Expression of IAP family proteins and its clinical importance in breast cancer patients.

Inhibitor of apoptosis (IAP) family proteins is involved in mechanisms of resistance to apoptosis in various cancer cells. The aim of this study was to assess the expression of selected IAP proteins such as XIAP, cIAP-1, cIAP-2 and survivin in breast cancer patients and evaluates their relationship with the prognostic and predictive factors and their impact to overall survival (OS) and progression free survival (PFS). The study was conducted with the use of tissue samples prospectively collected from 92 previously untreated female breast cancer patients. The control encompassed 10 fibroadenoma patients. The expression of XIAP, cIAP-1, cIAP-2 and survivin was assessed using flow multicolor cytometry. XIAP expression was present in 99 % of the breast cancer patients (91/92) with the median expression 13.65% (range 1-66.8%). Expression of XIAP in breast cancer was significantly higher compared to the control group (p=0.006). Median expression of cIAP-1, cIAP-2 and survivin in the study group was 25.95% (range 0.8-83.7%), 16.7% (range 1-53.2%) and 4.6% (range 0-43%) respectively. In the rank Spearman test, strong correlations (p<0.001) were seen among the expressions of XIAP, cIAP-2 and survivin, in all combination. Additionally, week correlation between XIAP and cIAP-1 was observed (p=0.02). The median expression of XIAP and survivin was significantly higher in more advanced tumors (stages pT2/pT3 vs. pT1). The median PFS and OS in breast cancer group were 46.15 and 47.1 months respectively. No significant correlations were observed among expressions of IAP family proteins and survival. However, low expression of XIAP in breast cancer showed trend to longer PFS (p=0.08). XIAP, cIAP-1 cIAP-2 and survivin participate in antiapoptotic mechanisms in breast cancer and XIAP and survivin seem to have the most significant prognostic importance. Further studies are needed to establish more complete prognostic and predictive values of IAP family proteins in breast cancer patients.

Ciprofloxacin prophylaxis for patients undergoing high-dose chemotherapy and autologous stem cell transplantation (ASCT) - a single-center experience.

PURPOSE: The objective of the study was to evaluate the efficacy of ciprofloxacin prophylaxis for patients undergoing high-dose chemotherapy followed by autologous stem cell transplantation (ASCT). MATERIALS AND METHODS: The data of 104 patients transplanted at the Department of Hematology Medical University of Lodz between 2005 and 2008 were analyzed. The cohort was divided into two groups depending on the administered ciprofloxacin prophylaxis. Conditioning regimens did not differ significantly among the groups. Multiple myeloma was the main indication for ASCT in both groups. RESULTS: Ciprofloxacin prophylaxis resulted in a statistically significant reduction of duration of intravenous (IV) antibiotic treatment in the group with prophylaxis (p=0.01). The trend has been observed towards lower prevalence of infectious episodes in the prophylaxis group. Positive blood cultures were similar in both groups with no significant resistance to ciprofloxacin. CONCLUSION: These data demonstrate that ciprofloxacin prophylaxis is beneficial for patients treated with ASCT following high dose chemotherapy regimen, in terms of the intravenous antibiotics usage. This advantage directly translates into economic benefit and may also induce less bacterial resistance due to less exposure to antibiotics.

Correlation of Smac/DIABLO protein expression with the clinico-pathological features of breast cancer patients.

Smac/DIABLO protein promotes caspase-dependent apoptosis by inhibition of inhibitor of apoptosis protein (IAP) family members. The role of Smac/DIABLO in breast cancer has not been yet established. Therefore, the aim of the study was to assess the expression of this protein in tumor cells from breast cancer patients. The expression of Smac/DIABLO was analyzed in 62 breast cancer patients by flow cytometry. The obtained results were compared with expression of this protein in benign breast tumor tissue, which served as the control (11 patients with fibroadenoma). Expression of caspase-3 proteins in breast cancer was also evaluated. Smac/DIABLO expression in breast cancer was correlated with clinical and pathological data. Although the expression of Smac/DIABLO protein was found in all examined samples of both the breast cancer and fibroadenoma patients, the median expression of Smac/Diablo in breast cancer was significantly lower than in the control (39.1% vs. 48.1%; p=0.0047). Smac/DIABLO expression correlated with expression of caspase-3 (p=0.000008). In pT1 breast cancer patients, expression of Smac/DIABLO protein was higher than in those with pT2-3 (p=0.02). Diffuse cancer infiltration significantly correlated with lower expression of Smac/DIABLO protein (p=0.02). Moreover, there was a loose correlation between low expression of Smac/DIABLO protein and cancer embolus in minor blood and lymphatic vessels (p=0.08). Our results indicate that expression of Smac/DIABLO inversely correlates with the tumor stage, which may suggest that this protein may play an important role in the breast cancer development.

Novel and emerging drugs for acute myeloid leukemia: pharmacology and therapeutic activity.

For the last twenty years, significant progress in Molecular and Cellular Biology has resulted in a better characterization and understanding of the biology and prognosis of acute myeloid leukemia (AML). These achievements have provided new opportunities for the development of innovative, more effective therapies. Novel agents potentially useful in the treatment of patients with AML include new formulations of established drugs, newer nucleoside analogs, molecular target drugs, monoclonal antibodies and other agents. Three newer nucleoside analogs, clofarabine, troxacitabine and sapacitabine have been recently investigated in patients with AML. Two methylation inhibitors, 5-azacyticline and decitabine are pyrimidine nucleoside analogs of cytidine which can be incorporated into RNA and/or DNA. Lower doses of these agents are active in AML and have been extensively investigated, especially in secondary AML and AML in elderly patients. Tipifarnib and lonafarnib are orally available farnesyltransferase inhibitors with in vitro and in vivo activity against AML. In recent years, FLT3 inhibitors, lestaurinib, tandutinib and PKC 412 have been developed and tested in AML. The preclinical observations and clinical studies indicate that FLT3 inhibitors are promising agents in the treatment of FLT3 mutated AML patients, especially when used in combinations with chemotherapy. Several newer MDR inhibitors, including valspodar (PSC-833) and zosuquidar trihydrochloride have been also tested for the treatment of relapsed AML. This article reviews the various classes of AML targets and drugs that are under early phase clinical evaluation, especially those that are likely to enter clinical practice in the near future.

Circulating endothelial cells and angiogenic proteins in patients with systemic lupus erythematosus.

The aim of our study was to assess absolute counts of different subpopulations of circulating endothelial cells (CEC) in patients with active and inactive systemic lupus erythematosus (SLE). We have also investigated a potential correlation of CEC numbers with serum levels of angiogenic proteins as well as with clinical and laboratory symptoms of the disease. For the first time in SLE, CEC were enumerated directly, by the 'single platform' method. Resting (rCEC), activated (aCEC) and progenitor (pCEC) endothelial cells were identified in patients with SLE and healthy volunteers using four-colour flow cytometry. Serum concentrations of angiogenic proteins (vascular endothelial growth factor, placental growth factor (PIGF), soluble vascular cell adhesion molecule and endoglin) were evaluated by ELISA. The SLE activity was scored according to the Systemic Lupus Activity Measure system. We found that total CEC number in patients with SLE was significantly higher (median 14.2/microL) than in the control group (median 3.3/microL) (P < 0.0001). Absolute counts of aCEC, rCEC and pCEC (medians 4.9/microL, 6.8/microL and 2.3/microL, respectively) were also higher in patients with SLE than in healthy persons (medians 0.9/microL, 1.6/microL and 0.1/microL, respectively), with P < 0.0001 for all comparisons. There was no correlation between CEC or their subpopulations and SLE activity. Strong positive correlations were found between CEC, rCEC and pCEC numbers and serum levels of PIGF. Moreover, aCEC, rCEC and pCEC counts were significantly higher in SLE patients with leukopenia. In conclusion, our results show that absolute CEC counts and angiogenic proteins levels are elevated in patients with SLE as compared with healthy controls. These data may suggest that angiogenic process is involved in the pathogenesis of this disease.

Outcome of treatment in adults with Philadelphia chromosome-positive and/or BCR-ABL--positive acute lymphoblastic leukemia-retrospective analysis of Polish Adult Leukemia Group (PALG).

Patients with Philadelphia chromosome-positive (Ph+) and/or BCR-ABL+ acute lymphoblastic leukemia (ALL) have extremely poor prognoses. Most of these patients have additional, heterogenous karyotype abnormalities, the majority of which have uncertain clinical significance. In this study we analyzed the clinical characteristics, karyotype abnormalities, and outcome of 77 patients with Ph+ and/or BCR-ABL+ ALL registered in Poland in 1997-2004. In 31/55 patients with known karyotype, the sole t(9;22)(q34;q11) abnormality had been diagnosed; in one patient, variant translocation t(4;9;22)(q21q31.1;q34;q11), and additional abnormalities in 23 (42%) patients, had been diagnosed. The characteristics of the patients with Ph chromosome and additional abnormalities were not significantly different when compared with the entire analyzed group. Out of 77 patients, 54 (70%) achieved first complete remission (CR1) after one or more induction cycles. The overall survival (OS) probability of 2 years was 63, 43, and 17% for patients treated with allogeneic stem cell transplantation (alloSCT), autologous SCT, and chemotherapy, respectively (log rank p=0.002). Median OS from the time of alloSCT was significantly longer for patients transplanted in CR1 compared with alloSCT in CR >1 (p=0.032). There were no significant differences in CR rate, disease-free survival (DFS), and OS for patients with t(9;22) and additional abnormalities compared with the whole group. Only WBC >20 G/l at diagnosis adversely influenced OS probability (log rank p=0.0017). In conclusion, our data confirm poor outcome of Ph+ and/or BCR-ABL+ ALL. Only patients who received alloSCT in CR1 had longer DFS and OS. We have shown that additional karyotype abnormalities did not influence the clinical characteristics of the patients; however, their influence on treatment results needs to be further assessed.

A multicenter, open, noncomparative, phase II study of the combination of cladribine (2-chlorodeoxyadenosine), cytarabine, granulocyte colony-stimulating factor and mitoxantrone as induction therapy in refractory acute myeloid leukemia: a report of the Polish Adult Leukemia Group.

Purine nucleoside analogues, cladribine (2-chlorodeoxyadenosine, 2-CdA) and fludarabine (FAMP) are active agents in acute myeloid leukemias (AMLs). Synergistic interaction between FAMP or 2-CdA with cytarabine (cytosine arabinoside, Ara-C) has been demonstrated in preclinical and clinical studies. The current multicenter phase II study was initiated to evaluate the efficacy and toxicity of induction treatment consisting of 2-CdA (5 mg/m2), Ara-C (2 g/m2), mitoxantrone (MIT, 10 mg/m2) and granulocyte colony-stimulating factor (G-CSF) (CLAG-M) in refractory AML. In case of partial remission, a second CLAG-M was administered. Patients in complete remission (CR) received consolidation courses based on high-dose Ara-C and MIT with or without 2-CdA. Forty-three patients from five centers were registered: 25 primary resistant and 18 relapsed. CR was achieved in 21 (49%) patients, 20 (47%) were refractory and 2 (5%) died early. Hematologic toxicity was the most prominent toxicity of this regimen. The overall survival (OS; 1 year) for the 42 patients as a whole and the 20 patients in CR were 43% and 73%, respectively. Disease-free survival (1 year) was 68.6%. None of the analyzed prognostic factors influenced the CR and OS probability significantly. We conclude that CLAG-M regimen has significant antileukemia activity in refractory AML, which seems to be better than the activity of many other regimens. The toxicity of the treatment is acceptable.

The inhibitor of apoptosis protein family and its antagonists in acute leukemias.

The Inhibitor of Apoptosis Protein family (IAP) functions as inhibitors of apoptotic pathways, both death receptor- and mitochondrial mediated. We detail the current body of knowledge for the IAP family with regard to their structure and function, their expression in normal and leukemic cells, and their prognostic importance in acute leukemia. Although there is some evidence that IAPs play an important role in the chemoresistance of leukemia cell lines, little is known about their influence on this phenomenon in acute leukemia cells of human origin. IAPs are also explored as a specific target for new antitumor strategies, including antisense oligonucleotides of XIAP (X-chromosome-linked IAP) or survivin and small molecules of polyphenylurea-based XIAP inhibitors. Several proteins negatively regulate the function of the IAP family. One of those antagonists is Smac/DIABLO. Short peptides of Smac were found to enhanced apoptosis, induced by chemo- or immunotherapy, in the leukemic cells in vitro. Moreover, small-molecule agents, resembling Smac/DIABLO in function, were shown to potentiate cytotoxicity of chemotherapy in different malignancies. IAPs, exhibiting downstream influence on both external and intrinsic pathways as well as on some caspase-independent mechanisms of apoptosis, are potentially attractive target for anti-tumor therapy, although their role in the pathology and prognosis of acute leukemia has to be further elucidated.

Addition of cladribine to daunorubicin and cytarabine increases complete remission rate after a single course of induction treatment in acute myeloid leukemia. Multicenter, phase III study.

To assess the efficacy of an original DAC-7 regimen: daunorubicine (DNR) 60 mg/m2/day, days 1-3; cytarabine (AraC) 200 mg/m2/day, days 1-7; cladribine (2-CdA) 5 mg/m2/day, days 1-5, 400 untreated adult acute myeloid leukemia patients (including 63 with preceding myelodysplastic syndrome), aged 45 (16-60) years were randomized to either DAC-7 (n=200) or DA-7 (without 2-CdA, n=200). The overall CR rate equaled 72% for DAC-7 and 69% for DA-7 arm (P=NS). After a single course of DAC-7 induction, the CR rate equaled 64% and was significantly higher compared to 47% in the DA-7 arm (P=0.0009). Median hospitalization time during the induction was 7 days shorter for DAC-7 compared to the DA-7 group (33 vs 40 days, P=0.002). Toxicity was comparable in both groups. The probability of 3-year leukemia-free survival (LFS) for DAC-7 and DA-7 group equaled 43 and 34%, respectively (P=NS). There was a trend toward higher LFS rate for patients aged >40 years receiving DAC-7 compared with DA-7 regimen (44 vs 28%, P=0.05). This study proves that addition of 2-CdA increases antileukemic potency of DNR+AraC regimen, thus resulting in a higher CR rate after one induction cycle when compared to DA-7, without additional toxicity. It shortens hospitalization time and may improve long-term survival in patients aged >40 years.

Acute lymphoblastic leukemia in elderly: the Polish Adult Leukemia Group (PALG) experience.

This is a retrospective, multicenter study to evaluate biological features and outcome of elderly patients diagnosed with acute lymphoblastic leukemia (ALL) during the last 10 years in ten hematological centers in Poland. Eighty-seven patients aged 60 years or older were studied. To our knowledge, this is one of the largest group of elderly patients with ALL evaluated. We have not observed differences in immunological subtypes and Ph chromosome incidence as compared with younger adult ALL presented in the literature. Induction chemotherapy was administered in 75 patients. We observed complete remission (CR) in 34 (45%, 95% CI: 33-56%) patients. Induction death occurred in 11 (15%) patients. Thirty patients (40%) showed primary resistance to chemotherapy. Median overall survival (OS) of all patients was 150 days. Median disease-free survival (DFS) of responding patients was 180 days. We observed four long-term survivors (DFS longer than 3 years) in our group of patients. Factors influencing OS were CR achievement, female gender, and WBC below 30 x 10(9)/l. Male gender was the only prognostic factor negatively affecting probability to achieve CR. We have not observed any differences in either biology or outcome between patients aged 60-69 years and those aged more than 70 years. ALL of the elderly is a rare disease with poor prognosis. Further clinical trials evaluating the disease features, outcome, and new therapeutic approaches are warranted.

A multicenter, open, non-comparative, phase II study of the combination of cladribine (2-chlorodeoxyadenosine), cytarabine, and G-CSF as induction therapy in refractory acute myeloid leukemia - a report of the Polish Adult Leukemia Group (PALG).

OBJECTIVES: To evaluate the efficacy and toxicity of cladribine (2-chlorodeoxyadenosine, 2-CdA), cytarabine (Ara-C), and granulocyte-colony stimulating factor (G-CSF) (CLAG) regimen in refractory acute myeloid leukemia (AML) in the multicenter phase II study. METHODS: The induction chemotherapy consisted of 2-CdA 5 mg/m2, Ara-C2 g/m2, and G-CSF. In the case of partial remission (PR), a second CLAG was administered. Patients in complete remission (CR) received two consolidation courses based on HD Ara-C, mitoxantrone or idarubicine, with or without 2-CdA. RESULTS: Fifty-eight patients from 11 centers were registered; 50 primary resistant and eight early relapsed (CR1 < 6 months). CR was achieved in 29 (50%) patients, 19 (33%) were refractory, and 10 (17%) died early. Forty of 50 primary resistant patients received daunorubicin (DNR) and Ara-C as the first-line induction therapy (DA-7), 10 received additional 2-CdA (DAC-7). The CR rates after CLAG were 58% and 10%, respectively in each group (P = 0.015). Five of six patients with myelodysplastic syndrome (MDS)/AML achieved CR. Hematologic toxicity was the most prominent toxicity of this regimen. The overall survival (OS, 1 yr) for the 58 patients as a whole, and the 29 patients in CR were 42% and 65%, respectively. Disease-free survival (DFS, 1 yr) was 29%. Only first-line induction treatment with DA-7 significantly influenced the probability of CR after CLAG. None of the analyzed factors significantly influenced DFS and OS. CONCLUSION: CLAG regimen has significant anti-leukemic activity and an acceptable toxicity in refractory AML. The addition of 2-CdA to the first-line induction treatment may worsen the results of salvage with CLAG. The high CR rate in patients with MDS preceding AML deserves further observation.

The treatment of acute myeloid leukemia with mitoxantrone, etoposide and low-dose cytarabine in elderly patients - a report of Polish Acute Leukemia Group (PALG) phase II study.

The common dilemma in the treatment of elderly patients with acute myeloid leukemia (AML) is whether to use intensive myelosuppresive therapy with higher risk of treatment related mortality (TRM), but a chance for complete remission (CR), or to treat less intensively in order to prolong survival time with a better quality of life. The aim of this prospective, phase II study was to assess the efficacy and toxicity of low dose combination induction treatment consisted of cytarabine at a dose of 10 mg/m2 every 12 h s.c. for 7 days, VP-16 at a dose of 100 mg/day p.o. for 7 days and mitoxantrone at a dose of 6 mg/m2 i.v daily on days 1-3. Two induction courses were planned. In the group of 44 patients 12 (27%) achieved CR, 4 (9%) patients were in PR and there were 9 (20%) early deaths (ED). Age, performance status, preceding myelodysplastic syndrome, karyotype, WBC and % of blasts in bone marrow were not significant prognostic factors for CR probability. The following initial factors appeared to be related to a shorter duration of survival time from the start of treatment: age >70 (p<0.03), poor performance status (p<0.03), and % of BM blasts 50 (p<0.05). We conclude that, despite promising results in the pilot study the efficacy of this induction treatment is not better than the efficacy of other regimens. The hematological toxicity of this treatment seems to be comparable with "3+7" regimen.

Circulating IL-6-type cytokines and sIL-6R in patients with multiple myeloma.

We investigated the serum concentration of interleukin-6 (IL-6) and four IL-6 family cytokines - oncostatin M (OSM), leukaemia inhibitory factor (LIF), interleukin-11 (IL-11) and ciliary neurotrophic factor (CNTF) as well as IL-6 soluble receptor (sIL-6R) - using an enzyme-linked immunosorbent assay (ELISA) in 67 patients with multiple myeloma (MM) and 24 healthy controls, for a possible association between the serum levels of these peptides with disease activity and known prognostic factors. sIL-6R was detectable in all 67 and IL-6 in 65 (97%) patients. Both peptides were measurable in all healthy controls. In contrast, OSM was detectable in 30 (44.8%) MM patients and in only four (16.6%) normal individuals. The serum levels of IL-6, OSM and sIL-6R were significantly higher in MM patients compared with control group (P < 0.001, P < 0.03 and P < 0. 001 respectively). The highest concentrations of these cytokines were found in patients with progressive disease and the lowest in MM patients with stable disease and in healthy persons. LIF was detectable in four (6%), CNTF in 28 (41.8%) and IL-11 in eight (11. 9%) of the patients with MM. In the control group LIF, CNTF and IL-11 were measurable in 8.3%, 33.3% and 8.3% respectively. The serum concentration of these cytokines did not correlate either with clinical stage or with the phase of disease and was similar to those in healthy individuals. We found significant positive correlation between IL-6 levels and OSM (P < 0.001). We also observed positive correlation between beta2-M concentration and serum levels of IL-6 (P < 0.002), sIL-6R (P < 0.02) and OSM (P < 0.04) as well as a positive relationship between CRP and IL-6 (P < 0.001) and OSM (P < 0.002). In conclusion, the serum levels of IL-6, OSM and sIL-6R, but not LIF, IL-11 and CNTF, may be useful markers of MM activity.

Serum levels of IL-6 type cytokines and soluble IL-6 receptors in active B-cell chronic lymphocytic leukemia and in cladribine induced remission.

We have investigated the serum concentrations of interleukin-6 (IL-6) and two IL-6 family cytokines-oncostatin M (OSM) and leukemia inhibitory factor (LIF)-in 63 patients with B-cell chronic lymphocytic leukemia (B-CLL) and 17 healthy controls using the enzyme-linked immunosorbent assay (ELISA) method. Simultaneously, we measured the serum levels of the soluble forms of two subunits of the IL-6 receptor complex-ligand binding glycoprotein 80 (sIL-6R) and glycoprotein 130 (sgp130). The cytokines and receptors were evaluated in 25 untreated patients and 38 patients treated with cladribine (2-CdA), as well as in 17 healthy controls. We have correlated the serum levels of these proteins with Rai's clinical stage of the disease, the response to 2-CdA treatment and some hematological parameters. We have also evaluated the correlation of the IL-6 serum level with the concentration of OSM and IL-6 soluble receptors. IL-6 was measurable in 62/63 (98.4%), OSM in 20/25 (80%) of untreated and 14/38 (37.8%) of the treated patients. sIL-6R and sgp130 were detectable in all 63 patients and LIF in none of the CLL patients. IL-6 serum level in untreated patients was not significantly different as compared to its concentration in the control group (P>0.05). However, in the patients treated with 2-CdA the IL-6 level was significantly lower (P<0.02), and the lowest concentration was found in the patients with complete remission (CR; median 1.4pg/ml; P<0.02). The concentration of sIL-6R was significantly higher in untreated (median 61.8 ng/ml) and treated (median 50.1 ng/ml) CLL patients when compared to normal persons (median 41.2 ng/ml; P=0.04; P<0.001, respectively). There was no difference between the sIL-6R levels in the patients with CR and the healthy controls. In non-responders sIL-6R concentration was the highest and similar to its level in the untreated patients. OSM level was higher in the untreated patients (median 1.8pg/ml) than in the normal controls (median 0.0pg/ml; P<0.001) and in the CR patients (median 0.0pg/ml; P<0.03). The serum concentration of sgp130 was similar in the untreated (median 480 pg/ml) and treated (median 470 pg/ml) patients, as well as in the healthy persons (median 420 pg/ml; P>0.05). We have found significant positive correlation between the levels of sIL-6R and the lymphocytes count in CLL patients (p=0.423; P<0.001). In addition, sIL-6R and OSM serum concentrations correlated also with CLL Rai stage. In conclusion, the serum level of IL-6, OSM and sIL-6R, but not LIF and sgp130, are useful indicators of CLL activity.