Anemia in children with cancer is associated with decreased erythropoietic activity and not with inadequate erythropoietin production

A defect in erythropoietin (EPO) production has been advocated as being the main cause of anemia presented at time of diagnosis or during treatment by adults with solid tumors. On the basis of this defect, anemic cancer patients, both adults and children, have been treated with recombinant human EPO (rHuEPO). To further elucidate the pathophysiology of anemia in children with cancer, we measured serum soluble transferrin receptor (sTfR), a quantitative marker of erythropoiesis, and serum EPO at time of diagnosis and during chemotherapy in children suffering from solid tumor or leukemia. We determined serum EPO in 111 children (55 leukemia, 56 solid tumors) at time of diagnosis. In the last 44 patients (23 leukemia and 21 solid tumors), sTfR levels were also measured. Serum EPO together with sTfR levels were also determined in 60 children receiving chemotherapy (29 leukemia, 31 solid tumors). These results were compared with those obtained from appropriate control groups. In all patients, we found a highly significant correlation between the logarithm of EPO (log[EPO]) and the hemoglobin (Hb) level. In all subsets of patients, sTfR levels were inappropriately low for the degree of anemia. Neither leukemic nor solid tumor groups showed a significant inverse relationship between log(sTfR) and the Hb level as would be expected in anemic patients with appropriate marrow response. Thus, in children with cancer, anemia is associated with a decreased total bone marrow erythropoietic activity which, in contrast to what has been reported in anemic cancer adults, is not related to defective EPO production.     

Erythropoiesis in chronic renal disease

The diminished erythropoiesis in the anemia of chronic renal disease has been attributed to three possible factors: (1) decreased erythropoietin production, (2) inhibition of erythropoietin activity, and (3) decreased bone marrow response to erythropoietin. In this report we isolated and evaluated these parameters in 19 patients with chronic renal disease, nine patients with iron-deficiency anemia, and seven control subjects. The results in patients with chronic renal failure were as follows: (1) erythropoietin enhanced heme synthesis in bone marrow cell cultures by 88 +/- 12 per cent in renal failure, as compared to 65 +/- 7 per cent in the control group; (2) plasma erythropoietin activity did not increase appropriately for the degree of anemia; and (3) erythropoietin inhibitor activity in renal failure was not greater than in a control group. In conclusion, the relative failure of erythropoiesis in chronic renal disease appears to be due primarily to decreased production of erythropoietin and not to diminished marrow response to erythropoietin.     

In vivo effects of recombinant human erythropoietin on bone marrow hematopoiesis in patients with chronic renal failure

Studies of hemopoietic progenitors and precursors in bone marrow before and after two months of recombinant human erythropoietin (rhEpo) therapy in 12 patients with uremic anemia are the subject of this investigation caried out in order to have a better insight into the effect of Epo in vivo. Eight patients were on hemodialysis and four others were predialysis patients with chronic renal failure. The starting dose of rhEpo was 30-50 U/kg bw and was increased by 50 percent every four weeks. The mean hemoglobin values rose from 6.08 +/- 1.03 to 9.8 +/- 1.98 g/dl at the time of study. The number of bone marrow derived erythroid colonies, both early (BFU-E) and late (CFU-E) were found to be higher than subnormal values, found before the therapy. The percentage of erythroid progenitors in cell cycle increased to higher than normal values for BFU-E and to normal values for CFU-E. At the same time granulocytic progenitors (CFU-GM) decreased to the range of normal values (67.3 per 10 superset5 cells). Slightly increased Epo levels (approx. 30mU/ml) during the replacement therapy were optimal for correction of anemia. The rhEpo therapy induced an increase of percentage of erythroblasts and the decrease of myeloid to erythroid ratio (M/E) in the bone marrow. Only in predialysis patients in whom the target hemoglobin values were achieved by rhEpo therapy at the time of the study the percentage of erythroblasts in the bone marrow increased to normal values. Increase of erythroblasts in bone marrow in patients under two months of substitutive therapy with rhEpo with the increase of both, early and late erythroid progenitors we have observed, is significant indicating the stimulative effect of rhEpo on all subsets of erythropoiesis leading to normalization of erythropoiesis at all levels. No stimulative effect of rhEpo replacement therapy on granulopoiesis was observed.     

Indium bone marrow scintigraphy as an aid in selecting marrow biopsy sites for the evaluation of marrow elements in patients with lymphoma

One hundred and two previously treated lymphoma patients were studied with 111Indium bone marrow scans and bone marrow biopsies. The biopsies were considered to represent sampling errors when the cellularity of the biopsy did not reflect the general state of the marrow organ cellularity as demonstrated by the scan. In each instance the accuracy of the scan was confirmed by either another biopsy or the subsequent clinical course of the patient. Sampling errors were infrequent (1/51) in patients with normal peripheral blood counts and whose marrow had never been involved with tumor. Errors were especially likely (17/51) in patients who had had marrow involvement or those who had anemia, leukopenia, or thrombocytopenia. The 111Indium bone marrow scan allows the clinician to avoid selecting a biopsy site with a high risk for sampling error.     

The hypereosinophilic variant of Ph''-positive chronic myeloleukemia

AIM: To confirm clonal nature of idiopathic hypereosinophilic syndrome (IHES), its relevance to Ph'-positive chronic myeloid leukemia. MATERIALS AND METHODS: 3 cases of idiopathic hypereosinophilic syndrome are reported with morphologic analysis of bone marrow cells and cytogenetic examinations. In one patient the presence of Ph'-chromosome was confirmed at fluorescent in situ hybridization (FISH) and molecular-genetic analysis (bcr/abl). Samples of bone marrow, spleen and liver were examined pathohistologically. RESULTS: The presence of chromosome anomaly t(9;22), i.e. Ph'-chromosome, associated with chronic myeloid leukemia (CML) was identified in all the 3 cases. There was also myeloid hyperplasia in the bone marrow (with primarily mature, eosinophilic granulocytes), spleen and liver, depression of megakaryocyto- and erythropoiesis. 2 patients had similar clinical symptoms which was not typical for CML in chronic phase: fever, elevated ESR, clear-cut anemia and thrombocytopenia. In the absence of hyperleukocytosis, blood and bone marrow eosinophils remained high (42.5, 21.5, 42.5% and 21.4, 7.1, 6.5%, respectively) due to "mature" forms. The number of blasts in the bone marrow was maximum 2.4%. CONCLUSION: The literature and the obtained data suggest closeness of idiopathic hypereosinophilic syndrome and Ph'-positive CML within myeloproliferative diseases.     

The effect of subcutaneous recombinant human erythropoietin (r-HuEPO) on anemia in cancer patients receiving platinum-based chemotherapy

Advanced cancer is commonly associated with significant anemia which worsens with the administration of cytotoxic drugs. Erythropoietin (EPO) levels in these patients are usually inappropriately low for the degree of anemia. We evaluated the effect of subcutaneous administration of recombinant human erythropoietin (r-HuEPO) on hematologic parameters and transfusion requirements in anemic cancer patients who were receiving platinum-based chemotherapy. Baseline studies included complete hemogram, reticulocyte count, serum iron, TIBC, ferritin and determination of performance status and quality of life (QOL). Twenty-three patients, 13 females, 10 males with mean age 52 years received 150 units/kg of r-HuEPO three times weekly for a minimum of 10 weeks. They also received supplemental iron. Ovarian cancer was the commonest underlying malignancy. Most of the patients received platinum-based combination chemotherapy. Mean duration of r-HuEPO therapy was 12.6 weeks. Average baseline reticulocyte count was 1.8% which increased to 7.0% after one week therapy. Eight patients had normalization of hemoglobin values. Another eight patients improved their hemoglobin by at least 2 g/dl, however, hemoglobin values remained below the normal range. Two patients had only slight increase in hemoglobin but never required blood transfusion. Three patients who were transfusion dependent had decrease in the transfusion requirements. Two patients had no significant benefit. In most patients response was evident within 2 weeks. All responders had improvement in QOL. No significant toxicity was observed. We conclude that r-HuEPO, given subcutaneously, is highly effective in amelioration of anemia and prevention of or reduction in transfusion requirements in cancer patients receiving platinum-based chemotherapy.     

Novel guide catheter for left coronary intervention via a right upper limb approach

A Standardbred gelding and a colt were examined because of poor performance and anemia. Each horse had been given recombinant human erythropoietin (rhEPO; 4,000 IU) at least twice within the preceding 2 to 4 months. The horses had an Het of 16 and 24%, serum iron concentrations of 210 and 304 micrograms/dl (reference range, 73 to 140 micrograms/dl), total iron binding capacities of 239 and 321 micrograms/dl (reference range, 266 to 364 micrograms/dl), values for the percentage saturation of transferrin by iron of 87.9 and 94% (reference range, 20 to 52%), and serum ferritin concentrations of 255 and 355 ng/ml (reference range, 43 to 261 ng/ml), respectively. There was no clinical or laboratory evidence of immune-mediated hemolysis or an infectious or inflammatory cause of the anemia. Examination of sternebral marrow biopsy specimens revealed generalized bone marrow hypoplasia; myeloid-to-erythroid ratios were 6.7 and 3.2. Moderate-to-marked erythroid hypoplasia was diagnosed in both horses. Compared with serum from a healthy control horse, serum from the affected horses inhibited rhEPO-induced proliferation of erythroid progenitors in vitro. Results suggested that the horses had developed anti-rhEPO antibodies that cross-reacted with endogenous erythropoietin, thereby inhibiting erythropoiesis. Horses were discharged with instructions that rhEPO administration be discontinued and that dexamethasone be administered. Five months later, both horses were back in training. For 1 horse, Hct had increased to 35%, and the other horse was not available for examination.     

Myelodysplastic syndrome with bone marrow eosinophilia: clinical and cytogenetic features

We investigated the hematological and clinical status of 145 patients with de novo myelodysplastic syndrome (MDS), 14 of whom (10%) had eosinophilia in the bone marrow (MDS-Eo). Most of these 14 patients had severe anemia. Their bone marrow cells exhibited trilineage dysplasia and some morphological abnormalities in the eosinophils, including disproportion of eosinophilic granules, basophilic granules, a ring-shaped nucleus, and vacuolation in the cytoplasm. However, these abnormalities were less prominent than those of acute myelomonocytic leukemia with eosinophilia (FAB: M4Eo). Three of the 14 MDS-Eo patients had refractory anemia (RA), seven had RA with excess of blasts (RAEB), and four had RAEB in transformation. Cytogenetic analysis revealed chromosomal abnormalities in 12 of 13 MDS-Eo patients (92%), in particular, there were major karyotypic abnormalities (MAKA) in eight patients (62%). Cytotoxic agents were not effective in the treatment of four patients after leukemic transformation occurred. These four patients died of the leukemic transformation while seven died of bone marrow failure. The other three MDS-Eo patients are still alive; two of them have already transformed to a leukemic phase. The duration of survival of these patients was significantly shorter than that of the other MDS patients. These findings suggest that bone marrow eosinophilia in MDS may be a poor prognostic factor that is strongly related to the existence of MAKA.     

Micrometastatic breast cancer cells in bone marrow at primary surgery: prognostic value in comparison with nodal status

BACKGROUND: Approximately 30% of the patients with primary breast cancer who have no axillary lymph node involvement (i.e., lymph node negative) at the time of surgery will relapse within 10 years; 10%-20% of the patients with distant metastases will be lymph node negative at surgery. Axillary lymph node dissection, as a surgical procedure, is associated with frequent complications. A possible alternative to nodal dissection in terms of prognosis may be the immunocytochemical detection of tumor cells in bone marrow. PURPOSE: In a prospective study, the value of tumor cell detection (TCD) in bone marrow was compared with axillary lymph node dissection in the prognosis of primary breast cancer after surgery. METHODS: Data from 727 patients with primary, operable breast cancer were included in the analysis. All patients had surgery, including axillary lymph node dissection, from May 1985 through July 1994 at the Women's Hospital of the University of Heidelberg (Federal Republic of Germany). Bone marrow aspiration at two sites on each anterior iliac crest was performed immediately after surgery while the patients were under general anesthesia. Most patients received some type of systemic adjuvant therapy. The monoclonal antibody 2E11, directed against the polymorphic epithelial mucin TAG12, was used to detect tumor cells in bone marrow samples. The association of TCD with recognized prognostic indicators was evaluated by means of chi-squared tests. Survival without the development of distant metastases (i.e., distant disease-free survival) and overall survival were estimated by use of the Kaplan-Meier method; the logrank test was used to compare survival curves. A multivariate Cox regression analysis with stratification according to adjuvant treatment type was used to assess the independent prognostic value of TCD in bone marrow in relation to other variables. Reported P values are two-sided. RESULTS: Tumor cells were detected in the bone marrow of 203 (55%) of 367 lymph node-positive patients and in 112 (31%) of 360 lymph node-negative patients. TCD was associated with larger tumors (P < .001), lymph node involvement (P = .001), and higher tumor grade (i.e., more undifferentiated) (P = .002). After a median follow-up of 36 months, patients with tumor cells in their bone marrow experienced reduced distant disease-free survival and overall survival (both P values < .001). TCD was an independent prognostic indicator for both distant disease-free survival and overall survival that was superior to axillary lymph node status, tumor stage, and tumor grade. Among patients with tumors less than 2 cm in diameter, TCD was the most powerful predictor of outcome. CONCLUSIONS AND IMPLICATIONS: TCD in the bone marrow of patients with breast cancer is a valuable prognostic tool associated with negligible morbidity. Prospective randomized studies should be performed to determine whether TCD might replace axillary lymph node dissection in a defined subgroup of patients with small tumors.     

Training of new coworkers in the intensive care unit as a means for quality assurance

Among patients with bone marrow failure, differentiating acquired aplastic anemia (AA) from hypocellular refractory anemia (hypo RA) can be a difficult and challenging task. Morphological, cytochemical, immunocytochemical, and cytogenetic studies may provide tools for discriminating between both entities. In addition, differences in the pattern of proliferation and apoptosis of bone marrow cells in AA and in the myelodysplastic syndrome have been reported. Because of the correlation between p53 and apoptosis, we examined the overexpression of p53 on bone marrow biopsies in RA and AA. Our study included 14 patients with hypo RA, 14 patients with hypercellular (hyper) RA, ten patients with classic acquired AA, and 37 hematologically normal individuals. p53 was overexpressed in eight (57%) hypo RA patients and 11 (79%) hyper RA patients. All normal individuals and patients with AA showed no overexpression of p53 in their marrow. These results were statistically significant:p < 0.01 (AA vs hypo RA), p<0.001 (AA vs hyper RA), while the difference between hypo RA and hyper RA was not statistically significant. We conclude that p53 overexpression in bone marrow biopsies is a valuable tool for studying bone marrow failure and may provide additional information to help differentiate hypo RA from acquired AA.     

Long-term outcome after marrow transplantation for severe aplastic anemia

We reviewed the records and reevaluated 212 patients with aplastic anemia transplanted at the Fred Hutchinson Cancer Research Center (FHCRC) between 1970 and 1993 who survived >/=2 years and who have been followed for up to 26 years. Parameters analyzed included hematopoietic function, chronic graft-versus-host disease (GVHD), skin disease, cataracts, lung disease, skeletal problems, posttransplant malignancy, depression, pregnancy/fatherhood, and the return to work or school, as well as patient self-assessment of physical and psychosocial health, social interactions, memory and concentration, and overall severity of symptoms. Survival probabilities at 20 years were 89% for patients without (n = 125) and 69% for patients with chronic GVHD (n = 86) (the status was uncertain in 1 surviving patient). All patients had normal hematopoietic parameters. Skin problems occurred in 14%, cataracts in 12%, lung disease in 24%, and bone and joint problems in 18% of patients. Eleven patients (12%) developed a solid tumor malignancy and 19% of patients experienced depression. Chronic GVHD was the dominant risk factor for late complications. Seventeen patients died at 2.5 to 20.4 years posttransplant; 13 of these had chronic GVHD and related complications. At 2 years, 83% of patients had returned to school or work; the proportion increased to 90% by 20 years. At least half of the patients preserved or regained the ability to become pregnant or father children. Patients rated their quality of life as excellent and symptoms as minimal or mild. In conclusion, marrow transplantation in patients with aplastic anemia established long-term normal hematopoiesis. No new hematologic disorders occurred. The major cause of morbidity and mortality was chronic GVHD. However, the majority of patients who survived beyond 2 years returned to a fully functional life.     

Comprehensive treatment of advanced neuroblastoma involving autologous bone marrow transplant

Encouraging results are reported with high-dose chemotherapy and total body irradiation followed by autologous bone marrow transplantation in the treatment of advanced neuroblastoma. However, relapse remains a significant problem. We used high-dose chemotherapy, surgery, intraoperative radiation and an autologous bone marrow transplant treated in vitro to remove tumor cells followed by 13-cis-retinoic acid to treat 36 children with advanced neuroblastoma. This comprehensive treatment appears to improve the survival rate of patients with advanced neuroblastoma, including those with N-myc amplification and bony involvement. The disease-free survival rate was 66% (95% confidence interval, 49-84%) at 3 years. All patients who received 13-cis-retinoic acid developed cheilitis, but no bone marrow depression occurred in these patients. Five patients developed hemolytic uremic syndrome (HUS) post-transplant. This may have been related to the procedure used for total body irradiation. Patients who had their kidneys shielded during this procedure did not develop this syndrome. Patients who received local irradiation at the primary site showed no evidence of relapse in this region, indicating that such therapy may help to prevent a relapse. These data suggest a high rate of 3 year disease-free survival with this treatment strategy. The nonrandomized nature of the study and use of multiple modalities precludes analysis of the specific contribution of each.     

Phase II trial of recombinant interferon-alpha-2a and eflornithine in patients with recurrent glioma

The aim of this study was to investigate the influence of interferon-alpha (IFN-alpha) on bone marrow fibrosis associated with chronic myelogenous leukemia (CML). Eighty-two bone marrow biopsies from 41 patients in chronic phase CML were stained with Snook's reticulin stain for argyrophilic fibers. Grading of reticulin fibrosis (scale of 1 to 4) was according to the quantity and pattern of distribution of reticulin. The interval between biopsies was a median of 25 months (range 12 to 40 months). Patients had been treated with IFN-alpha for at least 12 months, were still on IFN-alpha therapy during the study, and had achieved at least a complete hematological response. Before the start of IFN-alpha therapy, reticulin fibrosis was grade 1-2 in 23 patients (56%) and grade 3-4 in 18 (44%). During IFN-alpha therapy, reticulin fibrosis in bone marrow did not worsen or was reduced in 33 patients (80%) and increased by one grade in eight patients (20%). Only five patients (12%) with limited fibrosis (grade 1-2) before start of IFN-alpha therapy showed an increase towards significant fibrosis (grade 3-4), while eight of the 18 patients (44%) with grade 3-4 fibrosis achieved a decrease of fiber content in bone marrow. In summary, IFN-alpha was not associated with an enhancing effect on myelofibrosis in patients with CML and may have prevented increasing fibrosis in patients who respond to therapy.     

Measurement of secondary colony formation after 5 weeks in long-term cultures in patients with myelodysplastic syndrome

Pancytopenia is a frequent manifestation of myelodysplastic syndromes (MDS). In the presence of an empty bone marrow, clinical distinction from aplastic anemia may be difficult. The hypoplastic marrow morphology seen in some cases of MDS raises questions about etiologic and pathophysiologic relationships between aplastic anemia and MDS. The goal of our study was to compare the degree of the hematopoietic failure in these diseases at the level of the most immature progenitor and stem cells that can be measured in vitro. In a systemic, prospective fashion, we have studied bone marrow (n = 45) and peripheral blood (n = 33) of patients with MDS for the number of long-term culture initiating cells (LTC-IC) in comparison to 17 normal controls and patients with new, untreated aplastic anemia (46 marrow; 62 blood samples). Due to the low numbers of cells available for the analysis, formal limiting dilution analysis could not be performed, instead secondary colony-forming cells (CFC) after 5 weeks of LTBMC were measured. As the number of these cells is proportional to the input number of LTC-IC, the number of secondary CFC per 10(6) mononuclear cells (MNC) initiating the LTBMC can be used as a measure of the content of immature stem cells in bone marrow and peripheral blood. The MDS group consisted of 34 RA, three RARS, eight RAEB and two RAEB-T patients with mean absolute neutrophil values of 1992, 1413, 1441, and 380 per mm3, respectively. The diagnosis was established based on bone marrow morphology and results of cytogenetic studies. In comparison to controls (147 +/- 38/10(6) MNC), significantly decreased numbers of bone marrow secondary CFC were found in MDS: in patients with RA and RARS, 21 +/- 7 secondary CFC per 10(6) bone marrow MNC (P < 0.001); patients with RAEB and RAEB-T: 39 +/- 12 CFC per 10(6) marrow MNC (P < 0.001). In all groups tested, the decrease in peripheral blood secondary CFC numbers was consistently less pronounced. In MDS patients with hypocellular bone marrow, secondary CFC were lower but not significantly different in comparison to MDS with hypercellular marrow (18 +/- 6 vs 35 +/- 11; NS; hypoplastic bone marrow also was not associated with significantly lower neutrophil counts). However, in 24% of patients with MDS, bone marrow secondary CFC were within the normal range, while in the aplastic anemia group only one of the patients showed secondary CFC number within normal range. Bone marrow and blood secondary CFC numbers in hypoplastic RA were significantly higher than those in severe aplastic anemia 919 +/- 5 in bone marrow, P < 0.01; 7 +/- 2 in blood, P < 0.05). This trend was even more pronounced in hypoplastic RA with chromosomal abnormalities. However, no significant differences were found between the secondary CFC numbers in hypoplastic RA and moderate aplastic anemia. We concluded that, although the deficiency in the stem cell compartment is less severe in MDS than in aplastic anemia, depletion of early hematopoietic cells is an essential part of the pathophysiology in both diseases.     

宫颈癌术后骨髓保护调强放疗研究

Objective: The aim of this study was to compare bone marrow-sparing intensity-modulated radiotherapy (IMRT)with IMRT without entering pelvic bone marrow as a planning constraint in the treatment of cervical cancer after hysterectomy. Methods: For a cohort of 10 patients, bone marrow-sparing IMRT and routine IMRT planning were designed. Theproscribed dose was 45 Gy/1.8 Gy/25f, 95% of the planning target volume received this dose. Doses were computed with a commercially available treatment planning system (TPS) using convolution/superimposition (CS) algorithm. Plans were compared according to dose-volume histogram (DVH) analysis in terms of planning target volume (PTV) homogeneity and conformity indices (HI and CI) as well as organs at risk (OARs) dose and volume parameters. Results: Bone marrow-sparing IMRT had an vantages over routine IMRT in terms of CI, but inferior to the latter for HI. Compared with routine IMRT, V5,V10, V20, V30, V40 of pelvic bone marrow of bone marrow-sparing IMRT reduced by 1.81%, 8.61%, 31.81%, 29.50%, 28.29%,respectively. No statistically significant differences were observed between bone marrow-sparing IMRT and routine IMRT in terms of small bowel, bladder and rectum. Conclusion: For patients with cervical cancer after hysterectomy, bone marrowsparing IMRT reduced the pelvic bone marrow volume irradiated at all dose levels and might be conducive to preventing the occurrence of acute bone marrow toxicity.     

Cytogenetic abnormalities in patients with severe aplastic anemia

BACKGROUND: Cytogenetic abnormalities have been described in a few patients with otherwise typical severe aplastic anemia (SAA), and the possible clonal nature of this disease is a controversial issue. MATERIALS AND METHODS: Sixty-nine patients with acquired severe aplastic anemia underwent cytogenetic examination on bone marrow cells at the time of diagnosis (n = 34) and/or at least twice after immunosuppressive therapy (IS) (n = 35). RESULTS: We identified 2 major groups. Group A: 51 patients (74%) were normal and remained normal. Group B: 18 patients (26%) had at least one abnormal cytogenetic analysis. This second group could be further subdivided as follows: (B1) chromosomal abnormalities not present at first examination and acquired in the course of the disease (n = 7); (B2) clonal cytogenetic abnormalities present at first examination and persisting (n = 3); (B3) reversible cytogenetic abnormalities (n = 8). The most frequent abnormality was trisomy 8 (n = 8) followed by monosomy 7 (n = 2); 82% of patients are alive in group A and 61% in group B. Three patients developed acute leukemia, all from group B. This represents 4% of all patients or 17% of those with at least one abnormal cytogenetic test. CONCLUSIONS: Thus the majority of SAA patients have normal karyotypes in marrow cells at presentation and at follow-up. Patients with abnormal karyotypes exist and can be further subdivided into those with reversible and those with persistent abnormalities. The latter are at risk of developing myelodysplasia or acute leukemia.     

Recombinant human erythropoietin resistance in hemodialysis. Effects of paired filtration dialysis

A percentage of hemodialysis (HD) patients are resistant to recombinant human erythropoietin (rHuEPO), a phenomenon that occurs less frequently in patients dialyzed with biocompatible membranes (M) and in peritoneal dialysis. The authors evaluated the effects of paired filtration dialysis (PFD)--a dialysis technique based on the use of an emophan M in conjunction with a polysulphone M--on erythropoiesis in HD patients resistant to rHuEPO. Twelve HD patients with anemia resistant to long-term therapy with rHuEPO (200.24 U/kg body weight three times per week intravenously for 10.2 months) were studied. Patients had been treated for an average of 46.9 months with bicarbonate HD, using cuprophan M (Phase A) and, successively, for 12 months by PFD (Phase B). The following parameters were evaluated monthly: 1) hemoglobin and hematocrit values; 2) serum levels of erythropoietin (EPO); and 3) serum levels of interleukin (IL)-3, IL-6, IL-10, IL-1 beta, tumor necrosis factor alpha (TNF-alpha), and interferon gamma (IFN-gamma). At the end of Phase A and Phase B, patients underwent bone marrow biopsies to evaluate 1) bone marrow burst forming unit erythroid (BFU-E) and colony forming unit erythroid (CFU-E) proliferative capacity, and 2) bone marrow mononuclear cell EPO-receptor (EPO-R) number. During Phase B, there was a progressive rise in hematocrit and hemoglobin values, so that within the sixth month, the rHuEPO dose was reduced to 80 +/- 15 U/kg body weight three times per week. At the same time, an increase in serum IL-3, IL-6, and IL-10 levels was seen, whereas serum IL-1 beta, TNF-alpha, and IFN-gamma levels decreased. This was accompanied by a rise in BFU-E and CFU-E growth and in bone marrow mononuclear cell EPO-R number. During Phase B, after the dialysis session, serum EPO levels increased by about 30% in comparison with pre dialysis values, whereas during Phase A they decreased by about 14%. In HD patients, EPO resistance may caused either by absorption of rHuEPO to the cuprophan M, or an increased release of cytokines that inhibit erythropoiesis, such as IL-1 beta, TNF-alpha, and IFN-gamma, and to a decrease in stimulatory cytokines such as IL-3, IL-6, and IL-10. These negative phenomena are reversed by the use of biocompatible dialysis techniques such as PFD.     

Amifostine (WR-2721) shortens the engraftment period of 4-hydroperoxycyclophosphamide-purged bone marrow in breast cancer patients receiving high-dose chemotherapy with autologous bone marrow support

4-Hydroperoxycyclophosphamide (4-HC), a commonly used marrow-purging agent, is active against many tumors, but is also toxic to normal marrow progenitors. Amifostine (WR-2721) is a sulfhydryl compound with chemoprotectant activity. Preclinical studies using suspensions of bone marrow and breast cancer cells demonstrated that ex vivo treatment with amifostine followed by 4-HC resulted in protection of marrow progenitors, with no compromise in the antitumor effect of 4-HC. This fact stimulated the development of a clinical trial. Bone marrow was harvested from 15 poor-prognosis breast cancer patients and randomly assigned to ex vivo treatment with amifostine followed by 4-HC (amifostine + 4-HC), or treatment with 4-HC alone. High-dose chemotherapy was then administered followed by infusion of the purged autologous bone marrow support (ABMS). Leukocyte engraftment, defined as a white blood cell count > or = 1 x 10(9)/L, was achieved in an average of 26 days for patients whose marrow was purged with amifostine + 4-HC versus 36 days for patients whose marrow was purged with 4-HC alone (P = .032). The average number of platelet transfusions (12 v 29; P = .017) and days of antibiotic therapy (28 v 40; P = .012) were significantly less for patients whose marrow was exposed to amifostine + 4-HC, compared with 4-HC alone. Unpurged backup marrow fractions were infused into three patients whose marrow was purged with 4-HC alone, because of inadequate marrow recovery. None of the patients who received amifostine + 4-HC-purged marrow required a backup marrow fraction. Complete remissions were achieved in 83% of patients with measurable disease, with no difference between the two cohorts. Forty-three percent of patients remained alive and progression-free at a mean of 13 months posttransplant. There was no significant difference in the rate or pattern of relapse for patients whose marrow was purged with amifostine + 4-HC compared with those whose marrow was purged with 4-HC alone. Ex vivo treatment of marrow with amifostine significantly shortens the time to marrow recovery, thereby reducing the risk of myelosuppressive complications in breast cancer patients receiving high-dose chemotherapy and 4-HC-purged ABMS. Since supportive care requirements are also significantly decreased, amifostine may reduce the cost of such therapy.     

Quantitative studies of splenic erythropoiesis in polycythaemia vera and myelofibrosis

A quantitative scanning method employing cyclotron-produced 52Fe has been developed to assess splenic erythropoiesis in patients with myeloproliferative disorders. In 12 patients with myelofibrosis splenic uptake of 52Fe was from 5.0% to 48% of the injected dose. Although a single patient with classical polycythaemia vera had a minor uptake of 2.8% of six other patients with this diagnosis showed no concentration of isotope in the splenic area. The fraction of 52Fe in the spleen of four patients with 'transitional' myeloproliferative disorders characterized by a high red cell mass, hypercellular bone marrow and a leucoerythroblastic blood film varied from 5% to 41%. No clear relationship was noted between the degree of splenic erythropoiesis as defined by this technique and the level of haemoglobin, the degree of splenomegaly, the effectiveness of erythropoiesis of traditional 59Fe surface counting. If splenectomy is considered in patients with myelofibrosis splenic 52Fe quantitation will provide more precise data on the contribution of splenic erythropoiesis than 59Fe surface counting alone.     

Removal of carcinoma cells from contaminated bone marrow using the lipophilic cation rhodamine 123

Autologous bone marrow transplants for solid tumor treatment are severely limited by the potential presence of residual cancer cells in the reinfused bone marrow and can lead to future tumor recurrence. This article presents a novel method of removing carcinoma cells from bone marrow with contaminating cancer cells. This method is based on our previous studies demonstrating that carcinoma cells have a higher uptake of lipophilic cations such as rhodamine 123 than their normal epithelial counterparts. When the relative differences in rhodamine 123 uptake are quantified, carcinoma cell lines demonstrated a 7.4-21 times greater uptake of rhodamine 123 than normal mouse bone marrow cells. More important, when normal bone marrow cells and carcinoma cell lines are mixed to simulate carcinoma-contaminated bone marrow, individual cell populations continue to exhibit characteristic and identifiable relative differences (10-20 times) in rhodamine 123 uptake. Differential sorting of bone marrow/carcinoma cell mixtures with respect to rhodamine 123 fluorescence intensity resulted in the removal of 95-99% of the "contaminating carcinoma cells." The recovered bone marrow cells were fully viable as ascertained by their ability to form splenic colonies. In our preliminary experiments, sorted bone marrow cells transplanted into lethally irradiated C57BL6 mice allowed the mice to survive for more than 8 months. In light of these promising results, we propose that lipophilic cations may play a role in the purification of autologous bone marrow used in transplants for patients with advanced solid tumors.