Repeat Bone Marrow Aspiration Research Articles

Liquid Biopsies as Non-Invasive Tools for Mutation Profiling in Multiple Myeloma: Application Potential, Challenges, and Opportunities.

Over the last decades, the survival of multiple myeloma (MM) patients has considerably improved. However, despite the availability of new treatments, most patients still relapse and become therapy-resistant at some point in the disease evolution. The mutation profile has an impact on MM patients' outcome, while typically evolving over time. Because of the patchy bone marrow (BM) infiltration pattern, the analysis of a single bone marrow sample can lead to an underestimation of the known genetic heterogeneity in MM. As a result, interest is shifting towards blood-derived liquid biopsies, which allow for a more comprehensive and non-invasive genetic interrogation without the discomfort of repeated BM aspirations. In this review, we compare the application potential for mutation profiling in MM of circulating-tumor-cell-derived DNA, cell-free DNA and extracellular-vesicle-derived DNA, while also addressing the challenges associated with their use.

Successful Use of Granulocyte Transfusion Therapy As a Bridge to Hematopoietic Stem Cell Transplant in a Child with Aplastic Anemia and Refractory Perirectal Abscess

Background: Idiopathic aplastic anemia (IAA) is a rare, life-threatening acquired form of bone marrow failure characterized by multilineage cytopenias and hypocellular bone marrow. Profound neutropenia associated with very severe aplastic anemia (vsAA) is an important risk factor for development of life-threatening fungal and bacterial infections. Perirectal cellulitis and perianal abscess are one such infection and are associated with high rates of morbidity. The ability to eradicate infections in IAA is limited by the lack of imminent neutrophil recovery prior to treatment with either immune suppressive therapy (IST) or hematopoietic stem cell transplant (HSCT). We describe the successful use of granulocyte transfusion to treat antibiotic refractory proctitis with perianal abscess in a profoundly neutropenic child with vsAA. Case: A 7yoF presented with fever and pancytopenia in the setting of Human Metapneumovirus and Parainfluenza. Bone marrow biopsy showed hypocellularity (60-70%) with granulocytic and megakaryocytic hypoplasia, favoring viral suppression. Ultimately, she progressed to transfusion dependence. Repeat bone marrow aspirate and biopsy ultimately revealed bone marrow aplasia, DEB chromosomes, telomere lengths, and PNH clones were negative, confirming the diagnosis of vsAA. She was subsequently admitted for febrile neutropenia (F&N) and was found to have mild respiratory COVID-19, but began to complain of rectal pain that was initially attributed to her history of constipation. She defervesced on IV antibiotics (abx) and was discharged home with negative cultures. Within a week she represented with hematochezia and F&N and was found to have proctitis with perianal fistula on MRI (figure 1); GI PCR panel was + Enteroaggregative E. coli, no resistant organisms were identified. She clinically improved and discharged on IV piperacillin-tazobactam (pip-tazo). Despite 2 weeks of IV abx, she developed new fever and imaging confirmed persistent proctitis with peri-rectal abscess. As she had limited response to abx alone we trialed a short course of granulocytes with improvement in anorectal inflammation by CT. She was discharged with a plan to continue IV pip-tazo until achieving radiographic cure; however, interval imaging after an additional 2 weeks of abx showed rebound of her proctitis and she was again admitted with F&N and rising inflammatory markers. Her failure to eradicate her peri-rectal infection despite prolonged antibiotics (>4weeks), along with her previous response to a short course of granulocyte transfusion prompted us to prescribe a prolonged 10-day course of granulocytes with continued antibiosis. Her post-granulocyte CT showed resolution of her perirectal abscess and proctitis (figure 2). With resolution of her infection, she was able to proceed to HSCT which she completed without recurrence of her anorectal infection. Discussion: Conventional granulocyte transfusions are considered when patients have severe neutropenia (<500/uL), bacterial or fungal infection refractory to antimicrobials, and reasonable expectation of neutrophil recovery, but evidence for efficacy is limited. In our case, there was no expectation of imminent bone marrow recovery and our patient's infection limited her ability to proceed to IST or HSCT for treatment of her vsAA. Thus, in conjunction with infectious disease and transfusion medicine, and given her known response to granulocytes, we successfully re-attempted a prolonged, but limited, 10-day course of granulocyte transfusions and were able to eradicate a case of antibiotic-refractory proctitis with peri-anal fistula. We suggest an expanded application for the use of granulocytes in profoundly neutropenic patients without imminent recovery for severe, anti-microbial refractory infection. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Acquired Aplastic Anemia in Immunocompetent Pediatric Patients with Asymptomatic COVID-19 Infection: A Case Series

Background: In pediatrics, acquired aplastic anemia (AA) is most commonly due to infection, particularly viruses, when a cause can be identified. Coronavirus disease 2019 (COVID-19) has affected more than 197 million people worldwide, and children typically experience a less severe disease course. COVID-19 is known to cause transient hematologic abnormalities, including leukopenia, lymphopenia, anemia and thrombocytosis or thrombocytopenia in severe cases.Objectives: Describe three cases of COVID-19 associated acquired aplastic anemia in immunocompetent pediatric patients.Design/Methods: Case series established by retrospective review of the electronic medical record.Results: Case 1: An 8-year-old Hispanic male presented with a three-week history of increased bruising and a one-week history of progressive exercise intolerance, shortness of breath, pallor and fatigue. Labs showed pancytopenia. Bone marrow aspirate and biopsy was markedly hypocellular at 5-10% consistent with aplastic anemia (Figure 1). Work-up for the etiology of his aplastic anemia was only significant for positive SARS-COV-2 antibodies and a SEC23B variant of unknown significance on a comprehensive bone marrow failure (BMF)/myelodysplastic syndrome (MDS)/leukemia panel from the Children's Hospital of Philadelphia (CHOP). He was treated with eltrombopag olamine and then proceeded to immunotherapy with cyclosporine (CsA) and horse antithymocyte globulin (ATG) when a sibling match was not identified for hematopoietic stem cell transplant (HSCT). Three months later, his peripheral blood counts have improved, and he is no longer transfusion-dependent. Repeat bone marrow aspirate and biopsy continues to show markedly hypocellularity at <5%.Case 2: A 5-year-old non-Hispanic white female presented with a two-week history of easy bruising, petechial rash, fatigue and bone pain. Labs showed pancytopenia, and bone marrow aspirate and biopsy showed marked hypocellularity at 5-10% consistent with aplastic anemia (Figure 2). Her aplastic anemia work-up was significant for positive SARS-COV-2 antibodies and subclinical RBC and WBC paroxysmal nocturnal hemoglobinuria (PNH) clones. She was started on eltrombopag olamine and then proceeded to immunotherapy with CsA and ATG when a matched sibling donor was not identified. Three months later, she continues to be severely neutropenic, anemic and thrombocytopenic requiring multiple transfusions. Repeat bone marrow aspirate and biopsy showed variable cellularity with some areas 10-20% and others 70% with an overall cellularity of 50%.Case 3: An 8-year-old non-Hispanic white female presented with a 10-day history of fatigue, bilateral leg pain and pallor. Labs showed pancytopenia, elevated inflammatory markers and elevated hemoglobin F. Bone marrow aspirate and biopsy demonstrated mild-moderate hypocellularity at 40-50%, left-shifted myelopoiesis and dyspoiesis in the erythroid and megakaryocytic cell lines (Figure 3). MDS and acute lymphoblastic leukemia (ALL) fluorescence in situ hybridization (FISH) panels were negative. Additional work-up revealed positive SARS-COV-2 antibodies. Her pancytopenia resolved within two weeks of her initial hospitalization. Four months later, she presented with increased bruising and fatigue. Labs showed leukocytosis, thrombocytopenia, anemia and circulating peripheral blasts. Bone marrow aspirate and biopsy was consistent with B-cell ALL. She is receiving chemotherapy on study COG AALL1732.Conclusion: Severe aplastic anemia (SAA) has high morbidity and mortality, and timely diagnosis is needed for appropriate treatment. Multiple different viral infections have been known to cause acquired aplastic anemia. Data on all the sequelae of COVID-19 infection is still emerging, but it is plausible that COVID-19 infection may cause SAA. All three patients were found to have positive COVID-19 antibodies but did not have any evidence of previous COVID-19 infection. Further research and follow-up is needed to determine if previous COVID-19 infection is indeed a risk factor for development of SAA. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Immunohistochemistry Aids in the Diagnosis of Blastic Plasmacytoid Dendritic Cell Neoplasm in a Patient with Multiple Cutaneous Plaques and Nodules

Abstract Introduction/Objective Blastic Plasmacytoid Dendritic Cell Neoplasm (BPDCN) is a rare hematologic malignancy of plasmacytoid dendritic cell precursors with an estimated incidence of 0.04 cases per 100,000 in the US. Given the cutaneous tropism of BPDCN, it should be differentiated from other CD56+ hematopoietic neoplasms with skin involvement such as CD56+ AML, extranodal NK/T-cell lymphoma, and other T-cell lymphomas. Methods/Case Report A 71-year-old male presented to the emergency department with asymptomatic, pink- violaceous plaques/nodules on the trunk for 3 months. One month prior, a dermatologist diagnosed epidermal inclusion cysts that were left untreated. The nodules progressed and a course of steroids prescribed by a primary care physician provided short term improvement before the lesions flared again. Review of systems and vitals were normal. Biopsies of representative lesions sent for histologic examination showed a diffuse dermal infiltrate of small to medium atypical cells with irregular nuclear contours, fine chromatin, one to several nucleoli and scant cytoplasm. Immunohistochemistry showed these cells were positive for CD2, CD4, CD56 and CD45 with strong expression of BCL2 and focal CD123. The cells were negative for CD3, CD20, CD79a, CD8, CD30, ALK-1, MUM-1, CD10, Cyclin- D1, C-MYC, EBER, BCL6, Langerin, Granzyme, TIA1, CD68, CD163, MPO, and Lysozyme. The histology and immunoprofile were consistent with BPDCN. A bone marrow biopsy showed cells with similar morphology and staining pattern, including expression of CD123.Treatment with chemotherapy and Tagraxofusp, was initiated. Within a week, the patient showed near resolution of cutaneous lesions. Repeat bone marrow aspirate and flow cytometry a month later showed no malignant cells. Results (if a Case Study enter NA) NA Conclusion We present this case as a rare hematologic malignancy with a challenging clinical and histopathologic diagnosis. The histologic findings suggested either a high grade myeloid or lymphoid malignancy. The combination of CD3-/CD56+/CD4+/CD123+ in the lesional cells helped establish the diagnosis of BPDCN, allowing for prompt treatment.

Abstract PD9-11: Identifying breast cancer survivors with dormant disseminated tumor cells: The PENN-SURMOUNT screening study

Abstract Background: Patients (pts) treated for early stage breast cancer have a 30% lifetime risk of developing incurable, distant metastatic disease. Current models suggest that this occurs through escape of cells from the primary tumor into the circulation and subsequent sequestration of “disseminated tumor cells” (DTCs), in the bone marrow and other sequestration sites, where they enter dormancy. DTCs identified by immunohistochemistry (IHC) are associated with poor prognosis in longitudinal studies and meta-analyses, increasing odds of recurrence by approximately 2 to 5-fold. However, little is known about the test characteristics of the DTC-IHC assay, clinical DTC detection rates over time, and patient and disease risk factors that can identify pts harboring these cells. Methods: The PENN-SURMOUNT Screening Study (NCT 02732171) is a prospective, longitudinal study examining bone marrow and blood biomarkers of recurrence among pts within 5 years of diagnosis who have completed therapy for primary breast cancer (with the exception of endocrine therapy). Pts with positive lymph nodes, triple negative receptors, ER-positivity with RS ≥ 25 and/or high-risk MammaPrint (MP), or residual disease after neoadjuvant chemotherapy were screened with bone marrow aspirate (BMA) for presence of DTCs. A positive DTC-IHC result is defined by the presence of at least one pancytokeratin-DAB positive cell utilizing the methods of Naume et al. Cytospin slides prepared from the BMA are independently reviewed by two pathologists with adjudication for the presence of DTCs; Pts who screen negative for DTCs are offered repeat screening annually. Pts who screen positive are referred to an interventional clinical trial (CLEVER, NCT 03032406). Results: A total of 194 pts screened eligible for enrollment on PENN-SURMOUNT between 6/2016 and 3/2020. Of these, 158 consented and 151 underwent BMA with successful IHC analysis on 100%. Pts came from 22 U.S. states; ≥ 1/3 traveled over 50 miles to the study center. At baseline BMA, 36/151 (24%) had at least 1 measurable DTC by IHC. Patient characteristics and DTC distribution among subpopulations are shown in Table 1. Of the 78/115 who were initially DTC negative and continued to be eligible for repeat screening, as of 3/2020, 46 (59%) returned for at least one repeat BMA. 13/46 (28%) had at least 1 detectable DTC on 1 of up to 3 subsequent follow up assessments for a total DTC positivity rate of 32.5% (49/151). 48 (98%) DTC+ pts have subsequently enrolled on the CLEVER trial. Conclusions: BMA assessment for DTCs is feasible in pts with high risk, early stage breast cancer. DTCs are detected in up to a third of breast cancer survivors with repeat assessment during the surveilance period. DTC positivity rates are relatively similar across all receptor subtypes, and after both neoadjuvant and adjuvant chemotherapy. Pts harboring DTCs are highly likely to enroll on interventional trials designed to reduce recurrence risk. Table 1. Patient characteristics and distribution of % DTC positivity among subpopulationsDTC+ (N=49)DTC- (N=102)Total (N=151)DTC+ Rate (Overall: 32.5%)DEMOGRAPHICSMedian Age at BMA (yrs)51.9 (43.9-60.6*)50.5 (42.9-58.1)50.5 (43.8-58.8)N/ARaceCaucasian44 (89.8%)91 (89.2%)135 (89.4%)32.6%African American5 (10.2%)9 (8.8%)14 (9.3%)35.7%Other0 (0%)2 (2.0%)2 (1.3%)0%Menopausal StatusPre-15 (30.6%)34 (33.3%)49 (32.5%)30.6%Post-34 (69.4%)68 (66.7%)102 (67.5%)33.3%BMI at BMA (kg/m2)24.2 (21.9-28.9*)26.9 (23.4-31.4)26.1 (22.8-30.4)N/ARECEPTOR STATUSER/PR+ HER2neg (by ASCO/CAP)24 (49.0%)51 (50.0%)75 (49.7%)32.0%HER2+ (any ER/PR)9 (18.4%)12 (11.8%)21 (13.9%)42.9%ER/PRneg HER2neg23 (46.9%)48 (47.1%)71 (47.0%)32.4%RISK CRITERIALymph Node Positive24 (49.0%)65 (63.7%)89 (58.9%)27.0%Triple Negative (ER/PR&amp;lt;10%)27 (55.1%)50 (49.0%)77 (51.0%)35.0%Non-pCR11 (22.4%)25 (24.5%)36 (23.8%)30.6%RS ≥ 25 and/or High Risk MP6 (12.2%)8 (7.8%)14 (9.3%)42.9%Median T size (cm) -excluding NACT2.1 (1.5-2.9*)1.8 (1.3-2.8)1.8 (1.3-2.9)N/APRIOR THERAPYAdjuvant Chemo25 (51.0%)60 (58.8%)85 (56.3%)29.4%Neoadjuvant Chemo22 (44.9%)41 (40.2%)63 (41.7%)34.9%Endocrine Therapy19 (38.8%)47 (46.1%)66 (43.7%)28.8%XRT29 (59.2%)75 (73.5%)104 (69.3%^)27.9%*Ranges represent interquartile range^ XRT data not available on 1 patient; n=150 was used to figure percentage Citation Format: Lauren Bayne, Isoris Nivar, Brooke Goodspeed, Shannon Deluca, E. Paul Wileyto, Natalie Shih, Anupma Nayak, Michael D Feldman, Joshua Edwards, Kevin Fox, Jennifer M. Matro, Susan Domchek, Hayley Knollman, Rachel Jankowitz, Angela Bradbury, Payal D. Shah, Jewell Graves, George Woodfield, Elizabeth Chislock, Jianping Wang, George Belka, Lewis A. Chodosh, Amy S. Clark, Angela DeMichele. Identifying breast cancer survivors with dormant disseminated tumor cells: The PENN-SURMOUNT screening study [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD9-11.

Jumping translocations of chromosome 1q occurring by a multi-stage process in an acute myeloid leukemia progressed from myelodysplastic syndrome with a TET2 mutation

BackgroundJumping translocations (JTs) are rare chromosome rearrangements characterized by re-localization of one donor chromosome to multiple recipient chromosomes. Here, we describe an acute myeloid leukemia (AML) that progressed from myelodysplastic syndrome (MDS) in association with acquisition of 1q JTs. The sequence of molecular and cytogenetic changes in our patient may provide a mechanistic model for the generation of JTs in leukemia.Case presentationA 68-year-old man presented with pancytopenia. Bone marrow aspirate and biopsy showed a hypercellular marrow with multilineage dysplasia, consistent with MDS, with no increase in blasts. Karyotype and MDS fluorescence in situ hybridization (FISH) panel were normal. Repeat bone marrow aspirate and biopsy after 8 cycles of azacitidine, with persistent pancytopenia, showed no changes in morphology, and karyotype was again normal. Myeloid mutation panel showed mutations in RUNX1, SRSF2, ASXL1, and TET2. Three years after diagnosis, he developed AML with myelodysplasia-related changes. Karyotype was abnormal, with unbalanced 1q JTs to the short arms of acrocentric chromosomes 14 and 21, leading to gain of 1q.ConclusionsOur patient had MDS with pathogenic mutations of the RUNX1, SRSF2, ASXL1, and TET2 genes and developed 1q JTs at the time of progression from MDS to AML. Our data suggest that the formation of 1q JTs involves multiple stages and may provide a mechanistic model for the generation of JTs in leukemia.

Acute Myeloid Leukemia (AML) Treated with Azacitidine: Survival Outcomes for Patients Who Complete More Than Six Cycles Are Similar to High-Risk Myelodysplastic Syndrome/Low Blast Count AML

INTRODUCTION: Despite accumulating evidence supporting the efficacy of hypomethylating agents in patients with AML and &gt; 30% bone marrow blasts as well as in relapsed/refractory AML, this therapy is not yet funded by National Health Plans / Healthcare Funding Agencies in a number of countries including Canada. The assistance of an industry-sponsored compassionate program has enabled provision of azacitidine for this group of patients at The Ottawa Hospital. We report here our local "real-world" experience of azacitidine efficacy in this diverse group of AML patients and identify a sub-group whose outcomes are equivalent to that of patients with higher-risk Myelodysplastic Syndrome (MDS) and AML with 20-30% blasts for whom azacitidine therapy has funding approval in Canada. METHODS: All patients who received azacitidine at The Ottawa Hospital between 2009 and 2016 were included in this single-center, retrospective analysis. Azacitidine was administered at a dose of 75mg/m2 subcutaneously daily for 7 consecutive days every 28 days. Response was evaluated with a repeat bone marrow aspirate and trephine biopsy after the 6th cycle. In those patients confirmed to have stable or responsive disease, azacitidine was continued until progression of disease, intolerable side-effects of the drug or the patient chose to discontinue therapy. Overall survival curves were generated using the Kaplan-Meier method and log-rank tests were used to compare subgroups of patients. Actuarial median survival months were calculated with 95% confidence intervals (CI). P-values less than 0.05 were considered statistically significant. RESULTS: During the study period, 109 patients received azacitidine: 54 had MDS /AML with 20-30% blasts (the 'funded' group) and 55 had either AML with &gt; 30 % blasts (n=23), AML relapsed post-intensive chemotherapy (n=14), AML relapsed post-allogeneic stem cell transplant (n=10) or primary refractory AML (n=8) (the 'unfunded' group). Median survival of the 'funded' group was 12.2 months while median survival of the 'unfunded' group was 5.6 months (95% CI 3.3-7.7; p=0.0058). Of the AML patients in the 'unfunded' group, 24% completed more than 6 cycles of azacitidine compared to 52% of patients in the 'funded' group. In both the 'funded' and 'unfunded' groups, patients who completed more than 6 cycles of azacitidine had similar survival outcomes (p=0.7277): the 'funded' group had a median survival of 19 months (95% CI 14.4-25.3) while the median survival of this sub-population of the 'unfunded' AML group was 22 months (95% CI 11.7-24.9). Patients in both groups who failed to complete more than 6 cycles of azacitidine also had a similar outcome (p=0.39), with a median survival of 5.7 months (95% CI 4.0-6.3) for patients with MDS/AML 20-30% blasts and 3.6 months (95% CI 2.2-5.1) for AML patients with &gt; 30% blasts or relapsed/refractory disease. Reasons for patients not completing at least 6 cycles of azacitidine included progression of disease (25%), bacterial infections most commonly pneumonia (53%) and patient preference (7%). CONCLUSION: A significant sub-population of AML patients with &gt; 30% blasts or refractory/relapsed AML can achieve a meaningful survival benefit with the hypomethylating agent, azacitidine. A higher proportion of this AML patient population discontinued azacitidine as a result of infective complications. The provision of routine prophylactic antibiotics may enable more patients with AML to receive an adequate amount of azacitidine to achieve therapeutic benefit and warrants further investigation. Our results add to the growing body of 'real-world' evidence that supports healthcare funding agencies to provide coverage of azacitidine for patients with AML who in some countries at present do not fulfill government funding criteria. Disclosures Bredeson: Otsuka: Research Funding. Maze:Pfizer Inc: Consultancy; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees. Sabloff:Novartis Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; ASTX: Membership on an entity's Board of Directors or advisory committees, Research Funding; Actinium Pharmaceuticals, Inc: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Astellas Pharma Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi Canada: Research Funding.

Type B Lactic Acidosis in Adults with Hemophagocytic Lymphohistiocytosis

Hemophagocytic lymphohistiocytosis (HLH) is a rare syndrome that can present with variable clinical and laboratory manifestations, and some distinct features are seen in adults. Type B lactic acidosis, which occurs in the absence of tissue hypoxia, has been reported in a few adult patients with HLH. We report a case of HLH presenting with type B lactic acidosis, with a literature review of this association. A 48-year-old Saudi woman with a past medical history of hypertension, dyslipidemia, and morbid obesity, presented with a 1-month history of intermittent high-grade fever up to 39°C daily. The patient was admitted for further evaluation of fever and was in our hospital for 70 days. Initial laboratory investigations revealed normocytic hypochromic anemia (102 g/L [normal 120-160]), lactic acidosis (2.36 mmol/L [0.5-2.2]), mild transaminitis, and elevated levels of lactate dehydrogenase (1761 U/L [125-220]), erythrocyte sedimentation rate (95 mm/hr [0-20]), and ferritin (1090.5 μg/L [4.6-204.0]). Further investigations failed to reveal any autoimmune or rheumatologic disease and extensive septic workup was negative. Lymphocyte subset by flow cytometry showed a low percentage of absolute natural killer cells (5% [10-20%]). Abdominal CT revealed splenomegaly with a span of 16.5 cm. Positron emission tomography-CT scan showed diffuse prominence of splenic activity, with a standardized uptake value of 4.5, and bone marrow activity throughout the axial skeleton. Liver biopsy and bone marrow aspiration were unremarkable. A month after admission, her anemia worsened with the hemoglobin level gradually dropping to 63 g/L prompting multiple transfusions. Lactic acid levels remained persistently elevated throughout this period [Figure 1]. A trial of intravenous steroids and immunoglobulins failed to decrease the lactic acid level [Figure 1; A to B]. The hospital course was complicated by Acinetobacter baumannii pneumonia requiring antibiotics [Figure 1; C to E]. Two months after the initial presentation, she developed pancytopenia, and repeated bone marrow aspiration and biopsy showed hemophagocytosis. Repeated ferritin level was more than 40,000 μg/L with hypertriglyceridemia (2.92 mmol/L [&lt;1.7]), and hypofibrinogenemia (0.9 g/L [1.5-4.1]). HLH-94 protocol with etoposide (150 mg/m2twice weekly) and dexamethasone (10 mg/m2) was commenced; however, the patient continued to deteriorate prompting elective intubation. She then developed acute acalculous cholecystitis and brain CT revealed a large central pontine hypodensity. Nine days after initiation, the patient was switched from the HLH-94 protocol to alemtuzumab, however, the patient arrested and passed away. To our knowledge, this is the first report of HLH presenting with persistent lactic acidosis lasting several weeks. Although no cause was found, this patient likely had HLH that was not detected early. We searched the PubMed and EMBASE databases for English-language reports describing adult patients (&gt;18 years) with HLH and lactic acidosis. Patients with type A lactic acidosis, associated with tissue hypoxia, were excluded. Five case reports describing patients with variable underlying diagnosis causing HLH and type B lactic acidosis were identified (Table I). Lactic acidosis started before HLH was diagnosed in all patients except one, in whom lactic acidosis developed two days after diagnosis. All patients had neither sepsis nor hypotension at the time of lactic acidosis. The mechanism of type B lactic acidosis in HLH is poorly understood and is likely multifactorial. As HLH in adults is more likely to be secondary to malignancies, Warburg effect, a high rate of glycolysis exhibited by tumor cells, has been proposed as the cause in these cases. Alternatively, acute cytokine hyperactivation in inflammatory states can enhance lactate dehydrogenase activity leading to excess lactic acid production. Inflammation, infection, or tumor cells can also induce stress and accelerate aerobic glycolysis-which does not depend on hypoxia-resulting in excess lactate production. Finally, acute hepatic dysfunction, a common finding in adult-onset HLH, can impair lactate clearance and further elevate levels. This report suggests that type B lactic acidosis may be a manifestation of HLH. Analyses of large HLH registries can better clarify the diagnostic and prognostic value of type B lactic acidosis in adult-onset HLH. Disclosures No relevant conflicts of interest to declare.

Low-dose Oral Ketamine as a Procedural Analgesia in Pediatric Cancer Patients Undergoing Bone Marrow Aspirations at a Resource-limited Cancer Hospital in India.

Aim:Many pediatric cancer patients undergo repeated bone marrow aspirations (BMAs) for diagnostic and treatment evaluation purposes. Full anesthesia is the standard of care during this procedure in high-income countries. At hospitals with low resources in low/middle-income countries many children undergo these painful procedures without sufficient pain relief. This study aimed to evaluate the usefulness of low-dose oral ketamine as a procedural analgesic in a low-resource pediatric cancer care department.Materials and Methods:Pediatric patients, 4–15 years of age, who underwent BMAs between September 31 and November 30, 2018, were invited to participate. The study was designed as a placebo-controlled, single-blinded trial with three trial groups. Group K received 1.0 mg/kg of ketamine and Group KM received 1.0 mg/kg ketamine with an addition of 0.2 mg/kg midazolam, mixed in juice 30 min before procedures. Group P received placebo consisting of plain juice. All three groups also received the hospital's current standard treatment for procedural pain in BMAs. Patients and caregivers assessed the procedural pain, as did the performing doctors. For the patients, Faces Pain Scale – Revised was used and the Numeric Rating Scale-11 for caregivers and doctors.Results:A total of 87 patients were included in the study distributed with 29 in Group K, 29 in Group KM, and 29 in Group P. Seven patients were excluded, one patient denied participation and the remaining did not meet the inclusion criteria. There was no significant difference between the pain reported by the groups. A total of 69% patients in Group KM and 35% in Group K had somnolence reported as a side effect compared to 14% in Group P.Conclusion:We found no significant effects on the procedural pain in any of the treatment groups compared to placebo. There were only mild side effects. The doses of ketamine might be insufficient for this painful and stressful procedure.

Molecular Residual Disease Monitoring Provides Insufficient Lead-Time to Prevent Morphologic Relapse in the Majority of Patients with Core-Binding Factor AML

IntroductionCore-binding factor (CBF) acute myeloid leukemias (AML) are characterized by the chromosomal abnormalities t(8;21) or inv(16)/t(16;16) and their fusion proteins RUNX1/RUNX1T1 and CBFB-MYH11, respectively. Despite their relatively favorable prognosis, it has been reported that up to 45% of patients with CBF-AML relapse. Current guidelines recommend monitoring the peripheral blood (PB) or bone marrow (BM) for molecular evidence of residual disease (RD) every 3 months for 2 years following remission. By monitoring patients for rising molecular transcripts, those at risk of impending relapse can be identified and treated prior to the emergence of overt disease. However, the relapse kinetics of CBF-AML are poorly-understood, and it is unknown if serial RD monitoring can detect molecular relapses with sufficient lead-time to intervene and prevent morphologic relapse.MethodsAfter local REB approval, we identified patients with CBF-AML treated at the Princess Margaret Cancer Centre in Toronto, Canada between 2000 and 2017. Patients underwent induction and consolidation chemotherapy according to standard protocols followed by RD monitoring with polymerase chain reaction (qPCR) of RUNX1/RUNX1T1 or CBFB-MYH11 transcripts in a CAP/CLIA certified lab every 3 months for a median of 1.2 years (range 0-5.3). RD assessment was performed on BM aspirates, but PB could be tested in patients who could not tolerate repeat BM aspirates. Morphologic relapse was defined as emergence of >5% blasts in the PB or BM, and molecular relapse as a positive transcript PCR (if previous PCR measurements were undetectable) or an increase by 1 log (if previous PCR measurements were detectable) on 2 successive samples without morphologic relapse. Rapid relapse was defined as <100 days from molecular to morphologic relapse (i.e. timeframe insufficient to administer pre-emptive treatments such as allotransplant to prevent morphologic relapse). We extracted relevant clinical and cytogenetic data and performed unpaired t-tests and Fisher's exact tests as appropriate using GraphPad statistical software and version 9.4 of the SAS System for Windows (SAS Institute, Cary, NC).ResultsWe included 114 patients with CBF-AML. Median age was 46.5 years (range 18-79). t(8;21) was present in 59% and inv(16)/t(16;16) in 41% of patients. All patients achieved remission with 7+3 induction chemotherapy. Over a median follow-up time of 3.7 years (range 0.2-14.3), RD measurements were performed a mean of 5 times per patient with mean sampling interval of 103±54 days. Remission was maintained in 71 (62%) patients but 43 (38%) developed morphological (n=34) or isolated molecular relapse (n=9), with median time to relapse of 4.4 months (range 1.4-31.4). Patients with relapsed disease were significantly less likely to have achieved ≥3 log reduction in RUNX1/RUNX1T1 or CBFB-MYH11 BM transcripts at the end of consolidation chemotherapy compared to patients who remained in remission (75% vs 90%, p=0.046). Of the 43 patients who relapsed, the majority (74.4%, n=32) had rapid relapse kinetics. Of these 43 patients, 25 received reinduction chemotherapy with achievement of CR2, 6 had refractory disease or death, 1 was lost to follow-up, and 17 went on to receive allotransplantation. RD monitoring enabled timely detection of impending relapse and permitted intervention prior to morphologic relapse in only 11 patients (25.6%). Among these 11 patients with slower relapse kinetics, 6 received reinduction chemotherapy with achievement of CR2, 2 received reinduction chemotherapy with refractory disease, and 8 went on to receive allotransplantation. Median overall survival was 20 months (range 3-128) for patients with rapid relapse kinetics and 28 months (range 14-166) for patients with slow relapse kinetics (p=0.02). Clinical features, BM vs PB RD measurements, additional molecular mutations, and cytogenetic abnormalities did not distinguish patients with rapid vs slow relapse kinetics.ConclusionsCurrent guidelines recommend molecular RD monitoring every 3 months for CBF-AML. However, in the majority of patients who relapsed at our institution, RD monitoring every 3 months provided insufficient lead-time to identify molecular relapses prior to morphologic relapse. Further research is warranted to identify the patients with CBF at the highest risk of relapse and the best strategies to monitor these patients over time. DisclosuresGupta:Novartis: Consultancy, Honoraria, Research Funding; Incyte: Research Funding. Maze:Novartis: Consultancy, Honoraria. Schuh:Celgene: Consultancy; Novartis: Consultancy; Pfizer: Consultancy; Jazz: Consultancy; Amgen Inc.: Consultancy; Shire: Consultancy; Teva: Consultancy; Otsuka: Consultancy. Yee:Celgene, Novartis, Otsuka: Membership on an entity's Board of Directors or advisory committees; Agensys, Astex, GSK, Onconova, Genentech/Roche: Research Funding. Schimmer:Otsuka Pharmaceuticals: Consultancy; Jazz Pharmaceuticals: Consultancy; Medivir AB: Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Abstract CT137: Biomarkers of response to leukemia stem cell targeted therapy with low dose daunorubicin in relapsed/refractory acute leukemia

Abstract Patients with acute leukemia harbor chemotherapy-resistant leukemia stem cells (LSC) which ultimately lead to disease relapse in many patients. Clinical trial design of LSC targeted therapy is a challenge. The majority of leukemia cells are differentiated leukemia cells, with LSC comprising &amp;lt;1%. Clinical benefit of LSC targeted therapy is best measured by overall survival and difficult to measure in the short-term using the conventional endpoint of remission. Response to LSC targeted therapy may be confounded by lack of response of the bulk population. Optimal treatment strategies may ultimately include combined or sequential therapy targeted at both the LSC and bulk leukemia cell populations. However, this approach makes it difficult to measure effects on LSC specifically. In order to isolate specific effects of targeted therapy on LSC, we designed a clinical proof of concept study using serial bone marrow aspirates to measure the effects of LSC-targeted therapy in adults with relapsed acute leukemia. In pre-clinical models, we demonstrated that AKT-β-catenin interaction, indicated by Akt phosphorylation of β-catenin at serine 552 (pS552) is critical for LSC self-renewal. High throughput screening identified daunorubicin (DNR) as a potent inhibitor of pS552. To better understand the role of DNR in specifically targeting LSC, we experimented with lower doses, recognizing that most leukemia patients receive high dose DNR, yet LSC persist and result in disease relapse. In our murine model, low dose anthracycline inhibited LSC number and function. We hypothesized that low dose DNR treatment inhibits LSC number in patients with acute leukemia. Our pilot trial includes relapsed acute myeloid (AML) and lymphoblastic leukemia (ALL) patients (NCT02914977). Bone marrow aspiration was performed at study entry. Patients received DNR at 6.75 mg/m2/day x 5 days followed by repeat bone marrow aspiration on day 8. The plasma pharmacokinetics of low dose DNR was also characterized. Samples from pre- and post-treatment were analyzed by flow cytometry for the frequency (%) of total cells that are pS552-β-catenin positive LSCs (defined as CD34+ CD38- cells that express TIM3, which distinguishes LSCs from normal stem cells). Nine patients (8 AML, 1 ALL) have been treated. Seven patients had previous DNR treatment. Two patients had prior allogeneic stem cell transplant. Five of 9 patients had reduction in LSC number as measured by flow cytometry; 3/5 had prior DNR treatment. In this pilot proof of concept trial, we demonstrate the feasibility of serial bone marrow aspiration to measure biomarkers of LSC response. In our small series, 5/9 relapsed leukemia patients had reduction in LSC number after one cycle of low dose DNR. Future plans include incorporating low dose DNR following conventional chemotherapy in newly diagnosed patients with AML. This clinical proof of concept trial design with serial bone marrow aspirations may also be used to rapidly assess the validity of other LSC markers and targeted agents. Citation Format: Tara L. Lin, John M. Perry, Xi He, Gregory Reed, Na Zhang, Scott Weir, Joseph P. McGuirk, Linheng Li. Biomarkers of response to leukemia stem cell targeted therapy with low dose daunorubicin in relapsed/refractory acute leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT137.

Hemophagocytic Lymphohistiocytosis Associated With Visceral Leishmaniasis

This is the report of a 2-year-old boy who presented with fever, cytopenia, and splenomegaly. The patient was diagnosed with hemophagocytic lymphohistiocytosis (HLH) and treated with HLH-2004 protocol. Repeated bone marrow aspiration showed amastigotes on follow-up. In endemic countries, visceral leishmaniasis should be considered in the differential diagnosis to avoid chemotherapy toxicity.

178 A Rare Case of Refractory Anemia With Ringed Sideroblasts and Chronic Myelomonocytic Leukemia: Myelodysplastic/Myeloproliferative Disease

We present an unusual case of myeloid neoplasm presenting with mixed features of refractory anemia with ringed side oblasts (MDS-RARS) and chronic myelomonocytic anemia (CMML). A 62-year-old man initially presented with macrocytic anemia and fatigue. CBC showed Hgb 11.9 g/dL, MCV100.9 fL, WBC 9.0 x 109/L, PLT 489 x 109/L. Bone marrow examination revealed a hypercellular marrow (80% cellular), erythroid hyperplasia with mild dyserythropoiesis, increased megakaryocytes with dysplastic forms. Storage iron was increased with 70% ringed sideroblasts. Cytogenetic studies were negative. A diagnosis of, MDS-RARS was rendered. The patient received Neupogen and Procrit. Over the next few years the patient became refractory to Procrit. The CBC showed macrocytic anemia and absolute monocytosis. A repeat bone marrow aspirate showed trilineage dyspoiesis, absolute monocytosis and 3% blast. Storage iron was increased with increased ringed sideroblasts (>15%). Flow cytometry demonstrated a population of myeloid blasts (CD34, CD117, CD13, HLA-DR, and CD33 positive), comprising 3% of total cells and a population of monocytoid cells (CD4, CD36, CD11b, CD14, CD36, and CD45) comprising 27%. A next-generation sequencing panel showed a mutation in FLT3 (seen in 3% of CMML), RUNX1 (seen in 10%-37% of CMML) and SF3B1 (seen in >80% of, MDS RARS). Based on the morphologic, immunophenotypic and molecular studies the diagnosis was revised to myelodysplastic/myeloproliferative neoplasm with overlapping features of RARS and CMML. Studies have found that CMML with more dysplastic features share common molecular features with, MDS-RARS, and this might represent a continuum in the morphologic expression of a molecular lesion. This entity is currently not recognized in the World Health Organization classification of myeloid neoplasms.

Proptosis in orbital myeloid sarcoma: A precursor to acute leukaemia

Myeloid sarcoma is a rare, aggressive presentation of acute myeloid leukemia which tends to affect bones and soft tissue. It is usually seen in children, and has a poor prognosis. We report a case of orbital myeloid sarcoma presenting with unilateral proptosis in a young adult who subsequently developed acute myeloid leukaemia. An 18-year-old male presented with sudden, progressively increasing right eye swelling over two weeks. Examination revealed right eye proptosis, with a positive relative afferent papillary defect and a right eye visual acuity of 6/60, pinhole 6/18. The right fundus showed optic disc hyperemia, tortuous veins and choroidal folds. The left eye was normal. On initial examination, computed tomography (CT) of the brain and orbit showed an enhancing mass at the lateral aspect of the right orbit measuring 3.8 × 2.1 cm, with optic nerve displacement. Right lateral orbitotomy and excisional biopsy yielded a tissue diagnosis of myeloid sarcoma. The peripheral blood smear was normal at this time. The patient defaulted treatment but presented again three months later with right eye massive proptosis and loss of vision. A repeated CT scan showed a large lobulated mass measuring 104.5mm × 87.21mmm × 77.25mm occupying the right orbit, with destruction of the bony orbital walls and intracranial extension. A repeat full blood picture and bone marrow aspiration showed 30% blast cells, consistent with the diagnosis of acute myeloid leukemia. Patient was started on chemotherapy and radiotherapy. The tumour reduced in size after five cycles, but unfortunately, patient passed away due to brain metastases.

A Single Center Retrospective Analysis of Adherence to Molecular Monitoring Guidelines in Patients with Chronic Myeloid Leukemia

Introduction In 2016, it is predicted that there will be 8,220 new cases of Chronic Myelogenous Leukemia (CML) in the United States, with an estimated 1,070 deaths from this disease (Siegel RL, et al, CA Cancer J Clin 2016, 66:7-30). Due to the prognostic significance of cytogenetic and molecular response to frontline Tyrosine Kinase Inhibitors (TKIs), expert panels such as the European LeukemiaNet and the National Comprehensive Cancer Network (NCCN) recommend strict guidelines to ensure adequate surveillance of patient's response to TKI therapy (Deininger M, et al, Blood 2009, 114: Abstract 1126). In this retrospective observational study, we studied adherence to current NCCN Guidelines in the management of CML. Methods Using both electronic and archived paper medical records, we identified 26 patients diagnosed with CML from 3/3/2003 to 12/9/2015 who were followed in a single community practice. Information at initial diagnosis regarding age, CML phase, spleen size, and initial TKI therapy selected were obtained. Monitoring parameters including frequency of follow-ups, complete blood counts, chromosomal analysis, molecular testing, and repeat bone marrow aspirates were reviewed. Evaluation for response to treatment with time to Complete Hematologic Response (CHR), time to Major Molecular Response (MMR), and time to Complete Molecular Response (CMR) were noted. Evaluations of treatment adherence and testing for BCR-ABL kinase mutations when milestones were missed or molecular progressions detected were recorded. Reasons for change in therapy and choice of subsequent treatment were also documented. Results The median age at diagnosis of CML was 53, with a male to female ratio of 1.6:1. In this group of 26 patients, only a single patient was diagnosed in the accelerated phase, the rest were in the chronic phase of CML. In 92%, clinical spleen size was logged. However, a single standardized method was not utilized which precluded calculation of Sokal scores for risk classification. In 88.5%, bone marrow aspirates were obtained to establish disease phase and enable bone marrow cytogenetic studies. Out of 26 patients, 16 (61.5%) were initially treated with imatinib, 6 (23%) were treated with dasatinib and 4 (15.4%) were treated with nilotinib. Attempts to achieve superior cytogenetic and deeper molecular response with dasatinib or nilotinib, coupled with lower rates of progression to advanced phases, were most frequently cited as reasons for choosing non-imatinib front line therapy. Only 50% of patients had complete compliance with periodic molecular testing at 3, 6 and 12 months during the first 2 years of diagnosis. The frequency of molecular and cytogenetic response assessments were often less than recommended. Only 2 of the patients studied underwent repeat bone marrow testing, with one enrolled in a clinical trial requiring periodic repeat bone marrow biopsies. The median time to achieve CHR was 27 days, median time to reach MMR was 26.5 weeks and median time to CMR was 7.9 months. Attempts to confirm patient compliance with oral TKI therapy were documented in 22 of the patients (84.6%). Three patients underwent testing for BCR-ABL kinase mutations at the time of molecular progression, however, none tested positive for resistance mutations. No patient had a hematological or cytogenetic relapse, and two patients switched from nilotinib to imatinib because of poor compliance due to toxicity. Conclusion Clinical practice guidelines promote evidence-based recommendations to guide clinicians in the management of CML. TKI therapy has evolved to be the primary treatment for chronic phase CML, and monitoring response to TKI therapy by setting cytogenetic and molecular milestones has improved quality of care and success with treatment. Unfortunately, real-world application of strict time lines for testing may not be feasible in all circumstances. Patient non-compliance with testing and marrow exams, interval illness and hospitalization, and errors in laboratory testing have served as barriers to adherence to NCCN guidelines. Despite a 50% compliance rate, there were no major untoward events (i.e. transformation to accelerated or blast phase) appreciated in this subset of patients. Disclosures No relevant conflicts of interest to declare.

Resolution of epoetin-induced pure red cell aplasia 2 years later, successful re-challenge with continuous erythropoiesis receptor stimulator

Epoetin-induced pure red cell aplasia (PRCA) is most commonly associated with epoetin-a; nevertheless, its occurrence has been reported in epoetin-β and darbepoetin-a. We report a young hemodialysis patient who developed PRCA 2 years after receiving intravenous epoetin-β. Epoetin- induced PRCA was confirmed by bone marrow aspiration, associated with markedly elevated anti-erythropoietin antibody. He was treated with prednisolone and cyclophosphamide for 3 months but continued to be transfusion-dependent. 17 months after the development of PRCA, he was started on intravenous continuous erythropoiesis receptor stimulator (CERA) in view of frequent transfusions. He tolerated the CERA injection well and the hemoglobin level stabilized 7 months later. Repeat bone marrow aspiration confirmed complete resolution of PRCA with disappearance of anti-erythropoietin antibody. To date, he maintained a stable hemoglobin level and has been transfusion-independent for the past 1 year. This is the first in the literature that reported the utilization of CERA in epoetin-induced PRCA. Very low or undetectable level of anti-erythropoietin antibody might be the key to the success of re-challenge strategy in cases of epoetininduced PRCA. Thus, routine checking of anti-erythropoietin antibody before the rechallenge with an alternative erythropoietin product is highly recommended.

Utility of peripheral blood for cytogenetic and mutation analysis in myelodysplastic syndrome

AbstractRecent studies have shown that more than 80% of bone marrow (BM) samples from patients with myelodysplastic syndrome (MDS) harbor somatic mutations and/or genomic aberrations, which are of diagnostic and prognostic importance. We investigated the potential use of peripheral blood (PB) and serum to identify and monitor BM-derived genetic markers using high-resolution single nucleotide polymorphism array (SNP-A) karyotyping and parallel sequencing of 22 genes frequently mutated in MDS. This pilot study showed a 100% SNP-A karyotype concordance and a 97% mutation concordance between the BM and PB. In contrast, mutation analysis using Sanger sequencing of PB and serum-derived DNA showed only 65% and 42% concordance to BM, respectively. Our results show the potential utility of PB as a surrogate for BM for MDS patients, thus avoiding the need for repeated BM aspirates particularly in elderly patients and those with fibrotic or hypocellular marrows.

Performance of a gravitational marrow separator, multidirectional bone marrow aspiration needle, and repeated bone marrow collections on the production of concentrated bone marrow and separation of mesenchymal stem cells in horses

Objective-To determine the efficiency of a novel point-of-care gravitational marrow separator and bone marrow aspiration needle for concentrated bone marrow production and bone marrow-derived mesenchymal stem cell (MSC) separation and assess the effect of repeated bone marrow collections in horses. Animals-8 healthy adult horses. Procedures-Bone marrow aspiration was performed twice (1 month apart) from sternebral bodies with a standard or prototype multidirectional needle. Concentrated bone marrow was obtained by gravitational marrow separation and evaluated for WBC and platelet counts, automated and cytomorphologic cell differential counts, MSCs, and cell viability. Results-Concentrated bone marrow samples obtained with the marrow separator had 5- to 19-fold bone marrow-derived MSC, WBC, and platelet counts, compared with original bone marrow samples. Use of a multidirectional needle increased the frequency of obtaining MSC-richer concentrated bone marrow. Repeating bone marrow aspiration at 1 month yielded greater MSC numbers but slightly lower cell viability after processing. Conclusions and Clinical Relevance-The gravitational bone marrow separator and multidirectional needle were used to effectively harvest bone marrow and improve the quality of concentrated bone marrow. Comparable, or even greater, numbers of bone marrow-derived MSCs were collected by repeated bone marrow aspiration after a 1-month interval from the same aspiration sites. Use of the marrow separator and multidirectional bone marrow aspiration needle can facilitate a 1-step, point-of-care, nonlaboratory method to obtain concentrated bone marrow as a mixture of bone marrow-derived MSCs and growth factors from platelets and plasma.

Induction Therapy for Gamma-Heavy Chain Disease with Bortezomib and Dexamethasone: A Case Report

Abstract 5051Heavy chain diseases (HCD) are rare B-cell lymphoproliferative disorders characterized by production of a monoclonal immunoglobulin without a light chain. Three subtypes are known based on the type of monoclonal immunoglobulin produced ie. alpha-heavy chain, gamma-heavy chain and mu-heavy chain disease in order of decreasing frequency. Here we report a case of gamma-heavy chain disease in a previously healthy 48 year old African American man, which demonstrated clinical and pathologic response to treatment with bortezomib and dexamethasone.The patient was transferred to our institution for work-up of persistent high fever, hypotension and eosinophilia after a thorough negative infectious and rheumatologic work-up spanning two outside hospital admissions. The patient had originally presented with two months of constitutional symptoms and cough refractory to antibiotics and prednisone. His outside hospital admissions were notable for an RCA NSTEMI (an unexpectedly large thrombus was noted on cardiac catheterization), eosinophilia and persistent fevers to 105 degrees Fahrenheit. Imaging studies included a CT torso and tagged WBC scan which were unrevealing but for mild splenomegaly of 15 cm and notably did not show lymphadenopathy, masses or evidence for infection. The fevers transiently abated with an empiric course of steroids but recurred accompanied by shock, which responded to a pulse of high dose methylprednisolone. Ultimately he was felt to have primary adrenal insufficiency without evidence of infiltrative disease on dedicated CT. On presentation to our hospital, his initial labs showed WBC 5. 3 K/uL (83% neutrophils, 10% lymphocytes, 3% monocytes, 4% eosinophils), Hb 11. 2 g/dL, platelets 173 K/uL. Comprehensive chemistry profile was unremarkable. SPEP/IFE of the serum and urine revealed a monoclonal gamma-heavy chain without associated light chain. The total IgG was elevated at 2522 mg/dL, IgA was mildly depressed at 57 mg/dL and IgM was normal at 147 mg/dL; the monoclonal protein itself could not be quantitated; hence, total serum IgG was followed. The patient underwent a bone marrow biopsy and aspiration which showed trilineage hematopoiesis, marked esosinophilia, no evidence of leukemia or lymphoma, 10% CD138+ plasma cells, with the overwhelming majority staining for IgG but not for kappa or lambda. A skeletal survey was negative for lytic lesions. The findings were felt to be consistent with a plasma cell dyscrasia associated gamma-heavy chain disease. The observed eosinophilia was felt to be reactive to the gamma-heavy chain disease as has been previously described.In the literature, gamma heavy chain disease is reported to most commonly present as a lymphoplasmacytic process and the finding of a plasma cell dyscrasia appears to be an unusual presentation of what is already a rare disease. We decided to tailor the treatment to the finding of a plasma cell dyscrasia. Therefore the patient was started on bortezomib 1. 3 mg/m2 SC D1, 4, 8, 11 and dexamethasone 20 mg PO D1, 2, 4, 5, 8, 9, 11, 12 of a 21 day cycle. His treatment was complicated by paroxysmal atrial tachycardia; a cardiac MRI did not reveal evidence of infiltrative cardiomyopathy. After 4 cycles of bortezomib and dexamethasone, the patient's total IgG had normalized at 1265 mg/dL and though a monoclonal gamma heavy chain was still detected by IFE of the serum and urine, a repeat bone marrow biopsy and aspiration was without morphologic evidence of the heavy chain plasma cell dyscrasia. The patient reported feeling subjectively improved with abatement of his malaise and fatigue and without further episodes of fever. Interestingly, the parameter that most seemed to correlate with his clinical status was eosinophilia which also decreased from a peak of 61% (absolute eosinophil count of 5063/uL) to 8–19% after 4 cycles with bortezomib and dexamethasone. Secondary eosinophilia has been reported in the literature in a subset of gamma HCD patients. The patient further underwent stem-cell mobilization in preparation for auto stem cell transplant with high dose Cytoxan, after which the eosinophilia completely resolved.There are only approximately 120 cases of gamma HCD described in the literature. To our knowledge, this is the first reported case of plasma cell dyscrasia associated gamma HCD responding to treatment with bortezomib and dexamethasone. Disclosures:No relevant conflicts of interest to declare.

Utility of Peripheral Blood for Cytogenetic and Mutation Analysis in Myelodysplastic Syndrome

Abstract 1707With the advent of high throughput and high resolution techniques, >80% of myelodysplastic syndrome (MDS) patients harbour somatic mutations and/or genomic aberrations, which provide diagnostic and prognostic utility; however, frequent bone marrow (BM) aspirates are required. In a significant minority, the BM is hypocellular and fibrotic with suboptimal in vitro growth and, additionally, the procedure causes discomfort particularly in the elderly. This led us to investigate the use of peripheral blood (PB) and serum to identify and monitor BM derived genetic markers using high resolution single nucleotide polymorphism array (SNP-A) karyotyping and 454 parallel sequencing (454-PS) of a 22 gene myeloid panel comprising of all the exons of DNMT3a, RUNX1, CEBPα, TP53, EZH2, TET2 and ZRSR2 and mutations ‘hotposts' for NPM1, FLT3, ASXL1, IDH1, IDH2, MPL, JAK2, BRAF, cCBL, NRAS, KRAS, C-KIT, SF3B1, SRSF2, and U2AF1.We selected 23 MDS patients with concurrent BM and PB samples and detected 45 mutations in TET2, SF3B1, ASXL1, TP53, DNMT3a, FLT3, U2AF1, NRAS, cCBL, JAK2 and IDH2 in BM and subsequently analysed their PB using 454-PS with independent validation performed by Sanger sequencing (SS). All the mutations identified in BM were detected in PB with the exception of a single NRAS (BM-11%). Nine patients had a single mutation in SF3B1, ASXL1, TP53, TET2, DNMT3a and U2AF1 with the remaining patients having multiple coexisting mutations. Overall there was no significant difference in the mutation burden between the PB (median 25%(1.5%-50%)) and the BM (median 33%(5%-68%)). Concurrent analysis of unamplified and whole genome amplified PB DNA from 3 patients with mutations in TET2, U2AF1, ASXL1 and NRAS showed no difference in their mutation profile. As expected, in three patients post therapy the mutation burden in the PB was lower than in the pre-treatment BM sample.The lower mutation burden and sequence context in the PB contributed significantly to the quality of SS analysis. Prior knowledge of the mutation site resulted in 98% concordance (smallest clone size - 1.5%) between BM and PB, however, a blind approach decreased this to 84% (smallest clone size - 15%). In addition, serum was available from 14 patients (22 known mutations in BM) and SS of serum DNA identified 12 mutations correctly (U2AF1, FLT3, SF3B1, TET2, TP53, ASXL1, DNMT3a and IDH2, 4 as wildtype (TET2, cCBL, ASXL1 and SF3B1) and 6 failed to amplify (ASXL1, TP53 and TET2) without any preference for specific genes and the failure attributed to the quantity of serum DNA.Karyotype aberrations in PB were assessed using Affymetrix SNP 6.0 arrays on 31 MDS patients; normal karyotype (n=11), del5q (n=9), del7q/-7 (n=5), trisomy 8 (n=2), complex (n=2), isodiXq13 (n=1) and t(2:4)(q33;q27). An overall karyotype concordance of 94% was observed in PB with the 2 discordant cases showing normal karyotype in PB and BM by SNP-A but having monosomy 7 (partial cytogenetic remission after 5-azacitidine) and t(2:4)(q33;q27) respectively in their BM by MC. The mean copy number (CN) in the PB was lower than BM (PB vs BM); deletions (CN of 1.8 vs 1.6) and gains(CN of 2.2 vs 2.4), implying a smaller abnormal clone in the PB. Concurrent SNP-A karyotype from BM from 9 patients was concordant with SNP-A karyotype from PB, although a patient with complex karyotype determined by SNP-A in the BM and having 30 aberrations had only 15 aberrations detected in the PB.To determine if the 5q deletion was lineage restricted, we enriched PB for CD3+, CD19+ and CD3-CD19- populations from 4 patients. FISH and SNP-A karyotyping showed the presence of the 5q deletion using both techniques in all three fractions, however at a lower level in PB lymphocytes indicating the presence of a smaller clone.In conclusion, our study showed an excellent concordance between BM and PB, both for karyotype and mutational analyses using high resolution SNP-A karyotyping and 454-PS suggesting its clinical utility as a surrogate for BM, thereby, avoiding the discomfort of repeated BM aspirates and help in monitoring response to therapy more frequently. Disclosures:No relevant conflicts of interest to declare.