scholarly journals Super Resolution Optical Microscopy for Analysis of Granules in B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5347-5347
Author(s):  
Rio Barrere-Cain ◽  
Sebastian Wachsmann-Hogiu ◽  
Denis M. Dwyre
Keyword(s):  
Bone Marrow ◽  
Surface Area ◽  
Bone Marrow Aspirate ◽  
Lymphoblastic Leukemia ◽  
Average Diameter ◽  
Maximum Intensity ◽  
Average Depth ◽  
Average Surface Area ◽  
High Resolution Images ◽  
Cell Acute Lymphoblastic Leukemia

Abstract An unusual case of B-cell acute lymphoblastic leukemia (B-ALL) was diagnosed at our institution. The B-ALL had unusually large prominent granules, but the diagnosis was confirmed by flow cytometry. Using structured illumination microscopy (SIM), we obtained high resolution images of the granules in the lymphoblasts. With these high resolution images, we acquired detailed quantitative information on the morphology of these unusual cells. We compared the morphology of this unusual B-ALL case with granular acute myeloid leukemia (AML) cases, standard B-ALL cases, and non-leukemic marrow aspirate specimens. Granular B-ALL can sometimes be misdiagnosed as AML, which would result in patients not receiving the optimal treatment. Our goal is to study the structure of these unusual B-ALL cases. More information characterizing the granules may assist in understanding how these leukemic lymphoblasts function. Granules in B-ALL are believed to be abnormal organelle formations or fusions. Electron microscopy (EM) reveals that the granules resemble membrane bound vesicles and it has been suggested that they are atypical mitochondria, lysosomes, or damaged chromosomes. These suggestions have been based on EM images and the cytochemical profile of the cells (Pitman et al., 2007; Cap et al., 2000). To our knowledge, granular B-ALL lymphoblasts have never been imaged with SIM. We took images of the same cells using both brightfield light microscopy and SIM. Figures 2 and 3 show the same cells imaged with both microscopes. We made maximum intensity Z projections (the pixels in stack of images with maximum intensity projected onto one image), cutting off Z-slices where the cell was not focused in order to avoid measuring granules outside the cell. Next, we randomly selected granules to measure in each cell's Z projection. Using Image J software, we measured the surface area, diameter, depth, intensity relative to background, and shortest distance to nucleus and plasma membrane for each individual granule. We then averaged the measurements from each type of sample (granular B-ALL, AML, non-leukemic). The granules in B-ALL had an average diameter of .285 micrometers, average depth of .506 micrometers, average surface area of .147 micrometers squared, and were on average 2.74 times more intense than the background. The granules in AML had an average diameter of .479 micrometers, average depth of .440 micrometers, average surface area of .2255 micrometers squared, and were on average 1.953 times more intense than the background. The B-ALL granules were smaller and more intense. The stain used for the cytoplasm, eosin, is acidic. The brighter B-ALL granules suggest that they have higher concentrations of basic proteins than the AML granules. This technique may be useful in separating granules in AML leukemia, ALL leukemias, and possibly leukemias of ambiguous lineage. Other techniques we are investigating include automated count of granules and granule characterization with Raman Spectroscopy to determine chemical composition. Evaluating the cells with fluorescent probes that specifically attach to either lysosomes or mitochondria may give additional biochemical information about the granules/leukemias. Matching fluorescent signals from these probes to granules in ALL may assist in determining whether the granules are lysosomal or mitochondrial in origin. Figure 1: Granular B-ALL bone marrow aspirate sample imaged with SIM and converted to a maximum intensity Z projection, arrow indicates a granule Figure 1:. Granular B-ALL bone marrow aspirate sample imaged with SIM and converted to a maximum intensity Z projection, arrow indicates a granule Figure 2: AML bone marrow aspirate sample imaged with SIM and converted to a maximum intensity Z projection, arrow indicates a granule Figure 2:. AML bone marrow aspirate sample imaged with SIM and converted to a maximum intensity Z projection, arrow indicates a granule Figure 3: AML bone marrow aspirate sample imaged with a brightfield light microscope and magnified 100 times, arrows indicate granules Figure 3:. AML bone marrow aspirate sample imaged with a brightfield light microscope and magnified 100 times, arrows indicate granules Disclosures No relevant conflicts of interest to declare.

Successful Treatment of Refractory Acute Lymphoblastic Leukemia with Tisagenlecleucel in the Setting of Life-Threatening Infection

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5125-5125
Author(s):  
Keith J August ◽  
Terrie Flatt ◽  
Erin Marie Hall ◽  
Doug Myers
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Bone Marrow Aspirate ◽  
Lymphoblastic Leukemia ◽  
Severe Neutropenia ◽  
Active Infection ◽  
Post Infusion ◽  
Life Threatening ◽  
Cell Acute Lymphoblastic Leukemia

Tisagenlecleucel is a CD19 directed immunotherapy approved for the treatment of young patients with relapsed or refractory precursor B-cell acute lymphoblastic leukemia (ALL). The most important toxicity related to tisagenlecleucel therapy is cytokine release syndrome (CRS). CRS is an exaggerated systemic inflammatory response that occurs frequently along with T-cell expansion following the administration of tisagenlecleucel. Tisagenlecleucel guidelines recommend delaying treatment when an active infection is present due to the concern that the pre-existing inflammatory response associated with infection may predispose patients to severe CRS. We describe two cases where tisagenlecleucel was successfully administered to patients in the setting of life-threatening infection. Patient 1 is a 23-year-old Caucasian male with refractory Philadelphia chromosome negative B-cell ALL who had received prior treatment with chemotherapy, blinatumomab, inotuzumab ozogamicin and a haploidentical stem cell transplant (SCT) followed by multiple Zalmoxis infusions. Five months following SCT he relapsed. At relapse, he underwent leukapheresis followed by bridging chemotherapy with ifosfamide and etoposide. He developed severe neutropenia and respiratory failure associated with a right lower lung consolidation. A biopsy demonstrated a mucormycosis infection and he required surgical debridement including resection of portions of the lung, diaphragm and liver. At this time, he had 92% blasts in the bone marrow. Eleven days after his surgery he received tisagenlecleucel despite being persistently febrile. Prior to the infusion, he received a modified lymphodepleting chemotherapy regimen including two days of fludarabine and cytarabine. Due to severe neutropenia, he was receiving granulocyte transfusions. These were discontinued prior to the infusion and resumed after 12 days. CRP and ferritin the day prior to infusion were 26.2 mg/dL and 18,419 ng/mL. He remained persistently febrile for 13 days post-infusion. He received a single dose of tociluzimab 7 days following his infusion due to high fevers. He did not require any treatment with corticosteroids for CRS. The absolute neutrophil count recovered to >500x103/µL at 31 days post infusion. A bone marrow aspirate done 26 days post-infusion did not show any evidence of leukemia by multicolor flow cytometry (MFC). He remains alive without evidence of disease 11 months after treatment with tisagenlecleucel. Patient 2 is a 4-year-old Hispanic female with refractory B-cell ALL found to have a TP53 deletion and t(1;19). She had received prior treatment with chemotherapy, blinatumomab and local radiation therapy to the site of extramedullary disease found in the left maxillary sinus at relapse. She underwent leukapheresis and received bridging chemotherapy with mercaptopurine and methotrexate. After 3 days of lymphodepleting chemotherapy she developed septic shock and E. Coli bacteremia. She became severely ill requiring continuous renal replacement therapy for 5 days and extracorporeal membrane oxygenation (ECMO) for 6 days. Shortly after ECMO decannulation she developed fever and was found to have multiple pulmonary opacities concerning for fungal infection. Blasts were noted in the peripheral blood. Sixteen days after presenting with septic shock and 11 days from ECMO she received tisagenlecleucel. CRP at the time of infusion was 22.9 mg/dL. She developed persistent fevers post-infusion for 17 days. She received two doses of tociluzimab 20- and 21-days post-infusion due to recurrence of high fever and reactive lymphadenopathy with third spacing and concern for renovascular compromise. She did not require any treatment with corticosteroids for CRS. Bone marrow aspirate done 32 days post-infusion did not show any evidence of leukemia by MFC. The absolute neutrophil count recovered to >500x103/µL at 59 days post infusion. The patient remained without evidence of disease for 7 months following treatment but died due to infectious complications with persistent pancytopenia. Tisagenlecleucel is a potentially life-saving treatment for relapsed and refractory B-cell acute lymphoblastic leukemia in children and young adults 24 years of age or younger. Tisagenlecleucel is an option for the treatment of patients with active infection and/or inflammation with progressive leukemia when no other therapeutic alternative exists. Disclosures August: Novartis Pharmaceuticals: Speakers Bureau. Myers:Novartis Pharmaceuticals: Consultancy, Speakers Bureau.

scholarly journals Negative initial bone marrow aspirate does not rule out acute lymphoblastic leukemia

2020 ◽  
Author(s):  
Monica Davini ◽  
Kevin Shiah ◽  
Anurag Agrawal
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
Bone Marrow Aspirate ◽  
Lymphoblastic Leukemia ◽  
Definitive Diagnosis ◽  
Abnormal Bone Marrow ◽  
Institutional Experience ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Marrow Cellularity ◽  
Rule Out

Patients with precursor-B-cell acute lymphoblastic leukemia (B-ALL) may initially present with a prodrome, cytopenia(s) with abnormal bone marrow cellularity, but without clonal abnormalities. Prior cases of “indolent ALL” report infections preceding B-ALL diagnosis. Here we describe our institutional experience, eight patients over a 15-year period with a prodrome (2% of B-ALL diagnoses) prior to definitive diagnosis. Patients ranged from 3-15 years of age (median 5 years), requiring a median 3.5 months from presentation to diagnosis, with a median 3 bone marrow aspirates (BMA) to reach definitive diagnosis. Practitioners must be aware that initial negative BMA does not rule out B-ALL.

scholarly journals Tumor staging in a Beagle dog with concomitant large B-cell lymphoma and T-cell acute lymphoblastic leukemia

2021 ◽  
pp. 104063872110110
Author(s):  
Alessandro Ferrari ◽  
Marzia Cozzi ◽  
Luca Aresu ◽  
Valeria Martini
Keyword(s):  
Bone Marrow ◽  
Lymph Node ◽  
T Cell ◽  
B Cell ◽  
Cell Lymphoma ◽  
Tumor Staging ◽  
Lymphoblastic Leukemia ◽  
B Cell Lymphoma ◽  
Large B Cell Lymphoma ◽  
Cell Acute Lymphoblastic Leukemia

An 8-y-old spayed female Beagle dog was presented with peripheral lymphadenomegaly. Lymph node cytology and flow cytometry led to the diagnosis of large B-cell lymphoma (LBCL). We detected minimal percentages of LBCL cells in peripheral blood and bone marrow samples. However, a monomorphic population of neoplastic cells different from those found in the lymph node was found in the bone marrow. T-cell acute lymphoblastic leukemia was suspected based on flow cytometric immunophenotyping. PCR for antigen receptor rearrangement (PARR) revealed clonal rearrangement of both B-cell and T-cell receptors, and the presence of both neoplastic clones in the lymph node, peripheral blood, and bone marrow. The dog was treated with multi-agent chemotherapy but died 46 d following diagnosis. Tumor staging and patient classification are needed to accurately establish a prognosis and select the most appropriate therapeutic protocol.

scholarly journals Co-culture model of B-cell acute lymphoblastic leukemia recapitulates a transcription signature of chemotherapy-refractory minimal residual disease

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Stephanie L. Rellick ◽  
Gangqing Hu ◽  
Debra Piktel ◽  
Karen H. Martin ◽  
Werner J. Geldenhuys ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Tumor Cells ◽  
Minimal Residual Disease ◽  
Residual Disease ◽  
Lymphoblastic Leukemia ◽  
Adherent Cells ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Minimal Residual

AbstractB-cell acute lymphoblastic leukemia (ALL) is characterized by accumulation of immature hematopoietic cells in the bone marrow, a well-established sanctuary site for leukemic cell survival during treatment. While standard of care treatment results in remission in most patients, a small population of patients will relapse, due to the presence of minimal residual disease (MRD) consisting of dormant, chemotherapy-resistant tumor cells. To interrogate this clinically relevant population of treatment refractory cells, we developed an in vitro cell model in which human ALL cells are grown in co-culture with human derived bone marrow stromal cells or osteoblasts. Within this co-culture, tumor cells are found in suspension, lightly attached to the top of the adherent cells, or buried under the adherent cells in a population that is phase dim (PD) by light microscopy. PD cells are dormant and chemotherapy-resistant, consistent with the population of cells that underlies MRD. In the current study, we characterized the transcriptional signature of PD cells by RNA-Seq, and these data were compared to a published expression data set derived from human MRD B-cell ALL patients. Our comparative analyses revealed that the PD cell population is markedly similar to the MRD expression patterns from the primary cells isolated from patients. We further identified genes and key signaling pathways that are common between the PD tumor cells from co-culture and patient derived MRD cells as potential therapeutic targets for future studies.

scholarly journals High cortactin expression in B-cell acute lymphoblastic leukemia is associated with increased transendothelial migration and bone marrow relapse

Leukemia ◽  
2018 ◽  
Vol 33 (6) ◽  
pp. 1337-1348 ◽  
Author(s):  
Martha Velázquez-Avila ◽  
Juan Carlos Balandrán ◽  
Dalia Ramírez-Ramírez ◽  
Mirella Velázquez-Avila ◽  
Antonio Sandoval ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Transendothelial Migration ◽  
Cell Acute Lymphoblastic Leukemia

scholarly journals Bone marrow niche-derived extracellular matrix-degrading enzymes influence the progression of B-cell acute lymphoblastic leukemia

Leukemia ◽  
2020 ◽  
Vol 34 (6) ◽  
pp. 1540-1552 ◽  
Author(s):  
Divij Verma ◽  
Costanza Zanetti ◽  
Parimala Sonika Godavarthy ◽  
Rahul Kumar ◽  
Valentina R. Minciacchi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Extracellular Matrix ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cell ◽  
Bone Marrow Niche ◽  
Significant Prolongation ◽  
Tnf Α ◽  
Cell Acute Lymphoblastic Leukemia

AbstractSpecific and reciprocal interactions with the bone marrow microenvironment (BMM) govern the course of hematological malignancies. Matrix metalloproteinase-9 (MMP-9), secreted by leukemia cells, facilitates tumor progression via remodeling of the extracellular matrix (ECM) of the BMM. Hypothesizing that leukemias may instruct the BMM to degrade the ECM, we show, that MMP-9-deficiency in the BMM prolongs survival of mice with BCR-ABL1-induced B-cell acute lymphoblastic leukemia (B-ALL) compared with controls and reduces leukemia-initiating cells. MMP-9-deficiency in the BMM leads to reduced degradation of proteins of the ECM and reduced invasion of B-ALL. Using various in vivo and in vitro assays, as well as recipient mice deficient for the receptor for tumor necrosis factor (TNF) α (TNFR1) we demonstrate that B-ALL cells induce MMP-9-expression in mesenchymal stem cells (MSC) and possibly other cells of the BMM via a release of TNFα. MMP-9-expression in MSC is mediated by activation of nuclear factor kappa B (NF-κB) downstream of TNFR1. Consistently, knockdown of TNF-α in B-ALL-initiating cells or pharmacological inhibition of MMP-9 led to significant prolongation of survival in mice with B-ALL. In summary, leukemia cell-derived Tnfα induced MMP-9-expression by the BMM promoting B-ALL progression. Inhibition of MMP-9 may act as an adjunct to existing therapies.

scholarly journals JAK3 mutants transform hematopoietic cells through JAK1 activation, causing T-cell acute lymphoblastic leukemia in a mouse model

Blood ◽  
2014 ◽  
Vol 124 (20) ◽  
pp. 3092-3100 ◽  
Author(s):  
Sandrine Degryse ◽  
Charles E. de Bock ◽  
Luk Cox ◽  
Sofie Demeyer ◽  
Olga Gielen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
Mouse Model ◽  
Lymphoblastic Leukemia ◽  
Marrow Transplant ◽  
Selective Inhibitor ◽  
Mouse Bone Marrow ◽  
Key Points ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Transplant Model

Key Points JAK3 pseudokinase mutants require JAK1 for their transforming potential. JAK3 mutants cause T-ALL in a mouse bone marrow transplant model and respond to tofacitinib, a JAK3-selective inhibitor.

scholarly journals Bone marrow sites differently imprint dormancy and chemoresistance to T-cell acute lymphoblastic leukemia

2017 ◽  
Vol 1 (20) ◽  
pp. 1760-1772 ◽  
Author(s):  
Xavier Cahu ◽  
Julien Calvo ◽  
Sandrine Poglio ◽  
Nais Prade ◽  
Benoit Colsch ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Lymphoblastic Leukemia ◽  
Key Points ◽  
Cell Acute Lymphoblastic Leukemia

Key Points BM niches differentially support T-ALL. BM niches differentially protect T-ALL cells from chemotherapy.

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