ALBERT

Description: Rationale: No published randomized trials have compared low molecular weight heparin (LMWH) to unfractionated heparin (UFH) for thromboprophylaxis in medical-surgical ICU patients. Objectives: A) Our Feasibility Objectives were to assess: 1) timely enrollment and complete, blinded study drug administration, 2) LMWH bioaccumulation in renal insufficiency, 3) twice weekly leg ultrasounds, and 4) recruitment rates for a future trial. B) Our Clinical Objectives were to assess 1) the effectiveness of LMWH vs UFH by estimating DVT, and pulmonary embolism (PE) rates, and 2) the safety of LMWH vs UFH regarding bleeding, thrombocytopenia and HIT. Methods: In this prospective randomized stratified concealed blinded multicenter trial, we included patients ≥18 with expected ICU stay ≥72h. We excluded trauma, orthopaedic, cardiac, or neurosurgery, severe hypertension, DVT, PE or hemorrhage within 3 mos, INR 〉2ULN, PTT 〉2ULN, platelets 99% of scheduled doses; every dose was blinded. 2) No LWMH bioaccumulation was observed as measured by anti-Xa levels when creatinine clearance decreased to

Description: Background: The Research on Adverse Events and Reports (RADAR) project, an independent pharmacovigilance project, has reported that post-marketing dissemination of clinical information on systemic adverse drug reactions (sADRs) in the FDA approved package labels of cancer drugs is often incomplete, delayed, or absent. Methods: We compared post-marketing clinical assessments describing one previously unreported sADR: zoledronate (Z) and pamidronate (P)-associated osteonecrosis (ON) of the jaw. Data sources included 1) information submitted to the FDA by the manufacturer in support of an FDA package insert revision, 2) RADAR investigators review of case reports of BPs associated ON of the jaw reported to the FDA MedWatch program and, 3) RADAR investigators review of medical and dental records of BP treated multiple myeloma (MM) patients with confirmed, clinically obvious ON of the jaw at a large NCI-designated comprehensive cancer center. Results: The manufacturers summary material indicated that ON occurs at a rate of 〈 1 per 10,000 BPs treated individuals; occurs 4x more frequently in cancer versus non-cancer patients; has multiple causes including trauma, infection, radiation therapy, and long-term corticosteroids; and causal relationship with BPs is unknown. Radar investigators found that not a single case of BPs associated ON of the jaw has been reported to the FDA’s MedWatch program as of to date. RADAR investigators, however, identified 7 patients (on BPs) with confirmed ON of the jaw among 600 myeloma patients seen between March 2001 and June 2004 at our institution. Characteristics of these patients included presentation with localized jaw symptoms (n=7) requiring surgical and/or medical treatment. Conclusions: Independent toxicity assessments from the RADAR program suggest that ON is a serious, previously unrecognized ADR which can occur at a rate 〉 1% in MM patients, appears to be causally associated with prolonged BP therapy, and is difficult to treat. In comparison, materials submitted by the manufacturer to the FDA in support of a package insert revision as well as the FDA’s MedWatch data (with 0 cases reported) indicate that this sADR is rare, has multiple etiologies, and its association with BP is unknown. The manufacturer has informed the FDA that it will conduct a retrospective single-site study to further evaluate this toxicity but this method is likely to be limited by underreporting. In contrast, our findings highlight the need for 1) a multi-site prospective study to identify the true extent of this problem in patients receiving BPs, 2) educating health care officials about the importance of reporting ADRs to the FDA MedWatch program and, 3) careful evaluation of the source of toxicity information when assessing discrepant reports of sADRs.

Description: FAS (TNFRSF6/CD95/APO-1) is a cell surface receptor involved in apoptotic signal transmission. Dysregulation of this pathway is believed to result in down regulation of apoptosis and subsequent persistence of a malignant clone. A single nucleotide polymorphism resulting in guanine-to-adenine transition in the FAS promoter region occurs at position −1377, affecting an SP1 transcription factor binding site; an adenine residue at position −1377 of FAS promoter significantly reduces SP1 binding compared to guanine residue causing a decrease in FAS expression. Recent data in adult leukemia indicate that a variant allele at this site increases susceptibility to AML (Morgan et al; Cancer Research 63, 4327–4330). We hypothesized that FAS genotype would also increase risk of childhood AML and, by altering susceptibility to apoptosis might impact outcome of AML therapy. Methods: 442 children treated for AML on CCG studies 2941 and 2961 (intensive induction with daunorubicin, idarubicin, Ara-C, 6-thioguanine, etoposide, dexamethasone, +/− fludarabine, Ara-C, idarubicin; consolidation with high-dose Ara-C and L-asparaginase or HLA-matched sibling BMT) were genotyped for the −1377 FAS promoter polymorphism using mismatch amplification coupled with real time PCR (MAMA assay). Genotype frequencies were compared with published normal control frequencies. In addition patient outcomes were analyzed according to genotype. Results: Comparison of gene frequencies in AML patients and reported normal controls showed similar frequencies (78.7% GG, 18.6% GA, 2.7% AA in patients vs. 78% GG, 20% GA and 2% AA in controls; p=0.6). Stratification of cases by age at diagnosis, white blood count at diagnosis, AML subtype, or cytogenetics revealed no difference in genotype frequencies. Outcome data showed no significant difference in OS (p=0.22), EFS (p=0.31), TRM (p=0.42) or relapse rate (p=0.57) between patients with 1377GG genotype vs. 1377GA/1377AA genotypes. However, there was trend towards improved survival (OS at 5 years 75% AA, 55% GA, 49% GG; p=0.16) and EFS (66% AA, 43% GA, 39% GG; p=0.19) in AA cases, but small numbers (n=12 AA) may have contributed to the results not being statistically significant. Conclusion: These data, representing the only data in pediatric AML, suggest that FAS genotype does not affect the etiology of childhood AML and that children with AML differ from adults in terms of biology of their disease. However, further studies are needed to evaluate a potentially important role of FAS genotype in outcome of AML therapy.

Description: Anti-CD20 monoclonal antibody (rituximab) has been used in autoimmune cytopenia with variable success.We report 13 patients with chronic refractory autoimmune hemolytic anemia (AIHA) and idiopathic thrombocytopenic purpura (ITP) who each received 4 cycles of rituximab 375mg/m2 weekly. All 8 AIHA patients (6 idiopathic,2 secondary to a lymphoproliferative disorder, 5 splenectomised) had warm antibody type. Response was seen in 4(50%) of 8 patients (3 CR,1PR) and 3 patients remain in CR at 5,7,14 months post-therapy. Median time to maximum response (TMR) in responders was 9 weeks(range 6 – 18 weeks). In 5 ITP patients (4 splenectomised), 4(80%) responded (3CR) and one continues in CR 50 weeks after completion of rituximab treatment Median time to maximum response was 4 weeks (range 4 – 12 weeks). No pre-treatment clinical or laboratory parameters that predict response could be identifird in the AIHA or ITP groups.Our data indicate that rituximab is a relatively safe and effective treatment option in patients with refractory autoimmune hemolytic anemia and thrombocytopenia. Table 1 Number of patients Mean age (Years) Sex Overall response Complete response No response Time to maximal response AIHA 8 47.75 (26– 73) 3M:4F 4 (50%) 3 (37.5%) 4 (50%) 9 weeks ITP 5 58.6 (28–89) 4M:1F 5 (80%) 3 (60%) 2 (40%) 4 weeks

Description: Matched unrelated donor hematopoietic stem cell transplantation (MUD HSCT) is a potentially curative treatment for patients with acute myeloid leukemia (AML). The graft-versus-leukemia (GVL) effect may be potent enough to overcome the otherwise poor prognosis associated with AML though its efficacy for high risk cytogenetic subgroups is uncertain. To test this hypothesis, we analyzed outcomes by cytogenetic risk group in 324 patients in first complete remission (CR1), and 440 in CR2 undergoing NMDP-facilitated MUD HSCT between 1988–2002. Using the SWOG/ECOG classification of cytogenetic risk groups (Slovak et al. Blood, 2000) cytogenetics were classified as favorable in 14% of patients, intermediate in 71% and unfavorable in 16%. 56% of the patients were male and 42% were 〉 35 years at HSCT. 76% of patients and donors were matched at HLA-A, -B and -DRB1, 17% were mismatched at one or more loci and 7% were potentially matched (serologically matched at HLA-A and -B and potentially allele matched at -DR). Disease Status N Kaplan-Meier Estimate for Survival at 5 years Kaplan-Meier Estimate for Disease-Free Survival at 5 years Cumulative Incidence for 100 Day Transplant-Related Mortality Cumulative Incidence for Relapse at 5 years * p-value indeterminate; ** p=0.01 CR1 324 32 ± 6% 32 ± 5% 32 ± 5% 18 ± 4%     Intermediate 227 33 ± 7% 32 ± 7% 31 ± 6% 16 ± 5%*     Unfavorable 85 31 ± 11% 31 ± 10% 29 ± 10% 23 ± 9%* CR2 440 36 ± 5% 35 ± 5% 25 ± 4% 16 ± 3%     Favorable 93 46 ± 10% 44 ± 10% 25 ± 9% 10 ± 6%**     Intermediate 313 33 ± 6% 32 ± 5% 27 ± 5% 16 ± 4%**     Unfavorable 34 37 ± 17% 38 ± 16% 15 ± 12% 32 ± 15%** These data suggest that with the exception of the 5-year relapse rate, results of cytogenetics have little apparent influence on the outcome for patients undergoing MUD HSCT for AML in CR1. In CR2, results in patients with favorable cytogenetics appear to be better than those with intermediate or unfavorable cytogenetics, but are not statistically significantly different. Effective GVL and protection against relapse is observed, even in high risk cytogenetic subgroups. In this retrospective study, other prognostic factors may influence the outcome, but overall survival for patients with unfavorable cytogenetics appears at least as good as previously reported for matched sibling HSCT.

Description: Standard allogeneic stem cell transplant (allo-SCT) regimens have been associated with a high transplant-related mortality (TRM) in multiple myeloma (MM). Nonmyeloablative therapy can establish stable engraftment after allo-SCT and maintain the antitumor effect with less toxicity, which is important in heavily pretreated and elderly patients. We report on 16 poor-risk MM patients receiving allo-SCT from an HLA-matched (n = 14) or mismatched (n = 2) sibling following conditioning with melphalan 100 mg/m2 (MEL-100). Ten patients had refractory relapse, 4 responsive relapse, and 2 patients were in near complete remission (nCR) with poor-prognosis disease. Patients had received 1 (n = 9) or 2 (n = 7) prior autotransplants. Donor lymphocyte infusions (DLIs) were given to 14 patients with no clinical evidence of graft versus host disease (GVHD) either to attain full donor chimerism (n = 4) or to eradicate residual disease (n = 10). Fifteen patients showed myeloid engraftment, and 12 patients were full donor chimeras at day +21. No TRM was observed during the first 100 days. Acute GVHD developed in 10 patients; 1 had fatal grade IV GVHD. Seven progressed to chronic GVHD, limited in 3 and extensive in 4 patients. At a median follow-up of 1 year, 5 patients achieved and sustained CR, 3 nCR, and 4 partial remission. Of 4 patients progressing after transplantation, 3 achieved a remission following further chemotherapy and DLI. Remarkable graft versus myeloma responses were seen in chemotherapy-refractory patients. Two patients died of progressive disease, and 3 died of GVHD complications without active disease. GVHD remains a major problem with this procedure.

Description: Malignant B-cells from a high proportion of chronic lymphocytic leukaemia (B-CLL) patients over express the multidrug resistance (MDR) -1 gene encoded transmembrane efflux pump P-glycoprotein (P-gp). Inhibition of glucosylceramide synthesis has been shown to correlate with the expression and function of P-gp and sensitise cells to cytotoxic agents. We analysed the ability of glucosylceramide synthetase (GCS) inhibitors N-butyl-deoxygalactonojirimycin (OGB-1, 500μM) and N-nonyl-deoxygalactonojirimycin (OGB-2, 100μM) to sensitise B-CLL cells to conventional cytotoxic drugs 2-chlorodeoxyadenosine (CdA), chlorambucil (Chl) and fludarabine (FdR) using the in vitro cytotoxicity MTT assay. The effect on P-gp activity was also analysed using the calcein-AM accumulation assay and the results expressed as multidrug activity factor (MAF), where a MAF of 〉10 in the presence of a P-gp inhibitor denotes P-gp functional activity. GCS inhibitors were cultured with B-CLL cells for 24-48h before the assays were performed. The P-gp negative cell line CEM-CCRF had no MAF activity with an IC50 for vincristine (a known P-gp substrate) of 10ug/ml to 55.5ng/ml in the presence of OGB-2. In peripheral blood mononuclear cells from 3 normal volunteers (all P-gp +ve), the mean MAF value for Z.3HCL was 23.86 and for OGB-2 was 16.2. In 9/13 B-CLL samples there was P-gp functional activity in the presence of Z.3HCL with a mean MAF value of 22.15 (range 11.27–37.3). P-gp was over expressed in10/13 B-CLL samples. However, when available samples from this cohort were assessed with OGB-1 (n=4) and OGB-2 (n=13) the MAF value was

Description: Twenty-eight patients with relapsed or refractory CD20+ NHL have been enrolled in an ongoing phase I trial of dose-escalated 90YZ followed by high-dose BEAM and autotransplant in which the 90YZ dose is patient-specific based on dosimetry. 90YZ doses are calculated to deliver cohort-defined radiation doses (100, 300, 500, ... cGy) to critical organs (liver, lung or kidney), with 3–6 patients per group. On D -22, rituximab (R) 250 mg/m2 is infused followed by the imaging dose of 111In Zevalin® (5 mCi). Imaging is performed immediately post-injection and at 4, 24, 72, and 144 hours; dosimetry is performed on D -15. On D -14, R 250 mg/m2 is administered followed immediately by 90YZ at the dose calculated to deliver the cohort-prescribed absorbed radiation dose to the critical organ. On D -6 through -1, patients receive high-dose BEAM. On D0, a minimum of 2.0 X 106 CD34+ cells/kg is infused and G-CSF 5 μg/kg SQ daily begun. The median age was 54 (range: 25–72) years. NHL histologic subtypes were as follows: mantle cell 5, diffuse aggressive 13, low grade 5, and transformed 5. Most had received 3 or more treatment regimens, including R. The toxicity profile was similar to that associated with high-dose BEAM and included a decrease in DLCO for most patients with one patient at the 500 cGy dose level experiencing a transient decline to below 50% of the predicted value corrected for hemoglobin. The most common grade III/IV toxicities were infection, fever, stomatitis, nausea, vomiting, diarrhea, hemorrhage, and edema. One patient experienced transient veno-occlusive disease at the 700 cGy dose level. Engraftment occurred at a median of 10 days (range:8–18) to granulocytes ≥ 500/μL, and 21 days (range:13–40days) to platelets ≥20,000/μL . With a median follow-up of one year, the 3 year overall and progression-free survivals are 60% and 50%, respectively. Figure Figure 90-Y Zevalin Dosing by Cohort (median; range) Cohort (cGy) Total Dose (mCi) mCi/kg 100 (n=3) 5 (2–14) .06(.05–.12) 300 (n=7) 22(14–57) .25(.18–.63) 500 (n=6) 31(16–48) .40(.14–.63) 700 (n=6) 37(26–55) .38(.27–.73) 900 (n=3) 28(27–37) .32(.27–.44) 1100 (n=3) 48(29–65) .57(.50–.75) The liver was the critical organ in nearly all cases. Patient-specific doses calculated to deliver a cohort-prescribed absorbed radiation dose to the critical organ were highly variable suggesting that dosing based on weight and not dosimetry is likely to result in a wide range of absorbed dose to critical organs. In the context of this study, 90YZ has been administered to eight patients at doses of .5 mCi/kg or greater. We conclude that with careful dosimetry, 90YZ doses higher than the conventional .4 mCi/kg may be safely combined with BEAM and autotransplant. Accrual continues at the 1300 cGy dose level.

Description: B-Chronic lymphocytic leukemia (B-CLL) patients whose malignant cells harbour unmutated immunoglobulin heavy chain variable region (IgVH) genes or express the zeta-associated protein tyrosine kinase ZAP-70 show a worse prognosis than do patients with mutated IgVH genes or ZAP-70−ve expression. The inability of malignant cells to activate the pro-apoptotic p53 pathway in response to ionizing radiation (IR) also correlates with a poor prognosis. We studied ZAP-70 expression and IgVH mutation status in 161 patients with B-CLL in order to determine the degree of concordance between these two prognostic criteria (M104/F57, wbc 2.44–576x109/l lymphocytes 0.56–287x109/l). We also studied the functional status of the p53 pathway and the apoptotic response to ionizing radiation in cells from a subset of patients from both prognostic categories. A human ZAP-70 antibody (clone 2F3-2) was conjugated to the Alexa Fluor 488 dye using a zenon mouse IgG labelling kit and used for a FACS based assay. FACS results were expressed as a ratio of B-cell mean cell fluorescence to T-cell mean cell fluorescence with a cut off at 〉 0.75 identifying a ZAP-70+ve sub-group. IgVH mutational status was studied by sequence analysis of FR1/JH polymerase chain reaction products. The ability of 5Gy ionizing radiation to augment levels of p53 and its transcriptional target p21CIP1 was quantified by western blot analysis. Cleavage of the caspase 3 target poly(ADP ribose) polymerase (PARP) was used as a measure of apoptosis induction. ZAP-70+ve expression was observed in 25% (41/161) of the samples with a median ratio of 0.85 (range 0.76–1.46) while the remaining 120 samples were ZAP-70−ve, with a median ratio of 0.56 (range 0.19–0.73). IgVH mutation status was analysed in 92 of these patients. Assignment of prognostic category by both criteria was concordant in 72/92 (78.2%) of the cases of which 54/92 (58.6%) were ZAP−ve/IgVH mutated (good prognosis) and 18/92 (19.5%) were ZAP+ve/IgVH unmutated (poor prognosis) patients. The remaining 21.7% were discordant, ie., either ZAP+ve/IgVH mutated (5.4%) or ZAP−ve/IgVH unmutated (16.3%). Isolates from 5/6 ZAP+ve/IgVH unmutated patients upregulated p53 in response to IR but nevertheless failed to initiate PARP cleavage, suggestive of a block in the apoptotic pathway distal to p53 induction. In 9 ZAP−ve/IgVH mutated isolates studied, 7 induced p53, p21 and PARP cleavage following IR. In conclusion, this large cohort of CLL patients demonstrated a good correlation between ZAP-70 expression and IgVH mutational status in identifying a poor prognosis sub-group. However, this prognostic category, as defined by both IgVH mutation status and ZAP-70 expression failed in some cases to predict the ability of B-CLL cells to induce an apoptotic response to DNA damage in vitro. Induction of the p53 pathway was not always sufficient to secure an apoptotic response, especially in the poor prognosis group. A combination of ZAP-70 and IgVH analysis with a functional assay for DNA damage-induced apoptosis will identify individuals in either prognostic category who are unlikely to respond to conventional cytotoxic drugs. Alternative therapeutic strategies independent of DNA damage-inducing agents may be of value in the treatment of these patients.