bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact of Chimerism Analysis and Kinetics on Haploidentical Stem Cell Transplantation.
Authors: Tang Xiaowen , Wu De Pei , Sun Aining , Yang Mingzhen , Wang Ying
Published: October 2019

Haploidentical transplantation (H-SCT) has been an alternative allogenetic transplant option for the high risk lukemia patients who do not have an HLA-identical related donor. The high dose of stem cell infusion and intensive GVHD prophylaxis treatment were applied in H-SCT to overcome the haploidentical major histo compatibility barrier and to achieve engraftment. We speculated that the dynamics of donor chimerism are different from conventional myeloablative allogeneic bone marrow transplantation(allo-BMT). Methods Quantitative analysis of chimerism were performed by multiplex PCR amplification of STR markers (STR-PCR) in unfractionated nucleated cells from bone marrow or whole blood of 30 high risk leukemia patients who received H-SCT(14 females,16 males; median age 25 years). Eighteen of 30 patients received G-CSF primed bone marrow transplantation and another 12 patients received G-CSF primed bone marrow and peripheral blood stem cell transplantation. The conditioning regimen consisted of modified BU/CY or modified total body irradiation(TBI) plus CY. One patient received cyclosporine (CSA), short-term methotrexate(MTX) and mycophenolate mofetil (MMF) as the regimen of prophylaxis of GVHD. Twenty-four patients received CSA, MTX and MMF plus ATG/ALG for GVHD prophylaxis. The combination of CSA, MTX, MMF, ATG/ALG and CD25 monoclonal antibody for preventing GVHD in 5 patients.

Results: All of the patients except one showed initial donor engraftment and just one patient presented primary graft failure. The early chimerism and its kinetics after transplantation showed that the donor chimerism became dominant (DC 84.1% >60%) by day +3, which preceded the detection of hematologic engraftment by a median of 9 days. On day +7, donor cells of majority patients had achieved stable engraftment. The median chimerism was 92.4%;The probability of achieving full donor chimerism (FDC) in PB was 56.3% on day +14, 61.5% on day +28;Comparing with the group of standard allo-BMT, median quantity of CD34+ cells and CD3+ T lymphocyte cells of H-SCT group were 2 and 3 times higher. Furthermore GVHD prophylaxis was more intensive with ATG/ALG and MMF in H-SCT group. The median percentage of donor chimerism of H-SCT was significantly higher than that of BMT on the day of +7d, +14d and +1month. Meanwhile there were difference in the rates of FDC among H-SCT and BMT(61.5% VS 20%, p<0.01)on day of +28.The median overall follow-up was 24 months(range, 1.5–34.5), and 12 (40%) patients were alive.Two of them maintained stable mix chimerism (DC>90%), and another 10 patients achieved stable FDC(DC ≥95%).

Conclusion: In the early stage after transplantation, the engraftment of donor cells of H-SCT was more rapidly and completely than that of conventional BMT owing to the high quantity of donor CD34+ cells, T lymphocyte cells and the more intensive immunosuppressive therapy.
Individual Treatment with Stem Cell Rescue in Patients with Germ-Cell Tumors. Results of One Centrum.
Authors: Nepomucka Jana , Abrahamova Jitka , Foldyna Martin , Donatova Zuzana , Kordikova Drahomira , Pagacova Ludmila , Kalanin Jan , Greplova Marcela , Bartova Markyta
Published: October 2019

Background: Treatment with high dose chemotherapy and autologous stem cell rescue in pacients with poor risk germ cell tumors is still controversial. Results of multicentric randomized EBMT study IT 94 presented at ASCO 2002 show benefit in 1-year EFS in high dose arm (52% versus 48%), 3-year EFS was the same in both arms (53%) in salvage treatment. Individual treatment with stem cell rescue as upfront treatment offers a survival benefit.

Methods:Autologous stem cell rescue was provided in our center, from September 1997 to May 2006 to 52 patients. High dose chemotherapy was indicated to 32 patients in salvage setting after 2nd line of treatment (VeIP) and to 20 patients as upfront treatment after 1st line treatment (BEP). Median age was 29 years and tumor markers were elevated: HCG in 9 pts, AFP in 13 pts.

Stem cell mobilization was performed after the 3rd cycle of VeIP or BEP in combination with G-CSF. The amount of CD34+ cell/kg b.w. was between 2,0 – 13.4×106. High - dose conditioning regimen CARBOPEC (carboplatin 1600 – 2 200 mg/m2, etoposide 1 800mg/m2, cyclophosphamide 6 400 mg/m2) was used. The treatment was well tolerated without transplant - related mortality.

Results: WHO criteria non - hematological toxicity was predominantly grade 2 to 3. Engraftment was rapid, recovery of hematopoiesis in neutrofils over 1.0×109/l and platelets over 50×109/l was reached an average on days +10 and +13 respectively. Additional post-transplant treatment for persistence, progression or relaps had 20 patients (8pts had 2nd line treatment VEIP, 12pts had 3nd line treatment with paclitaxel+gemcitabine and 5 pts had retroperitoneal lymfadenectomy).

The follow - up period ranges from 3 to 99 months, at present 38 (73 %) patients are alive, 14 (27 %) pts died. Median TTP of all pts is 10 months, median OS of all pts is 39 months. Median DFS of surviving pts is 38 months.

Conclusion: high-dose chemotherapy with autologous stem cell rescue in patients with poor risk germ cell tumors is feasible and beneficial method of the individual treatment. High-dose chemotherapy as upfront treatment for poor prognosis germ cell tumors and as salvage treatment in good risk pts seems to be good possibility of the individual treatment.
Fully Retargeted Oncolytic Measles Virus for Multiple Myeloma Therapy.
Authors: Hummel Horst D. , Kuntz Gaby , Nakamura Takafumi , Greiner Axel , Russell Stephen J. , Einsele Hermann , Topp Max S.
Published: October 2019

Multiple Myeloma (MM) is a disseminated plasma cell malignancy with approximately 14,600 new cases diagnosed in the USA annually. Despite recent progress in current therapeutical options the median survival is 3 to 5 years and cure is extremely rare. Therefore the evaluation of new treatment modalities for MM is highly warranted. An attractive approach to treat Myeloma with a minimum of undesired side effects is the use of a tumour antigen specific for MM cells. Wue-1, a monoclonal antibody binds very selectively normal and malignant plasma cells (50 of 51 MM samples, 14 of 15 immunocytoma and 13 of 13 MALT type lymphomas with plasma cell differentiation were Wue-1 positive, normal tissue including hematopoietic cells were negative) and offers the possibility to define MM cells as targets. The tool for selective killing of MM cells recognized by Wue-1 monoclonal antibody is in this study the measles virus vaccine strain Edmonston B in an ablated variant (MV-Wue) which no longer binds the usual measles receptors CD46 and CD150 (SLAM) expressed on almost every human cell type displaying a single-chain antibody (scFv) derived from the monoclonal Wue-1-antibody which has been tethered to the C-terminus of the H protein to restrict and retarget its interaction to malignant plasma cells especially MM cells. In addition, MV-Wue encodes EGFP facilitating the read out of infected cells. To determine if the fully retargeted MV-Wue would be able to infect MM cell lines and primary MM cells selectively an array of infection assays were performed using the MM cell lines U266 as well as primary CD138 positive MM cells expressing the Wue-1 antigen as expected targets and CD138 negative cells and normal B cells as controls negative for Wue-1. In these experiments selective infections of the MM cell line and primary MM cells were observed whereas the control cells were not infected with MV-Wue. In all cell types GFP expression indicating replicative infection correlated with the expression of the Wue-1 antigen determined by FACS. Infection experiments performed in the presence of monoclonal Wue-1 antibody showed a decreased GFP expression of about 78% in CD138 positive MM cells demonstrating specificity of the infection by MV-Wue. These results indicate that the engineered virus can be a safe and potential curative oncolytic agent to face the main problem in Multiple Myeloma which is responsible for frequent relapses, the minimal residual disease (MRD).
Intravenous Injection of Manganese Superoxide Dismutase Plasmid/Liposome Complexes (MnSOD-PL) Protects the Bone Marrow from Irradiation Damage.
Authors: Epperly Michael W. , Franicola Darcy , Smith Tracey , Greenberger Joel S.
Published: October 2019

Radioprotection of tissues including lung, esophagus, oral cavity and bladder has been demonstrated by local administration of MnSOD-PL. One potential use of the MnSOD-PL is protection of bone marrow as well as other tissues against total body irradiation (TBI) perhaps applicable for first responders who enter areas of high irradiation. To determine if systemically delivered MnSOD-PL protects bone marrow from TBI, groups of C57BL/6NHsd mice were injected intravenously with MnSOD-PL (100 μg of plasmid DNA in 100 ul volume) and irradiated 24 hrs later to 9, 9.5 or 9.75 Gy whole body irradiation. The mice were followed for bone marrow failure. Mice injected with MnSOD-PL showed improved survival at all irradiation doses. After 9 Gy TBI, 93% of the mice injected with MnSOD-PL survived past day 30 compared to 80% of control irradiated mice. At 9.5 Gy, 87% of the MnSOD-PL treated survived past day 30 compared to 53% of control irradiated mice (p = 0.0402). Following 9.75 Gy, the control irradiated mice had a median survival of 16 days with 12.5% survival at 30 days compared to 75% of the MnSOD-PL treated mice (p = 0.0016). To determine the efficiency of marrow transfection mice were injected with a plasmid containing the HA (hemagglutinin) epitoped tagged MnSOD transgene. The marrow was harvested 24 hr later, cytospun, and immunohistochemically stained for HA. The bone marrow demonstrated that 22.1 ± 3.7% of the bone marrow cells expressed HA-MnSOD protein. The results demonstrate that systemically delivered MnSOD-PL protects against TBI induced murine bone marrow damage and improves survival.
VELCADE/Dexamethasone (Vel/Dex) Versus VAD as Induction Treatment Prior to Autologous Stem Cell Transplantation (ASCT) in Newly Diagnosed Multiple Myeloma (MM): An Interim Analysis of the IFM 2005-01 Randomized Multicenter Phase III Trial.
Authors: Harousseau Jean-Luc , Marit Gerald , Caillot Denis , Casassus Philippe , Facon Thierry , Mohty Mohamad , Maloisel Frederic , Maisonneuve Herve , Chaleteix Carine , Benboubker Lofti , Esseltine Dixie-Lee , Attal Michel
Published: October 2019

ASCT is considered the standard of care for patients up to 65 years of age with MM. The optimal induction treatment prior to ASCT is still unknown. VAD is a commonly used regimen but is associated with a number of toxicities, administrative inconvenience, and a complete remission (CR) rate of typically less than 10%. Recently the combination thalidomide/Dex was reported to induce a higher overall response rate than Dex alone in newly diagnosed patients, but with significantly more toxicities, especially deep vein thrombosis, and a 4% CR rate (Rajkumar et al, J Clin Oncol 2006;24:431–6). In a phase II trial we have evaluated the combination of Vel 1.3mg/m2 D1, 4, 8, 11 with Dex 40mg/D D1-4, D9-12, for 4 consecutive 21-D cycles (Dex only on D1-4 during cycles 3–4). This combination was well tolerated and yielded a 21% CR/nCR rate with successful stem cell harvest and engraftment (Harousseau et al, Haematologica 2006). In 2005, the IFM initiated a multicenter randomized phase III trial comparing Vel/Dex with VAD as induction treatment of patients with newly diagnosed MM up to the age of 65. The primary objective of the study is the CR (negative immunofixation) plus nCR (CR but positive immunofixation) rate after 4 cycles. A secondary objective is to evaluate the role of consolidation with 2 courses of DCEP (Dex, cyclophosphamide, etoposide, cisplatin). Patients were randomized at diagnosis among 4 arms (A1: 4 cycles of VAD; A2: 4 cycles of VAD + 2 cycles of DCEP; B1: 4 cycles of Vel/Dex; B2: 4 cycles of Vel/Dex + 2 cycles of DCEP); stratification was by cytogenetics and β2 microglobulin. Stem cell collection was performed between cycle 3 and cycle 4 after priming with G-CSF (10μg/kg/D × 6 D). At the end of induction (± DCEP consolidation) treatment, patients received melphalan 200mg/m2 plus ASCT. Enrollment of 480 patients is planned, to detect a 10% higher CR/nCR rate with Vel/Dex (20% vs 10%) with 80% power (two-sided test, significance level 0.05). An interim analysis is planned when 222 patients have been evaluated for response at the end of induction. As of July 2006, 304 patients have been enrolled. Randomization of the 222 patients to be considered for interim analysis was completed on May 19th. Baseline characteristics include: median age 55y; 140M, 82F; β2 microglobulin >3mg/L in 54%; del13 by FISH in 45%. The occurrence of serious adverse events (SAEs) was similar between the VAD and Vel/Dex arms (31% with VAD, 33% with Vel/Dex). The most frequent SAEs were pneumopathy (5.3% VAD, 4.0% Vel/Dex), thrombosis (2.7% VAD, 3.3% Vel/Dex), and peripheral neuropathy (1.3% VAD, 3.3% Vel/Dex). We intend to present results from the interim analysis.
Hyperchidam Verona897 Regimen Is Effective in Treatment of Advanced Peripheral T Cell Lymphoma (PTCL): The GITIL Experience in 33 Cases.
Authors: Todeschini Giuseppe , Corradini Paolo , Cortelazzo Sergio , Rambaldi Alessandro , Gallamini Andrea , Benedetti Fabio , Ferreri Andres , Olivieri Attilio , Tecchio Cristina , Bonifacio Massimiliano , Frungillo Niccolo , Castellino Claudia , Rossi Andrea , Toldo Chandrakala , Crocchiolo Roberto , Attolico Immacolata , Pizzolo Giovanni
Published: October 2019

Background. Peripheral T cell Lymphoma (PTCL) represent a major therapeutic challenge. In the previous Verona experience (Todeschini G et al. Cancer2005;104:555–560), the novel intensive regimen HyperCHiDAM Verona897 (high-dose IV MTX 2 g/sqm 400 sqm bolus, 1600 sqm CI day 1 with IV leucovorin rescue; hyperfractionated IV CTX 300 mg/sqm Q12h, dd 2–4; high-dose IV AraC 2g/sqm Q12h, dd 2-4; plus G-CSF) achieved salutary results in refractory/relapsed aggressive lymphomas, in particular in PTCL. Supportive measures included hydratation 3000 ml/sqm/day, antimicrobial prophylaxis comprising oral ciprofloxacin, itraconazole, trimethoprim/sulfamethoxazole. CMV antigenemia (pp65) was monitored 2 times a week. Patients. Following these results, 7 centers belonging to the Italian co-operative group GITIL (Gruppo Italiano Terapie Innovative Linfomi) treated with 2 cycles of HyperCHiDAM Verona897, 33 patients affected by PTCL (17 upfront, 16 refractory/relapsed) followed in the majority of cases by stem cell transplant. Patients: M/F 21/12, median age 49 (19–63) years; histology: PTCL-NOS 15, ALCL ALK-negative 9, EALTC 4, AIL 3, T-NK nasal 1, Angiocentric 1; stage IV 19/33 (57.5%), bone-marrow positive 10/33 (30.3%), extranodal involvement 24/33 (72.7%), high LDH 18/33 (54.5%). Seven patients needing urgent treatment were treated before HyperCHiDAM with 1 cycle of Campath-CHOP (4 patients) or CHOP + L-ASE (3 patients).

Results. Upfront patients: after 2 cycles of HyperCHiDAM, CR were 82.3% (14/17), early toxic deaths 0 (1 late toxic death occurred after SCT, due to CMV pneumonia), relapses 3/14. With a median follow-up of 21 months (3–90+), 11/17 (64.7%) patients are disease-free, 10 in first CR, 1 after rescue with stem cell transplant. Three of the CCR patients received HyperCHiDAM alone. Refractory/relapsed patients: CR were 5/16 (31.2%), CCR 4/16 (25%), with a median follow-up of 24 months (5–64+). The progression-free survival was significantly superior in upfront patients (p=0.036). Overall, toxic deaths were 3/33 (9%), 1/17 (5.8%) in upfront and 2/16 (12.5%) in refractory/relapsed patients. One patient had major cerebellar toxicity.

Conclusions. The intensive regimen HyperCHiDAM Verona897 is effective in inducing CR in aggressive PTCL, in particular as upfront therapy. The intensiveness of this treatment requires a careful supportive therapy. Following these results, HyperCHiDAM has been included in a national trial for treatment of PTCL, after 2 cycles of Campath-CHOP and before stem cell transplant (allogeneic or autologous depending on the availability of an HLA-matched sibling). The co-operative study is recruiting.
Phase 1 Clinical Trial of a Novel Proteasome Inhibitor (NPI-0052) in Patients with Lymphomas and Solid Tumors.
Authors: Kurzrock Razelle , Hamlin Paul , Younes Anas , Hong David , Gordon Michael , Spear Matthew A. , Palladino Michael A. , Lloyd G. Kenneth , Longenecker Angie M. , Neuteboom Saskia T.C. , Cropp Gillian F. , Hannah Alison , Aghajanian Carol
Published: October 2019

NPI-0052 is a novel proteasome inhibitor that produces prolonged inhibition of all three catalytic activities (C-L, T-L, CT-L) of the 20S proteasome. Preclinical data suggest NPI-0052 may demonstrate an improved therapeutic ratio, with activity in multiple myeloma, lymphoma, leukemia and solid tumor models. Phase 1 dose escalation studies are ongoing in patients with myeloma, lymphomas and solid tumors. Data from the Phase 1 study in patients with lymphomas or solid tumors are presented herein to examine the endpoints of pharmacodynamics (proteasome inhibition), pharmacokinetics and safety at doses below the MTD. Patients in this study were treated with NPI-0052 administered as a weekly IV bolus, for 3 weeks in 4-week cycles. The dose of NPI-0052 was escalated in cohorts of 3 patients dependent on observed adverse events utilizing a 3+3 design. In addition to weekly safety monitoring, proteasome inhibition and pharmacokinetics were assayed after the 1st and 3rd dose and with intrapatient dose escalations. Sixteen patients have been treated at doses ranging from 0.0125 mg/m2 to 0.112 mg/m2 for up to 7 cycles without reaching an MTD. Although, one SAE of MRSA sepsis and renal failure was reported at a dose of 0.1 mg/m2, drug related adverse events have not been reported at 0.112 mg/m2. Proteasome inhibition in whole blood has been assayed from 0.0125 mg/m2 through 0.075 mg/m2, with results ranging from no inhibition to 63% inhibition of CT-L activity and indication of a correlation of inhibition with dose, inclusive of intrapatient dose increases. Preliminary pharmacokinetic data demonstrate a rapid elimination half-life (estimated to be 3–4 minutes) with clearance between 8–21 mL/min and Vz of 44–99L. No responses have been confirmed, however, 4 patients have had stable disease for 4 or more months, including patients with hepatocellular carcinoma (6 months), adenoid cystic carcinoma (4 months and 4 months), and cervical carcinoma [7 months with investigator reported complete resolution of target (lymph node) lesion(s) to palpation]. These data suggest that NPI-0052 is affecting parameters relevant to pharmacodynamics, pharmacokinetics and potentially clinical benefit at doses below the MTD. Dose escalation continues to define a recommended phase 2 dose and further investigate the relevance of these outcomes in larger Recommended Phase 2 Dose Cohorts.
Prognostic Significance of Molecular Cytogenetics in Chinese Patients with Chronic Lymphocytic Leukemia: A Prospective Unicentric Analysis.
Authors: Xu Wei , Li Jianyong , Li Li , Wu Yujie , Yu Hui , Qiu Hongxia , Fan Lei , Ruan Changgeng
Published: October 2019

B-cell chronic lymphocytic leukemia (CLL) is the most common type of adult leukemia in the Western countries, however, infrequent in the Eastern. It is characterized by a highly variable clinical course; some patients survive more than 20 years, whereas others die within a few months of diagnosis. The characteristics of Chinese patients with CLL compared with the Western countries have not yet been clarified. The aim of this study was to prospectively explore characteristics and prognostic significance of molecular cytogenetic aberrations in Chinese patients with CLL. Interphase fluorescence in situ hybridization (FISH) and a panel of probes (LSI D13S319,LSI p53,LSI ATM,CEP 12,LSI MYB,LSI IGHC/IGHV) were used to detect cytogenetics abnormalities in 95 patients with CLL. Cytogenetics aberrations and their association with some other prognostic factors were analyzed. Kaplan-Meier was used for survival time. Out of the 95 CLL patients, molecular cytogenetic aberrations were found in 69 (72.6%) cases and 25 (26.3%) patients showed more than two kinds of abnormalities. The most frequent abnormalities detected were del(13q14) in 46 cases (48.4%), followed by trisomy of chromosome 12 in 22 patients (23.2%), 14q32 rearrangement in 21 patients (22.1%), del(17p13) in 16 patients (16.8%), del(11q22) in 9 patients (9.5%) and del(6q23) in 5 patients (5.3%). There were no significant differences of molecular cytogenetic aberrations in sex, age, Binet stages, peripheral lymphocyte count, and the levels of lactate dehydrogenase (LDH), β2-microglobulin (β2-MG), and ZAP-70. The TP53 and ATM gene deletion rates were higher in the group of CD38 high expression than that in the group of low expression (P=0.047 and P=0.001). No patient with TP53 and ATM gene deletion achieved complete response (CR) among 41 patients received treatment with fludarabine. The survival time was shorter in patients with high levels of LDH (P=0.028), β2-MG (P=0.012), and CD38 (P=0.000), and with TP53 gene deletion (P=0.000). Patients with sole del(13q14) had longer survival time than those with other abnormalities (P=0.044). It was showed that panel FISH has greatly increased the sensitivity of cytogenetic analyses and del(13q14) was the most frequent abnormality in CLL. Detection of molecular cytogenetic aberrations with FISH had important prognostic significance in CLL. The patients with sole del(13q14) had favorable outcome, and with del(17p13) had poor outcome.
Alemtuzumab (ALZ) Is an Effective Treatment for Both Primary and Relapsed Severe Paraneoplastic Pemphigus (PNP) Associated to B-Cell Chronic Lymphocytic Leukemia (CLL).
Authors: Bech Rikke , Baumgartner-Nielsen Jane , Peterslund N.A. , Steiniche Torben , Bang Karen , Deleuran Mette , d’Amore Francesco
Published: October 2019

ALZ is a monoclonal antibody used in the treatment of a variety of lymphoproliferative diseases, primarily CLL. PNP is a severe mucocutaneous disease often associated with B-cell malignancies. Besides distinct clinical and histopathological features, a correct PNP diagnosis relies on the demonstration by direct immunofluorescence of intercellular and basement membrane IgG deposits in the affected tissue. PNP is often refractory to immunosuppressive drugs and has frequently a fatal outcome. We report three cases, where sustained remissions of both PNP and CLL were induced by ALZ. In two of these cases, ALZ was able to reinduce a sustained PNP remission at the reappearence of the disorder years after the primary treatment. In all cases, the PNP diagnosis was confirmed by immunofluorescence. A 68-year old male with a 4-year CLL history developed blisters and ulcers on the extremities and trunk and severe stomatitis. Despite immunosuppression with steroids, fludarabine, cyclophosphamide, ciclosporin A, and azathioprin, PNP exacerbated. A standard 12-weeks ALZ schedule was initiated. PNP lesions started to regress under the last weeks of treatment and resolved completely within a few further weeks. A bone marrow biopsy showed a morphological and flowcytometric CLL remission. Now, 30 months after ALZ, the patient is still in remission without evidence of either PNP or CLL activity. Almost a year after this first case, a 77-year old male with a 3-year CLL history, presented with severe mucocutaneous lesions of the lips, oro-esophageal cavity, penis and extremities. PNP was diagnosed and a standard 12-weeks ALZ schedule was initiated. As in the first case, the lesions began to regress during the last weeks of treatment and normalized within the subsequent 8 weeks. At present, 1½ year after ALZ, the patient is well displaying only discrete residual PNP activity (left tongue margin, and praeputium). In the third case, a 80-year old woman with a B-CLL diagnosed 4 years previously, a severe PNP involving skin (upper trunk and extremities) and mucosae (anal and vagina) developed in association with marked CLL progression. After some unsuccessful immunosuppressive attempts, alemtuzumab treatment was initiated. All PNP lesions disappeared within a few weeks.

Conclusion: ALZ should be considered as a treatment option in severe CLL-associated PNP.
Prognostic Impact of Cancer Testis Antigens Expression in Advanced Stage Multiple Myeloma Patients.
Authors: Andrade Valéria C.C. , Vettore André L. , Almeida Manuella S.S. , Oliveira José S.R. , Chauffaille Maria de Lourdes L.F. , Yamamoto Mihoko , Andriolo Adagmar , Felix Roberta S. , Carvalho Fabrício , Caballero Otávia L. , Araujo Amélia G. , Panepucci Rodrigo A. , Zago Marco A. , Colleoni Gisele W.B.
Published: October 2019

Background: Cancer testis antigens have become the most extensively studied antigen group in the field of tumor immunology.

Aims: This study aims to analyze global expression of 14 CT (cancer/testis) antigens in MM to identify possible prognostic markers and therapeutic targets.

Patients and Methods: The expression of MAGEA1, MAGEA2, MAGEA3/6, MAGEA4, MAGEA10, MAGEA12, BAGE1, MAGEC1/CT7, GAGE family, LAGE-1, PRAME, NY-ESO-1, SPA17 and SSX1 was studied by RT-PCR in: 15 normal tissues, one pool of 10 normal bone marrow samples, three normal tonsils and bone marrow aspirates from six normal donors, three monoclonal gammophaties of undetermined significance (MGUS), five solitary plasmacytomas, 39 MM samples (95% advanced stage) and MM cell line U266. CodeLink Human UniSet I Bioarrays 10,000 genes was used for arrays analyses.

Results: SPA17 was positive in all normal tissues and was excluded for further analyses. MAGEC1/CT7 was positive in bone marrow aspirates from one MGUS and in one plasmacytoma. U266 cell line was positive for all CT antigens, except SSX1. The frequencies of CT antigens expression in MM patients were: MAGEC1/CT7 = 30/39 (77%); LAGE-1 = 19/39 (49%); MAGEA3/6 = 16/39 (41%); MAGEA2 = 14/39 (36%); GAGE family = 13/39 (33%); NY-ESO-1 = 13/39 (33%); BAGE-1 = 12/39 (28%); MAGEA1 = 10/39 (26%); PRAME = 9/39 (23%); SSX-1 = 10/39 (26%); MAGEA12 = 8/39 (20.5%); MAGEA4 and MAGEA10 = 0%. Cox’s regression model showed that GAGE family positivity and number of positive CT antigens > 6 were independent prognostic factors when all patients were analyzed. However, MAGEC1/CT7 expression was the only independent prognostic factor when non-transplanted patients where analyzed. Three samples predominantly positive (> 6) and three samples predominantly negative (0 or 1) for the 13 analyzed CT antigens were submitted to microarrays analyses. 147 genes were overexpressed in predominantly positive CT antigens samples.

Conclusions: Based on our findings, MAGEC1/CT7, MAGEA3/6 and LAGE-1 seem good candidates for immunotherapy, since together they are overexpressed in 85% of our MM cases. Besides, GAGE family expression, number of CT antigens > 6 and MAGEC1/CT7 seem to have impact on MM prognosis. Also, the results of arrays analyses corroborate the hypothesis that MM can be separate in two groups: predominantly positive and predominantly negative for CT antigens, meaning that these antigens may have important role for MM biology.
Therapy Response by Decrease of Free Light Chain Concentration in Multiple Myeloma Patients Treated with Bortezomib/Doxorubicin/Dexamethasone as First-Line Therapy.
Authors: Zinke-Cerwenka Wilma , Stojakovic Tatjana , Scharnagl Hubert , Wassertheurer Sabine , Linkesch Werner
Published: October 2019

Incorporation of new drugs such as the proteasome inhibitor bortezomib in the treatment of multiple myeloma (MM) has increased response rates. However, the challenge is to find the optimal combination of drugs and to change therapy as early as possible in case of suboptimal response. Therefore, we analyzed retrospectively the concentrations of serum free light chains (FLC) before, during and after therapy with bortezomib/doxorubicin/dexamethasone. In order to evaluate their prognostic value we compared the FLC kinetics to therapy response according to the International Uniform Response Criteria. Sixteen patients with newly diagnosed symptomatic MM (IgG/kappa: 9, IgG/lambda: 2, IgA/lambda: 1, IgA/kappa: 1, IgD/lambda: 1, IgD/kappa: 1, light chain kappa: 1) who received 4 cycles of bortezomib/doxorubicin and dexamethasone as initial therapy from November 2005 to June 2007 were analyzed. All patients showed a decrease in the FLC and/or a change of the quotient kappa/lambda towards the normal value after therapy. Reduction of FLC concentration ranged from 2.4- to 840-fold. Marked reduction could be observed after the first cycle of therapy. Seven patients revealed good response (sCR, CR or VGPR). In six of them, the decrease in FLC concentration was at least 20-fold, in one patient 2.9-fold. This patient had only a moderate increase of FLC concentration before therapy (below four times upper limit of normal). Two patients did not achieve good response (SD: 1, PD: 1); they had only a slight reduction in FLC (2.4- and 2.5-fold, respectively) although their initial FLC concentration was high. In parallel to the reduction of FLC, the serum concentration of immunoglobulins decreased. However, lowering of FLC occurred earlier and was more pronounced. Measurement of FLC concentration provides more rapid information for treatment monitoring than immunoglobulins and thus may be helpful to change therapy as early as possible in case of suboptimal response.
Preliminary Results of the Combination Rituximab, Fludarabine, Cyclophosphamide and Mitoxantrone (R-FCM) Followed by Rituximab Maintenance in Previously Untreated Chronic Lymphocytic Leukemia (CLL).
Authors: Bosch Francesc , Muntañola Ana , Villamor Neus , Terol Maria Jose , Gonzalez-Barca Elena , Ribera Josep Maria , Gonzalez Marcos , Abella Eugenia , Delgado Julio , Carbonell Felix , Garcia-Marco Jose A. , Escoda Lourdes , Ferrer Secundino , Monzo E. , Gonzalez Yolanda , Abrisqueta Pau , Salamero Olga , Gine Eva , Montserrat Emili
Published: October 2019

Immuno-chemotherapy (rituximab + combination chemotherapy) is emerging as the new gold standard for CLL treatment. However, the best combination chemotherapy to be given along with rituximab has not yet been determined. In addition, there is some indication that rituximab maintenance could be of benefit in CLL, as it has already been demonstrated in follicular lymphoma. In patients with previously untreated CLL, FCM results in a response rate of 90%, including a high proportion of MRD-negative CRs. These results suggested that FCM could be an ideal companion for rituximab in CLL therapy. Against this background, we have recently conducted a prospective clinical trial in which patients with untreated CLL receive R-FCM followed by maintenance with rituximab. Patients are eligible for therapy if they are younger than 70 yrs, have active disease (NCI-WG criteria), and adequate performance status. R-FCM consists of rituximab 500 mg/m2 on day 1 (375 mg/m2 the first cycle), fludarabine 25 mg/m2 i.v. on days 1 to 3, cyclophosphamide 200 mg/m2 on days 1 to 3, and mitoxantrone 6 mg/m2 i.v. on day 1, given at a 4-week intervals up to six courses. All patients receive G-CSF and cotrimoxazole. Patients achieving response (CR or PR) are subsequently treated with rituximab 375 mg/m2 every three months up to two years. Response is assessed three months after R-FCM treatment and includes MRD evaluation by four-color flow cytometry. Out of 69 patients included in the study, 38 (74% male, median age. 59 years) are evaluable for response to the first part of the treatment (R-FCM induction therapy). At study entry, 83% of the patients were in advanced (B and C) Binet’s clinical stage and 64% had increased (>20%) ZAP-70 expression. Ninety per cent of the patients have received the entire planed treatment. Overall response rate is 92%. MRD-negative CR is 36%, MRD-positive CR 41% (CR rate, 77%), nPR 7%, and PR 8%. Two out of 4 PR cases are MRD-negative. Toxicity has been manageable, with grade III–IV neutropenia being observed in 8% of the cases. Six serious adverse events have been documented (4 infections, 1 coronary disease, 1 CMV reactivation), two of them unrelated to the treatment. In conclusion, R-FCM is a well tolerated regimen that induces a high CR rate, including an important proportion of MRD-negative CRs. Whether these already extremely promising results are improved by maintenance therapy with rituximab remains to be seen.
Interim Results of a Phase I/II Study of Ocrelizumab, a New Humanised Anti-CD20 Antibody in Patients with Relapsed/Refractory Follicular Non-Hodgkin’s Lymphoma.
Authors: Morschhauser Franck , Marlton Paula , Vitolo Umberto , Linden Ola , Seymour John , Crump Michael , Coiffier Bertrand , Pulsoni Alessandro , Belch Andrew , Lerch Erika , Kimby Eva , Arnljots Kristina , Wassner Elisabeth , Mendila Myriam
Published: October 2019

Background: Ocrelizumab is a new humanised anti-CD20 antibody with the potential for enhanced efficacy in non-Hodgkin’s lymphoma (NHL) compared with rituximab due to increased binding affinity for the low-affinity variants of the FcγRIIIa receptor. An open-label, multicentre, dose-escalation study was conducted to evaluate the safety, efficacy, pharmacokinetics and pharmacogenetics of ocrelizumab in patients (pts) with relapsed/refractory follicular NHL following prior rituximab-containing therapy.

Methods: A total of 47 pts with a response (complete response [CR], unconfirmed CR [CRu], partial response [PR]) or stable disease (SD) of ≥6 months’ duration following prior rituximab-containing treatment were enrolled into 3 sequential dose cohorts. Pts received infusions of ocrelizumab q3w at 200 mg/m2 (cohort A), 375 mg/m2 (cohort B) and 750 mg/m2 (cohort C) for up to 8 doses. Response was assessed after 4 doses and after the end of treatment.

Results: Fifteen pts were enrolled in cohort A, 16 in cohort B and 16 in cohort C; 60% were male; mean age was 57 years; 78% had stage III/IV disease. At study entry 30%, 50% and 67% had intermediate or high FLIPI score in cohorts A, B and C, respectively. Median number of prior therapies was 2 (range 1–7); time to progression after last rituximab-containing therapy of <12 months: 47% (cohort A), 31% (cohort B) and 62% (cohort C) of pts. The majority of pts reported ≥1 adverse event (AE) [80–100%, cohorts A-C]; most of these were grade 1/2, with only 6 pts experiencing a grade 3 AE and no grade 4 AEs observed. The most common AEs were infusion-related reactions (IRR; 73%, 75% and 69% in cohorts A, B and C, respectively); only one was grade 3. Two pts discontinued treatment due to toxicity (dyspnoea, IRR). A total of 17 pts responded to ocrelizumab for a response rate (RR) of 36% (13% CR/CRu) across all cohorts. RR by cohort was 27% (13% CR/CRu) for A, 50% (25% CR/CRu) for B and 31% (no CR/CRu) for C. In pts relapsing after prior response (CR/PR) to rituximab-containing therapy (n=40), RRs were 23% (15% CR/CRu), 62% (31% CR/CRu) and 33% (no CR/CRu) in cohorts A, B and C, respectively. Of 6 pts with SD after prior rituximab-containing therapy, 1 pt had a PR, 4 pts had SD and 1 pt progressed.

Conclusion: Ocrelizumab is well tolerated at doses up to 750 mg/m2 given q3w; AEs consist mainly of grade 1/2 IRRs. Severe IRRs (grade 3/4) following ocrelizumab occur less frequently than with rituximab. In this heavily and rituximab-pretreated pt population, the RR of 36% is encouraging.
Ratio of Mutant JAK2-V617F to Wild Type Jak2 Determines the MPD Phenotypes in Transgenic Mice.
Authors: Tiedt Ralph , Hao-Shen Hui , Sobas Marta A. , Looser Renate , Dirnhofer Stephan , Schwaller Jürg , Skoda Radek C.
Published: October 2019

The reason why the JAK2-V617F mutation is associated with several phenotypic manifestations of human myeloproliferative disorders (MPD), i.e. polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF), is currently unknown. We established an inducible transgenic mouse model for MPD using a bacterial artificial chromosome (BAC) containing the human JAK2-V617F gene under the control of the JAK2 promoter. The sequences encoding the kinase domain were placed in the inverse orientation and flanked with antiparallel loxP sites to make the construct inducible by Cre-recombinase. A transgenic strain (FF1) containing 9 copies of the JAK2-V617F transgene was analyzed in detail. In this strain, Cre activity can lead to activation and/or excision of the multiple transgene copies. Depending on the number of actively rearranged transgene copies, we observed graded levels of expression of the JAK2-V617F mRNA and different MPD phenotypes. Crossing FF1 mice with transgenic mice expressing Cre-recombinase under the control of the hematopoiesis specific Vav promoter (VavCre) led to reduction of the FF1 copy number and low levels of JAK2-V617F expression (approximately 40% of endogenous wild type Jak2). These FF1;VavCre mice developed a phenotype resembling ET with strongly elevated platelet counts and moderate neutrophilia (Figure 1A). In contrast, induction of the JAK2-V617F transgene with the interferon-inducible MxCre resulted in less excision and higher JAK2-V617F transgene expression (approximately equal to wild type Jak2). These MxCre;FF1 mice displayed a PV phenotype with increased hemoglobin, thrombocytosis and neutrophilia (Figure 1B). The highest expression levels of JAK2-V617F were achieved by retroviral transduction (approximately 300% of wild type Jak2). Transplantation of these bone marrow cells into irradiated recipients caused a PV-like phenotype without thrombocytosis. Thus, the phenotype correlated with the ratio of mutant to wild type JAK2 mRNA. In patients with MPD, we found a similar correlation between the ratio of mutant to wild type JAK2 mRNA and the ET, PV and PMF phenotypes. In contrast to our transgenic mice, which display graded levels of JAK2-V617F with wild type JAK2 being present in every cell, each individual blood cell from patients with MPD can only be homozygous or heterozygous for the mutation, or normal. Therefore, the molecular mechanism determining the phenotype in humans may be more complex than in our mouse model and appears to be linked to the transition of the JAK2-V617F mutation to homozygosity.

Figure Figure
Chromosome 21-Encoded miR-125b and Its Role in the Development of Myeloid Leukemia in Children with Down’s Syndrome.
Authors: Klusmann Jan-Henning , Janikova Katarina , Li Zhe , Orkin Stuart H. , Reinhardt Dirk
Published: October 2019

Children with Down’s syndrome (DS) are at high risk to develop transient leukemia (TL) or acute myeloid leukemia (ML-DS), primarily the megakaryoblastic subtype (AMKL). Recently, acquired mutations of the hematopoietic transcription factor GATA1 - encoded on chromosome X - have been identified in leukemic blasts from virtually all patients with ML-DS or TL leading to the expression of a shorter isoform of GATA1 (GATA1s). During embryonic/fetal development mice carrying the GATA1s mutation show a hyperproliferation of a transient population of yolk sac/fetal liver megakaryocytic progenitors. However, GATA1s mice as well as GATA1s mice bred to mouse DS models (Ts65Dn and Ts1Cje) have normal adult hematopoiesis and do not develop leukemia. Hence, the factors on chromosome 21 that predispose to TL or ML-DS in cooperation with oncogenic GATA1s remain largely unknown. To date, four miRNAs (miR-99a, miR-125b-2, miR-155 and let-7c) are known to be encoded on chromosome 21. Three out of four miRNAs (miR-99a, miR-125b-2, let-7c) are not trisomic in the DS mouse model. To elucidate their role in the development of TL and ML-DS, we first measured their expression level in sorted cells from patient samples [ML-DS (n = 4), TL (n = 4), non-DS AMKL (n = 4)], and in megakaryocytes from healthy donors (n = 3) by qRT-PCR. The expression of miR-125b was clearly elevated in ML-DS (10-fold), TL (5-fold) and non-DS AMKL (3-fold) compared to normal megakaryocytes (ppatients vs. control <0.01). Using Gene Set Enrichment Analysis (GSEA), we confirmed the downregulation of putative target genes of miR-125b - predicted by Targetscan and Pictar - in the gene expression profile of ML-DS in comparison to non-DS AMKL. By retroviral overexpression using the Mouse Stem Cell Virus (MSCV), we could show that ectopic expression of miR-125b markedly enhanced proliferation of myeloid progenitor cells (MPC, CD34+CD33+) (MPCmiR-125b) in vitro. In colony-forming assays, the number of MPCmiR-125b derived colonies exceeded the number of MPCvector derived colonies (empty vector) by 4-fold (χ2 < 0.01). All MPCmiR-125b derived colonies contained a mixture of immature cells (CFU-GEMM) indicating an affected differentiation. This could also be confirmed by immunophenotyping of MPC cultured in liquid medium containing various cytokines (percentage CD34+CD33+ 26% vs 9%). In GATA1s fetal liver progenitors retroviral overexpression of miR-125b-2 could further enhance the proliferation rate and mediate the formation of large megakaryocytic colonies. These data support the hypothesis that miR-125b is involved in development of trisomy 21 associated leukemia by mediating the proliferation of MPC while repressing their differentiation. Therefore, miR-125b might be potential target in the therapy of ML-DS.
A Phase II Open-Label Study Evaluating Hemostatic Activity, Pharmacokinetics and Safety of Recombinant Porcine Factor VIII (OBI-1) in Hemophilia A Patients with Alloantibody Inhibitors Directed Against Human FVIII.
Authors: Mahlangu Johnny , Andreeva Tatiana A. , Macfarlane Donald , Reding Mark T. , Walsh Christopher , Ritchie Bruce , Ewing Nadia , Kessler Craig M. , Kempton Christine , Libby Edward , Zozulya Nadezda , Shapiro Amy D. , St-Louis Jean , Warrier Indira , Hoots W. Keith , Gruppo Ralph A. , Mueksch Josef N.
Published: October 2019

Development of inhibitors to human FVIII (hFVIII) is a significant complication in the reversal and prevention of bleeding events in hemophilia A patients. Porcine FVIII (pFVIII) possesses low cross reactivity to anti-hFVIII antibodies. OBI-1, a recombinant B-domain deleted pFVIII, has recently been tested in a Phase II trial in patients with congenital hemophilia A and inhibitors experiencing a non-life/non-limb threatening bleed. In patients with a measurable anti-pFVIII antibody titer, dosing was initiated with a loading dose (LD) followed by up to 8 doses of 50 to 150 U/kg of OBI-1 administered at 6 hour intervals until the bleed was controlled. The PK profile of OBI-1 was assessed for the first OBI-1 infusion of each patient. FVIII levels were measured 30 minutes after each infusion. Inhibitor titers were evaluated for a minimum of 6 months after the first OBI-1 infusion. A total of 25 bleeding episodes in 9 patients were treated successfully with OBI-1. The median time from bleeding onset to treatment was 7 hours (range: 3 20 hr). OBI-1 showed a cumulative efficacy of 72% after 1 injection, 84% after 2 injections or less, 92% after 3 injections or less, and 100% after 8 or less injections. In over 40 infusions, OBI-1 was well tolerated and no drug related SAEs were observed. One case of pruritus rated as mild was easily controlled with diphenhydramine. FVIII levels measured 30 minutes after each OBI-1 infusion ranged from < 0.5% to 226%. They were generally lower with higher anti-pFVIII titers although all bleeds were successfully controlled. Follow up inhibitor titers are still being monitored and final results will be presented and discussed. Other investigative parameters, including vital signs and laboratory variables did not show treatment related abnormalities. OBI-1 can be given as a short infusion. It was effective in controlling all bleeds which occurred in this study and was well tolerated. The results suggest that a LD is not needed with OBI-1 treatment, and in some instances patients may have been over treated. Given the promising results in this study, additional studies are planned to optimize dose range for OBI-1 and to confirm the long term safety and efficacy of OBI-1 in the treatment of bleeds in a larger cohort of individuals with hemophilia A complicated by the presence of hFVIII inhibitors.
The Novel Aurora Kinase Inhibitor AS703569 Shows Potent Anti-Tumor Activity in Acute Myeloid Leukemia (AML).
Authors: Sarno Samantha , Shaw Jamie , Spooner Edward , Ma Jianguo , Clark Ann , Dumontet Charles , Romanelli Angela
Published: October 2019

Aurora kinases have been implicated in the onset of several human cancers. They are overexpressed in different tumor types including colon, breast, and pancreatic cancers, as well as in leukemias. AS703569 (formerly known as R763) is an orally bioavailable inhibitor of all three Aurora kinase isoforms (A, B and C), and is currently in Phase I clinical studies for both solid tumors and hematological malignancies. AS703569 also exhibits potent inhibition of the receptor tyrosine kinase FLT3. FLT3 is postulated to play an important role in the pathogenesis of AML. Somatic mutations and internal tandem duplications in FLT3 resulting in constitutive kinase activation are the most commonly acquired genomic abnormalities found in this disease. The potential therapeutic benefits of AS703569 as a dual Aurora kinase/FLT3 inhibitor is being explored in AML. The anti-proliferative effect of AS703569 has been tested in a panel of AML cell lines harboring different genetic defects. All cell lines were potently inhibited by AS703569 with IC50s between 0.7nM and 1 μM. The MV4-11 cell line possessing the FLT3 ITD was the most sensitive. Examination of cell cycle profiles demonstrated that AS703569 induced a block in the G2/M phase with a subsequent accumulation of cells in sub G (indicative of apoptosis). Dose-dependent induction of endoreduplication was observed in a subset of the cell lines tested. The pro-apoptotic role for the compound was further supported by the observation that increasing concentrations of AS703569 led to a significant increase in Caspase-3 cleavage and Annexin-V staining. AS703569 treatment led to a dose-dependent inhibition of FLT3 and Histone H3 phosphorylation, suggesting that modulation of both FLT3 and Aurora kinase B are implicated in the observed anti-proliferative, pro-apoptotic activity of AS703569 in these cell lines. To extend our investigation from cell lines to primary patient samples, AML blasts isolated from 15 patients were exposed ex vivo to increasing concentrations of AS703569 or Ara-C. In every case, AS703569 induced a dose-dependent decrease in viability that was more potent than Ara-C. We also tested the anti-tumor activity of AS703569 in two leukemia xenograft mouse models: MV4-11 and HL-60. In the MV4-11 model, tumors were grown to different sizes: 230, 750, and 1100 mm3, and in all three experimental conditions, tumor regression was seen after one dose (75 mg/kg) of AS703569. We found that a weekly dosing schedule is best tolerated by mice and reproducibly shows potent anti-tumor activity. Consistent with our in vitro data, AS703569 was less potent in the HL-60 xenograft model where the FLT3 mutation is absent. In this model, AS703569 only partially inhibited tumor growth (data not shown). Taken together, our data suggest that AS703569 shows particularly promising therapeutic activity in AML, especially cases presenting FLT3 mutations.