[Endoscopic removal of an intraluminal duodenal diverticulum in an adult with upper abdominal pain]. / Endoskopische Spaltung eines intraluminalen Duodenaldivertikels beim Erwachsenen wegen Oberbauchbeschwerden.

We report here on a 44-year-old previously healthy patient with a two-year history of intermittent upper abdominal pain. In the outpatient gastroduodenoscopy and X- ray examinations of the small intestine an intraluminal duodenal diverticulum was suspected. Clinical examination and laboratory tests did not show any abnormal findings. In order to exclude other causes for the patient's complaints coloscopy, ERP and MRCP were performed. The latter was done because the bile duct could not be intubated in the ERCP due to the altered anatomy. By use of endoscopic ultrasound a mucosal duplication was demonstrated and thus the diagnosis confirmed. Subsequently, the diverticulum sac was sliced by argon plasma coagulation. The postinterventional course was without complications and the patient was without symptoms afterwards. The intraluminal duodenal diverticulum is a rare differential diagnosis of pain in the upper abdomen. The diverticulum should be endoscopically removed if other causes for abdominal pain have been ruled out and possibly associated malformations have been excluded.

[Posthepatic icterus caused by a solitary granular cell tumor of the common bile duct]. / Posthepatischer Ikterus durch einen solitären Granularzelltumor des Ductus choledochus.

We report a case of a 26-year-old female patient who was referred to our hospital with a painless jaundice. By means of an ERCP we found a nearly complete occlusion of the common bile duct caused by a tumor. After stenting and normalization of the cholestasis parameters, additional investigations such as endosonography, cholangioscopy and MR angiography no longer showed any manifestation of the tumor. The histological and cytological specimens did not confirm a malignant process. As there was apparently only a solitary tumor a resection according to Whipple was performed. The histological examination showed a granular cell tumor. This case report demonstrates that a posthepatic icterus may rarely be caused by a granular cell tumor. Furthermore, it clearly shows the difficulty to correctly diagnose a tumor in the extrahepatic common bile duct as one may often have to rely solely on imaging procedures.

Carcinoma of the pancreas with neuroendocrine differentiation and nodular panniculitis.

BACKGROUND: On rare occasions tumours of the pancreas produce high amounts of pancreatic lipase. The enzyme activity in the blood and in different tissues causes a syndrome called nodular panniculitis by focal necrosis of lipids and a concomittant inflammatory reaction. CASE REPORT: A 72-year-old man was admitted to the dermatology clinic with the diagnosis of erythema nodosum. The patient had been well until 3 months earlier when painful red nodes developed on the skin of both shanks. He complained of profuse night sweating and a weight loss of 10 kg within that time but did not have fever. He also had noticed a painful swelling of his right index finger, left middle finger and the third toe on his left foot. Biopsy of the nodes revealed a focal necrosis of fatty tissue. Laboratory examinations showed a highly elevated concentration of serum pancreatic lipase. Further investigations showed a tumour in the pancreas and several osteolytic lesions. Tumour biopsy revealed a neuroendocrine carcinoma. After tumour resection serum lipase level immediately fell to almost normal values, and all skin and bone manifestations disappeared quickly. CONCLUSION: Due to its clinical appearance the panniculitis syndrome is most often mistaken for either erythema nodosum or rheumatoid arthritis. A resection of the tumour after correct diagnosis should always be considered because the widespread manifestations in the skin and bones do not represent distant metastasis and have a very good chance to dissolve completely.

Procalcitonin: a useful discriminator between febrile conditions of different origin in hemato-oncological patients?

Plasma concentrations of procalcitonin (PCT) have been shown to be elevated in bacterial and fungal infections. In contrast to C-reactive protein (CRP), PCT is not elevated in inflammations of noninfectious origin. Febrile inflammatory conditions are frequent in patients with hemato-oncological diseases. A reliable marker to discriminate infectious inflammations from drug-related and tumor-associated fever is still lacking. To evaluate the impact of PCT in this setting, PCT and CRP were prospectively measured in 95 febrile hemato-oncological patients. Infections could be identified in 40 of 95 patients: 38 of 95 had fever of unknown origin (FUO), 9 patients were suspected to suffer from drug-related fever, and 8 patients from tumor-associated fever. In the noninfection group (drug-related and tumor-associated fever), PCT levels were significantly lower than in patients with infections (P<0.001) or FUO (P<0.001). Differences were still highly significant comparing patients with suspected drug-related or tumor-associated fever alone with the infection or the FUO cohort. All eight patients with tumor-associated fever as well as eight of the nine patients with drug-related fever had PCT levels within the normal range (<0.5 micro g/l). CRP values only partially allowed discrimination between the various subgroups. Differences were significant between patients with drug-related fever and the infection (P=0.001) or FUO group (P=0.004). However, as CRP levels were far above the normal range also in the patients with drug-related fever, the significance of individual values was rather limited. In conclusion, PCT may provide useful additional information to assess the clinical significance of febrile conditions. PCT may facilitate the decision on when to initiate antimicrobial or cytotoxic therapy.

Effects of hexadecylphosphocholine on thrombocytopoiesis.

Hexadecylphosphocholine (HePC) is the first representative of the alkylphosphocholines, a novel group derived from the cytotoxic etherlysophospholipids. HePC shows a broad spectrum of antiproliferative effects in neoplastic cells in vitro and in vivo. HePC has been tested successfully in several clinical studies. One of the remarkable features of this compound has been the induction of a leucocytosis and a thrombocytosis in most of the patients receiving HePC systemically. In this paper, we have investigated the biological and molecular mechanisms by which HePC exerts this interesting effect. We found that HePC acts as an unspecific costimulator on human megakaryocytic proliferation in a soft agar assay system predominantly together with thrombopoietin (TPO). Furthermore, HePC leads to the synthesis and secretion of several haematopoietic growth factors in monocytes and bone marrow fibroblasts, determined by the direct measurement of growth factors in cellular supernatants and by the measurement of growth factor mRNA in cell extracts. Thus, HePC seems to produce the increase of blood platelets in tumour patients by two different mechanisms.

Cytotoxic etherphospholipid analogues.

1. Alkyllyso-derivatives of physiologic cell membrane phospholipids show remarkable cytostatic and cytotoxic activity on many malignant tumor cell lines and tumors in vitro and in vivo. Three of these etherphospholipid analogues have already been tested in clinical phase II studies and one of these compounds, hexadecylphosphocholine (HePC), is now commercially available as a drug for the treatment of mammary carcinoma in Germany. 2. Etherphospholipid analogues possess a variety of interesting biological characteristics like induction of cellular maturation, inhibition of tumor cell invasion or modulation of the immune response with high potential value for tumor therapy. 3. Though there have been extensive investigations on the biochemical mode of action of these substances, the precise mechanism responsible for the majority of biological effects has not yet been identified. 4. In recent years growing evidence has been accumulated that etherphospholipid analogues substantially interfere with intracellular signal transduction.

Synergistic cytotoxic effects of ether phospholipid analogues and ionizing radiation in human carcinoma cells.

BACKGROUND AND PURPOSE: There is growing evidence in recent years that the antiproliferative effects of ionizing radiation may not be exclusively mediated via DNA damage but also by interactions and alterations of cell membrane associated processes. Here, we tested the hypothesis that membrane active cytotoxic ether lipids and analogues may interact with ionizing radiation, enhancing its antiproliferative effects. MATERIALS AND METHODS: The two epithelial tumor cell lines HTB 43 and KB, and the ether lipid resistant subline KBr were treated by a combination of radiation and ether lipids. Cytotoxic effects were measured by colony forming assays and the effects on membrane phospholipids were determined by quantitative thin-layer chromatography of cell lipid extracts. RESULTS: We present evidence that some ether lipids show supra-additive cytotoxic effects with ionizing radiation. These effects seem to depend on the same structural properties of ether lipids that determine their intrinsic cytostatic and cytotoxic activity. Identical growth inhibitory results were achieved when cells were treated before, or 30 min after irradiation. Analysis of major membrane phospholipids revealed no statistically significant differences of phospholipid distribution pattern in cells exposed to both treatment modalities. CONCLUSION: Our data indicate that changes of overall membrane phospholipid composition do not seem to be the mechanism of synergistic antiproliferative activity of ether lipids and ionizing radiation.

Differential regulation of phospholipase A2 in human leukemia cells by the etherphospholipid analogue hexadecylphosphocholine.

Hexadecylphosphocholine (HePC) is the main representative of a new group of antineoplastic agents, the alkylphosphocholines, which were originally derived from cytotoxic etherlysophospholipids. HePC shows antiproliferative action against a whole variety of tumor cells and tumors in vitro and in vivo. Furthermore, it also induces differentiation in some hematologic cell lines and prevents invasive growth of neoplastic cells in vitro. To date, the precise molecular mechanisms mediating the biological effects of HePC have not been identified yet. As etherlysophospholipids seem to inhibit some pathways of lipid-dependent intracellular signalling, similar effects may be relevant for HePC. We therefore investigated the influence of HePC on phospholipase A2 (PLA2-EC 3.1.1) in the human leukemia cell line U 937. HePC seems to inhibit enzyme activity independently of protein kinase C (PKC) in differentiated U 937 cells stimulated by tumor necrosis factor alpha (TNFalpha). Inhibition of purified secretory PLA2 from snake venom (EC 3.1.1.4) in vitro shows characteristics of a non-competitive mode. In contrast, HePC leads to an enhancement of PLA2 activity in immature cells which cannot be explained by changes in membrane composition. Our data suggest that PLA, inhibition is most probably not the mechanism by which HePC mediates its antiproliferative effects.

Expression of two 50 kDa proteins is associated with sensitivity towards etherlipid analogues in the human leukaemia cell line HL 60.

Hexadecylphosphocholine (HePC) is a new etherphospholipid derived substance with pronounced antineoplastic activity. So far the mode of action of this compound has not been resolved. Therefore, we decided to approach this problem by generating HePC resistant sublines of susceptible cells. The human leukaemia cell line HL 60 was successfully adapted to high concentrations of HePC over a period of 14 months. The resistant cell line HL 60 R shows similar functional characteristics as the original HL 60. Both lines can be induced to terminal differentiation into a granulocytic phenotype by DMSO. In this process, normal HL 60 cells also become resistant towards HePC. Determinations of cellular membrane lipid composition did not show significant changes, which would explain the resistance mechanism. Analysis of cellular proteins by 2D-gelelectrophoresis revealed two 50 kDa proteins expressed in HL 60 and differentiated HL 60 cells, which were not expressed in HL 60 R. Reversion of resistance of HL 60 R after prolonged cultivation without HePC led to re-expression of the two proteins, indicating at a possible involvement of these proteins in HePC sensitivity.

The influence of 1-beta-D-arabinofuranosylcytosine on the metabolism of phosphatidylcholine in human leukemic HL 60 and Raji cells.

We report that high-dose 1-beta-D-arabinofuranosylcytosine (Ara-C) treatment leads to substantial changes of membrane lipid composition in human leukemic cell lines. HL 60 cells are at least 10- to 20-fold more sensitive to Ara-C than Raji cells. After 4 h incubation with 50 microM Ara-C, both cells show deviations in their phosphatidylcholine (PC) and triglyceride (TG) contents, starting as early as 8 h after treatment. After 24 h, the Ara-C-induced changes in lipid metabolism are accompanied by a severe loss of viability in HL 60 cells but not in Raji cells. At this time point the HL 60 cells show a 20% depletion of PC with a concomitant increase in TG of 25%, whereas in Raji cells both PC and TG are increased 20 and 22%, respectively. The addition of lysophosphatidylcholine (lysoPC) antagonizes Ara-C-induced cell death in various leukemic cell lines and primary AML blasts from patients. Since lysoPC is a direct precursor for PC and increases the PC content of the membrane, we assume that the loss of PC in the sensitive cell line HL 60 and in other cells plays a role in Ara-C-induced toxicity. Further evidence for this mechanism is presented by the observation that hexadecylphosphocholine, an inhibitor of PC synthesis shows synergistic antiproliferative effects with Ara-C. We conclude that the rapid cell lysis described during high-dose Ara-C treatment seems to be mediated by reduction of cell membrane PC content.

Induction of resistance in the human leukemia cell line HL60 towards hexadecylphosphocholine and other ether phospholipid analogues.

Hexadecylphosphocholine (HePC) is a new ether lipid analogue with remarkable antineoplastic activity in vitro and in vivo. As the precise molecular mechanism by which this substance and probably other ether lipids exert their biological effects is still not defined, we tried to approach this problem by generating a cell line resistant to the antiproliferative properties of HePC. This was successfully accomplished by slow adaptation over a period of 14 months, of the very sensitive human leukemia cell line HL60. HePC resistant HL60 cells (HL60R) tolerate 8- to 10-fold higher doses of HePC and are continuously cultured in medium containing 10 micrograms/ml of HePC. An immunophenotypic and karyotypic characterization of HL60 and HL60R cells showed only marginal differences between the two cell lines. Total phospholipids, total cholesterol, protein and vinyl ether lipid content were equal in both cells. A down-regulation of the ether lipid mass in HL60R of about 40% could reflect one mechanism of tolerance induction. Though HePC uptake in HL60R cells was significantly lower than in the parental line, steady state measurements of cellular HePC content revealed similar HePC content in the membranes at HePC concentrations that were cytotoxic for HL60 but did not affect HL60R. This observation indicates that uptake and cellular accumulation of HePC do not determine HePC resistance. The resistant HL60R cells also showed a considerable degree of cross-resistance to ether phospholipids ET-18-OCH3 and BM 41.440, suggesting a common mode of action for HePC and other ether lipid analogues.

Hexadecylphosphocholine inhibits phosphatidylinositol and phosphatidylcholine phospholipase C in human leukemia cells.

Hexadecylphosphocholine (HePC) is the main representative of a new group of antineoplastic agents, the alkylphosphocholines. Besides remarkable antiproliferative properties on tumor cells in vitro and in vivo, HePC also induces differentiation and inhibits invasive growth of neoplastic cells. Knowledge of the molecular mechanisms by which HePC mediates its biological effects is poor. The observation that analogous substances, the alkyllysophospholipids, may interfere with lipid dependent intracellular signaling suggested similar mechanisms for HePC. We therefore investigated the effects of HePC on phospholipase C (PLC) activation in intact human leukemia cell lines. HePC inhibited fMLP induced phosphatidylinositol-specific PLC activation in HL60 cells and TNF-alpha induced activation of phosphatidylcholine-specific PLC in U937 cells. HePC reduced the number of TNF-alpha receptors on the surface of U937 cells by about 60%. Receptors for fMLP were not affected. Inhibition of TNF-alpha induced PC-PLC activation, however, seemed to be regulated at a post-receptor level as PLC inhibition and receptor occupancy did not correlate.

Induction of resistance to hexadecylphosphocholine in the highly sensitive human epidermoid tumour cell line KB.

Hexadecylphosphocholine (HePC, Miltefosine) is a representative of the group of alkyl-lysophosphocholines showing remarkable antitumoral activity in in vitro experiments and in experimental animal tumour models. The epidermoid tumour cell line KB, which is highly sensitive to HePC (half-maximal growth inhibiting concentration, IC50: 1.2 microM; half lethal concentration, LC50: 2.8 microM), was slowly adapted to increasing concentrations of HePC. After 14 months, the adaptation process was stopped at a concentration of 10 micrograms/ml (23.5 microM). At this point, the KB cells tolerated high doses of HePC (IC50: 41.2 microM; LC50: 87.1 microM). The resistant cells (KBr) also showed crossresistance to the other well studied ether-lysophospholipids, Edelfosine (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine, OMG-3PC; ET18OCH3) and Ilmofosine (1 S-hexadecyl-2-methoxymethyl-rac-(1-thio-3-hydroxy)propyl-3-phosphocho lin e, BM 41.440). Comparison of the KB and KBr cells showed that total lipid phosphate, ether-lipid content, vinyl-ether-lipid content, protein content as well as cholesterol content were unchanged. Furthermore, no changes were observed in the lipid composition between KB and KBr cells. Uptake of choline was also unchanged in both cells, but the uptake of D-myo-inositol was lower by a factor of two in the KBr cells. However, in KB cells, the addition of HePC induced a 50% reduction of D-myo-inositol-uptake, whereas in KBr cells inositol uptake was unchanged. Differences in HePC uptake and HePC metabolism were apparent between the KB and KBr cell lines. KBr cells showed a 3-fold lower uptake for HePC and a 3- to 4-fold faster metabolism of HePC than KB cells. However, the amount of non-metabolised HePC after 2 days of incubation with 1 microgram/ml HePC (LC50: 1.2 microgram/ml) in KB cells was 3- to 4-fold lower than the amount of HePC in KBR cells at 10 micrograms/ml (LC50: 37 micrograms/ml), indicating that KBr cells can incorporate higher amounts of HePC than KB cells without adverse effects for cell growth and viability. This seems to indicate that mechanisms other than slower uptake and faster metabolism are involved in the induction of resistance to HePC in KBr cells.

Effects of hexadecylphosphocholine on membrane phospholipid metabolism in human tumour cells.

Hexadecylphosphocholine (HePC) is an analogue of the antiproliferative alkyllysophospholipids (ALP). As these lipid-like compounds interfere with membrane lipid metabolism at several sites, we studied the effects of HePC on uptake and metabolism of inositol and choline, two important phospholipid precursor molecules in two sensitive cell lines, Raji and KB, and in a resistant variant of KB cells, KBr. HePC substantially inhibited the membrane uptake of inositol and of choline in KB and Raji. Inositol uptake of KBr cells was constitutively low and was not further decreased by HePC. In all three cell lines, uptake inhibition of choline was less pronounced. Uptake inhibition showed characteristics of a non-specific effect, probably due to the physicochemical properties of HePC as a "lyso" structure. Decreased uptake of inositol did not affect phosphoinositide synthesis. Cellular phosphatidylcholine (PC) metabolism seemed to be affected through inhibition of choline incorporation and enhancement of PC degradation in the two sensitive cells. In KBr cells, these effects were not observed.

dic(9;20): a new recurrent chromosome abnormality in adult acute lymphoblastic leukemia.

Loss of chromosome 20 and rearrangement of the short arm of chromosome 9 were identified by banding analysis of three adult patients with acute lymphoblastic leukemia (ALL). The G-banding pattern suggested an identical deletion of 9p, but, also, an unbalanced translocation with chromosome 20 was taken into consideration. Dual-color chromosome painting with probes for chromosomes 9 and 20 revealed the presence of material from chromosome 20 at the short arm of the abnormal chromosome 9 in all three cases. Centromeric alpha-satellite DNA of both chromosome 9 and chromosome 20 was demonstrated by fluorescence in situ hybridization and indicated the presence of a dicentric chromosome. The hybridization of a YAC clone of the short arm of chromosome 20 proved that the dicentric chromosome contained the short arm of chromosome 20, which had been suspected from the G-banding pattern. Thus, the rearrangement was interpreted as dic(9;20)(p11;q11.?1). Because this was the sole chromosome abnormality in two patients, dic(9;20) may be a primary chromosome aberration in ALL. In one case, a 9q+ chromosome derived from a Philadelphia (Ph) translocation was involved in the formation of the dicentric chromosome. Immunophenotyping revealed CD10+ B-cell precursor ALL in all three cases. Whereas the two patients in whom dic(9;20) was the sole cytogenetically detectable change are in continuous complete remission for 10 and 45 months, respectively, the Ph+ patient relapsed with leukemia and died 8 months after diagnosis.

Inhibition of calcium-dependent protein kinase C by hexadecylphosphocholine and 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine do not correlate with inhibition of proliferation of HL60 and K562 cell lines.

We investigated the hypothesis that the antiproliferative effect of hexadecylphosphocholine (HePC) and 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) is mediated through the inhibition of cellular protein kinase C (PKC). In the sensitive HL60 cell line, ID50 and LD50 values of 5.6 and 5.3 microM, respectively (HePC), and of 3.8 and 4.2 microM, respectively (ET-18-OCH3) were obtained. In the more resistant K562 cell line, these values were 69.1 and > 97 microM, respectively (HePC) and 7.8 and 76.8 microM, respectively (ET-18-OCH3). Treatment of both cell lines with HePC and ET-18-OCH3 (25 microM) for 2 h did not lead to PKC translocation. However, a 30% reduction of PKC activity, mainly due to a decrease in the cytosolic compartment, was found. Half maximal stimulation of PKC translocation by phorbolester (TPA) in HL60 and K562 cells, which were pretreated for 2 h with 25 microM of the lipids, resulted in a 20-30% decrease of membrane-bound PKC, whereas the cytosolic form was found to be unchanged. In the same experimental setting, dioctanoylglycerol (DIC8)-stimulated PKC translocation was not affected by HePC or ET-18-OCH3. However, a 10-20% reduction of PKC enzyme activity in the membrane and in the cytosolic fraction was obtained. These findings indicate that HePC and ET-18-OCH3 do not interfere with PKC translocation but rather mediate a general decrease of the enzyme activity in the membrane and cytosol of the cells. Since the extent of PKC inhibition was somewhat similar in the sensitive HL60 and the resistant K562 cell line, inhibition of PKC is probably not a prerequisite for the antiproliferative action of HePC and ET-18-OCH3.

Investigations on the cellular uptake of hexadecylphosphocholine.

The uptake of [(9,10)-3H]hexadecylphosphocholine (HePC) in six tumor cell lines was studied. All cell lines incorporated HePC in similar amounts, with the exception of the epidermoid cancer cell line KB, which took up higher amounts of HePC. The uptake of HePC at 37 degrees C was shown to be time and concentration dependent. At 20 degrees C, uptake was drastically reduced and at 4 degrees C it was blocked completely. Binding of HePC, at 4 degrees C, was not saturable at concentrations between 5 micrograms/mL (11.8 microM) and 100 micrograms/mL (235.3 microM), indicating that cell surface binding is not receptor-mediated. Furthermore, the effects of inhibitors of endocytosis were investigated. We observed a pronounced inhibitory effect by monensin and cytochalasin B. Colchicine was somewhat less effective whereas chloroquine was almost without effect. From these data we conclude that uptake of HePC is most probably mediated via a receptor-independent endocytotic mechanism.

TNF activates NF-kappa B by phosphatidylcholine-specific phospholipase C-induced "acidic" sphingomyelin breakdown.

In this paper, we describe a phospholipid transmission pathway mediating tumor necrosis factor (TNF) activation of the nuclear transcription factor kappa B (NF-kappa B). Central to this TNF signaling route is the second messenger-like molecule ceramide, which is generated by sphingomyelin (SM) breakdown catalyzed by a sphingomyelinase (SMase). SMase activation is secondary to the generation of 1,2-diacylglycerol (DAG) produced by a TNF-responsive PC-specific phospholipase C (PC-PLC). The functional coupling of these two C type phospholipases is revealed by D609, a selective inhibitor of PC-PLC. SMase itself, or SMase-inducing regimens such as exogenous PLC or synthetic DAGs, induces NF-kappa B activation at pH 5.0, suggesting the operation of an acidic SMase. A model is proposed in which a TNF-responsive PC-PLC via DAG couples to an acidic SMase, resulting in the generation of ceramide, which eventually triggers rapid induction of nuclear NF-kappa B activity.