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Symptoms of anaemia due to clinically significant extravascular haemolysis can affect patients with paroxysmal nocturnal haemoglobinuria (PNH) treated with C5 inhibitors (ravulizumab or eculizumab). The aim of this study was to assess the efficacy and safety of danicopan (ALXN2040), an investigational, first-in-class, oral complement factor D inhibitor, as add-on therapy to ravulizumab or eculizumab in patients with PNH and clinically significant extravascular haemolysis.

In this article, we discuss the dielectric relaxation of water in the hemoglobin (Hb) aqueous solutions in different states [methemoglobin (MetHb), oxygenated hemoglobin (OxyHb), and deoxygenated hemoglobin (DeoxyHb)]. The interpretation of the results was performed by the 3-D trajectory approach, which considers dielectric parameters of water relaxation and protein concentration. It has been shown that the interaction of amino acid dipole residues, located on the surface of a protein macromolecule, with water dipoles determines the protein hydration. In addition, ions of the buffer, in which proteins are dissolved, are also hydrated. The transition from a dipole–ion interaction of water molecules to a dipole–dipole interaction of water with an increase in protein concentration is observed. We proposed a new approach to calculate Hb hydration shells. The theoretical model defines the change in the ratio between the content of free and bound water molecules (BWMs) as a function of MetHb concentration in ion-free and ion-containing aqueous solutions. It considers the number of positive and negative charges at the surface of the protein molecule, the number of BWMs in its hydration shells, and the partition of water molecules bound to MetHb and the inorganic ions. The theoretical evaluation of the ratio of free-to-bound water reveals that, in the absence of ions, MetHb binds about 1400 water molecules, which is in good agreement with experimental data. At MetHb concentrations close to physiological, Hb is dominating in binding water over the inorganic ions, and the amount of BWM remains at the level of ~20% of total cytosol water as the concentration of Hb increases above 15 g/dL. These results suggest that in the physiological concentration diapason (29–35 g/dL), molecules of MetHb are so close together that their hydration shells interact and shrink from four to one layer of water molecules. In contrast, Hb molecules aggregate to neutralize their surface charges mutually.

Alopecia areata is a common autoimmune disorder leading to hair loss. It usually affects individuals under the age of 40, and first appearance in older subjects is considered uncommon. Here, we report 2 cases of rapidly progressing alopecia areata, which appeared for the first time in adults. Patient 1 had alopecia universalis, which preceded the identification of hepatosplenic T-cell lymphoma, a rare form of lymphoma. Patient 2 suffered from the ophiasis type of alopecia areata, presenting for the first time following chemotherapy for non-Hodgkin B-cell lymphoma. These 2 cases highlight the need to screen for malignancies in patients who present with rapidly progressing alopecia areata for the first time after the age of 40.

Goldschmidt: Abbvie Inc: Consultancy, Research Funding.

Introduction:Primary central nervous lymphoma (PCNSL) is a rare type of non-Hodgkin lymphoma that may involve the brain, spinal cord, leptomeninges and vitreous. The median age at diagnosis is in the fifth decade and most patients (pts) present with significant neurologic deficit and a low performance status. The optimal treatment of PCNSL is controversial. High dose methotrexate (HDMTX) is a standard treatment of PCNSL and is more effective when given in combination with other CNS-penetrating medications, which, however, add to the toxicity of the treatment. We aimed to evaluate the effectiveness and the safety of the combination of rituximab, HDMTX, procarbazine and lomustine (R-MPL) in pts with PCNSL.

Wolach: Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Speaker; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Speaker. Rowe:BioSight: Consultancy.Venetoclax for relapsed/ refractory AML

Introduction:On November 2018 the FDA approved the treatment of venetoclax together with hypomethylating agent (HMA) or low dose cytarabine (LDAC) for newly diagnosed AML patients who cannot tolerate intensive chemotherapy. There is limited data regarding the efficacy and safety of venetoclax in the setting of relapsed/refractory patients who received intensive chemotherapy for their disease.

OBJECTIVE: To report a case of neutropenia and thrombocytopenia secondary to use of zuclopenthixol in a schizophrenic patient.CASE SUMMARY: A 66-year-old white man with chronic schizophrenia was referred to the hospital due to neutropenia and thrombocytopenia that developed shortly after initiation of zuclopenthixol therapy. Prior to zuclopenthixol administration, his white blood cell and platelet counts were 8.5 times 109cells/L3and 305 cells x 109/L, respectively. Progressive reduction in leukocyte and platelet counts occurred, reaching a nadir of 2.9 times 109cells/L3(granulocytes 18.9%) and 109 cells x 109/L, respectively. Zuclopenthixol was discontinued on admission, resulting in complete recovery within the next five days.DISCUSSION: Neutropenia and thrombocytopenia are well-known complications of antipsychotic drug therapy. Zuclopenthixol, a well-established antipsychotic agent, has relatively few adverse effects. The rapid decrease of white blood cell and platelet counts following the initiation of zuclopenthixol, as well as the rapid recovery, implicate zuclopenthixol as the predominant cause for neutropenia and thrombocytopenia in this patient.CONCLUSIONS: Although neutropenia and thrombocytopenia are rare complications of zuclopenthixol therapy, monitoring blood counts in patients receiving this agent seems to be justified.

Salogub: Affimed: Research Funding. Mayer:Novartis: Research Funding; Roche: Research Funding; Eisai: Research Funding; Johnson & Johnson: Research Funding; Affimed: Research Funding. Folber:Affimed: Research Funding. Grosicki:Affimed: Research Funding. Skotnicki:Affimed: Research Funding. Prochwicz:Affimed: Research Funding. Myasnikov:Affimed: Research Funding. Gural:Affimed: Research Funding. Schoenborn-Kellenberger:Affimed: Consultancy. Brindley:Affimed: Consultancy. Knackmuss:Affimed: Employment. Schwarz:Affimed: Employment. Schmich:Affimed: Employment. Choe-Juliak:Affimed: Employment. Strassz:Affimed: Employment. Alland:Affimed: Employment. Doubek:AbbVie: Consultancy, Research Funding; Gilead: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria; Roche: Consultancy, Honoraria; Affimed: Research Funding; Novartis: Consultancy.

Background

Introduction:Progression-free (PFS) and overall survival (OS) rates for Primary mediastinal large B-cell lymphoma (PMBL) have risen to 84% and 92%, respectively, with the addition of rituximab to standard CHOP. Despite general acceptance of RCHOP as standard of care for Diffuse Large B-Cell lymphoma (DLBCL), many centers recommended alternative regimens for PMBL such as RMACOPB, RVACOPB, RCHOP-RICE and DA-EPOCH-R. The latter was adopted with great worldwide enthusiasm, despite its lack of proven superiority in randomized controlled trials (RCT). The benefits of DA-EPOCH-R include the omission of consolidation radiotherapy (RT), an attractive option in PMBL patients (pts), given their demographic profile-mainly females in their 3rd decade. We aimed to evaluate the PFS, the OS, the number of hospitalization days for treatment, and complications and the need for consolidation RT in a single center in the Rituximab era; where since 2007 over 80% of our pts were treated with the RCHOP-RICE regimen consisting of 4 courses of RCHOP followed by 3 courses of RICE.

Lavie: Pfizer: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees.

Abstract 1340This icon denotes a clinically relevant abstract

Drug metabolism/disposition genes have substantial impact on drug pharmacology. Single nucleotide polymorphisms (SNPs) may affect gene activity. For CML, the human organic cation transporter (hOCT1) transports imatinib mesylate (IM) into cells. The multidrug resistance gene MDR1 effluxes IM. MDR1 SNPs are correlated with response to IM (Dulucq et al, 2008). We studied hOCT1 and MDR1 SNPs and correlated them with plasma levels and clinical response to IM. We studied an exon 2 hOCT1 SNP (480C>G, Phe160Leu) for which homozygosity (GG) is associated with IM failure (Kim, 2009), and an exon 7 SNP (1222A>G, Met408Val) which correlated with major molecular remission (MMR) (Takahashi, 2010). Both are prevalent in Caucasians (exon 2 SNP: 22%, exon 7 SNP: 60%) (Kerb, 2006).We studied 84 chronic phase CML patients (pts) aged 17–89 years, followed from 16 months (mos) to 20 yrs from diagnosis (mean 94 mos). 19% were diagnosed during or prior to 2000. Time to initiating IM ranged from <1 to 108 mos (median <1 mo). IM therapy duration ranged 12–142 mos (mean 75). 61 pts took 400 mg/day, 14 pts took >400 mg (mean 621 mg), and 9 pts took <400 mg (mean 283 mg). Trough IM levels were performed on 81 pts by Novartis Pharmaceuticals. Treatment response was defined as per European Leukemia Net criteria. Until 2004, MMR was defined as PCR negativity using RT-PCR, and thereafter, using real time PCR (BCR/Abl<0.1% on an international scale).The overall mean IM trough level was 1,168 ng/ml (+/–477), and varied by dose. Pts on 400 mg had a mean level of 1145 (+/– 468), pts on >400 mg had 1392 (+/– 472), and pts on <400 mg had 909 (+/– 425). Table 1 shows genotype frequencies.hOCT1 exon 2 GG homozygotes had higher IM levels than CG/CC genotypes (1231 +/− 644 versus 1162 +/−463). hOCT1 GG patients were on slightly higher doses (mean 457 +/− 162 mg compared to 422 +/− 96 mg for CC/CG pts). hOCT1 exon 7 AA homozygotes had the same IM levels as did GG/AG pts (1178 +/− 477 for AA, 1177 +/− 476 for GG/AG). This is despite the fact that the AA homozygotes were on higher doses (500 +/− 134 mg for AA, 413 +/− 93 mg for GG/AG). Notably, 20% (2/10) AA patients failed IM compared to 11% (8/73) GG/AG pts.MDR1 2677 TT had lower than average IM levels (964 +/− 557) on a mean dose of 414 +/− 90 mg. Pts with genotypes 2677GG/GT had IM levels of 1195+/−473 on a slightly higher dose of 428 +/− 104 mg.MDR1 3435 TT homozygotes had similar IM levels to CC/CT pts (TT: 1127 +/− 434; CC/CT: 1188 +/− 434). TT patients were on slightly lower doses (404 +/− 77 compared to 434 +/− 111 mg).82/84 of the pts are still alive and 82% are still on IM. 15 pts discontinued IM (10 treatment failures, 5 due to intolerance). Only two pts developed accelerated phase or blast crisis.Time to MMoR is seen in Table 2, as related to genotype.20 pts lost molecular remission (LMR) at some point. Some of these had received IM as second line therapy (average 17.7 mos from diagnosis to starting IM). Half of the LMR events occurred when no second like TKIs were available so IM dose was increased. Half the pts with LMR switched to a newer TKI. Of treatment failures, there was a tendency to more A alleles in hOCT1 Exon 7 (AG or AA) and more T alleles in both MDR1 SNPs (GT/TT for 2677 and CT/TT for 3435).1. IM levels are associated with clinical response in CML. Treatment failures may be caused by suboptimal IM dosing. Alternate TKIs may overcome this problem. 2. Failures were more likely in pts for whom IM was not front line therapy. 3. Genotypes seem to correlate with IM levels and with treatment response for both SNPs. The hOCT1 AA genotype was associated with a higher dose requirement and more treatment failures. TT genotype for both MDR1 SNPs correlated with longer time to MMR (Table 2).Rund: Novartis Corporation: Honoraria, Research Funding.

Drug metabolism and disposition genes have a substantial impact on the pharmacology of many medications. For CML, the human organic cation transporter (hOCT1) actively transports imatinib mesylate (IM) into cells, thus is a crucial factor in the response to IM therapy, without a known effect on plasma levels. A number of single nucleotide polymorphisms (SNPs) are believed to affect the activity of hOCT1 and therefore may influence IM accumulation in cells. We therefore studied hOCT1 SNPs and correlated with IM plasma levels and clinical response to IM. We studied a SNP in exon 2 of hOCT1 (480C>G, Phe160Leu, rs683369), which, in the homozygous state (GG), has been associated with treatment failure on IM (DH Kim, Clin Cancer Res., 15:4750, 2009), and a SNP in exon 7 (1222A>G, Met408Val) which has been hypothesized as having increased activity (A Giannoudis, Haematologica 96(s2):87, 2011). Both of these are prevalent SNPs; the exon 2 SNP is found in 22% of Caucasians and the exon 7 SNP, in 60% of Caucasians (R Kerb, Cancer Letters 234:4, 2006).We studied 48 patients with CML in chronic phase (M:F 21:27 aged 17–89 years. Time since CML diagnosis was between 10 months and 18 years. IM therapy was between 10–128 months' duration. Most of the patients were on a dose of 400 mg/once daily, with the exception of 11 patients who were on doses >400 mg/once daily (mean 609.1 mg/day +/− 70.1 mg), and 9 patients, whose dose was <400 mg/once daily (mean 281 mg/day +/− 35.4 mg). Trough IM levels were performed by Novartis Pharmaceuticals on samples taken 24 hours after last dose, reported in ng/ml. Compliance was verified by a patient diary during the trough drug testing period, but not throughout the entire period of IM therapy. Genotyping for hOCT1 SNPs was performed on 36 patients. The exon 2 SNP was detected by PCR amplification and Dde I restriction enzyme analysis. The exon 7 was detected by PCR and restriction with BmgB1. Detailed molecular data was available on 36 patients, and partial molecular data on 8 others. Complete molecular remission (CMR) was defined as PCR negativity using RT-PCR until 2004, and thereafter, using a highly sensitive real time PCR assay.The mean IM trough level for all 48 patients was 1,153.0 (+/−493.1). There was much variation between IM levels in individual patients and levels also varied by dose. Patients who were on 400 mg/day had a mean IM level of 1064.1 (+/− 399.2). Patients on higher IM doses (>400 mg) had IM levels averaging 1397.2 (+/− 493.1). Patients on doses <400 mg/day had mean IM levels of 1057.3 (+/− 667.0), which was nearly identical to that of patients on 400 mg/day. Genotyping for the hOCT1 exon 2 SNP demonstrated that 66.7% of the 36 patients analyzed were homozygous for the wild type allele, 13.9% were heterozygous for the polymorphism and 19.4% were homozygous polymorphic. GG homozygotes had higher IM levels than CG/CC genotypes (1562.75 +/− 560 compared to 1310.6 +/− 470). This is because the patients were on doses greater than 400 mg/day (mean 483 mg/day compared to 405 mg/day). These patients took an average of 8.8 months to achieve a molecular CR and one transiently lost molecular CMR. Genotyping for the hOCT1 exon 7 SNP demonstrated that 47% of the 36 patients analyzed were heterozygous AG for the polymorphism and 53% of the patients were GG homozygotes. IM levels for the GG homozygotes were higher than for AG heterozygotes (1413 +/− 549.6 compared to 1254.0 +/− 393.9 ng/ml), despite the fact that they were on a slightly lower average dose (mean: 390.1 mg/day compared to 436.7 mg/day). For 13 of these GG homozygous patients, detailed molecular followup was available. Two of these patients never achieved CMR and 2 transiently lost CMR. The time to achieve complete molecular remission was generally long, 17.6 months (range 3–83 months). Importantly, 12 out of 13 of these patients with unsatisfactory clinical responses to IM were CC or CG at the exon 2 SNP, which are considered favorable genotypes for IM response (DH Kim, Clin Cancer Res., 15:4750, 2009).These data may suggest that the GG genotype for exon 7 in hOCT1 (rs628031), independent of the exon 2 (rs683369) genotype, is an adverse prognostic parameter despite adequate IM levels. These patients may be candidates for an alternate tyrosine kinase inhibitor. Further studies are necessary on larger groups of patients to confirm these results, which are important in view of the high frequency of the polymorphic rs628031 allele.Cohen: Novartis Pharmaceutical Company: Research Funding. Rund:Novartis Pharmaceutical Company: Honoraria, Research Funding.

Abstract 3487

No relevant conflicts of interest to declare.

Abstract 1588

Although the transcriptional product of the (8;21) chromosomal translocation, the AML1-ETO protein, has numerous well established effects on the behavior of human hematopoietic cells, the specific cellular pathways perturbed by it are only partly known. Recently AML1-ETO has been shown to bind E-proteins (members of the class I basic helix-loop-helix family) via its NHR1 (nervy homology region one) domain, thus causing a replacement of the co-activator p300/CBP complex by the co-repressor hystone deacytelase complexes and abrogating the E-protein induced transcriptional activation (Zhang et al, Science 2004). The removal of amino acids 93 to 189 from the NHR1 domain (the ?NHR1 mutant form) has completely reversed this inhibitory effect, and thus we have studied the biological effects of this deletion. To assess the difference in self-renewal capacity between cells transduced with the full length and the ?NHR1 forms of AML1-ETO, CD34+ cells were plated into methylcellulose culture medium. On days 14 and 28 myeloid and erythroid colonies were scored, and the cells harvested and re-plated. The scoring on day 14 showed no significant difference in either the overall number or the type of colonies, with both the full-length AML1-ETO and the ?NHR1 mutant showing a decreased erythroid colony formation, thus indicating a preserved negative effect on hematopoietic cell differentiation. However, on days 28 and 42 cells expressing the full-length AML1-ETO scored a significantly higher number of colonies than cells expressing the ?NHR1 mutant, whereas cells transduced with the empty MIGR1 vector were unable to form any colonies after second re-plating. To assess the difference in growth potential, we have plated cells containing each of the constructs in IMD medium supplemented with 20% BIT and cytokines (SCF, FLT-3L, IL-6, TPO). Following one week in culture the cells were harvested, counted and then serially re-plated for five weeks. We observed a significantly higher number of cells expressing the full-length AML1-ETO as compared to the ?NHR1 mutant at each weekly time point. Cells growing in the liquid culture were also weekly re-plated into methylcellulose culture medium. In this assay the loss of NHR1 domain significantly diminished the ability of AML1-ETO to maintain colony-forming progenitors in culture. To assess the difference in transcriptional activity of the different constructs, we have measured the expression of p21, and several other potential AML1-ETO target genes, by Real Time PCR. The expression of p21 was increased 14-fold by the full-length AML1-ETO, but only 2-fold by the ?NHR1 mutant. We conclude that partial deletion of the NHR1 lowers the self-renewal capacity of transduced hematopoietic cells, decreases the proliferative advantage conferred by AML1-ETO, undermines its ability to maintain colony-forming units, and affects the transcriptional activation of AML1-ETO target genes. As the loss of NHR1 abolishes the E-protein induced transcriptional activation without affecting the binding of AML1-ETO to DNA, our findings identify an additional mechanism of action for AML1-ETO, independent of its dominant-negative effect on AML1 target genes.

Although the transcriptional product of the (8;21) chromosomal translocation, the AML1-ETO protein, has numerous well established effects on the behavior of human hematopoietic cells, the specific cellular pathways perturbed by it are only partly known. Recently AML1-ETO has been shown to bind E-proteins (members of the class I basic helix-loop-helix family) via its NHR1 (nervy homology region one) domain, thus causing a replacement of the co-activator p300/CBP complex by the co-repressor hystone deacytelase complexes and abrogating the E-protein induced transcriptional activation (Zhang et al, Science 2004). The removal of amino acids 93 to 189 from the NHR1 domain (the ΔNHR1 mutant form) has completely reversed this inhibitory effect, and thus we have studied the biological effects of this deletion.

ABVD is widely considered as the gold standard treatment for advanced Hodgkin’s lymphoma (HL), although about 40% of patients relapse or do not respond to initial treatment. Recently, the HD9 trial of the German Hodgkin’s Lymphoma Study Group has shown that escalated (esc) BEACOPP regimen can achieve better disease control than ABVD, but has a high incidence of acute and long-term toxicities including high occurrence of AML/MDS. In an attempt to decrease toxicity while preserving the potential benefit of upfront intensive therapy, we conducted a pilot study, which tested the feasibility, toxicity and efficacy of combined escBEACOPP-ABVD regimen as therapy for newly diagnosed patients with high risk (IPS≥3) stage III–IV HL. Patients initially received 2 cycles of escBEACOPP followed by reevaluation with CT and gallium or PET/CT-FDG scans. When complete response (CR) or partial response (PR) was achieved, patients continued to receive 4 cycles of ABVD, while those failing to achieve a response were withdrawn from the study. Since August 2001, 21 patients entered the study and at the time of present analysis four of them are still receiving therapy. Three (18%) of 17 patients, who completed chemotherapy, received consolidative radiotherapy. Median age at diagnosis was 26 years (range 18–56) and 11 (57%) were males. Stage IV and B symptoms were evidenced in 19 (90%) and 17 (81%), respectively, and 7 (33%) had bulky mediastinal mass. Histology included nodular sclerosis in 18 patients (86%), mixed cellularity in 2 (10%) and lymphocyte predominance in 1 (5%). Following the first 2 cycles of escBEACOPP the overall response rate (CR+PR) was 100%. At the end of all therapy 15 patients (88%) were in CR, one patient in PR and only a single patient had progressive disease. With a median follow-up of 20 months no patient relapsed or died. Toxicity included WHO grade III–IV granulocytopenia in 15 patients (88%) and grade III–IV infection in one patient. Hospitalization for intravenous antibiotics was necessary in 10 patients (59%). Almost all of these events occurred during the first two cycles of escBEACOPP, while acute toxicity during the ABVD phase was mild. In conclusion, the combined escBEACOPP-ABVD regimen is well tolerated and is associated with a high CR rate when used in advanced HL patients with high IPS scores. Further follow-up is obviously required in order to determine long-term survival and late complications of this regimen.

Platelet anti-aggregants play a major role in the treatment of cardiovascular disorders. Recently it has been suggested that a significant proportion of patients fail to respond to these agents according to different testing methods. We have developed a new method for evaluating the response to platelet anti-aggregants, the modified Cone and Plate(let) Analyzer (CPA) test. The method is based on the observation that pre-activation of platelets by an agonist will result in reduced platelet deposition on a polystyrene surface under arterial flow conditions (the phenomenon of platelet adhesion refractoriness). In vitro studies demonstrated that the basic platelet adhesion (surface coverage, SC 12.3±6.8%) was significantly reduced in response to pre-incubation of the sample (for 1 min) with arachidonic acid (AA, SC 2.1±1.5%), adenine diphosphate (ADP, SC 1.3±0.6%) and epinephrine (EPI, 2.9±0.9%). This effect was selectively inhibited by aspirin (for the response to AA, SC 8.1±3.8%), and by 2-Methylthio-AMP triethylammonium (2-MeSAMP), a selective inhibitor of P2Y12 (for the response to ADP, SC 4.8±2.0%), p<0.001. The potential application of this method for monitoring the effect of aspirin therapy (daily doses of 100, 300 and 500 mg), among 20 healthy volunteers was investigated in comparison to turbidimetric aggregometer. All volunteers responded to aspirin in the three doses according to the aggregometer test (reduced max. aggregation from 88±8% to 12.4±7%), with no evidence for a dose response manner in this dose range. Basic platelet adhesion as tested by the CPA (SC 9.8±2.2%), was unaffected by aspirin in the three tested doses. However, the response to pre-incubation with AA was significantly inhibited by aspirin in all patients (SC of 0.6±0.3% before aspirin therapy and 3.5±1.3%, 4.4±1.7%, and 4.1±2.0% following therapy with 100, 200 and 500 mg aspirin per day, respectively, p<0.001 for all doses), again with no dose response effect. This effect was specific for AA, since no difference in SC was observed when ADP was used as an agonist. A significant correlation between the effect of aspirin as tested by the modified CPA and by the turbidimetric aggregometer using AA as an agonist (R2=0.55) was observed. In conclusion, the CPA was found useful point-of-care method for testing the response of platelets to aggregating agonists as well as for monitoring the effect of anti-platelet drugs.

ABVD is widely considered as the gold standard treatment for advanced Hodgkin's lymphoma (HL), although about 40% of patients relapse or do not respond to initial treatment. Recently, the HD9 trial of the German Hodgkin's Lymphoma Study Group has shown that escalated (esc) BEACOPP regimen can achieve better disease control than ABVD, but has a high incidence of acute and long-term toxicities including high occurrence of AML/MDS. In an attempt to decrease toxicity while preserving the potential benefit of upfront intensive therapy, we conducted a pilot study, which tested the feasibility, toxicity and efficacy of combined escBEACOPP-ABVD regimen as therapy for newly diagnosed patients with high risk (IPS≥3) stage III–IV HL. Patients initially received 2 cycles of escBEACOPP followed by reevaluation with CT and gallium or PET/CT-FDG scans. When complete response (CR) or partial response (PR) was achieved, patients continued to receive 4 cycles of ABVD, while those failing to achieve a response were withdrawn from the study. Since August 2001, 21 patients entered the study and at the time of present analysis four of them are still receiving therapy. Three (18%) of 17 patients, who completed chemotherapy, received consolidative radiotherapy. Median age at diagnosis was 26 years (range 18–56) and 11 (57%) were males. Stage IV and B symptoms were evidenced in 19 (90%) and 17 (81%), respectively, and 7 (33%) had bulky mediastinal mass. Histology included nodular sclerosis in 18 patients (86%), mixed cellularity in 2 (10%) and lymphocyte predominance in 1 (5%). Following the first 2 cycles of escBEACOPP the overall response rate (CR+PR) was 100%. At the end of all therapy 15 patients (88%) were in CR, one patient in PR and only a single patient had progressive disease. With a median follow-up of 20 months no patient relapsed or died. Toxicity included WHO grade III–IV granulocytopenia in 15 patients (88%) and grade III–IV infection in one patient. Hospitalization for intravenous antibiotics was necessary in 10 patients (59%). Almost all of these events occurred during the first two cycles of escBEACOPP, while acute toxicity during the ABVD phase was mild. In conclusion, the combined escBEACOPP-ABVD regimen is well tolerated and is associated with a high CR rate when used in advanced HL patients with high IPS scores. Further follow-up is obviously required in order to determine long-term survival and late complications of this regimen.

Platelet anti-aggregants play a major role in the treatment of cardiovascular disorders. Recently it has been suggested that a significant proportion of patients fail to respond to these agents according to different testing methods. We have developed a new method for evaluating the response to platelet anti-aggregants, the modified Cone and Plate(let) Analyzer (CPA) test. The method is based on the observation that pre-activation of platelets by an agonist will result in reduced platelet deposition on a polystyrene surface under arterial flow conditions (the phenomenon of platelet adhesion refractoriness). In vitro studies demonstrated that the basic platelet adhesion (surface coverage, SC 12.3±6.8%) was significantly reduced in response to pre-incubation of the sample (for 1 min) with arachidonic acid (AA, SC 2.1±1.5%), adenine diphosphate (ADP, SC 1.3±0.6%) and epinephrine (EPI, 2.9±0.9%). This effect was selectively inhibited by aspirin (for the response to AA, SC 8.1±3.8%), and by 2-Methylthio-AMP triethylammonium (2-MeSAMP), a selective inhibitor of P2Y12 (for the response to ADP, SC 4.8±2.0%), p<0.001. The potential application of this method for monitoring the effect of aspirin therapy (daily doses of 100, 300 and 500 mg), among 20 healthy volunteers was investigated in comparison to turbidimetric aggregometer. All volunteers responded to aspirin in the three doses according to the aggregometer test (reduced max. aggregation from 88±8% to 12.4±7%), with no evidence for a dose response manner in this dose range. Basic platelet adhesion as tested by the CPA (SC 9.8±2.2%), was unaffected by aspirin in the three tested doses. However, the response to pre-incubation with AA was significantly inhibited by aspirin in all patients (SC of 0.6±0.3% before aspirin therapy and 3.5±1.3%, 4.4±1.7%, and 4.1±2.0% following therapy with 100, 200 and 500 mg aspirin per day, respectively, p<0.001 for all doses), again with no dose response effect. This effect was specific for AA, since no difference in SC was observed when ADP was used as an agonist. A significant correlation between the effect of aspirin as tested by the modified CPA and by the turbidimetric aggregometer using AA as an agonist (R2=0.55) was observed. In conclusion, the CPA was found useful point-of-care method for testing the response of platelets to aggregating agonists as well as for monitoring the effect of anti-platelet drugs.