bluuebunny/crossref_metadata_2025_split huggingface dataset:
An In Vitro and In Vivo Evidence That Downregulation of Leukemia Inhibitory Factor (LIF) Receptor (LIF-R) Decreases the Metastatic Potential of Human Rhabdomyosarcoma (RMS) Cells.
Authors: Wysoczynski Marcin , Miekus Katarzyna , Marcinkowska Anna , Janowska-Wieczorek Anna , Ratajczak Mariusz Z. 
Published: October 2019

Abstract
Rhabdomyosarcoma (RMS) and skeletal muscle-derived tumors frequently infiltrate bone marrow (BM). We have demonstrated that the stromal-derived factor (SDF)-1-CXCR4 receptor (Blood2002;100:2597) and hepatocyte growth factor (HGF)-c-Met receptor (Cancer Res. 2003;63:7926) play an important role in RMS metastasis to BM. Leukemia inhibitory factor (LIF) is a well known factor that plays an important role in skeletal muscle development/regeneration and similarly as SDF-1 and HGF is secreted by BM stroma. This prompted us to examine whether the LIF-LIF receptor (LIF-R) axis affects the biology/metastasis of RMS cells. We employed in our studies, human established RMS cell lines, as well as RMS samples isolated from patients and noticed that LIF-R was expressed not only on established human RMS cell lines (7/7) but more importantly, it was also detectable in patient samples (23/23). We also found that in RMS cells LIF stimulatesphosphorylation of MAPKp42/44, AKT and STAT3,chemotaxis and adhesion andincreases resistance to cytostatics (e.g., etoposide).
These LIF-mediated effects were inhibited after downregulating the LIF-R by siRNA. To learn more on the biological significance of the LIF-LIF-R axis in vivo we employed two models. First, human RMS cells (RH-30) were exposed or not exposed to LIF-R siRNA and subsequently injected into SCID™-Beige immunodeficient mice. To estimate the number of RMS cells that seed to BM and liver in these animals, we isolated DNA and using real- time RT-PCR, amplified human a-satellite sequences and murine b-actin. The number of human cells present in murine organs was subsequently calculated from a standard curve derived from mixing varying numbers of human cells with a constant number of murine cells. We noticed that downregulation of LIF-R by siRNA significantly decreased the number of human RMS cells in murine BM and liver (x4 and x2 respectively). In a second model, the RH30 cell line was selected by repetitive chemotaxis for cells that are highly responsive to LIF (RH-30 L) and subsequently the cells from parental RH-30 cell line and RH-30 L cells were injected intramuscularly. Six weeks after tumour inoculation, we detected more metastasis in bone marrow and lungs in mice injected with RH-30L cells as compared to parental RH-30 clone (x6 and x3 respectively). In conclusion, we present evidence for the first time that the inhibition of LIF-LIF-R axis may decrease the invasive potential of human RMS both in vitro and in vivo. Hence, molecular targeting of LIF-LIF-R axis could possibly become a more effective new strategy to control the progression and metastasis of RMS.
Co-Transplantation of Autologous Umbilical Cord Matrix Mesenchymal Stem Cells Improves Engraftment of Umbilical Cord Blood in NOD/SCID Mice.
Authors: Friedman Robb , Betancur Monica , Tuncer Hande , Boissel Laurent , Cetrulo Curtis , Klingemann Hans-Georg 
Published: October 2019

Abstract
Umbilical cord blood (UCB) is a viable source of hematopoietic stem cells for transplantation of children and adults undergoing treatment for hematological malignancies. However only 4% of adults 70kg and over have a UCB unit available which contains the widely accepted minimum cell dose of 1.5x107 mononuclear cells per kilogram. Co-transplantation of hematopoietic stem cells with mesenchymal stem cells may enhance engraftment and therefore decrease transplant-related morbidity and mortality from delayed leukocyte recovery associated with a low pre-transplant cell dose. Umbilical cord matrix (UCM) cells, found in the Wharton’s Jelly, were easily and reliably extracted from minced pieces of cord by culture in RPMI + 20% fetal bovine serum at 37° and 5% humidified CO2. UCM expand best in 20% FBS but can also be expanded in human serum, autologous serum, and X-VIVO10. Small (1–3mm) minced pieces of umbilical cord can be cyropreserved at the time of delivery in 10% DMSO solution. UCM cells exhibit a fibroblast morphology and express markers common to mesenchymal stem cells: CD73 (SH3), CD105 (SH2), CD 29, CD44, CD49b, CD117, CD166, STRO-1 and HLA-DR. UCM are negative for CD14, CD 19, CD34, and CD45.
Morphology and cell surface marker expression is stable after greater than fifteen passages. UCM cells grown in culture were shown to produce more GM-CSF and G-CSF than similar numbers of adult bone marrow mesenchymal stem cells, GM-CSF 178 pg/mL versus 77 pg/mL and G-CSF 82.6 pg/mL versus 7.9 pg/mL. NOD/SCID mice treated with anti-NK 1.1 antibodies and irradiated with 350 cGy were injected with suboptimal (1x104) numbers of cord blood CD34+ cells with and without 1x106 autologous UCM cells, extracted from the same umbilical cord as the cord blood CD34+ cells. Bone marrow was harvested at six weeks post transplant from both femurs and tibias and peripheral blood obtained via cardiac puncture. The percentage of human CD45+ cells in the bone marrow and the peripheral blood was assessed by flow cytometry. NOD/SCID mice transplanted with 1x104 cord blood CD34+ cells alone had 3.0% human CD45+ cell engraftment in the bone marrow and 3.6% human CD45+ cells in the peripheral blood, while NOD/SCID mice transplanted with 1x104 CD34+ cells and 1x106 UCM cells had an average of 27.3% human CD45+ cell engraftment in the bone marrow and 3.9% human CD45+ cells in the peripheral blood. These results indicate a trend towards improved engraftment in vivo with co-transplantation of suboptimal numbers of umbilical cord blood CD34+ cells and autologous umbilical cord matrix cells versus transplantation of suboptimal numbers of umbilical cord CD34+ cells alone.
Secondary Bortezomib-Resistance Is Characterized by Upregulation of Proteasomal Subunits and Their Insufficient Inhibition by Bortezomib, Which Can Be Overcome by Alternative Proteasome Inhibitors Combined with Induction of the ER Stress Response.
Authors: Ruckrich Thomas , Gogel Jeannette , Kraus Marianne , Ovaa Huib , Driessen Christoph 
Published: October 2019

Abstract
Resistance towards proteasome inhibition by Bortezomib (Velcade®) represents a challenge for myeloma therapy. Its biology has not yet been characterized in detail. We have demonstrated that Bortezomib-sensitive malignant haematopoetic cells can acquire secondary resistance to Bortezomib in vitro. We here present the first analysis of proteasome biology and activity, alternative proteolytic pathways, ubiquitin-specific proteases (USP) and the ER stress response (unfolded protein response, UPR) upstream of the proteasome, as well as in vitro cytotoxicity of conventional cytotoxic drugs, alternative proteasome inhibitors and agents that target the UPR in Bortezomib-resistant (BR) cells, compared to wild type (WT) controls. BR cells had higher activities of all subunits of the constitutive and the immunoproteasome, as deferred from turnover of fluorogenic substrates as well as affinity-labelling of active proteasome subunits in intact cells. This was mirrored by increased levels of proteasomal β1 and β2, but especially β5 polypeptides, implicating a homeostatic system that senses and corrects low proteasome activity in cells chronically exposed to Bortezomib. While the vinylsulfone-type proteasome inhibitor NLVS abrogated detectable proteasome activity in both BR and WT cells, Bortezomib at therapeutic concentrations eliminated proteasomal β1 and β5-type activity only in WT cells, while BR cells retained residual activity. These changes in proteasome biology appear to be the molecular hallmark of required Bortezomib resistance, since no changes were observed between WT and BR cells in alternative cytosolic or lysosomal proteolytic pathways, UPR activity as well as the gross activity pattern of USP. As expected, this translated into sensitivity against cytotoxic drugs in vitro: BR cells were less sensitive towards alternative proteasome inhibitors. However, while the IC50 for pan-proteasome inhibitors was only roughly doubled in BR cells, it was nearly tenfold elevated for the β5-preferring vinylsulfone inhibitor NLVS. By contrast, sensitivity towards anthracyclines or cytotoxicity induced by ER stressors as well as the synergy between proteasome inhibitors and UPR-activators remained unaffected in BR cells. Based on our data, proteasome inhibitors with activity profiles different from that of Bortezomib, alone or in combination with induction of the UPR, may represent an appropriate concept to overcome secondary Bortezomib resistance.
Effective Targeting of Leukemic Cells in Children with B-Precursor Acute Lymphoblastic Leukemia Treated with Anti-CD22 (Epratuzumab). A Children’s Oncology Group (COG) Study.
Authors: Borowitz Michael J. , Carroll William L. , Adamson Peter C. , Cairo Mitchell S. , Goldenberg David M. , Wegener William A. , Raetz Elizabeth 
Published: October 2019

Abstract
Identifying new approaches to treating relapsed ALL is a top priority because these patients fare poorly with current retrieval strategies. COG protocol ADVL04P2 is a phase I/II feasibility pilot study for children with relapsed B-precursor ALL that uses epratuzumab, a humanized anti-CD22 monoclonal antibody, in combination with conventional re-induction chemotherapy. Children whose blasts are CD22 positive first receive 4 doses of 360 mg/m2 epratuzumab as a single agent on days minus(−)14, −10, −6 and −2. Subsequently, they receive epratuzumab weekly in combination with standard re-induction chemotherapy starting on day 0. We assessed the effectiveness of epratuzumab targeting by by assaying CD22 expression on residual leukemic blasts. Peripheral blood was obtained from 15 patients 24 hours after administration of the first dose (day −13), and again at day −6 and day 0 and stained with the combination of CD10/CD22/CD45/CD19 in four color flow cytometry to permit assessment of CD22 on analytically isolated leukemic blasts. We determined quantitative expression of CD22 with a calibration kit (Quantibrite, BDBiosciences, San Jose, CA) using two PE-conjugated anti CD22 antibodies directed against different epitopes of the CD22 molecule: clone RFB4 (Caltag, Burlingame CA), directed against the same epitope as epratuzumab, and clone SHCL-1 (BDBiosciences), directed against a non-cross-reacting epitope of CD22. RFB4 binding was decreased by more than 99% within 24 hours after administration of the first dose in all patients, indicating rapid targeting of epratuzumab to leukemic cells. In all but 4 patients, levels remained low at all subsequent time points; in 2 patients expression was restored to about 15% of baseline levels by day 0, and 2 additional patients had very small (1–2%) subsets of blasts with significant expression, one at day −6 and one at day 0. In contrast to RFB4, SHCL-1 identified residual CD22 antigen expression, but SHCL-1 binding was decreased by an average of 70–75% in 14 of 15 patients at all three post-epratuzumab timepoints. The one patient with no change in SHCL-1 binding was a patient with MLL-rearranged ALL with very low levels of CD22 expression at diagnosis, and poor response to therapy. Because binding of epratuzumab to CD22 has been shown to result in antigen internalization in vitro, we interpret our findings as demonstrating in vivo internalization of CD22 with epratuzumab binding, though we cannot exclude either shedding of antigen or conformational change to block SHCL-1 sites. We conclude that epratuzumab rapidly targets CD22 on ALL blasts in vivo, though some blasts in some patients may escape over time. Targeting is associated with a physical change in CD22 antigen expression in most patients, most likely through internalization.
AML Progenitor/Stem Cells Overexpress XIAP and Are Sensitive to the Small Molecular XIAP Inhibitor 1396-11.
Authors: Carter Bing Z. , Kim Tae Kon , Mak Duncan H. , Hu Wei , Pinilla Clemencia , Reed John C. , Andreeff Michael 
Published: October 2019

Abstract
Chemotherapy is the primary treatment modality for AML patients. Although most patients show initial responses to chemotherapeutic agents, the majority of patients relapse, suggesting that more primitive cells in AML are resistant to chemotherapy-induced apoptosis. XIAP, a potent cellular caspase inhibitor, is highly expressed in various tumor cells and leukemic cells and contributes to chemoresistance. However, the expression levels of XIAP in early progenitor/stem cells of AML patient are unknown. To address this question, we collected blasts from bone marrow or peripheral blood of AML patients (n=11) and separated them into CD34+/CD38+ and CD34+/CD38− populations by fluorescent-activated cell sorter after immunostaining. RNA was isolated and XIAP mRNA levels were determined by Taq-Man quantitative RT-PCR in these two cell populations and compared with levels from normal CD34+/CD38+ and CD34+/CD38− cells (n=13). Our results demonstrated that XIAP is highly expressed in both progenitor (CD34+/CD38+) and stem cell (CD34+/CD38−) compartments in primary AML blasts, with XIAP mRNA levels higher in these two cell populations in AML than in the corresponding normal bone marrow progenitor/stem cells (p=0.05 and 0.08 for CD34+/CD38+ and CD34+/CD38− compartments, respectively). Our previous studies showed that inhibition of XIAP expression by antisense oligonucleotide and its suppression by small molecule chemicals induce apoptosis of AML cells. 1396–11 and 1396–34, polyphenylurea-based small molecule XIAP inhibitors bind to and inhibit the BIR2 domain of XIAP. 1396–34 was previously tested in AML (Carter, et al. Blood105: 4043, 2005). Previous studies of 1396–11 demonstrated anti-tumor activity against human solid tumor xenografts in mice with no apparent hematopoietic or liver toxicities (Schimmer, et al. Cancer Cell5: 25, 2004; Berezovskaya, et al. Cancer Research65: 2378, 2005). We treated OCI-AML3, HL-60, Jurkat, and U937 leukemia cells with 1396–11 and found it to be more potent than 1396–34 in all leukemia cell lines tested. In addition, we treated AML blasts from 5 patients with 1396–11 and its inactive control 1396–28. Apoptosis was determined in total blasts and in both the CD34+/CD38+ and CD34+/CD38− compartments. Our results showed that 1396–11 induced cell death equally in bulk blasts, CD34+/CD38+ progenitor, and CD34+/CD38− stem cells. Four out of 5 AML samples showed significant apoptosis with ≤ 10 μM 1396–11. The control compound 1396–28 had minimal effect on cell survival in all 5 patient samples tested. Our studies illustrated that XIAP is overexpressed in AML progenitor/stem cells and that these cells are sensitive to a small molecule XIAP inhibitor, suggesting XIAP is a target for AML therapy, with the potential of eradicating primitive leukemic progenitor/stem cells.
A BMT Model Mice for Myelodysplastic Syndromes (MDS) and Transformation to AML.
Authors: Watanabe-Okochi Naoko , Kitaura Jiro , Harada Hironori , Ono Ryoichi , Nakajima Hideaki , Nosaka Tetsuya , Inaba Toshiya , Kitamura Toshio 
Published: October 2019

Abstract
To understand myelodysplastic syndromes (MDS) at the molecular level, we used a mouse bone marrow transplant (BMT) model. Several leukemogenic fusion proteins were demonstrated to induce leukemia in mouse BMT models. However, analysis of molecular basis of MDS has been hampered by the lack of mouse MDS models. Mutations of a transcription factor AML-1/Runx-1 were identified in 15–40% of MDS-RAEB and MDS/AML and 5–10% of de novo AML. To test transforming abilities of the mutant AML-1, we transduced AML-1 with different mutations (#5 and #27) into mouse bone marrow cells using our highly efficient retrovirus-mediated gene transfer, transplanted the transduced cells to irradiated mice, and found that most mice developed leukemia within 4–13 months after the transplant (table1 and figure1). The leukemic mice exhibited marked hepato-splenomegaly, and morphological abnormalities of myeloid and erythroid lineages were frequently observed. Some mice developed to leukemia after a long latency with MDS-like symptoms, either with pancytopenia or leukocytosis with anemia, thus mimicking the human disease. To clarify the differences between the leukemic mice with early onsets and those that developed leukemia after a long period of MDS-like symptoms in the mutant AML-1-induced mouse diseases, the integration sites of the transduced retroviruses were investigated using a polymerase chain reaction (PCR)-based technique bubble PCR. Our preliminary results suggested that the integration sites of the retroviruses harboring the mutant AML-1 contribute to the early onset of leukemia. In conclusion, we have developed a useful mouse in vivo model of MDS/AML that should help understand the molecular basis of MDS and progression of MDS to leukemia.
penetrance and latency mutation number position form penetrance latency #5 D171N point mutation in RHD 69.6 % 4–13 months #27 S291fsX300 truncation type 100 % 4–9 months Figure Figure
Evaluation of Cardial Iron Deposition by T2* MRI in Transfusion Dependent Patients with Myelodysplastic Syndromes.
Authors: Deplano Simona , Di Tucci Anna , Matta Gildo , Agus Annalisa , Gabbas Attilio , Murru Roberta , Angelucci Emanuele 
Published: October 2019

Abstract
Cardiac T2* Magnetic Resonance Imaging (MRI) has been recently used to evaluate myocardial iron deposition in patients with transfusion dependent beta-thalassemia major. No comparable studies have been published for patients with myelodysplastic syndromes receiving chronic red blood cell transfusion. Therefore we measured cardiac-MRI T2* in 16 patients (10 male, 6 female) with myelodysplastic syndromes (aged 54 – 82 years, median age 67). All of them were transfusion dependent having received a median number of 60 (range 16–225) packed red blood cell transfusion equivalent to 3.2 – 45 (median 12) grams of iron. Nine have been irregularly and sporadically chelated by deferoxamine, seven were unchelated. Serum ferritin levels ranged from 1163 to 6241 mg/dl (median value 2086). None of the patients presented signs or symptoms of cardiac dysfunction at the time of the study.
Cardiac-MRI T2*values obtained ranged from 5.6 to 80 (median value 46.5) milliseconds (ms). Correlation between serum ferritin and cardiac T2* value was weak ( r= 0.43, r2 =0.18). According to D. Pennel we considered as significant of myocardial iron deposition a relaxation time ≤ 20ms. Cardiac T2* was < 20ms in 3 patients who had never used iron chelators (5.6, 12.4 and 8.5 ms, respectively). They had received 39, 101 and 200 units of red blood cell transfusion, corresponding to 7.8, 20 and 40 grams of iron, respectively. Of relevance 2 of them died within few months after the end of the study and one showed early signs of left ventricular dysfunction. None of the patients with a cardiac T2* value >20 ms showed instrumental nor clinical signs of cardiac deterioration in six months follow up. No patient who had received less than 39 transfusions presented cardiac T2* value ≤20 ms. Evaluation of myocardial iron deposition by T2* cardiac MRI could be recommendable in myelodyplasia patients who had received more than 30 packed red blood cells transfusisions.
Efficacy and Safety of Denileukin Diftitox (Ontak®) in the Treatment of Cutaneous T-Cell Lymphoma (CTCL) According to CD25 Status.
Authors: Foss Francine , Duvic Madeleine , Geskin Larisa , Anderson Joseph , Porcu Pierluigi , Hohl Raymond J. , Cooper Maureen , Zain Jasmine M. , Zic John , Hibberd Patrica , Acosta Mark 
Published: October 2019

Abstract
Denileukin diftitox, an interleukin-2 (IL-2)-diphtheria toxin fusion protein, binds to and intoxicates cells expressing the medium (CD122, CD132) and high affinity (CD25, CD122, CD132) IL-2 receptor. Because its role in patients whose tumors tested negative for the CD25 component of the receptor had not been prospectively studied, we initiated a prospective, open label, multi-center study to evaluate the safety and efficacy of denileukin diftitox in CTCL patients according to CD25 status (CD25+ and CD25-). Patients with pathologically proven, persistent or recurrent CTCL Stages IB-IVA were treated with denileukin diftitox at a dose of 18 mcg/kg/day x 5 days every 21 days for up to 8 cycles. Expression of CD25 by tumor cells in skin biopsies was determined by IHC or flow cytometry at a central laboratory and investigators were blinded to CD25 status. Response was based on improvement in skin involvement by weighted skin assessment for lesion type (patch, plaque, tumor) and blood involvement based on quantitation of Sezary cells. Safety was evaluated based on reports of Grade 3 and 4 toxicities (NCI CTC version 2.0). Sixty-one pts were enrolled and treated with denileukin diftitox. Disease stages were: I-IIA (n=14), IIB (n=22), III (n=11) and IV (n=14). Eighteen (29.5%) had blood involvement. Fifty-seven pts completed at least one cycle of treatment and were evaluable for response. Four pts were excluded because they had Stage IVB (visceral) disease (n=2), did not have a post baseline skin assessment recorded (n=1), or did not complete the first cycle (n=1). The median number of cycles of denileukin diftitox was 4 (range 1–13). The overall response rate (CR+PR) was 53% (30/57, 95% CI 40–66%), with 2 CR. Of the 57 evaluable pts, tumor was CD25+ in 34 (70%), CD25- in 14 (25%) and unknown in 9. Response rate was similar in patients whose tumor tested CD25+ (56%, 95% CI: 39–73%), and CD25- (43%, 95% CI: 17–69%) (p=0.41). For the 61 patients the most common grade 3/4 toxicities included constitutional symptoms (11%) and metabolic/laboratory abnormalities (36%). One patient with significant cardiopulmonary disease died of myocardial infarction while on therapy. The frequency of grade 3/4 toxicities was similar in CD25+ 64% (23/36), vs CD25- 75% (12/16) (p=0.62). In conclusion, we found that 70% of the CTCL pts evaluated for this study tested CD25+ and that the response rates and safety profile of denileukin diftitox in CD25+ and CD25- patients were similar. These data support a role for denileukin diftitox in the treatment of CTCL regardless of CD25 status.