bluuebunny/crossref_metadata_2025 huggingface dataset:
Imbalance in Alternative Splicing of MCL-1 Inhibits Apoptosis in Diffuse Large B-Cell Lymphoma
Authors: Rekers Nicolle H , Moesbergen Laura M , Hijmering Nathalie J , Vos Wim , Oudejans Joost , Ossenkoppele Gert J. , Meijer Chris JLM , Cillessen Saskia AGM
Published: October 2019

Diffuse large B-cell lymphoma (DLBCL) remains eventually fatal in 30-40% of the patients, despite intensive chemotherapy (CHOP) in combination with rituximab. A subgroup of chemotherapy-refractory DLBCL is characterized by high expression levels of both pro- and anti-apoptotic genes, including MCL-1. Alternative splicing of the MCL-1 gene results in a Bcl-2-like anti-apoptotic MCL-1L protein and a BH3-only pro-apoptotic MCL-1S protein. In the present study, we investigated if a switch in alternative splicing of MCL-1 is involved in apoptosis-resistance in primary lymphoma cells of 20 DLBCL patients and 5 DLBCL cell lines.

RT-MLPA analysis revealed that MCL-1L and MCL-1S are both expressed in all tested DLBCL samples and DLBCL cell lines, however expression levels varied strongly. An imbalance between the expression levels of MCL-1L and MCL-1S to an anti-apoptotic status was observed in DLBCL patient cells and DLBCL cell lines, especially in activated B-cell like (ABC)-DLBCL, compared to tonsillar germinal center B-cells. MCL-1 mRNA expression was confirmed at protein level using immunohistochemistry and western blot analysis. Co-immunoprecipitation demonstrated that MCL-1L inhibited apoptosis by binding of Bak in MCL-1L positive DLBCL cell lines. Knockdown of MCL-1L with siRNA analysis resulted in induction of apoptosis in both GCB- and ABC-DLBCL cell lines and also in increased sensitivity to the conventional chemotherapeutical drugs etoposide. Downregulation of MCL-1L using flavopiridol induced apoptotic cell death of MCL-1L-positive DLBCL cells with low Bcl-2 expression.

In summary, a switch in alternative splicing of MCL-1 occurs in a subgroup of DLBCL leading to an increase in the level of anti-apoptotic MCL-1L that contributes to therapy-resistance. These preclinical data suggest that targeting of MCL-1L might be a therapeutic option for MCL-1L positive DLBCL.

Disclosures

No relevant conflicts of interest to declare.
Impact of FDG-PET/CT in Staging, Response Evaluation and Decision of Radiotherapy for Pediatric Hodgkin Lymphoma
Authors: Khan Safia , Riaz Shazia , Ahmecd Mehoob , Kazmi Ather , Bashir Humayun
Published: October 2019

PURPOSE: To review the role of FDG-PET/CT in staging, assessment of treatment response, and decision of radiotherapy among Pediatric Hodgkin Lymphoma patients.

MATERIALS & METHOD: 125 hodgkin lymphoma patients up to 18 years of age who were evaluated with FDG-PET/CT for staging workup, treatment response and planning for radiatherapy during 2010-2013 in Shaukat Khanum Memorial Cancer Hospital & Research Center, Lahore, Pakistan were retrospectively reviewed for this study. 104 out of 125 patients were evaluated in mid treatment with FDG-PET/CT by giving F18 FDG dose 0.21mg/Kg intravenously in resting condition followed by imaging over 25-30 min; contrast enhanced CT was required as a part of PET/CT protocol. Remaining 21 patients were evaluated for treatment response with isolated contrast enhanced CT due to non-availability of FDG-PET/CT.

RESULTS: Majority (82%) of the patients fall under the age group of 5 to 15 years. However, ratio of male patients was greater than female i.e. about 3:1. From Chi square analysis it is revealed that there is substantial association between staging with FDG-PET/CT and conventinal CT. From cross tabulation it is concluded that FDG-PET/CT downstaged one patient and up staged eight patients. On the bases of interim CT, radiotherapy was given to 7 patients of which 5 relapsed. Based on FDG-PET/CT 19 patients received radiotherapy, out of these 19 patients 10 relapsed.

The 48% of 21 patients evaluated by conventional CT relapsed and 15% of 104 patients evaluated by FDG-PET replaped.

CONCLUSION:Based on our data , initial staging FDG-PET/CT upstaged more patients than convential CT, while at mid-treatment response evaluation less patients received radiation in FDG-PET/CT group. Relapse rate was lower in FDG-PET/CT group.

Disclosures

No relevant conflicts of interest to declare.
Flow Cytometric Detection of Liposomes in the CSF of Patients with NHL/ALL, Treated with Intrathecal Liposomal Cytarabine: Synonymous of Toxicity?
Authors: Peñalver Francisco Javier , Pilar Martínez-Barranco , Villalón Lucía , Barón Manuel , Martín L , Ricard Pilar , Arribalzaga Karmele , Garcia Bueno María José , Orfao Alberto
Published: October 2019

Objectives

Patients with aggressive non-Hodgkin’s lymphoma (NHL) and acute lymphoblastic leukemia (ALL) with high risk of CNS infiltration or recurrence may receive prophylaxis or be treated with intrathecal injections (IT) of liposomal cytarabine (LC) once every 2 weeks (due to its sustained release of cytarabine).

Sequential flow cytometric studies in patients with CNS involvement and treated with LC, allow to detect tumor cells as well as LC liposomes (LCL) in the CSF. The true meaning of the presence of these liposomes during the treatment with LC is still unknown.

Patients and Methods

In this study, we aimed to investigate the presence of LCL detected by Flow Cytrometry (FC) among 166 CSF samples of 69 patients treated with LC. Samples were taken before each administration of intrathecal LC and centrally processed.

Results

Results are shown in the below table. Abstract 5444. Table 1Patients with NHL/ALL, treated with Intrathecal Liposomal CytarabineNPatients69Samples166Liposomal Cytarabine Liposomes (LCL) in CSFN%Patients1319%DLBCL646%LLT215%ALL215%MCL18%MM18%CLL18%Samples2616%Males862%2616%Age66 years[33-84]LCL in CSF of patients receiving Liposomal CytarabinePROPHYLAXISNToxycityTotal nº of IT LC dosesPatients7nº of IT LC doses*3 (2-5).LCL7After 1st dose5After 2nd dose21 LLT patient: arachnoiditis, high intrathecal pressure, papillitis, without sequelae5After 3rd dose2After 5th dose1LCL in CSF of patients receiving Liposomal Cytarabine leptomeningeal disease TREATMENTNTumor: ToxycityTotal nº of IT LC dosesPatients6nº of IT LC doses*6 (4-7)LCL6After 1st dose31 DLBCL patient: Headache, dizziness and instability51 MM patient: Headache, dizziness and instability4After 3rd dose2After 5th dose11 ALL patient: Headache, dizziness and instability7 * Median [range]

The thirteen patients were simultaneously treated with LC and IT + IV dexamethasone. It was found no relationship between the amount of LCL and toxicity.

Conclusions:

Presence of LCL in CSF is detected in 15-20% of patients receiving IT LC as prophylaxis / treatment and it does not show a direct relationship with presence of toxicity.

Although the true clinical significance of the presence of LCL in CSF remains unknown, their monitoring could be useful to individualize the treatment with CL IT (dose and time between doses).

Disclosures

Off Label Use: Liposomal cytarabine is not approved for prophylaxys of lymphomatous meningitis..
Pediatric Hodgkin Lymphoma-Treatment Outcome with COPDac/ABVD Combination Chemotherapy – Single Institution Experience
Authors: Ahmed Mehboob , Riaz Shazia , Abbas Syed Nasir , Rehman Fahim
Published: October 2019

Background:

We describe single center experience of using ABVD/COPDac . This regimen was adapted due to perceived excessive toxicity with OEPA/COPDac by treating physicians. Hodgkin lymphoma is being treated with multiple treatment regimens single agent or in combination. MOPP (mechlorethamine, vincristine, procarbazine and prednisolone) has been standard treatment for almost two decads. But it had significant toxic effects including infertility in both genders, myelosuppression and second malignancies . ChlvPP( chlorambucil, vincristine, procarbazine and prednisolone ) has comparable efficacy and toxicities to MOPP . ABVD( Adriamycin, bleomycin, vincristine and doxorubicin ) was mostly used as second line with high activity without permanent male sterility, myelodysplasia or risk of leukemia.

Objective :

We describe single center experience of using ABVD/COPDac . This regimen was adapted due to perceived excessive toxicity with OEPA/COPDac by treating physicians.

Results

62 patients out of 301( 20.6%) with age range of 2-18 years were treated with COPDac/ABVD alternating courses depending on the treatment group from 2012 to date. Out of this 62, 49 (79%) are on follow up and are free from disease relapse or recurrence. Mean duration of follow up is 12 months with a range of 8-16 months. No patient lost to follow up. No death reported due to any cause during or after completion of treatment till last follow up. 9 out of 62 has had progression of disease while on this treatment protocol (15%) and 4 out of 62(6%) has had relapse after completion of the therapy.

Conclusion

In our single center experience of ABVD/COPDac as first line therapy seems effective and safe when compared with historical published literature.

Disclosures

No relevant conflicts of interest to declare.
FCR (Fludarabine, Cyclophosphamide, Rituximab) Followed By 90yttrium Ibritumomab Tiuxetan for the Treatment of Relapsed Grades 1 and 2 Follicular Lymphoma. Long Term Efficacy and Safety Results of 9 Cases
Authors: Pisani Francesco , Dessanti Maria Laura , Giannarelli Diana , Kayal Ramy , Marchesi Francesco , Sciuto Rosa , Rea Sandra , Maini Carlo Ludovico , Marino Mirella
Published: October 2019

Background: This retrospective analysis is focused on the efficacy and safety of radioimmunotherapy (RIT) with Zevalin® in nine patients with recurrent follicular lymphoma (FL) who were treated in a consolidation setting after having achieved complete remission or partial remission with FCR.

Methods: The median age was 63 years (range 46-77), all patients were relapsed with histologically confirmed CD20-positive (grade 1 or 2) FL, at relapse they received FCR every 28 days: F (25mg/m2x 3 days), C (1gr/m2 day 1) and R (375mg/m2 day 4) for 4 cycles. Who achieved at least a partial remission, with<25% bone marrow involvement, was treated with 90Yttrium Ibritumomab Tiuxetan 11.1 or 14.8 MBq/Kg up to a maximum dose 1184 MBq, at 3 months after the completion of FCR. The patients underwent a further restaging at 12 weeks after 90Y-RIT with total body CT scan, FDG-PET/CT and bilateral bone marrow biopsy.

Results: Nine patients have completed the treatment: FCR followed by 90Y-RIT (6 patients at 14.8 MBq/Kg, 3 patients at 11.1 MBq/Kg). After FCR 7 patients obtained CR and 2 PR; after 90Y-RIT two patients in PR converted to CR 12 weeks later. With median follow up of 86 months (range 20-107) since FCR and 79 months (range13-97) since RIT, 3 deaths were observed not related to lymphoma: one for acute renal failure, one for ictus cerebri and one for sepsis; all three deceased patients obtained CR before 90Y-RIT and died still in CR. The median overall survival (OS) and progression free survival (PFS) have not been reached , current analysis has shown that either OS or PFS are 67% at 7 year. The most common grade 3 or 4 adverse events were hematologic, one patient developed herpes zoster infection after 8 months following valacyclovir discontinuation; another patient developed fungal infection.

Conclusions: These results confirm the long term efficacy and safety of 4 cycles of FCR followed by consolidation with 90Y-RIT in patients relapsed with grades 1 and 2 FL and suggest that this regimen could be an option to be used for treatment in this setting of patients, specially at age of 60-75 with no unexpected toxicities.

Figure 1. Progression free survival from RIT Figure 1. Progression free survival from RIT

Disclosures

No relevant conflicts of interest to declare.
Clinical Research on Burkitt Lymphoma—a Single-Center Report in China
Authors: Jiang Meng , Niu Ting
Published: October 2019

Objectives

To analysis the clinical characteristics and discuss the factors that influence the treatment efficacy and prognosis of the patients with Burkitt lymphoma (BL).

Patients and Methods

A retrospective study was performed to collect the clinical characteristics and follow-up data of the patients with BL from January 2004 to December 2014 in West China Hospital.

Results

A total of 42 patients with BL included 32 male patients and 10 female ones,with a median age at diagnosis of 45 yr (range: 13 to 74 yr) old. In the whole cohort, 66.67% of the patients had an Ann Arbor stage III`W. Patients with B symptoms accounted for 50.00%. Gastrointestinal discomfort and axillary, jugular and mandibular mass were the main complaints.

Twenty patients received Hyper-CVAD/HD-MTX-Ara-C regimen (Hyper regimen), while the other 22 patients received non-Hyper regimen, which included CHOP/CHOP-like, CODOX-M/IVAC regimen and others. The 42 cases showed an overall response of 59.52%, with a complete response (CR) rate of 57.14% (24/42).

With univariate analysis: the CR rates of the patients received Hyper regimen and non-Hyper regimen were 75.00% and 40.91%, respectively, indicating a statistical significance between the two groups (P=0.027); the CR rate of the patients with elevated LDH was higher than the patients with normal LDH; the CR rate of patients with ECOG PS of 0`2 was higher than the patients with PS 3`4; the CR rate of the patients with IPI of 0`2 was higher than the patients with IPI of 3`5; for the patients whose LDH increased, the CR rate of the patients received Hyper regimen was higher than the patients with non-Hyper regimen. The OS of the patients who received Hyper regimen was better; the OS of the patients with ECOG of 0`2 was superior to that with ECOG of 3`4; the OS of the patients with IPI of 0`2 is better than that of 3`5; the OS of the patients with Ann Arbor stage of ±`II was higher than that of III`W; the OS of the patients who acquired CR was better than that did not; the OS of the patients without bone marrow involvement was superior to that with it.

In multivariate analysis: treatment regimen was of significance in predicting the probability of CR, whereas treatment regimen and IPI score were confirmed as being highly predictive for OS.

The main side reactions of the 20 cases who accepted Hyper regimen were hematologic toxicity of III`W degree, gastrointestinal discomforts and infections, and there was no treatment-related mortality occurred.

Conclusions:

Hyper regimen could improve the CR rate and prolong the survival of BL patients, and the adverse reactions could well be tolerated.

Disclosures

No relevant conflicts of interest to declare.
Bing-Neel Syndrome: A Rare Complication of Waldenstrom’s Macroglobulinemia
Authors: Sanchez-Guerrero Sergio A. , Treon Steven P. , Guevara-Arnal Luis , Darwich Akram , Cardenas-Ortega Andrea , Castillo Jorge J.
Published: October 2019

A sixty-one year-old Hispanic female with Waldenstrom’s Macroglobulinemia diagnosed in 2011 and successfully treated with 6 monthly cycles of Cyclophosphamide, Rituximab and Dexamethasone (CDR) from 12/11 through 5/12 was then put on a two-year maintenance scheme with Rituximab every three months. In February, 2014 (six months before the end of the planned treatment), she came to the ER complaining with severe headache, aphasia and blurred vision. A stroke was initially ruled out and she received Paracetamol with partial improvement. Nonetheless, symptoms re-appeared accompanied with disorientation and agitation. Antipsychotic medication was given with no improvement. On PE she was disoriented with aphasia, paraparetic and neck stiffness suggestive of meningitis. Blood tests, a MRI and lumbar puncture were performed showing leptomeningeal hyperintensity with no signs of encephalitis (Figure 1).

Figure 1 Leptomeningeal reinforcement as seen in MRI. Figure 1. Leptomeningeal reinforcement as seen in MRI.

CSF analysis showed WBC 64 cells/µL, (95% MNC), glucose= 9.8 mg/dL and proteins= 110 g/dL. Gram dye was negative. A geneXpert for Tuberculosis was negative. CSF cytology showed an infiltration of lymphoid neoplastic cells confirmed by cytochemistry (Figures 2a and 2b).

Figure 2a: CD 20+ and 2b: kappa + neoplastic cells in CSF Figure 2a:. CD 20+ and 2b: kappa + neoplastic cells in CSF Figure 3 Figure 3.

With these results a Bing Neel syndrome was diagnosed and IT Methotrexate was given for a total of 6 doses resulting in a nice reduction of the neoplastic cells. However, she relapsed in April/2014 and IV Fludarabine was started. We are planning to add IT liposomal Cytarabine. Additionally, MYD 88 gene mutation was detected.

DISCUSSION: There are only 33 reported cases of Bing-Neel syndrome in the medical literature for the last 80 years and this one has been confirmed with the newest tools such as: MRI, cytochemistry and gene mutation.

CONCLUSION: Bing-Neel syndrome should be suspected in every patient with Waldenstrom’s Macroglobulinemia and CNS impairment.

Disclosures

No relevant conflicts of interest to declare.
Establishment of a Therapeutic Anti-Pan HLA-Class II Monoclonal Antibody That Directly Induces Lymphoma Cell Death Via Large Pore Formation
Authors: Matsuoka Shuji , Ishii Yasuyuki , Nakao Atsuhito , Masutani Hiroshi , Takahashi Satoshi , Nakauchi Yusuke , Hino Okio
Published: October 2019

To develop a new therapeutic monoclonal Antibody (mAb) for Hodgkin lymphoma (HL), we immunized a BALB/c mouse with live HL cell lines, alternating between two HL cell lines. After hybridization, we screened the hybridoma clones by assessing direct cytotoxicity against a HL cell line not used for immunization. We developed this strategy for establishing mAb to reduce the risk of obtaining clonotypic mAb specific for single HL cell line. A newly established mouse anti-human mAb (4713) triggered cytoskeleton-dependent, but complement- and caspase-independent, cell death in HL cell lines, Burkitt lymphoma cell lines, and advanced adult T-cell leukemia cell lines. Intravenous injection of mAb 4713 in tumor-bearing SCID mice improved survival significantly. mAb 4713 was revealed to be a mouse anti-human pan-HLA class II mAb. Treatment with this mAb induced the formation of large pores on the surface of target lymphoma cells within 30 min. This finding suggests that the cell death process induced by this anti-pan HLA-class II mAb may involve the same death signals stimulated by a cytolytic anti-pan MHC class I mAb that also induces large pores formation. This multifaceted study supports the therapeutic potential of mAb 4713 for various forms of lymphoma.

Disclosures

No relevant conflicts of interest to declare.
Using HDAC Inhibitors to Eradicate Adult T-Cell Leukemia-Lymphoma
Authors: Pimentel Agustin , Reis Isildinha , Toomey Ngoc , Diaz Luis , Ruiz Phillip , Taylor Graham , Ramos Juan Carlos
Published: October 2019

Adult T-cell leukemia-lymphoma (ATLL) is an aggressive malignancy with a poor prognosis caused by HTLV-I, which is endemic in Japan, the Caribbean, and South America. ATLL cannot be cured with conventional chemotherapy thus urging the development of new therapeutic strategies. Histone deacetyalse (HDAC) inhibitors are broadly active anti-neoplastic agents, which have shown efficacy in T-cell lymphomas. At our institution we encounter a relatively high number of ATLL cases as compared to other regions in the U.S. We have conducted a pilot trial using AZT/interferon-α(IFNα) plus the HDAC inhibitor valproic acid (VPA) during maintenance therapy in acute (leukemia-type) ATLL. We hypothesized that VPA would reactivate HTLV-I leading to a cytotoxic T-cell immune response followed by reduction of residual blood circulating ATLL clones that normally persist during AZT/IFNα therapy despite its suppressive effects. Supporting this notion, the addition of VPA to AZT/IFNα resulted in reduction of HTLV-I proviral loads in treated patients and a sustained molecular response in one subject who is still alive >4.5 years later. More recently, we have tested newly available and more potent HDAC inhibitors using fresh primary leukemic ATLL cells and low-passage cell lines. Through HDAC inhibition, these agents reactivated HTLV-I Tax gene expression and induced apoptosis in a dose-dependent manner. Belinostat, which is currently FDA-approved for the treatment of relapsed or refractory peripheral T-cell lymphoma, augmented ATLL cell death when added to AZT/IFNα at nanomolar concentrations. Therefore, we have proposed a new pilot clinical trial using belinostat for the treatment of ATLL during post-induction therapy. The role of HDAC inhibitors in ATLL, pre-clinical laboratory studies involving these agents, and clinical trial design will be discussed at the meeting

Disclosures

Off Label Use: The use of HDAC inhibitors in adult T-cell Leukemia-lymphoma.
Induction of Na+-H+ Ion Exchanger 1 Protein By Heme Oxygenase-1 Plays a Crucial Role in Imatinib-Resistant Chronic Myeloid Leukemia Cells Via p38/MAPK Signaling Pathway
Authors: Wang Jishi , Ma Dan
Published: October 2019

Background: Resistance toward imatinib (IM) and other BCR/ABL tyrosine kinase inhibitors(TKI) remains an increasing clinical problem in the treatment of advanced stages of chronic myeloid leukemia (CML). Heme oxygenase-1 (HO-1), as a Bcr/Abl-dependent survival molecule in CML cells, is important to resist against TKI in patients with freshly diagnosed CML or IM-resistant CML. However, the resistant mechanism on HO-1 up-regulation was still unclear.

Objective: In this study, our findings indicated that over-expression of Na+-H+ion exchanger 1 protein (NHE1) companied with HO-1 up-regulation in CML patients’ mononuclear cells who resisted with IM, while not in whom was sensitive to IM. Therefore, we explored the difference in the signaling pathway related to HO-1 and NHE1 in IM-resistant or sensitive CML cell lines.

Method: CCK-8 kits was used to determine the proliferation inhibition of cells. Flow cytometry was used for analyzing cell proliferation rate and apoptosis. Real-time PCR Array was used to search the relative pathway. Apoptosis relative factors expression were detected by real-time transcription and Western blot.

Result: As a result, we found that up-regulation of NHE1 after transfection of K562 cells line with lentivirus vector, lenti-TOPO-EGFP-HO-1. However, this effect disappeared when HO-1 expression was inhibited by transfection with lenti-U6.2-EGFP-siHO-1. Then, the relationship between HO-1 and NHE1 was detected by qPCR array, activation of p38/MAPK pathway mediated by protein kinase C-β II(PKC-β II) was found. NHE1 wasn’t up-regulated as PKC-β was blocked as well as p38/MAPK pathway was inhibited by SB203580 in K562/IM cells. Meanwhile, the apoptotic rate of the IM-resistant cells line K562 was increased by silencing HO-1, or treatment with SB203580, Enzastaurin, and Cariporide alone.

Conclusion: In this study, we demonstrated that HO-1 relative IM-resistance in CML was involved in the regulation of NHE-1 phosphorylation, which mediated by p38/MAPK signaling pathway activated by PKC-β.

Disclosures

No relevant conflicts of interest to declare.
“FLAG” Regimen Induces Deep, Although Short-Term, Responses in Patients Resistant to Tyrosine Kinases Inhibitors in BP CML.
Authors: Romanova Ekaterina , Girshova Larisa , Siordia Nadia , Goriunova Elena , Motorin Dmitry , Lomaia Elza , Konopleva Marina
Published: October 2019

Background:

Currently the main potential treatment for chronic myeloid leukemia (CML) blast phase (BP) are tyrosine kinase inhibitors. The aim of our study is to try to find any therapy in TKI resistant patients in CML BP.

Material and methods:

The patients were 5 male and one female (age range from 28 to 45 years old). 4 patients progressed to BP from chronic phase (CP), there was rapid progression from AP in other two pts (4 lymphoid, 1 myeloid, 1 undifferentiated). The pts were unresponsible to one or two second generation TKI treatment in BP (dasatinid, nilotinib). Two patients were also failed to respond to high dose cytarabine and methotrexate treatment after TKI failure. Chemotherapy «FLAG» (fludarabine, cytarabine high dose, G-CSF) was used to induce of CP.

Results:

Five of six pts achieved the second CP and deep response (1- CcyR, 3- MMR, and one BCR/ABL reduction to 0,371%).Two of these pts obtained deep response (CCyR, and bcr/abl reduction to 0.371% after unsuccessful treatment with dose mono cytarabine and methotrexat treatment. The response was short, the longest MMR duration is 48 days (patient is being prepared for ASCT).

Conclusion:

FLAG can induce deep molecular responses in BP pts resistant to TKIs. This regimen seems to be more effective than HDAC and HD MTX. The response is of short duration. Thus, in case of absence of relative sibling, haploidentical ASCT with unmanipulated bone marrow and posttransplantation cyclophosphamide could be the optimal treatment choice. The protocol for FLAG induced remission followed haploidentical ASCT is prepared.

Disclosures

No relevant conflicts of interest to declare.
A Phase 2 Study of Bendamustine Plus Melphalan Conditioning for Second Autologous Stem Cell Transplantation in "De-Novo" Multiple Myeloma Patients in a Tandem Transplant Strategy
Authors: Martino Massimo , Giuseppe Messina , Fedele Roberta , Irrera Giuseppe , Giuseppe Console , Gallo Giuseppe Alberto , Vincelli Iolanda Donatella , Ronco Francesca , Gentile Massimo , Morabito Fortunato
Published: October 2019

Background:

High-dose melphalan (HDM) is the standard therapy for autologous stem cell transplantation (ASCT) in multiple myeloma (MM), although the optimal conditioning regimen remains yet to be identified. Bendamustine (BENDA) has a proved activity in hematological malignancies including both first line and relapsed MM.

Methods:

We conducted a phase II trial, adding BENDA to HDM before second ASCT, in a tandem ASCT strategy, in 32 patients with "de-novo" MM. All patients received a bortezomib-based induction therapy. High-dose cyclophosphamide (CY) and G-CSF were used to mobilize stem cells. Four to 6 weeks after the administration of CY, patients received HDM (200 mg/mq), followed by ASCT. Three to 6 months after the first transplantation, patients received a second ASCT with BENDA (200 mg/m2) to HDM (140 mg/m2) as conditioning regimen (BM).

Results: The median age was 56 years (range 40 to 66). Overall, there was no transplant related mortality. The incidence of neutropenic fever and mucositis (grade 1-2) was 46.9% and 81.2%, respectively. No mucositis grade 3-4 was observed. Median number of days to neutrophil and platelet engraftment were 11 and 12, respectively. After the second transplantation, the complete response (CR) improved to 62.5%. Overall response rate was 90.6%. After a median follow-up of 18,2 months, 4 patients had progressed and 1 died. Median progression free survival (PFS) was not reached and actuarial 2-year PFS and OS was 78% and 90%, respectively.

Conclusion: Bendamustine plus melphalan is feasible as conditioning regimen for second ASCT in MM and should be explored for efficacy in a phase III study. Longer follow-up is needed to evaluate conversion rate and survival.

Disclosures

No relevant conflicts of interest to declare.
The Association of Iron Deficiency and Fatigue in Non-Anemic Females of Childbearing Age: An Nhanes Analysis
Authors: Akkad Mohamed , Shaik Mohammed , Al-Janadi Anas
Published: October 2019

Introduction:

Iron deficiency (ID) is the most common nutritional deficiency in the world and remains relatively common in at-risk groups in the United States. The development of (ID) is dependent upon the individual's iron stores which is a balance between iron absorption and loss. Commonly associated with (ID) is iron deficiency anemia (IDA) which has a well known association with fatigue. Fatigue is a common complaint in general, with a prevalence in population-based surveys in Great Britain and the United States of 6.0 and 7.5 % respectively. However it is not uncommon to have (ID) without subsequent anemia in females childbearing age group. Our study aims to evaluate the prevalence of ID in non anemic females of childbearing age and its association with fatigue.

Methods:

Data was obtained from continuous NHANES database, a nationally representative health survey conducted from 2005-2010. The data including age, race, complete blood count (CBC), ferritin and fatigue was obtianed from laboratory and questionnaire datasets, respectively. Subjects were further divided based on ferritin levels into two groups, Group A; ferritin ≤20 ng/mL and Group B; ferritin >20 ng/mL. The question regarding fatigue- "feeling tired or having little energy for past two weeks" was obtained from PHQ-9 scale. Univariate analysis was done comparing Group A and Group B using chi-square test and t-test for categorical and numerical variables, respectively. The effect of iron deficiency on fatigue was obtained using logistic regression to calculate odds ratio (OR) after adjusting for age and race.

Results:

Of 3230 female subjects( age 18yrs -49yrs), there were 809 with ID with no anemia. The weighted prevalence of iron deficiency in non-anemic women of childbearing age was 22.5% (95% CI 20.76-24.22). The weighted prevalence of fatigue in Group A and Group B was 61.8% (95% CI 56.4-67.2) and 59.7 % (95%CI 57-62.4) respectively (p=0.45)(table 1). In multivariate analysis, after adjusting for age and race, the odds ratio (OR) for fatigue with iron deficiency was 1.1 ( 95% CI 0.8-1.4).

Conclusion:

The prevalence of ID in non anemic females of childbearing age in the United States was 22.5 % .The percentage of subjects who reported fatigue was significantly high in both iron deficient and non-iron deficient subjects in this age group. There was no statistical difference in self-reported fatigue in both iron deficient and non-iron deficient women of childbearing age. Further studies are needed to validate our findings.

Table 1. Comparision of GpA and GpB Variables Ferritin≤ 20 ng/mL (GpA) Ferritin > 20 ng/mL Fatigue - Yes 512 1452 Fatigue - No 297 986 Total 809 2421

Disclosures

No relevant conflicts of interest to declare.
The Appropriateness of Heparin Antibody Testing. A Single Institution Retrospective Analysis
Authors: Landau Danny A. , Kholodnaya Nadezhda , CuestaFernandez Ana , PerezPerez Ariel
Published: October 2019

Background: Heparin-induced thrombocytopenia (HIT) is a drug-induced, immune-mediated prothrombotic disorder associated with thrombocytopenia and venous and/or arterial thrombosis.Up to 5% of patients exposed to heparin for at least one week develop HIT, and approximately 50% of them will have thrombosis. Diagnosis of HIT is suspected from the clinical picture based on the ''4 T's'' (Thrombocytopenia, Timing, Thrombosis, no other cause of platelet fall) or the HIT Expert Probability (HEP) scoring system. However, often these "4 T's" are ignored and testing is inappropriately ordered in low risk patients. This could lead to possible morbidity and increase length of hospital stay. We opted to look back at the appropriateness of testing done within our academic center.

Methods/ Design:

All hospitalized patients with thrombocytopenia who underwent HIT antibody testing (HIT ELISA) during February 2013 were screened using our internal electronic medical record.

Patients were subdivided to low, intermediate or high pretest probability group according to the 4Ts scoring system.

Appropriateness of testing was determined according ASH 2013 Clinical Practice Guideline on the Evaluation and Management of Adults with Suspected Heparin-Induced Thrombocytopenia (HIT). HIT Ab test is recommended to be tested when there is intermittent to high probability of HIT (4T's score above or equal to 4).

The percentage of appropriate/inappropriate testing was calculated.

The results were then subdivided by ordering physician group.

Results:

120 HIT ab tests were performed during February 2013 in our institution. Only 13 patients tested had a positive HIT Ab. Internal medicine ordered the majority of these tests. Of the 120 patients tested, 7 patients had high risk and another 50 had intermediate risk as per 4T's scoring. 61 patients were low risk and should not have been tested. (Table 1)

Table 1. HIT antibody test ordered per Service and probability of HIT. Service ordering test Low probability of HIT Intermediate/high probability of HIT TOTAL Internal Medicine 28 28 56 Cardiothoracic Surgery 13 11 24 Hematology Oncology 7 5 12 Cardiology 3 1 4 Surgery 1 4 5 Emergency medicine 0 1 1 Infectious disease 4 0 4 Pulmonary 0 1 1 Critical care 1 6 7 Nephrology 1 2 3 Ob Gyn 1 0 1 Family Medicine 2 0 2 TOTAL 61 59 120

Conclusions:

HIT testing has been overused within our institution. Low platelets, without other signs or symptoms of typical HIT were used as a trigger for testing the antibody. This has potential to cause harm due to increased bleeding risk, increased length of hospital stay, and the potential to utilize extended anticoagulation when not necessary. Better education on the appropriateness of the test can limit the potential harm of its use.

Disclosures

No relevant conflicts of interest to declare.
To Push or Not to Push: The Benefit of Administering Anti-Emetics By Intravenous Push
Authors: Arumugam Raajasekar Arun Kumar , Barola Sindhu , Tehrani Lynne , Chandra Abhinav B
Published: October 2019

Background

Prevention of chemotherapy-induced nausea and vomiting is crucial for maintaining the quality of life of cancer patients. Granisetron and dexamethasone are commonly used medications prior to chemotherapy to minimize these adverse effects. There are no well-defined guidelines on the route of administration of medications that are administered prior to chemotherapy. With the ever increasing cost of cancer care, innovative methods need to be employed to maximize utilization of resources.

Methods

A retrospective chart review was performed to identify patients who received granisetron and dexamethasone prior to emetogenic chemotherapy. The primary study site was the cancer center of a tertiary care community hospital. Over a two month period there were 1552 instances when patients received dexamethasone or granisetron prior to emetogenic chemotherapy. Patients either received medications by intravenous piggyback (IVPB) or intravenous push (IVP). The time spent on delivering medication by the two routes and the rate of occurrence of nausea and vomiting requiring the administration of rescue medications were compared.

Results

Patients who received granisetron and dexamethasone by IVPB had a 2.8% rate of adverse effect rate requiring rescue medications (n=806) and patients who received the medication by IVP had a 2.9% rate of adverse effects requiring rescue medications (n=746). There was no statistical significance in the occurrence of nausea and vomiting requiring rescue medications between the two groups (p>0.80). The total average duration for administration of granisetron and dexamethasone by IVPB was 30 minutes and two minutes by IVP. The average cost of an IVP is $57 as compared to $161 for IVPB based on national average costs.

Conclusions

An average of 28 minutes of chemotherapy chair time is saved on each patient for each session of chemotherapy by administering granisetron and dexamethasone by IVP. Approximately $167,000 would have been saved if the above IVPB group received medications by IVP. This sum excludes nursing costs, chemotherapy chair costs and other disposable charges. The route of administration of the medications did not affect the rate of occurrence of adverse effects requiring rescue medications. The utilization of the intravenous push route will save the patient time and reduce nursing hours without compromising on the effectiveness of the drugs while adding to sizeable cost savings.

Disclosures

No relevant conflicts of interest to declare.
Molecular Diagnosis of Hereditary Spherocytosis By Multi-Gene Target Sequencing in Korea
Authors: Choi Qute , Kim Jung Ah , Im Kyong Ok , Park Si Nae , Park Yoomi , Kim Ju Han , Jung Hye Lim , Kim Seon Young , Shin Hee Young , Lee Dong Soon
Published: October 2019

Background: Hereditary spherocytosis (HS) is the most common cause of hereditary hemolytic anemia. Spherocytes formed by defective membrane proteins are selectively captured in the spleen and destroyed, leading to hemolytic anemia. Current tests used to diagnose HS focus on the detection of hemolysis or indirectly assess protein defects. Direct methods to detect protein defects are complicated and difficult to implement. Recent next-generation sequencing (NGS) methods enable large-scale gene mutation analyses to be used for such diagnoses. In this study, we investigated the patterns of genetic variation associated with HS to determine the molecular mechanisms underlying the condition. Specifically, we analyzed mutations in red blood cell membrane protein-encoding genes in Korean HS patients using NGS.

Methods: In total, 60 patients with HS were enrolled in this study. Targeted sequencing of 43 genes (17 membrane protein-encoding genes, 20 enzyme-encoding genes, and 6 additional candidate genes) was performed using the Illumina HiSeq platform and variants were called according to a data-processing pipeline.

Results: Of the 60 patients, 50 (83%) had one or more significant variants in a membrane protein gene. A total of 54 significant variants (8 previously reported and 46 novel) were detected in 6 membrane protein-encoding genes, i.e., SPTB, ANK1, SPTA1, SLC4A1, EPB41, and EPB42. The most variants (28) were detected in SPTB. Four significant variants, all of which were previously reported, were detected in genes encoding enzymes (ALDOB, G6PD, GAPDH, and GSR). Additionally, 5 previously reported variants were detected in UGT1A1. These results suggest 35 primer sets that can be used to diagnose HS.

Conclusion: This was the first large-scaled genetic study of Korean HS patients. These results clarify the pattern of genetic variation associated with HS in Korean patients. They will enable easier, more rapid diagnosis of HS.

Disclosures

No relevant conflicts of interest to declare.
Gene Editing with Crispr-Cas9 for Treating Beta-Hemoglobinopathies
Authors: Lee Ciaran , Bao Gang , Porteus Matthew H. , Cornu Tatjana , Miccio Annarita , Cradick Thomas , Cathomen Toni , Lundberg Ante , Mavilio Fulvio
Published: October 2019

CRISPR/Cas9 systems provide a powerful tool for precision genome editing due to their highly efficient targeting of specific DNA sequences in a genome. We have designed CRISPR single-guide RNAs (sgRNAs) containing the guide and tracr RNAs to selectively edit the human globin locus for potential therapeutic benefit in treating sickle cell anemia and beta-thalassemia. Multiple sgRNAs have been identified and tested across key sites in the human globin locus, and the gene modification frequencies quantified in 293T, K562 and CD34+ hematopoietic progenitor cells. The nuclease cutting efficiency was determined by the rate of insertions, deletions or mutations (indels) from NHEJ-directed repair. The indel rates for these sgRNAs ranged from <1% to ~70%. We found that there was only a weak correlation between indel rates and deletion frequencies, and the gene modification efficiency varied between cell lines/cell types. Although bioinformatics tools are available for sgRNA design, determining the optimal sgRNAs is still largely empiric and system-dependent, thus better bioinformatics tools are needed. Our results demonstrate that, with careful design and optimization of sgRNAs, efficient genome editing can be achieved with high specificity, which promises effective treatment of beta-hemoglobinopathies.

Disclosures

Porteus: CRISPR Therapeutics: Consultancy, Equity Ownership. Cornu:CRISPR Therapeutics: Research Funding. Miccio:CRISPR Therapeutics: Research Funding. Cradick:CRISPR Therapeutics: Employment. Cathomen:CRISPR Therapeutics: Consultancy, Research Funding. Lundberg:CRISPR Therapeutics: Employment. Mavilio:CRISPR Therapeutics: Consultancy, Research Funding.
Successful Therapy of a Patient with a Novel STAT1 Gain of Function Mutation and Life-Threatening Cytopenias with Janus Kinase Inhibitor Ruxolitinib
Authors: Weinacht Katja G. , Charbonnier Louis m , Plant Ashley , Torgerson Troy , Rosenzweig Sergei , Fleisher Thomas , Notarangelo Luigi Daniele
Published: October 2019

Monoallelic gain of function (GOF) mutations in the human Signal Transducer and Activator of Transcription 1 (STAT1) manifest in immunodeficiency and autoimmunity. The immunodeficiency is caused by impaired IL-17 immunity and typically presents with chronic mucocutaneous candidiasis, variably associated with other opportunistic infections. The autoimmune manifestations due to exaggerated responsiveness to interferon are more diverse and include SLE, thyroiditis, hepatitis, and alopecia areata. The phenotypic spectrum of STAT1 GOF mutations is extremely wide, ranging from intermittent localized disease to chronic systemic illness and fatality. Allogeneic bone marrow transplantation has been attempted in severely affected patients but outcomes have been poor, leaving a void for alternate long-term strategies to control the disease and maintain remission. Recently, we diagnosed a patient suffering from chronic mucocutaneous candidiasis and life-threatening autoimmune cytopenias with a novel monoallelic mutation in the linker domain of STAT1 (c.1633G>A; p.E545K). The mutation caused a profound increase in STAT1 phosphorylation in response to type 1 and 2 interferon without affecting dephosphorylation kinetics, which is mechanistically distinct from all reported STAT1 GOF mutations to date. The potential of naïve patient CD4+ T cells to become IFN- γ or IL-17 producing cells was investigated under TH0 (anti-CD3/28), TH1 (anti-CD3/28, IL-12) and TH17 (anti-CD3/28, IL6, IL-23, TGF-β1) conditions and revealed that patient CD4+ T cells are biased to produce IFN-γ irrespective of polarizing conditions, and resistant to TH17 induction upon stimulation with IL6, IL-23 and TGF-β1. In addition, the patient's proportion of T follicular helper cells (TFH) relative to regulatory T cells (Treg) was increased. Treatment with the Janus Kinase (JAK) 1/2 inhibitor Ruxolitinib reduced the hyperresponsiveness to interferon, normalized the TH1 response, enabled naïve T cells to differentiate into TH17 cells and decreased the TFH to Treg ratio. Under therapeutic dose Ruxolitinib the patient maintained clinical remission with respect to both autoimmunity and mucocutaneous candidiasis.

Conclusion: Clinical vigilance for an underlying immune dysregulation syndrome due to abnormal JAK-STAT signaling is critical when evaluating patients with autoimmunity combined with opportunistic infections as JAK-inhibitors represents an effective targeted therapy for long-term disease control even in severely affected patients for whom hematopoietic stem cell transplantation is not available.

Disclosures

Off Label Use: Ruxolitinib use for STAT1 GOF mutation.
Standardized Ultrasonography (HEAD-US) of Joints and First Correlation with Function in Haemophilic Arthropathy: Results of a Clinical Trial and Potential Importance of Joint-Ultrasound for an Individualized Prophylaxis in Haemophiliacs
Authors: Sigl-Kraetzig Michael , Bauerfeindt Stefan , Wildner Amanda , Seuser Axel
Published: October 2019

Background: Sonography is applied in patients with bleeding disorders. By ultrasonography the extent of synovitis is detected as a sign of the activity of an existing hemophilic arthropathy, as a sign of progression are defects of articular cartilage and bony structures quantifiable. In 2013 an easy-to-use standardized ultrasound protocol (HEAD-US) for examination of early joint changes in Hemophilic Arthropathy was published. Correlation between functional and structural changes in Haemophilic Arthropathy was not published before.

Aims: The sonographic quantification of the arthropathy by haemophilia-treaters could be the basis for better individualized haemophilia therapy in young patients.

Methods: For this purpose, highly reproducible sonographic standard section planes and an examiner-independent ultrasound score (HEAD -US) were published by C. Martinoli in 2013. After a preliminary study of 27 patients now we have included 177 young german patients (children and young adults) with haemophilia A, haemophilia B or von-Willebrand- disease from different German hemophilia treatment centers in the HaemarthroSonoPilot study (DRKS00004483).

After informed consent sonography of the elbow, knee and ankle joints with a Zonare " z.one ultra " ultrasound device (Linear probe L14 -5w ) was performed. Simultaneously an orthopedic clinical examination with clinical scoring and 3D motion analysis of the lower limbs for detecting early function defects were performed with an ultrasonic topometer (developed by the University of Bonn, Germany).

Results: Already in the previous investigation of the pilot study on a small cohort yielded promising results depending on age. The investigations in the presented pilot study showed correlation of the sonographic diagnostics with the measurement of a clinical orthopedic examination in haemophilic arthropathy. Through the joint sonography changes were even partially already seen before that stood out in the clinical investigation.

Conclusion: It may be useful if haemophilia treaters in future apply this method to individualize the therapy under close control and evaluation of joint changes.

Disclosures

Sigl-Kraetzig: Pfizer Deutschland GmbH: Research Funding. Bauerfeindt:Pfizer Deutschland GmbH: Honoraria, Research Funding. Seuser:BAXALTA Deutschland GmbH: Honoraria, Research Funding.
Molecular Basis for Distinct, Antagonistic, Regulatory Functions of Ribosomal Protein Paralogs Rpl22 and Rpl22-Like1 (Like1) in Hematopoiesis
Authors: Wang Minshi , Zhang Yong , Rao Shuyun , Rhodes Jennifer , Rhodes Michele
Published: October 2019

The prevailing view of ribosomal proteins (RP) is that they act to facilitate the core function of the ribosome in synthesizing proteins. However, emerging evidence suggests that some RP perform regulatory functions outside of the context of the ribosome ("extra-ribosomally"). Indeed, our laboratory demonstrated that ribosomal protein, Rpl22 and its paralog Rpl22l1 (Like1), are dispensable for ribosome assembly and translation, yet they perform critical regulatory functions in hematopoiesis that are mediated by binding selected RNA targets and controlling their translation. Despite their high degree of homology, Rpl22 and Like1 antagonistically control the emergence of hematopoietic stem cells (HSC) by regulating the translation of the key transcription factor, Smad1, with Rpl22 repressing its translation and Like1 acting to oppose that translation. Here, we seek to establish the molecular basis for how such highly homologous proteins perform distinct and opposing functions. Since both Rpl22 and Like1 share highly conserved RNA-binding domains in the central core of the proteins, we hypothesize that their unique regulatory functions are likely to reside in their termini. To systematically test this hypothesis, we employed a domain-swapping approach, creating a series of Rpl22:Like1 chimeric proteins to assess which changes would confer on Like1, the Rpl22 function of blocking HSC emergence. In doing so, we utilized zebrafish as a model organism. During embryonic zebrafish development, the enforced expression of Rpl22, but not Like1, represses Smad1 expression and blocks HSC) emergence. Interestingly, our analysis of the Rpl22:Like1 chimeras revealed that transfer of the N-terminal amino acids of Rpl22 to Like1 was sufficient to confer upon Like1 the Rpl22 function of blocking HSC emergence. Moreover, this Rpl22:Like1 N-terminal chimera also was able to restore the block in T lymphocyte development observed upon loss of Rpl22, indicating that Rpl22 function in another hematopoietic process, supporting T lymphocyte development, can be conferred upon Like1 by the N-terminal sequences of Rpl22. Together, these data suggest that the unique and opposing functions of Rpl22 and Like1 reside in their N-termini. How these newly identified N-terminal amino acid sequences confer distinct biological functions on these two paralogs is unclear at present, but may involve the association with distinct co-factors. This possibility is currently under investigation. This study demonstrates that relative minor changes in amino acid sequence can have profound effects on the functions of these RP paralogs, enabling them to play divergent regulatory roles in controlling hematopoiesis.

Disclosures

No relevant conflicts of interest to declare.