Initial Bone Marrow Biopsy Research Articles

Vogt‐Koyanagi‐Harada‐like findings as the first manifestation of neuroleukaemia

Aims/Purpose: To report a well‐documented case of a 62‐year‐old woman presenting with VKH‐like findings as the first manifestation of neuroleukaemia.Methods: Case study.Results: A 52‐year‐old woman who had received a diagnosis of acute myeloid monoblastic leukaemia (AML) one month prior and was in remission after the induction phase presented with floaters and gradual visual loss in both eyes. Visual acuity was 20/200 and 20/25, respectively. Anterior segment exam was normal in both eyes. Funduscopy showed reduced foveal reflex. On spectral‐domain OCT, subretinal fluid was found in both eyes. Fluorescein angiography showed hypofluorescent areas compatible with multifocal serous retinal detachments and multiple hyperfluorescent dots surrounding the detached areas. Days after, the patient debuted with facial palsy and binocular diplopia due to bilateral VI cranial nerve (CN) palsy, left VII CN palsy and right complete III CN palsy. An MRI was performed with no signs of infiltration. A cerebrospinal fluid sample was drawn by lumbar puncture and its analysis denoted infiltration of AML cells with similar immunophenotype to that on initial bone marrow biopsy. Our patient presented with an atypical case of Vogt‐Koyanagi‐Harada (VKH) with multiple leakages along the periphery of the detached retinal area. This sparing of the macula should be considered a suspicious pattern, and combined with the finding of CN palsies was enough to suspect a relapse in the form of CSF infiltration, making the diagnosis of neuroleukaemia possible.Conclusions: Exudative retinal detachment can be a primary presentation of AML or denote a relapse. VKH is a diagnosis of exclusion, as one of the main criteria for its diagnosis is the absence of other systemic diseases. This becomes especially important in atypical presentations and older patients. So as not to miss life‐threatening underlying conditions, leukaemia and lymphoma should be considered in the differential diagnosis of VKH.

Rapid Progression of Myelodysplastic/Myeloproliferative Neoplasm with Neutrophilia to Acute Myeloid Leukemia: A Case Report

Abstract Introduction/Objective Myelodysplastic/myeloproliferative neoplasms (MDS/MPN) with neutrophilia, formerly atypical chronic myeloid leukemia, is a subtype of MDS/MPN that is characterized by peripheral blood neutrophilia with frequent left-shifted myeloid cells, dysplastic granulocytes, and an absence of the Philadelphia chromosome or BCR- ABL1 fusion gene. Its clinical course is variable, ranging from a chronic indolent disease to an aggressive leukemia. Herein, we report a case of who was diagnosed with MDS/MPN with neutrophilia and had rapid progression to acute myeloid leukemia. Methods/Case Report A 63-year-old male patient with no significant medical history presented with incidental findings of abnormal peripheral blood counts: leukocytosis of 153 × 109/L, platelets of 82 × 109/L, and Hb of 6.7g/dl. Flow cytometry (FCM) on peripheral blood was significant for increased granulocytes and a population of aberrant blasts (~7.3%) expressing CD5, CD11c, CD13, CD34, CD38, CD56 (partial), CD117, and HLA-DR. The initial Bone marrow biopsy showed a hypercellular marrow (90-100%), 3% blasts, dysgranulopoiesis, and dysmegakaryopoiesis. Fluorescence in situ hybridization (FISH) analysis for BCR-ABL1 fusion and MPN with Eosinophilia panel, including FIP1L1-CHIC2-PDGFRA; PDGFRB/5q33; FGFR1/8p12, were all negative. Next Generation sequencing (NGS) detected at least one variant of strong clinical significance (Tier I) including: ASXL1 c.1900_1922del23, NRAS c.35G>C, U2AF1 c.470A>C, potential clinical significance (Tier II) GATA2 c.1160_1165delCCATGA and unknown clinical significance (Tier III) SETBP1 c.2620G>A. The chromosomal analysis showed trisomy 14. The patient received Decitabine for 2 months, but follow-up FCM on the subsequent bone marrow examination revealed 40% blasts, with marked erythroid hyperplasia. The patient was diagnosed with acute myeloid leukemia. Despite receiving Venetoclax and Azacitidine, the patient deteriorated and expired in 10 months after the initial diagnosis of MDS/MPN with neutrophilia. Results (if a Case Study enter NA) NA Conclusion This case highlights the poor prognosis associated with MDS/MPN with neutrophilia. Optimal management strategies for these patients are still evolving, and new targeted therapies are needed to improve outcomes.

CUX1mut Acute Myeloid Leukemia As a Distinct Biological Entity: An Analysis of Clinical Outcomes and Implications

1. Introduction CUX1 is a tumor suppressor gene located on the long arm of chromosome 7. CUX1 mutations induce apoptosis evasion and are more commonly seen in therapy-related myeloid neoplasms. It is frequently mutated in myelodysplastic syndrome and less frequently in acute myeloid leukemia (AML). Preclinical investigations are ongoing to explore therapeutic approaches in CUX1-deficient leukemia cell lines. However, data on the clinical outcomes of patients with AML and mutated CUX1 are sparse. We aimed to characterize the biological and clinical outcomes of CUX1 mut AML. 2. Methods We analyzed next-generation sequencing from 318 patients in the Project ERIS database with newly diagnosed AML treated from January 1, 2013 to April 18, 2023 at VCU Massey Comprehensive Cancer Center. We identified 12 patients with CUX1 mut AML and compared them to a cohort of 306 CUX1 wild-type historical controls. We recorded baseline patient-related and disease characteristics, including age, ECOG, Charlson comorbidity index (CCI) scores, molecular profiling and ELN 2022 cytogenetic risk, dates of regimen initiation, and survival. We used the D'Agostino & Pearson method for normality testing and the t-test or Mann-Whitney (as applicable) tests for between-group comparisons. Categorical comparisons used Fischer's exact test. We applied the Bonferroni correction if multiple comparisons were made. The Wilson-Brown method was used for 95% confidence intervals. We analyzed survival by the Kaplan-Meier method with significance determined by the log-rank test. The event for calculating the overall survival (OS) was the date of death. Patients were otherwise censored at the date of last contact. 3. Results CUX1 mutations are found at a frequency of 3.9% (95% CI, 2.2-6.5) in AML. The median variant allele frequency of CUX1 mutations was 40.3% (range: 3.8-48.7), consistent with a heterozygous presentation. The median age at diagnosis of patients with CUX1 mut who received any anti-leukemic therapy was 57 years (range: 30-78 years), which was not significantly different from CUX1 wt at 65 years (range: 19-88, p = 0.312). Thirty-three (95% CI, 13.8-60.9) percent of patients had AML with myelodysplasia-related changes (AML-MRC) on the initial diagnostic bone marrow biopsy, compared with 29.2% (95% CI, 24.2-34.7, p=0.752) for CUX1 wt. CUX1 mut AML was found in cytogenetically normal specimens at a frequency of 30.0% compared with 49.3% for CUX1 wt (p = 0.337). The most frequent cytogenetic abnormality in CUX1 mut AML was -7 (20.0%), followed by +8 (10.0%) and complex cytogenetics (10.0%). The most frequent cooperating molecular lesions were RUNX1, SRSF2, STAG2, FLT3-ITD, and NRAS (25.0% each). Patients with CUX1 mutations were nonsignificantly more likely to have ELN 2022 adverse-risk disease (odds ratio: 2.013, p = 0.379). In patients treated with intensive cytarabine-based inductions, the median overall survival was 26.3 months in the CUX1 mut group and 17.6 months in the CUX1 wt cohort, which was not significantly different (p = 0.501). Two patients with CUX1 mut AML were treated with first-line venetoclax and a hypomethylating agent. The overall survival was 12.7 months in one patient and not reached at a follow-up time of 20.2 months in the second. 4. Discussion While definitive conclusions are limited by the rarity of the mutation, mutations in CUX1 are heterozygous and appear to be non-significantly more frequently associated with myelodysplastic-related changes and secondary-type mutations. These findings may explain a higher proportion of patients harboring CUX1 mutations that have ELN 2022 adverse-risk disease. While we did not detect any significant survival differences between patients with and without CUX1 mutations, larger cohorts are needed to confirm the prognostic impact of these rare mutations in AML.

La relación entre los parámetros inmunohistoquímicos, la fibrosis de la médula ósea y la captación de 18F-FDG en la médula ósea en pacientes con mieloma múltiple sometidos a examen PET/TC

AimThe aim of this study was to determine the power of the SUVmax value obtained from 18F-FDG PET/CT in multiple myeloma patients to be able to predict immunophenotype characteristics (CD20, CD44, CD56, CD117, and CD138 antigen expressions), bone marrow fibrosis, cyclin D1 oncogene, and M-protein subtypes which play a role in diagnosis-treatment and prognosis of the disease. Material and methodThe study included 54 patients with multiple myeloma who underwent PET/CT for initial staging and bone marrow biopsy. The relationship was examined in these patients between the SUVmax value measured from the iliac bone region and the immunohistochemical and bone marrow fibrosis data of the biopsy taken from the iliac bone. The Mann–Whitney U-test was used in the comparisons of dependent paired groups, and the Kruskal–Wallis H test in the comparisons of three or more groups. ResultsThe median SUVmax value was 4.5 (1.9-15.6) in patients with CD117 antigen positivity, which was statistically significantly higher than the value in the patients with CD117 negativity (P=0.031). When patient grouping was made according to the reticulin level: We found that the median SUVmax value was 4.9 (3.0-14.8) in the group with increased fibrosis and 3.6 (1.6-15.6) in the group with low fibrosis. The median SUVmax was statistically significantly higher in the group with increased fibrosis compared to the group with low fibrosis (P=0.004). No statistically significant difference was determined in the comparisons of the SUVmax values when the patients were grouped according to the immunoglobulin heavy chain and light chain, CD20, CD44, CD56, and cyclin D1 characteristics (P>0.05). ConclusionIn multiple myeloma patients who underwent PET/CT for initial staging, significant relationships were determined between FDG uptake in the bone marrow (SUVmax) and CD117 antigen and bone marrow fibrosis, which is an important prognostic factor. Higher SUVmax values were determined in the bone marrow of patients with increased fibrosis and CD117 positivity.

The relationship between immunohistochemical parameters, bone marrow fibrosis and bone marrow 18F-FDG uptake in multiple myeloma patients undergoing PET/CT examination

PurposeThe aim of this study was to determine the power of the SUVmax value obtained from 18F-FDG PET/CT in multiple myeloma (MM) patients to be able to predict immunophenotype characteristics (CD20, CD44, CD56, CD117, CD138 antigen expressions), bone marrow fibrosis, cyclin D1 oncogene, and M-protein subtypes which play a role in diagnosis-treatment and prognosis of the disease. Material and methodThe study included 54 patients with multiple myeloma who underwent PET/CT for initial staging and bone marrow biopsy. The relationship was examined in these patients between the SUVmax value measured from the iliac bone region and the immunohistochemical and bone marrow fibrosis data of the biopsy taken from the iliac bone. The Mann Whitney U test was used in the comparisons of dependent paired groups, and the Kruskal Wallis H test in the comparisons of three or more groups. ResultsThe median SUVmax value was 4.5 (1.9−15.6) in patients with CD117 antigen positivity, which was statistically significantly higher than the value in the patients with CD117 negativity (p = 0.031). When patient grouping was made according to the reticulin level; we found that the median SUVmax value was 4.9 (3.0−14.8) in the group with increased fibrosis and 3.6 (1.6−15.6) in the group with low fibrosis. The median SUVmax was statistically significantly higher in the group with increased fibrosis compared to the group with low fibrosis (p = 0.004). No statistically significant difference was determined in the comparisons of the SUVmax values when the patients were grouped according to the immunoglobulin heavy chain and light chain, CD20, CD44, CD56, and cyclin D1 characteristics (p > 0.05). ConclusionIn MM patients who underwent PET/CT for initial staging, significant relationships were determined between FDG uptake in the bone marrow (SUVmax) and CD117 antigen and bone marrow fibrosis, which is an important prognostic factor. Higher SUVmax values were determined in the bone marrow of patients with increased fibrosis and CD117 positivity.

Clinicopathological Features of Isolated Thrombocytopenia Associated with SRSF2 Mutations: A Case Series

Introduction: Mutations of SRSF2, a gene involved in RNA splicing, are identified across a heterogenous group of myeloid neoplasms including MDS, CMML, and MPN. SRSF2 mutations have been associated with thrombocytopenia in MDS, but not in the case of MDS/MPN overlap conditions. When co-mutated with TET2, SRSF2 is associated with monocytosis and a myeloproliferative phenotype. SRSF2 mutations are also identified in normal individuals and in those with clonal cytopenias of undetermined significance (CCUS). Studies have examined genotype-phenotype correlations in myeloid malignancies, but similar analysis of the heterogeneity amongst patients with CCUS are lacking. Although dysplasia is not a defining feature of CCUS, sub-diagnostic dysplasia can be seen in CCUS and this subset notably had a higher proportion of spliceosome mutations. Here we describe the clinical and pathological features of a series of CCUS patients with prolonged, isolated thrombocytopenia and a SRSF2 mutation. Methods: We conducted a retrospective review of the electronic medical records of patients seen in the outpatient Hematology Clinic at the University of Southern California Norris Comprehensive Center between May 2014 and December 2021. Patients with persistent (> 6 months) isolated thrombocytopenia, a SRSF2 pathogenic mutation identified on a myeloid molecular panel, and absence of WHO-defined dysplasia or hematologic neoplasm on initial bone marrow biopsy were included. A work-up for alternative etiologies of thrombocytopenia was completed based on the clinical judgment of the treating physician. Bone marrow biopsies were reviewed by 2 independent hematopathologists. This study was approved by the Institutional Review Board of the University of Southern California. All patients signed informed consent. Results: Five patients met criteria for inclusion. All patients were male. Median age at diagnosis of thrombocytopenia was 61 years (56-74). Two patients were Asian and 3 were Caucasian. The median time from diagnosis of thrombocytopenia to the identification of a mutation on a myeloid molecular panel was 4 years (3-12). No patients had a family history of hematologic malignancy. Initial bone marrow biopsies did not meet WHO-defined thresholds for morphologic dysplasia. However, all patients had megakaryocytic hyperplasia and megakaryocytes with hyperchromasia and high nuclear-cytoplasmic ratio in 20% of megakaryocytes (Figure 1). Two patients who had CD34 staining showed positive staining in 30% of megakaryocytes. Four patients had normal cytogenetics and 1 patient had monosomy 7. Median bone marrow cellularity was 65% (30-80%). Two patients had intermittent peripheral monocytosis but did not meet the diagnostic criteria for CMML. No monocyte proliferation or immature monocytes were seen on bone marrow. SRSF2 mutations were P95H in 3 patients and P95L in 2 patients. Average variant allele frequency was 37.3% (range 28 - 50.2). Four patients had a co-occurring TET2 mutation; other co-mutated genes included RUNX1 (3 pts.), ASXL1 (1 pt.), DNMT3A (1 pt.), and NF1 (1 pt.). Median number of co-occurring mutations was 2 (0-3). Three patients were treated empirically for immune thrombocytopenia and had no response to IVIG and/or steroids. Three patients were treated with thrombopoietin receptor agonists (2 eltrombopag, 1 romiplostim) and had minimal transient platelet count increases not meeting the IWG MDS criteria for response. One patient was treated with hypomethylating agents for 4 months before progressing to MDS or AML. Four patients required as-needed platelet transfusions. To date, 4 have progressed to MDS or AML with a median time from diagnosis to progression of 6.3 years (range 5-10). Three patients have received allogeneic hematopoietic stem cell transplant. Conclusion: This series describes a subset of CCUS patients with isolated thrombocytopenia associated with SRSF2 mutations. We also describe atypical megakaryocytic findings which do not meet current WHO diagnostic criteria for dysmegakaryopoiesis; however, the uniform presence of these findings in this cohort suggests an association with thrombocytopenia and CCUS. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Defining Primary Vs Secondary AML: Correlation between Pathology and NGS/FISH

Introduction: The treatment of acute myelogenous leukemia (AML) is often determined by factors that include an antecedent dysplasia. At presentation it is often unknown whether a patient has had dysplastic bone marrow features or not. We opted to look back at our leukemia patients to determine the frequency of antecedent dysplasia and to determine if next generation sequencing (NGS) or Florescent in situ hybridization (FISH) findings correlate with the pathologic review. Methods: We chart reviewed 43 charts from a period of 9/21/2021 to current from Orlando Health records. We focused on 29 charts that we could gather complete information from. We reviewed initial bone marrow biopsies and NGS/ FISH findings. We determined if a patient had a known history of myelodysplasia from previous records. If there was no known history of myelodysplastic syndrome (MDS) but dysplasia was seen on the bone marrow biopsy, we looked at NGS/FISH. While there can be overlap with AML, findings that suggest MDS are as follows: NGS: TET2, NRAS, U2AF1, RUNX1, TP53, SF3B1 FISH: 5q-, -5 (5p15, 5q31, 5q33), 7q-, -7 (Cen 7, 7q22, 7q31), Trisomy 8 (Cen 8), MLL (11q23), 20q- (20q12, 20qter) We then examined what treatments were used based on these findings. Results: Overall: 11/29 patients had dysplasia mentioned in their initial pathology report. 4/11 had a known preexistent dysplasia. Of the 7 who didn't have known dysplasia, 6/7 had an NGS or FISH suggesting MDS. Age: Of the patient who had dysplastic findings as well as NGS/FISH to support those findings, the patients had an average age of 57. The one patient without MDS/FISH findings has an age of 69. The average age of all patients with dysplastic findings on their bone marrow was 60. Of the patients who did not have any dysplastic findings on the bone marrow, the average age was 58. Treatment: Vyexos (liposomal daunorubicin and cytarabine), a therapy generally used for secondary leukemias was only used in two patients. Age and performance status was cited as a reason to use alternative therapy. Although one patient did receive 7+3 (idarubicin and cytarabine) instead of Vyxeos. One patient also had severe COVID pneumonia at diagnosis as the reason for not getting induction with Vyexos and instead getting venetoclax/azacitidine. Four patients were either treated with hypomethylating agent of either azacitidine or decitabine, again this was normally chosen due to performance status, age, or active COVID pneumonia as above. Conclusion: These results show that pathology findings of underlying dysplastic changes seem to correlate with NGS or FISH findings. Of the 7 patients with these bone marrow changes but no known MDS, 6 of them had NGS or FISH to support an underlying disease. Factors such as age did not help to predict antecedent MDS. Based on this information it may be appropriate to start with induction therapies such as Vyxeos when dysplastic changes are seen on the bone marrow while FISH and NGS are still pending, if their performance status and age allow to do so. Further larger retrospective studies to compare underlying dysplastic findings versus FISH/NGS findings should be done to further support this concept.

P163 The cytokine horror: Kawasaki disease shock syndrome complicated with macrophage activation syndrome

Abstract Background/Aims Kawasaki disease (KD) is an acute febrile systemic vasculitis of unknown aetiology, affecting medium-sized vessels. It often occurs in children less than 5 years old. During the acute phase of the disease, less than 5% of children develop hypotension and shock. This severe form is called Kawasaki Disease Shock Syndrome (KDSS). Macrophage activation syndrome (MAS) is a heterogenous syndrome complex encompassing a broad spectrum of clinical features. MAS associated with KDSS is an extremely rare life threatening entity that warrants a high index of clinical suspicion, prompt and proper management. Methods We present our experience in the dilemmas and challenges faced in the diagnosis and management of KDSS complicated with MAS in a poor resource setting. It is our first experience of encountering MAS in KDSS. Results A 9-month-old baby with fever and a maculopapular rash involving palms and soles for 5 days duration was transferred from a peripheral hospital. A preliminary diagnosis of acute Kawasaki disease was made with the classical features that included the rash, cracked lips, pedal oedema, BCG scar inflammation, perianal excoriation and irritability. The fever did not settle following standard therapy that included three doses of intravenous immunoglobulins 2g/kg, aspirin and low dose dexamethasone. The child quickly began to decompensate. She developed cold clammy skin with hypotension on the 8th day of the illness. There was evidence of hepatosplenomegaly with transaminitis. 2D Echo revealed normal coronary vasculature without any dilatations or strictures. However, there was evidence of myocarditis supported by diffuse T inversions in the anterolateral leads and a highly positive troponin titre. Investigations revealed aa leucocytosis (22,100/dl), an abrupt drop in platelet count (350,000/dl to 80,000/dl), red blood cells, haemoglobin (11g/dl to 8g/dl) and erythrocyte sedimentation rate 95mm/hr to 50mm/hr. Hyperferritinemia, hypertriglyceridemia and hypofibrinogenemia supported the diagnosis of MAS. Blood culture was sterile. Initial bone marrow biopsy was inconclusive. High dose dexamethasone 10mg/m2/day and cyclosporine 2mg/kg/day was initiated. The circulatory collapse was managed with fluid boluses and a low dose adrenaline infusion. Subsequently there was dramatic clinical improvement and her cell counts and transaminases normalised on day 15. Conclusion We emphasise the importance of early recognition of refractory MAS in pediatric populations with rheumatic diseases and we suggest early initiation of cyclosporin as a rapid and effective treatment option in the poor resource settings where the ideal management with anakinra is beyond the realms of possibility. Disclosure L. Rajagopala: None. J. Jagoda: None. M.L. Gunatilaka: None.

CD19-Targeted CAR T Cell Therapy for Concomitant Diffuse Large B Cell Lymphoma and Myeloma

While DLBCL and myeloma comprise a large fraction of annual hematologic malignancies, the co-occurrence of these malignancies is particularly rare. The FDA has approved three CD19-targeted CAR T cell therapies for B-cell lymphoma (Kymriah, Yescarta, and Brenyazi) and more recently a single CAR T cell therapy for myeloma (Abecma). We present a case of concurrent multiply r/r DLBCL and non-r/r myeloma treated exclusively with CD19-targeted CAR T cell therapy.A 77-year-old female was admitted to an outside hospital with a chief complaint of one week of constipation, incidental labs demonstrating hypercalcemia and AKI, and a bone marrow biopsy consistent with myeloma (phenotype lamda, CD138-, MUM01+, CD56- and CD117, KITE-). There were 20% lambda-restricted plasma cells with no evidence of lymphoma. She also had concomitant anemia with a hemoglobin of 9.6 g/dL best attributed to the plasma cell dyscrasia, thus fulfilling criteria from the International Myeloma Working Group.Beta-2-microglobulin was 6.8 mg/L, consistent with stage 3 disease. She demonstrated two M proteins at time of diagnosis; each was 0.40 g/dL. Interestingly these monoclonal proteins were IgG-Kappa and an IgM protein. The patient was transferred to our institution for management after receiving a single cycle of bortezomib, cyclophosphamide and dexamethasone. A right inguinal lymph node biopsy of an existing right thigh mass revealed atypical cells with findings consistent with non-GC DLBCL [CD20+, PAX5+ (weak), MUM1+, BCL2+, CD10-, CD5-, BCL6-, BCL1-, cMYC-; R-IPI 4, CNS-IPI 4, Stage IV]. Chemotherapy with R-CHOP for four cycles resulted in a mixed response. Upon subsequent hospitalization, a second right inguinal lymph node biopsy was consistent with refractory DLBCL [positive for CD79a, CD20 (weak), MUM1, BCL2, BCL6, MIB1 Ki67 60-70% proliferative fraction, CD30 (1-2%), negative for CD10, CD3, CD5, BCL1, MYC]. Radiation followed by RICE given as second line therapy had no meaningful response on imaging. However, monoclonal protein resolved after a second cycle of RICE. Pathology indicated a CD19+ specimen on review. Polatuzumab vedotin was given as bridging therapy, followed by lymphodepletion regimen and ultimately CD19-targeted CAR T cell infusion. A partial response for lymphoma was seen at 90-day evaluation (Deauville 3). Immunofixation was positive as late as 16 days post infusion with no abnormality detected at subsequent testing 32 days post infusion. At three months, the M protein level was 0.00 g/dL with no detectable monoclonal protein on immunofixation or electrophoresis testing. Flow cytometry peripheral blood smear was negative for plasma cells by CD38 and CD138 labeling seven months post CAR T infusion.CD19-targeted CAR T cell therapy is established as an accepted standard of care for multiply r/r DLBCL when it is available. However, the role of HSCT in r/r DLBCL is standard of care after second line salvage. In our patient, however, she failed to adequately respond to salvage chemotherapy. Though there is data supporting the use of HSCT in chemorefractory patients, it is not as successful as in chemosensitive disease. Several trials have investigated BCMA-targeted CAR T cell therapy in r/r myeloma and have reported high response rates, however there is a lack of data supporting these immunologic therapies in non-r/r myeloma. Data regarding CD19 CAR T cell's effect on myeloma is limited as secondary malignancies typically exclude subjects from enrollment; however, CD19-mediated responses have been reported in vitro and CD19-targeted clinical trials are underway in r/r myeloma. Based on emerging data regarding the effectiveness of CAR T, our team elected to pursue CD19 CAR T cell therapy. One critique is that the monoclonal protein detected in the blood was different from the lambda-restricted plasma cell population that was noted in the initial bone marrow biopsy. However, given the lack of detectable myeloma based on labs and imaging, we have elected to defer a bone marrow biopsy at this time.Our patient case shows that directing toward the more aggressive malignancy, typically the lymphoma with CAR T cell therapy, can target the myeloma as well. This has significant implications on guiding treatment decisions in future patients presenting with concomitant malignancies. Furthermore, new data from studies investigating CD19-targeted therapies' effect on myeloma will have amplified clinical relevance. DisclosuresCiccolini: Currant Insights, LLC: Consultancy; Techspert.io.: Consultancy; Guidepoint: Consultancy; Atheneum: Consultancy; First Thought: Consultancy; GLG: Consultancy; Kite Pharma: Membership on an entity's Board of Directors or advisory committees; JADPRO: Honoraria. Kalac: Astra Zeneca: Consultancy; Kyowa Kirin: Consultancy; Gilead: Consultancy; Jannsen: Research Funding; Guidepoint: Consultancy; GLG: Consultancy.

Epidemiology Unknown: Developing an Approach to Identify Potential Smoldering Multiple Myeloma Cases

BackgroundSmoldering Multiple Myeloma (SMM) is an asymptomatic clonal plasma cell disorder that identifies patients at risk for progression to Multiple Myeloma (MM). The standard of care for SMM has traditionally been observation, but some cancer centers are now treating high-risk SMM before progression to MM. The diagnostic criteria for SMM have also changed in recent years, and current estimates of SMM are derived from large MM databases and observations from tertiary centers. The goal of this study was to develop an approach for identifying SMM cases in a large integrated healthcare delivery system to better characterize the epidemiology of SMM in community-based populations.MethodsThis retrospective, observational study was conducted in Kaiser Permanente Northern California (KPNC) using KPNC SEER-based Cancer Registry data and information from the electronic health record (EHR). Potential SMM cases from 1/1/2010 to 12/31/2018 were identified using three approaches: Group 1 - identified via the KPNC Cancer Registry based on indicators of ‘asymptomatic myeloma’, ‘evolving myeloma’, and ‘smoldering myeloma’; Group 2 - identified via the KPNC Cancer Registry as MM cases who had a physician visit note containing the word ‘smoldering’ but did not begin treatment within 1 year of diagnosis; Group 3 - identified via the KPNC Cancer Registry as MM cases who had a physician visit note containing the word ‘smoldering’ but did begin treatment within 1 year of diagnosis. Chart review was performed for these potential SMM cases (Groups 1-3) to document initial bone marrow biopsy results (bone marrow plasma cell percentage, BMPC) and skeletal findings (presence or absence of lytic bone lesions) around the time of biopsy. When BMPC was reported as a range, the highest value was captured. Patient demographics (age, sex and race/ethnicity) were obtained from the EHR. Bivariate analyses were performed using the chi-squared test and the Wilcoxon-Mann-Whitney nonparametric test. For binomial comparisons by mode of potential SMM case identification, Groups 1 and 2 were combined and compared to Group 3.ResultsA total of 471 potential SMM cases were identified, including 178 (37.8%) via Group 1, 35 (7.4%) via Group 2, and 258 (54.8%) via Group 3 (Figure). The median age was 71 years (interquartile range, IQR 62-78) and 40.0% were female. The racial/ethnic distribution included 57.1% White, 17.6% Black, 10.8% Hispanic, 13.6% Asian, and 0.9% other/unknown race. There were no significant differences across groups (Group 1+2 vs Group 3) with respect to age (p=0.07), sex (p=0.85), or race/ethnicity (p=0.81).There were 442 (93.8%) who underwent bone marrow biopsy. Among those with BMPC data, the median BMPC for Group 1 was 20.0% (IQR 10.0%-28.0%); for Group 2 was 25.0% (IQR 12.5%-50.0%), for Group 3 was 28.0% (IQR 15.0%-50.0%) (p<0.001 comparing Groups 1+2, combined median 20.0%, IQR 10.0%-30.0%, vs Group 3, 28.0%, IQR 15.0%-50.0%). The proportion with BMPC ≥60% was 4%, 13%, and 22% for Groups 1, 2, and 3, respectively (Figure).There were 413 (87.7%) who had skeletal imaging (n=405, 86.0% with available results) within 6 months of diagnosis. Of those with imaging results available to view, n=68 were found to have lytic lesions; 7.6% among Group 1, 6.9% among Group 2, 24.9% among Group 3 (p<0.001 comparing Groups 1+2, 7.5%, vs Group 3, 24.9%).DiscussionThis study used a multifaceted approach to identify potential SMM cases from a large real-world clinical population in an integrated health system. We used an approach similar to prior SMM epidemiological studies and also included those with physician visit notes specifically containing the word ‘smoldering’ within the text. The vast majority of our cohort had BMPC between 10 and 60%, but those who received treatment within 1 year had greater BMPC and a higher proportion of lytic lesions. This suggests those who received treatment, Group 3, may have actually had a MM diagnosis, and physician visit notes containing the word ‘smoldering’ may have been intended to communicate something other than a SMM diagnosis. Further analyses will determine the effectiveness of each approach by confirming SMM cases according to the International Myeloma Working Group diagnostic criteria with incorporation of laboratory data and additional clinical findings. Among confirmed cases of SMM, the rate of progression to MM and the severity of end-organ damage at time of progression will be assessed. [Display omitted] DisclosuresLo: Novartis: Research Funding; Bristol-Myers-Squibb: Research Funding; CSL-Bering: Research Funding.

Prognostic Value of Tumor Associated Macrophage Markers CD163 and CD68 Immunohistochemistry in Classical Hodgkin Lymphoma.

Tumor associated macrophages have been implicated in the pathogenesis of classical Hodgkin's lymphoma and have been suggested to have a negative impact on outcome. The aim of this study is to determine the expression and the prognostic impact of CD163 and CD68 markers of the tumor associated macrophages, in the initial positively infiltrated bone marrow biopsy specimens of our subjects by immunohistochemistry and to corre¬late their expression with other clinical and laboratory prognostic factors. This study was conducted on fifty-one patients with de novo classical Hodgkin's lymphoma, presenting to the Clinical Pathology Department at the National Cancer Institute, Cairo University. CD163 and CD68 were detected in the initial bone marrow biopsy specimens from our subjects by immunohistochemistry. The present study included 51 patients with CHL. They comprised 24 males (47.1%) and 27 females (52.9%) with an age of 32.9 ± 14.5 years. After treatment, 33 patients (64.7%) achieved complete remission while 18 patients (35.3%) failed. Comparison between patients with CR and patients without revealed significantly lower CD68 expression [median (IQR): 30.0 (15.0 - 47.5%) versus 55.0 (43.8 - 55.0%), p = 0.003] and CD163 expression [25.0 (10.0 - 37.5%) versus 45.0 (0.35 - 55.0%)] in CR patients. Binary logistic regression analysis identified CD68 and CD163 expressions as significant predictors of CR in univariate and multivariate analyses. The expressions of both tumor-associated markers, CD68 and CD163, are significant predictors of CR in patients with CHL.

A Diagnostic Puzzle: New Liver Dysfunction with Severe Thrombocytopenia

Autoimmune hepatitis is a chronic disease characterized by inflammation and hepatocellular necrosis causing varying degrees of clinical and biochemical manifestations. A rare complication is the development of aplastic anemia defined by inability of hematopoietic stem cells to repopulate the bone marrow thus resulting in severe hypocellularity without other morphologic abnormalities. Treatment using immunosuppressive therapy drastically improves outcome. We present this case of a young adult with autoimmune hepatitis leading to aplastic anemia. A previously healthy 20 year old male presented with abdominal pain and vomiting. Workup revealed markedly elevated liver enzymes, hyperbilirubinemia, seronegative acute viral hepatitis panels, severe thrombocytopenia and moderate neutropenia. Initial bone marrow biopsy showed hypocellularity and mildly decreased megakaryocytes.Liver biopsy disclosed periportal and lobular inflammation with patchy necrosis. A course of steroids and IVIG resulted in improvement of transaminitis, but he developed progressive severe pancytopenia.5 weeks later, repeat bone marrow biopsy showed virtually acellular marrow confirming the diagnosis of hepatitis associated aplastic anemia. He was treated with antithymocyte globulin, steroids and cyclosporine. About 7 months later, he continued to have moderately severe neutropenia and mild thrombocytopenia without anemia. Currently the patient is a potential candidate for stem cell transplant. Hepatitis associated aplastic anemia (HAAA) is a rare complication of autoimmune hepatitis occurring mostly in children and young adults.Onset of aplastic anemia is usually within a few months of diagnosis of hepatitis and it evolves independently of successful treatment of hepatitis. Diagnosis criteria of HAAA is a combination of pancytopenia and bone marrow biopsy showing severe hypocellularity without other morphologic abnormalities in the cellular and stromal elements. Pathophysiology of HAAA is not well defined but it is widely believed that a decreased CD4:CD8 ratio, proliferation and activation of cytotoxic T-lymphocytes and release of cytokines all play important roles. If untreated, HAAA has a very high mortality rate. It must be promptly treated using immunosuppressive therapy and, in some cases of persistent severe cytopenia, allogeneic bone marrow transplant. Such an approach has been demonstrated to improve survival.

Urinary Exosomes Detect Amyloidogenic Light Chain in Patients Who Have Renal Progression Despite a Hematologic Complete Response

Background: Immunoglobulin light chain (AL) amyloidosis is a fatal complication of B-cell clonal proliferation. Currently, the best biomarker for treatment monitoring is serum free light chain (FLC) assay. Despite its higher sensitivity, it cannot distinguish clonality thus ultralow levels of monoclonal gammopathy can be missed. Urinary exosomes (UExs) are the smallest members of the extracellular vesicle family that are excreted in the urine. They exhibit different characteristics in patients with AL amyloidosis vs multiple myeloma. This study was undertaken to determine if urinary exosomes have different characteristics in patients AL amyloidosis who progress despite being in complete response (CR) vs those who have a renal response.Methods: UEx were extracted from 4 patients at different stages of AL (Table 1). Immunoglobin light chains (LC) were identified in the UEx using western blotting. Intact immunoglobulin light chains were identified in plasma, UEx, and kidney biopsy amyloid deposits using mass spectrometry (MS). cDNAs from bone marrow plasma cells (BMPC) collected at the time of diagnosis from patient 4 were sequened.Results: Oligomeric LC (250 kd) were identified in the UEx of patient 1 (newly diagnosed, Fig 1a) and 4 (renal progression in CR, Fig 1d) but only monomeric LC (25 kd) were detected in patient 2 (Fig 1b) and 3 (CR and renal response, Fig 1c). MS of the UEx and plasma of patient 4 detected 2 monoclonal λ LC. The mass of one of the LC was consistent with a lambda 6a LC which is also the most common cDNA (IGLV-6-57) found in the BMPC. Amino acid sequences derived from the tryptic digestion of the amyloid deposits matched the predicted sequence of the cDNA. The calculated weight of the peptide produced by the cDNA (23,304 Da ) was within margin of error of the mass of the lambda 6a LC (23,306 Da). The mass of the second LC (23,092 Da) was consistent with a λ4/λ5 LCs. A search of the amino acid and cDNA sequences failed to identify any similarity. The density ratio of the oligomeric LC to monomeric LC in the UEx was 0.47 which was similar to the intensity ratio by MS of the lambda 6a LC (23,306 Da) to the 23,092 Da LC in the blood (0.43).Discussion: Urinary EXs identified oligomeric LC in a patient with progressive renal disease despite achieving a CR. The mass one of the λ LC matched the predicted peptide product of the cDNA obtained from BMPC. The amino acid sequence predicted by the cDNA matched the trypsin digested sequences from the amyloid deposits. This strongly suggests the oligomeric LC represents the amyloidogenic λ6a LC detected in the blood which was being produced by the BMPC clone which is being deposited as amyloid in the kidney. The origin of the second λ LC is unknown but is likely a new monoclonal gammopathy of undetermined significance that developed after the initial bone marrow biopsy. The blood and urine samples were obtained 3 years after the BMPC. The development of a new monoclonal gammopathy in patients with plasma cell dyscrasia is not uncommon. Unfortunately, bone marrow biopsy collected at the time of the urine and blood sample had insufficient number of plasma cells (CR) for analysis.Conclusion: UEx can identify amyloidogenic LC even when current standard methods could not. The persistent presence of the amyloidogenic LC helps explain why patients can progress despite being in CR. The presence of a second λ LC which was not part of the amyloid deposit also has clinical implication. It may explain why some patients do well despite having a persistent monoclonal protein. If these results are confirmed, UEx may have a powerful role in the determination of organ and hematologic response in patients with AL amyloidosis. [Display omitted] [Display omitted] DisclosuresDispenzieri:Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Prothena: Membership on an entity's Board of Directors or advisory committees; Alnylam: Research Funding; GSK: Membership on an entity's Board of Directors or advisory committees; pfizer: Research Funding; Jannsen: Research Funding; Celgene: Research Funding.

Standard of Care of Patients with CML Treated in Community Based Oncology Group Practices in Rhineland-Palatinate (Germany)

Introduction: Significant progress has been made in CML-therapy since the introduction of imatinib and other tyrosine kinase inhibitors (TKI) into clinical care. The aim of this study was to assess diagnosis, treatment and outcome of CML-patients who received their treatment in community based oncology practices in Rhineland-Palatinate and whether European LeukemiaNET-guidelines were followed.Methods: All Ph-/BCR-ABL-positive CML-patients who were treated between 12/2001-12/2015 in 9 oncology group practices were analyzed retrospectively concerning diagnosis, treatment and outcome according to European LeukemiaNET-guidelines. Data were collected from patient files into a central data base and analyzed statistically with SPSS.Results: 264 patients (pts) with a median age of 60 (18-90) were analyzed. 126 (48%) were female, 138 (52%) were male. At initial diagnosis bone marrow biopsy was performed in 213 pts (81%). Cytogenetics was applied in 204 pts (77%) (38% in blood, 56% in bone marrow). FISH-analysis was used in 155 pts (59%) (33% in blood, 36% in bone marrow). PCR-testing to detect a BCR-ABL1-rearrangement was applied in 200 pts (76%) (52% blood, 37% bone marrow). 258 pts (98%) were in chronic phase, 5 (2%) in accelerated phase and 1 (0.4%) in blast crisis at diagnosis. EUTOS score could be calculated in 131 pts (50%). 20% were high risk, 80% low risk. 252 pts (95%) received some form of TKI-therapy. Out of 416 TKI-therapies 308 (74%) were PCR-based monitored, 148 (36%) were monitored by cytogenetics. First line treatment was imatinib in 201 pts (80%), 51 pts (20%) received a second generation TKI. Second line treatment consisted of dasatinib in 59%, nilotinib in 32%, imatinib in 6% and bosutinib in 3%. Third line treatment was nilotinib in 56%, dasatinib in 35%, ponatinib in 6% and imatinib in 3%. 62 pts (23%) were treated within a study protocol. 13 pts (5%) received an allogeneic transplantation. Overall survival probability was 88% after 5 years and 72% after 10 years. Disease specific survival was 95% after 5 years and 86% after 10 years.Conclusion: The overwhelming majority of CML-patients treated in oncology group practices receive standard of care as suggested by European LeukemiaNET-guidelines. Overall survival in routine care is comparable to international studies. DisclosuresNo relevant conflicts of interest to declare.

A Striking Reduction of Monoclonal Protein in a Patient with Concurrent Plasma Cell Dyscrasia and CMML-2, after Treatment with Rigosertib (01910.Na)

Rigosertib (ON 01910.Na) is a member of a broader class of unsaturated sulfone kinase inhibitors capable of inducing profound mitotic spindle abnormalities, abnormal centrosome localization, G2-M cell cycle phase arrest and mitotic catastrophe, culminating in apoptosis. Rigosertib is a Ras mimetic that interferes with phosphoinositide 3-kinase (PI-3K)/Akt, reactive oxygen species and Ras/Raf/polo-like kinase (PLK) signaling pathways. Although broadly cytotoxic against malignant cells, it is remarkably non-toxic for non-neoplastic cells. For this reason, this is a particularly attractive compound to test against neoplastic diseases of the bone marrow such as MDS and acute leukemia.This is a report of an unexpected reduction in monoclonal IgG, during a subject participation in a Phase III, randomized study of rigosertib, in patients with MDS who have either failed to respond, or progressed after receiving hypomethylating agents (ONTIME Trial).A 75-year-old man with CMML-2 had a CBC on day 1 of the trial that demonstrated leukocytosis, with absolute monocytosis, 7% blasts in the peripheral blood, Hgb of 9.4 gm/dl, and platelets of 7 K. He was transfusion dependent for both pRBCs and platelets. His chemistry panel demonstrated a high total protein of 9.9 (NL: 6.0 - 8.2 G/DL) with low albumin at 2.4 (NL: 3.5 - 5.0 G/DL); therefore, an SPEP/IPEP was performed, reporting the presence of monoclonal IgG kappa. Quantitative immunoglobulins showed an elevated IgG of 3594 mg/dl (NL: 596 - 1584 MG/DL). Serum free light chains were remarkable for an elevated Kappa fraction at 38.94 (NL: 0.33 - 1.94 MG/DL).On day 1 of cycle 5 of rigosertib, he was started on pulse decadron for 2 months, after which his disease progressed to AML, and he died shortly thereafter. Neither his bone marrow biopsies, nor his hematological parameters demonstrated a response to treatment with rigosertib.In contrast and interestingly, his total protein, serum kappa light chain load, and total IgG, all were drastically reduced shortly after initiation of rigosertib, as can be seen in the graph below depicting a substantial drop in his kappa light chain as well as the kappa/light chain ratio. Importantly, reduction in the monoclonal protein was noted prior to initiation of pulse decadron.Even though his initial bone marrow biopsy did not note a monoclonal plasma cell population, a subsequent bone marrow reported a low-level involvement with a plasma cell dyscrasia, with kappa light chain restriction. His final bone marrow biopsy confirmed progression to AML, but the previously seen plasma cell dyscrasia was no longer present.Conclusion: We are not aware of prior reports describing a similar effect of rigosertib on M-proteins. However, in vitro studies with rigosertib have demonstrated antitumor effects and induction of apoptosis in myeloma cell lines1. This observation merits further exploration of this agent in multiple myeloma.

Platelet-derived growth factor receptor β (PDGFRβ) immunohistochemistry highlights activated bone marrow stroma and is potentially predictive for fibrosis progression in prefibrotic myeloproliferative neoplasia.

Myelofibrosis is the result of aberrant stromal activity which is determined routinely by reticulin staining in bone marrow biopsies. As matrix fibres are the product of activated fibroblasts, we analysed fibre accumulation compared to stromal cell activity during myelofibrosis progression using the fibroblast activation marker platelet-derived growth factor receptor β (PDGFRβ) by immunohistochemistry. Initial and follow-up bone marrow biopsies from 84 patients with myeloproliferative neoplasia, including 55 cases with primary myelofibrosis, were evaluated from five haematopathology centres. The stromal mass was measured by conventional reticulin staining [myelofibrosis (MF) grade, 0-3] and PDGFRβ-positive cells using a novel PDGFRβ scoring system (0-3). Results were correlated for prediction of progression. The MF grade and the PDGFRβ score showed excellent correlation (Spearman's r = 0.83, P < 0.0001). Elevated PDGFRβ scores (higher than MF-grade) predicted myelofibrosis progression in total with 43% sensitivity and 57% specificity, and short-term (within 1 year) progression with 82% sensitivity and 53% specificity. Progression of prefibrotic disease to manifest myelofibrosis could be forecast with 90% sensitivity and 75% specificity. PDGFRβ highlights stromal cell activation in marrow fibrosis, which is closely related to matrix accumulation, indicating a direct clinical impact especially in prefibrotic myeloproliferative disorders.

MLL Exons 4-8 Duplication in a Patient with Calr-Mutated Essential Thrombocythemia

Background: About 25% of individuals with essential thrombocythemia (ET) harbor a somatic mutation of the cal reticulin (CALR) gene which is thought to be the primary driving factor for the myeloid clone in those cases. Different secondary mutations have been described in association with CALR defects, however to our knowledge mixed-lineage leukemia (MLL) gene alterations have not been described before in this setting before progression to MDS or AML. Case Report: A 68 year old woman with a past medical history significant for thyroid cancer in remission following treatment with radioactive iodine was initially noted to have a platelet count of 629 K/mcL at the occasion of a routine CBC (normal=160-400 K/mcL). Total white blood cell count, differential and hemoglobin levels were normal. Platelet count had been normal at 377 K/mcL about one year prior to presentation. Testing for mutations of the JAK2 and MPL genes came back negative. There was no splenomegaly; initial bone marrow biopsy showed mild hypercellularity with maturing trilineage hematopoiesis and atypical megakaryocytosis without an increase in blasts. A very small population of clonal B-cells was detected (&lt;2% of total). Cytogenetic analysis did not reveal any chromosomal rearrangement. The patient was started on hydroxyurea (HU) 500 mg PO daily one month after initial presentation. There was evidence of mild iron deficiency, which was corrected with administration of IV iron sucrose at a total dose of 400 mg. Four months after starting HU the platelet count had decreased to 543 K/mcL. Repeat bone marrow biopsy around that time showed mild hypocellularity (10-20%) along with persisting normal maturing trilineage hematopoiesis and atypical megakaryocytosis. There was no increase in reticulin fibers and blast count was normal. Stainable iron was present but no ring sideroblasts were noted. Karyotype was still normal and FISH revealed no evidence of deletion 5q, monosomy 5, deletion 7q, monosomy 7, trisomy 8, 11q23 translocation or 20q deletion. Given the absence of clonal defect and the lack of symptoms, HU was withheld following which the patient was observed closely. The platelet count then increased progressively, reaching a maximum of level of 977 K/mcL about 2 months after discontinuing cytotoxic treatment. The drug was then resumed and testing with the FoundationOne HemeTM panel was obtained, looking for a select list of base substitutions, insertions, deletions, copy number alterations and other DNA rearrangements for more than 400 genes known to be somatically altered in hematologic malignancies. This assay revealed the presence of an acquired anomaly of the MLL gene, consisting of a duplication of exons 4-8. Subsequent testing specific for the CALR gene also revealed an exon 9 insertion/deletion. HU was progressively increased, up to a dose of 1000 mg alternating with 1500 mg daily. With this treatment, the platelet count has decreased to 396 K/mcL one year after presentation. Total white blood cell count, differential and hemoglobin have remained normal. The patient is also taking aspirin 81 mg daily and remains clinically stable with no B symptoms, bleeding or thrombotic manifestations. Discussion: Since mutations of the CALR gene were found to be associated with ET and myelofibrosis, several secondary genetic defects were demonstrated. Those are thought to represent clonal evolution from a primary subset of cells carrying either the JAK2 V617F mutation or a CALR exon 9 alteration. To our knowledge, MLL gene mutations have not been described as early “hits” in myeloproliferative neoplasms. However they have been abundantly documented in myelodysplastic syndrome and acute myeloid leukemia (AML), where they are often a marker of poor prognosis. Conclusion: This is the first report of MLL exons 4-8 duplication in an individual with CALR-mutated ET. Interestingly, the patient presented here did not exhibit any neutrophilia, dysplasia or increase in bone marrow blasts. It remains unclear if in this setting such a genomic alteration confers an increased risk of progression to MDS or AML. Disclosures Levine: Foundation Medicine: Consultancy. Rampal:Foundation Medicine: Consultancy.

Significance of Bone Marrow Plasma Cell Percentage in Patients with Monoclonal Gammopathy of Unknown Significance Developing Multiple Myeloma

Introduction: Monoclonal gammopathies are characterized by the detection of a monoclonal immunoglobulin in the serum or urine and underlying proliferation of a plasma cell/B lymphoid clone. (1) Patients with monoclonal gammopathy of undetermined significance (MGUS) have a clonal plasma cell population in the marrow (<10%) and secrete a monoclonal protein in the serum (<3g/dL) and/or urine. However, they lack clinical features of overt Multiple Myeloma (MM) (lytic bone lesions, anemia, renal impairment and hypercalcemia). In a study from the Mayo Clinic, 59 of 241 patients with MGUS (24%) developed MM over a period of 22 years. (2) The interval from recognition of monoclonal protein to diagnosis of MM ranged from 2-29 years, indicating that patients with MGUS need to be followed indefinitely. Many risk factors have been looked at to identify those with MGUS who are at the highest risk to progress into MM. We hypothesize that a higher number of plasma cells would correlate with a greater risk of progression to MM and sought to find out if this could be documented by arbitrarily dividing patients between < or ≥5% plasma cells seen on initial bone marrow biopsy.Methods: We retrospectively reviewed patients diagnosed with MGUS at Henry Ford Hospital between 1999-2013 who underwent a bone marrow biopsy for documenting plasma cell percentage. In addition to this, we also recorded serum hemoglobin, calcium, creatinine, monoclonal protein type and amount, serum free light chains, beta-2 microglobulin and urine for monoclonal protein at the time of diagnosis of MGUS as well as last completed values. For patients that had skeletal surveys we noted if lytic lesions were present at diagnosis, as well as cytogenetics and karyotype evaluations on bone marrow biopsy samples, if completed.Results: 120 patients with bone marrow biopsies were reviewed. Out of this 17 patients were noted from initial bone marrow biopsy to have ≥10% plasma cells. The remaining 103 patients were categorized as having MGUS. While we were not able to complete full statistical analyses, we did note that 14 of these 103 (13.6%) patients went on to develop overt MM. Further evaluation of these patients revealed that 8 of 14 (57%) had bone marrow biopsies showing ≥5% plasma cells. Interestingly the average time to progression into MM in this subgroup was 1,879 days whereas in the 6 of 14 (43%) with bone marrow biopsy showing <5% plasma cells had average time to progression into MM of 1,965 days. Abnormal cytogenetics and karyotypes of the bone marrow biopsy were also seen in 37.5% of the subgroup of patients with ≥5% plasma cells whereas it was only seen in 16.7% of the subgroup of patients with <5% plasma cells. With statistical data analyses we hope to prove significance in the above collected data as well as make further correlations in regards to risk factors in patients with MGUS.Conclusion: While we have not been able to complete full statistical analyses of the collected data yet, basic review of the above patients with MGUS and ≥5% plasma cells in the bone marrow biopsy showed a trend to develop MM faster by an average of 86 days than those that had <5% plasma cells. These same patients also were more likely to have abnormal cytogenetics and karyotypes of their bone marrow biopsies. There is a need for further investigations to be done in patients with MGUS and higher risk features. It is important that hematologists be able to recognize a high risk MGUS patient as this would lead to closer monitoring and consideration for earlier aggressive treatment to potentially delay progression into overt MM. DisclosuresNo relevant conflicts of interest to declare.

Translocation (8;21) acute myeloid leukemia presenting as severe aplastic anemia

We report a case of t(8;21) acute myeloid leukemia presenting as severe aplastic anemia. While initial bone marrow biopsy lacked any cytogenetic abnormalities in 20 analyzed metaphases, repeat bone marrow biopsy eight days later demonstrated this translocation. Initial cytogenetic analysis of 20 metaphases was therefore insufficient to make the diagnosis of hypocellular acute myeloid leukemia. We discuss that further complementary molecular tests, such as CGH, would likely provide a more robust diagnosis of hematopoietic diseases.

Microrna Expression In Patients With Myelodysplastic Syndromes Treated With Demethylating Agents

IntroductionMyelodysplastic syndromes (MDS) comprise a heterogeneous group of hematopoietic stem cell disorders characterized by bone marrow failure and dysplasia of one or more blood cell lineages resulting in peripheral blood cytopenias with frequent progression to AML. Treatment with azanucleosides improves survival and quality of life without the toxicities associated with intensive chemotherapy programs. The azanucleosides are DNA demethylating agents that may improve hematopoiesis in MDS by releasing tumor suppressor genes from silencing due to promoter hypermethylation. MicroRNAs (miRNAs) play key roles in cell growth and differentiation and oncogenic transformation. Dysregulation of miRNA expression may contribute to the development of MDS, as expression of several miRNAs has been reported to be downregulated in bone marrow cells from MDS patients. We performed the current study to test the hypothesis that treatment with demethylating agents modulates miRNA expression in hematopoietic tissue from MDS patients and to characterize the specific changes in miRNA expression. MethodsThis was a two-center, prospective cohort study to evaluate miRNA expression signatures in hematopoietic progenitor cells isolated from the bone marrow of subjects undergoing evaluation for peripheral blood cytopenias. Subjects were included for analysis if RNA of sufficient quality was available and the final pathologic diagnosis was one of the following: 1) normal (served as controls), 2) dyspoiesis without meeting criteria for MDS and without cytogenetic abnormalities, 3) MDS and 4) acute myeloid leukemia (AML) with myelodysplasia-related changes. Subjects who underwent treatment with a demethylating agent had bone marrow collected after treatment for before and after comparisons of miRNA signatures. Mononuclear cells were isolated from bone marrow aspirate specimens by density gradient centrifugation using a Ficoll-Paque technique. MicroRNA was isolated using the miRNeasy kit (Qiagen) and was analyzed using miScript miRNA PCR Array Human Cell Development & Differentiation (MIHS-103Z, Qiagen). Isolated microRNA was checked for quality control using miScript miRNA QC PCR array. Data were analyzed using Qiagen analysis software for microRNA, and miRNA levels are expressed as 2-ΔΔCT. P-values<0.05 were considered statistically significant. ResultsForty subjects were enrolled in the study. Eight subjects were excluded after the initial bone marrow biopsy for poor quality RNA (n = 3) or inappropriate diagnoses (lymphoid (n = 3) or myeloproliferative (n = 2) neoplasms). The remaining 32 subjects comprised 5 groups: 1) normal (n = 6), 2) dyspoiesis but not meeting criteria for MDS and normal cytogenetics (n = 5), 3) low-intermediate risk MDS (n = 8), 4) high-risk MDS (n = 6) and 5) AML with myelodysplasia-related changes (n = 7). Bone marrow samples were obtained from 5 patients after treatment with azanucleosides. The miRNA expression pattern in subjects with normal bone marrow and in those with no clear morphological evidence of MDS (groups 1 & 2) was similar. Conversely, subjects with low-intermediate risk MDS showed decreased expression of multiple miRNAs compared to controls (> 2 fold decrease in 22/84 miRNAs), most notably miR-126-3p (p = 0.02) and miR-20a-5p (p = 0.04). In contrast, high-risk MDS patients demonstrated a significant overall increase in microRNA expression (>2-fold increase in 33/84 miRNAs), especially miR-20b-5p (p = 0.04) and miR-125b-5p (p = 0.01). Treatment with demethylating agents led to decreased expression of most miRNAs (>2-fold decrease in 14/84 miRNAs), including miR-181a and let-7a, but an increase in others, such as miR-22 (2.3-fold increase). We obtained similar results in experiments conducted with primary bone marrow samples that were treated with demethylating agents ex vivo. SummaryOur study is the first to characterize the changes in miRNA expression patterns in patients with MDS treated with demethylating agents. We also identify novel miRNA candidates to be further evaluated as biomarkers for assessing response to treatment with demethylating agents (e.g., miR-22, which has been described as possibly having a role in regulating TET2). We also demonstrate the striking difference in the pattern of miRNA expression between low-risk and high-risk MDS patients, a feature that may one day be exploited to assist with making treatment decisions. Disclosures:No relevant conflicts of interest to declare.