConAR) in cytoplasm demonstrated the presence of two subtypes of malignant histiocytes, ie, S100 beta+/NCA-/ConAR+ (4 cases) and S100 beta-/NCA+/ConA R+ (three cases). The results of lysozyme, alpha 1 AT, and S100 alpha staining were inconsistent. A survey of the literature disclosed that the incidence of S100 protein-positive cases in children was higher than in adults (12/21 v 5/19; chi 2, P less than .05). Further large scale investigation is necessary to confirm the independence and significance of these two subtypes of histiocytes in malignant histiocytosis.

Two human neuroblastoma cell lines, NCG and GOTO, were used to study the cytotoxic effect of gamma linolenic acid (GLA). The cell growth inhibition of these culture cells by GLA was found to be associated with striking membrane fatty acid modification. When culture cells were exposed to 20 micrograms/ml and 60 micrograms/ml GLA for 48 hr, polyenoic acids in cell membrane phospholipids (PC, PE, PI, PS) and triglyceride significantly increased; 1.8-21.0 fold for NCG and 1.04-11.5 fold for GOTO, in association with decreased monoenoic acids. The most remarkable changes were; increase of C18:3, C20:3, C20:4 and decrease of C18:1. CoQ10 (50 micrograms/ml) and vitamin E (10 microM) shown to protect against the cytotoxic effect of GLA did not modify the incorporation of GLA into tumor cells. These results indicate that the antitumor effect of GLA is probably due to cellular dysfunction caused by fatty acid modification after GLA incorporation.

Data on 28 patients with malignant histiocytosis (MH), fourteen patients with virus-associated hemophagocytic syndrome (VAHS) and two patients with familial erythrophagocytic lymphohistiocytosis (FEL) were collected from 21 hospitals in Japan to study the serum ferritin levels and clinical features. At diagnosis, the serum ferritin values were a median of 3,000 ng/ml (range, 59-270,000 ng/ml) in MH and 10,500 ng/ml (range, 44-68,600 ng/ml) in VAHS/FEL. Clinical signs and symptoms were not substantially different between MH and VAHS/FEL. Thus, serum ferritin markedly increased in the majority of MH/VAHS/FEL patients and should be a useful marker of disease activity in either neoplastic or reactive histiocytic proliferative disorders.

Serum ferritin is known to be one of the tumor markers for neuroblastoma. Serum ferritin is elevated in most children with neuroblastoma who are in stages III or IV, but it is not elevated in those in stage I or II. It has also been observed that iron load caused by blood transfusion shows a greater effect on serum ferritin levels than tumor activity due to neuroblastoma. Thus, serum ferritin increases in a linear fashion in children given repeated blood transfusions but the levels increase exponentially in children who have bulky neuroblastomas. Thus, serum ferritin values must be interpreted with caution in patients with advanced stages of neuroblastoma who inevitably require blood transfusions. The authors propose that the pretransfusion and posttransfusion serum ferritin values be in comparison with other tumor markers such as urinary VMA, urinary HVA, and neuron specific enolase. The serum ferritin level should not be used as a sole indicator of tumor activity in neuroblastoma.

gamma-linolenic acid (GLA) was found to suppress the cell growth of 4 human neuroblastoma cell lines. GOTO was the most sensitive, followed by SK-N-DZ and NKP, while NCG was much less sensitive to GLA. In terms of GLA cytotoxicity, neither cyclo-oxygenase inhibitors nor a lipoxygenase inhibitor showed any effect. On the other hand, 4 antioxidants (Coenzyme Q, (D) alpha-tocopherol, (DL) alpha-tocopherol, butylated hydroxytoluene) reduced the growth inhibitory effect of GLA, but not in proportion to the decrease of GLA-stimulated lipid peroxidation. Accordingly, prostaglandins and leukotrienes probably do not play a role, and lipid peroxide may only be partly involved in the GLA effect.

Involvement of platelets and coagulation systems in the hematogenous metastasis of tumor cells has been suggested from in vivo and in vitro studies, however, there is still controversy about the exact role of hemostasis in metastasis. To date, at least three types of platelet aggregating mechanisms and three types of tumor cell procoagulants have been reported in different tumor cells.We investigated platelet aggregating activity (PAA), procoagulant activity (PCA) and the relationship between these two activities, using eight human neuroblastoma cell lines, three human leukemia cell lines and human mature lymphocytes. PCA in tumor cells was measured by the single stage recalcification time and the assay with chromogenic substrate S2222. PAA was determined turbidometrically with an aggregometer by adding cell suspensions of tumor cells to platelet rich plasma (PRP). The effects of protease inhibitors, enzymes and thrombin inhibitors on PAA and PCA were also studied.Neuroblastoma cell suspensions showed high PCAs which were reduced in Factor VII deficient human plasma, indicating a tissue factor-like activity. NCG line possessing the highest PCA also showed a high PAA, which was inhibited by pretreatment of cell suspensions with phospholipase A2 and abolished in the presence of heparin, hirudin or MD805 in the assay system. Human leukemia cell lines and mature lymphocytes had weak to moderate PCAs without showing PAA, but became active to express PAA after being removed of cell surface sialic acid by neuraminidase. These results suggest that in neuroblastoma, PCA closely linked with PAA may play a role in the hematogenous metastasis. In hemopoietic cells, PAA expressed when cell surface sialic acid is removed does not correlate with PCA, and sialic acid in these cells possibly prevents direct interaction with platelets in the hemostatic homeostasis.

delta 12-prostaglandin(PG)J2 (7.5 micrograms/ml) significantly inhibited protein synthesis and cell growth in a human neuroblastoma cell line (NCG), decreasing these factors by 31.5% and 78.2% of the control values, respectively. Two protein synthesis inhibitors, cycloheximide (CHM) and emetine, exhibited a dose-dependent protective effect for neuroblastoma cells against delta 12-PGJ2 cytotoxicity. At a concentration of 15 micrograms/ml CHM, the number of viable cells increased from 21.8% to 36.7% of the control value (p less than 0.01). The sodium dodecyl sulfate-polyacrylamide gel analysis of [35S]methionine-incorporated proteins revealed an increased synthesis of 86k, 70k and 66k proteins in the delta 12-PGJ2-treated NCG cells under the condition that delta 12-PGJ2 exerts cytotoxicity. Of these proteins, the amount of 66k protein was particularly increased in cell cytosol; however, its synthesis did not occur when CHM prohibited the delta 12-PGJ2 cytotoxic effect. When emetine was used instead of CHM, similar results were obtained. These results strongly suggest that the 66k protein plays a critical role in the delta 12-PGJ2 cytotoxicity.

To determine the role of adenosine 3′5′-cyclic monophosphate (cAMP) in the antineoplastic effect of prostaglandins (PGE 1 , PGE 2 , PGD 2 and PGA 1 ), we studied 2 cell lines of human neuroblastoma; i.e. GOTO and SK-N-DZ. PGE 1 or E 2 at 30 μg/ml and PGD 2 or PGA 1 at 5 μg/ml were cytotoxic to these neuroblastoma cells. In both cell lines, increase of intracellular cAMP was closely associated with E-type PGs cytotoxicity, however, in PGD 2 , or PGA 1 cytotoxicity, cAMP increase was observed only in GOTO cells. Pretreatment of GOTO cells with 5 μg/ml PGE 2 for 24 hr caused a desensitization of cAMP responses to PGE 1 , PGD 2 or PGA 1 only in association with a reduced cytotoxicity of PGE 1 . On the other hand, PGE 2 -pretreated SK-N-DZ cells resulted in a desensitization in response to PGE 1 , but not to other PGs, without affecting the cytotoxicity of these PGs. A decreased [ 3 H]PGE 1 binding similarly occurred in either the PGE 2 -pretreated GOTO or SK-N-DZ cells. However, cholera toxin- or forskolin-induced cAMP production was suppressed only in the pretreated GOTO cells. cAMP response by forskolin rather increased in the pretreated SK-N-DZ cells. These results indicate that antineoplastic effect of E type PGs mediates through cAMP, but not that of PGD 2 and PGA 1 and that PGE 2 pretreatment may cause a down regulation of PGE 1 receptor site in both cell lines. It is also suggested that PGE 2 pretreatment results in a heterologous desensitization in GOTO and a homologous desensitization in SK-N-DZ cells.

Twenty-six studies by meta-(131I)-iodobenzylguanidine scintigraphy (131I-MIBG), 26 studies by 67Ga-citrate and 33 99mTc-hydorxymethylene diphosphate (99mTc-HMDP) scintigraphic studies were performed for 10 patients with abdominal neuroblastoma. Comparing the 131I-MIBG images obtained at 24, 48 and 72 h, the 48-h image was the most distinctive for the tumor. Intrabdominal primary lesions, which ranged from bean to fist-size, were visualized in 7/7 cases (100%) by 131I-MIBG, 4/7 cases (57%) by 67Ga-citrate and 4/8 cases (50%) by 99mTc-HMDP before surgery and at diagnosis. In serial follow-up of these patients after starting chemotherapy, 131I-MIGB detected 100% of regressing primary tumors. Studies of 5 postoperative patients showed negative images for the primary tumor in all 3 scintigraphies except one in whom 131I-MIBG was positive, but not 67Ga-citrate or 99mTc-HMDP, for an unresectable residual tumor. 131I-MIBG also detected metastatic lesions not predicted by 67Ga-citrate or 99mTc-HMDP and reflected tumor progression more sensitively than known tumor markers such as urinary vanillylmandelic acid (VMA), homovanillic acid (HVA), serum neuron-specific enolase (NSE) and ferritin. These findings indicate that the 48 hr 131I-MIBG scintigraphy is superior to 67Ga-citrate or 99mTc-HMDP images and to other biochemical markers in monitoring the effect of treatment on neuroblastoma.

Severe hypercalcemia (serum calcium, 4.25-5.25 mmol/l), in association with osteolytic bone lesions, was found in a girl aged 2 yr 7 mo with common acute lymphoblastic leukemia (ALL). Hormonal studies excluded the possibility of the hypercalcemia being caused by primary hyperparathyroidism or ectopic parathyroid hormone secretion. Increased plasma prostaglandin E2 (PGE2) levels (130 ng/l), probably produced by leukemic cells, were considered to be one of the pathogenic mechanisms responsible for the occurrence of hypercalcemia in this patient. Both the hypercalcemia and the abnormal plasma PGE2 level returned to normal after chemotherapy.

It has been known for more than 2 decades that amino acid changes accompany various forms of infectious diseases. In some of these infections, an elevation in the phenylalanine/tyrosine ratio has been reported (1–4). Similar amino acid changes have also been described in protein calorie malnutrition (5) and chronic renal insufficiency (6). So far, however, such changes have not been clearly described in patients with hematologic neoplasias, especially for proliferative disorders of mononuclear macrophages. Nor has the actual occurrence of hyperphenylalaninemia in association with infectious or neoplastic disorders been reported. Recently, we encountered 2 patients with malignant histiocytosis (MH) (7) and a case of virus-associated hemophagocytic syndrome (VAHS) (8). In all three patients the disease took a very rapid and fatal course with multiple organ dysfunction associated with hyperphenylalaninemia (>400 μmol/l) and an increased serum phenylalanine/tyrosine (Phe/Tyr) ratio (>4.00) (Table 1).

Tissue content of cyclic AMP was as much as 10 times greater in ganglioneuroma than in neuroblastoma. This high cyclic AMP in ganglioneuroma was not significantly different from that of sympathetic ganglia.

A vanillylmandelic acid spot test for screening infants for neuroblastoma was developed in cooperation with 11 health centers in Kyoto, Japan. In this system, the parent sends a filter paper spotted with the infant's urine at 6 or 7 months of age. From 78,331 infants screened during the six years from July 1973 to September 1979, four asymptomatic infants with neuroblastoma were discovered. Three of them have already completed their treatments and have recovered. The cost of screening 13,000 infants per year was $7,500. One false-negative test was encountered, as were a number of false-positive tests.