Citations

Abstract Abstract 1909 AL amyloidosis is the most common type of systemic amyloidosis. It is caused by extracellular deposition of circulating immunoglobulin light chains (LC) thought to be secreted by a primarily bone marrow based clonal plasma cell proliferative disorder. Although, invariably, the type of LC secreted by the plasma cell clone is identical to the LC deposited in the amyloid plaque, a comprehensive analysis of the re-arranged clonal LC gene coding secreted LC protein, the protein sequence of the serum and urine monoclonal band and the LC protein deposited in the amyloid plaques have not been previously performed. In this study, we compared the protein sequence predicted by the clonal LC gene sequence with the protein sequence identified in the serum and urine monoclonal bands and protein plaques by mass spectrometry (MS) based proteomics. 10 cases with systemic AL amyloidosis were studied. Bone marrow samples containing the clonal plasma cells were obtained and clonal LC variable gene sequences were determined by PCR amplification, cloning and sequencing as described previously (Abraham et al, Blood 2003, 101: 3801). For protein analysis, monoclonal protein bands identified in serum and/or urine protein electrophoresis and immunofixation were isolated from archival gels. For amyloid plaque protein analysis, the plaques were microdissected from paraffin embedded tissue. To identify the protein amino acid sequence of serum and urine bands and amyloid plaques, the samples were digested into peptides by trypsin and analyzed by MS based proteomic analysis. (see Vrana et al. and Blood 2009; 114: 4957 for methods). The results for amino acid sequence predicted from the clonal LC gene sequence, determined directly by MS from the serum and/or urine monoclonal band and amyloid plaques were then aligned and compared. In all 10 cases, the LC amino acid sequences predicted by sequencing of the clonal LC gene from bone marrow plasma cells, the amino acid sequence obtained directly from the monoclonal serum and/or urine protein and the amino acid sequence obtained from the amyloid plaque were identical. In most cases, both the complete variable region and constant regions were deposited as amyloid plaques. Some amyloid plaques showed cleavage of first 16 amino acids of the variable region suggesting that this part was not required for amyloid deposition. All LC variable region gene sequences were affected by somatic hypermutation, and identical mutations were present in both the serum and urine bands and in the amyloid plaques. In conclusion, these results, for the first time, comprehensively show that the monoclonal protein secreted by the neoplastic plasma cells is the pathogenetic protein causing the amyloid deposits in AL amyloidosis. Amyloid deposits contain virtually all of the LC protein identified in the serum or urine including both variable and constant regions, suggesting that the presence of the constant region may also play a role in amyloidogenesis. The technological capability to determine the complete protein sequence of the amyloidogenic monoclonal serum and urine LC protein isolated from archived gels will create new opportunities to develop diagnostic, predictive and prognostic serum or urine based tests for amyloidosis. Disclosures: Theis: Mayo Clinic: Patents & Royalties. Vrana:Mayo Clinic: Patents & Royalties. Dogan:Mayo Clinic: Patents & Royalties.

Abstract Background: Classical Hodgkin lymphoma (HL) is characterized by the presence of CD30-positive Hodgkin Reed-Sternberg cells. The standard of care for patients (pts) with relapsed or refractory (R/R) HL is salvage chemotherapy followed by high dose chemotherapy and autologous stem cell transplant (auto-SCT). Approximately 50% of these pts will experience relapse or progression of HL after auto-SCT. For these individuals, outcomes have historically been poor, with median overall survival (OS) from time of relapse ranging from 10.5 to 27.6 mos (Crump, 2008; Fanale, 2013). A pivotal phase 2 study evaluated brentuximab vedotin, a CD30-directed antibody-drug conjugate, in pts with R/R HL after auto-SCT (ClinicalTrials.gov #NCT00848926). Primary results and 3-year follow-up data have been previously reported (Younes, 2012; Gopal, 2015). Here, we summarize final data from the 5-year follow-up period. Methods: Pts received 1.8 mg/kg brentuximab vedotin once every 3 wks as a 30-min outpatient IV infusion for up to 16 cycles. The primary endpoint was the objective response rate (ORR) per independent review according to the Revised Response Criteria for Malignant Lymphoma (Cheson 2007). Assessments of response and durability of response per an independent review facility (IRF) have been previously reported. Following a protocol amendment that removed the requirement for routine CT scanning during the follow-up period, disease status was only assessed per the investigator. Survival and disease status were assessed every 3 mos for 2 yrs and then every 6 mos through Year 5. CT scans were required if progression was suspected clinically. At the time of the amendment, 18 patients were still being assessed for progression; these patients had been in long-term follow-up for a median of over 30 months. Results: The enrolled population of 102 pts (53% female) was heavily pretreated and had a median age of 31 yrs (range, 15-77 yrs). Pts received a median of 9 cycles (range, 1-16) of brentuximab vedotin. Per investigator, the ORR to brentuximab vedotin was 72% and complete remission (CR) rate was 33%. The most common treatment-related adverse events were peripheral sensory neuropathy, nausea, fatigue, neutropenia, and diarrhea. Adverse events of Grade 3 or higher that occurred in ≥5% of pts were neutropenia, peripheral sensory neuropathy, thrombocytopenia, and anemia. At study closure, which occurred approximately 5 yrs after the last patient's end-of-treatment visit, the median observation time for all enrolled patients from first dose was 35.1 mos (range, 1.8 to 72.9). The estimated 5-year overall survival rate was 41% (95% CI: 31 %, 51%) and the median OS was 40.5 mos (95% CI: 28.7, 61.9 [range, 1.8 to 72.9+]). Median OS by best clinical response was CR (n=34): median not reached; partial remission (PR, n=39): 39.4 mos; and stable disease (SD, n=28): 18.3 mos. The median PFS was 9.3 months overall, but was not reached in CR pts. Of the 102 enrolled patients, 15 remained in follow-up and in remission at study closure. Among these 15 pts, 6 received consolidative allo-SCT and 9 have received no further therapy since completing brentuximab vedotin. Conclusions: These end-of-study results demonstrate that single agent brentuximab vedotin can induce durable remissions and long-term survival in a subset of heavily pretreated patients with relapsed/refractory HL, particularly in pts that achieve CR. A randomized phase 3 study is being conducted to evaluate brentuximab vedotin in combination with AVD (doxorubicin, vinblastine, and dacarbazine) versus ABVD (A, bleomycin, VD) for frontline treatment of advanced HL (ECHELON-1 trial, ClinicalTrials.gov #NCT01712490). Figure 1. Overall Survival Figure 1. Overall Survival Disclosures Chen: Seattle Genetics: Consultancy, Research Funding; Genentech: Consultancy; Gilead: Consultancy; Janssen: Consultancy. Gopal:BMS: Research Funding; Piramal: Research Funding; Emergent/Abbott: Research Funding; Millenium: Honoraria, Research Funding; Janssen: Consultancy; Spectrum: Consultancy, Research Funding; BioMarin: Research Funding; Seattle Genetics: Consultancy, Honoraria; Gilead: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Sanofi-Aventis: Honoraria; Merck: Research Funding. Smith:Seattle Genetics: Research Funding; Celgene: Consultancy, Speakers Bureau. Ansell:Bristol-Myers Squibb: Research Funding; Celldex: Research Funding. Rosenblatt:Seattle Genetics: Research Funding. Savage:Seattle Genetics: Honoraria, Speakers Bureau; BMS: Honoraria; Infinity: Honoraria; Roche: Other: Institutional research funding. Connors:Roche: Research Funding; Seattle Genetics: Research Funding. Engert:Takeda Millenium: Honoraria, Research Funding; Seattle Genetics: Honoraria, Research Funding. Larsen:Seattle Genetics Inc.: Employment, Equity Ownership. Huebner:Takeda Pharmaceuticals International Co.: Employment, Equity Ownership. Fong:Seattle Genetics, Inc.: Employment, Equity Ownership. Younes:Janssen: Honoraria; Novartis: Research Funding; Incyte: Honoraria; Takeda Millenium: Honoraria; Celgene: Honoraria; Bristol Meyer Squibb: Honoraria; Curis: Research Funding; Bayer: Honoraria; Seattle Genetics: Honoraria, Research Funding; Sanofi-Aventis: Honoraria; Johnson and Johnson: Research Funding.

Abstract INTRODUCTION: Hemophagocytic lymphohistiocytosis (HLH) is a rare disorder caused by the pathologic activation of the immune system. In children, either a molecular diagnosis consistent with HLH or five out of the following eight criteria are considered necessary for a diagnosis of HLH (HLH-04 criteria): 1) fever; 2) splenomegaly; 3) cytopenia in two or more cell lines; 4) hypertriglyceridemia (≥265 mg/dL) or hypofibrinogenemia (≤150 mg/dL); 5) hemophagocytosis in the bone marrow, spleen, or lymph nodes; 6) hyperferritinemia (≥500 mcg/L); 7) impaired NK cell function; and 8) elevated soluble CD25 (sCD25). These criteria have been extrapolated to diagnose HLH in adults; however, it's unclear if these same criteria are applicable in the adult population. METHODS: We reviewed the Mayo Clinic electronic medical record for all adult (≥18 years) hospitalized patients with an admission serum ferritin of ≥500 mcg/L from January 2012 through December 2014. Patients' charts were reviewed and those who met the HLH-04 criteria were considered to have HLH. For the remainder of the patients, the etiology for hyperferritinemia was determined based on chart review and discharge diagnoses. Logistic regression models were used to assess the ability of these values in predicting a diagnosis of HLH. The Mayo Clinic IRB approved this study. RESULTS: We identified 1,329 patients with a serum ferritin ≥500 mcg/L. Of these, HLH was diagnosed in 28 (2.1%) patients (malignancy-associated HLH in 11 patients, infection-associated HLH in 4 patients, autoimmune-associated HLH in 7 patients, and idiopathic HLH in 6 patients). Table 1 describes the etiology of hyperferritinemia in the remaining 1,301 patients. In contrast to pediatric hospitalized patients (Allen, Pediatr Blood Cancer, 2008), adults are more likely to have malignancy (28.1% vs 7%; p<0.05); bacterial infection (21% vs. 13%; p=0.001); liver disease (9.9% vs. 2.7%; p<0.05); and cardiac disease (7.2% vs. 1.5%; p=0.0001) as the etiology of hyperferritinemia during their hospitalization. Among all patients in the study, the following variables were associated with higher odds of having HLH compared to hyperferritinemia due to an alternate cause: elevated ferritin (odds ratio [OR] 35.97, p<0.01); thrombocytopenia (OR 15.22, p<0.01), cytopenias defined by HLH-04 criteria (OR 8.04, p<0.01); elevated admission serum bilirubin (OR 1.05, p=0.02); and peak serum bilirubin (OR 1.05, p<0.01). After stepwise selection in multivariate analysis, serum ferritin ≥2,600 mcg/L and platelets ≤100 x 109/L were independently associated with HLH diagnosis (OR 24.9 and 7.8 respectively; p<0.01 for both). The area under the curve (AUC or c-statistic) ranges from 0.5 for no ability to discriminate, to 1.0 for perfect discrimination; this model has an AUC of 0.91, which shows very good discrimination between cases and controls. An adult hospitalized patient with a combination of serum ferritin ≥2,600 mcg/L and platelets ≤100x109/L predicts a ~200 fold increased likelihood of being diagnosed with HLH as compared to hospitalized adult meeting neither of these criteria (Figures 1 and 2). CONCLUSION: In contrast to hospitalized pediatric patients, the etiology of hyperferritinemia in adults is more likely to be malignancy, bacterial infection, liver disease, and cardiac disease. We conclude that a combination of serum ferritin ≥2,600 mcg/L and platelet count ≤100 x 109/L can be used as screening criteria to help identify adult patients most likely to have HLH. The traditional 5/8 criteria to diagnose HLH in pediatric patients do not appear necessary for establishing a diagnosis of HLH in adult patients. These observations need replication in an independent data set prior to broader applicability. Table 1.Patients with ferritin > 500 mcg/L with underlying cause for elevation.Number Diagnosed (%)Cardiac Disease95 (7.2%)Liver Disease131 (9.9%)Renal Disease87 (6.6%)Infectious343 (25.8%)Malignancy373 (28.1%)Autoimmune118 (8.9%)Solid Organ Transplant34 (2.6%)Stem Cell Transplant16 (1.2%)Bone Marrow Failure9 (0.7%)Shock39 (2.9%)Idiopathic53 (4.0%)Hemoglobinopathies3 (0.2%) Figure 1. ROC curve for the multivariable model with ferritin > 2,600 (yes/no) and platelets below 100,000 (yes/no) in the model with a very good discrimination c-statistic of 0.91. Figure 1. ROC curve for the multivariable model with ferritin > 2,600 (yes/no) and platelets below 100,000 (yes/no) in the model with a very good discrimination c-statistic of 0.91. Figure 2. Figure 2. Disclosures No relevant conflicts of interest to declare.

Abstract Background: Haploinsufficiency of GATA2 results in familial immunodeficiency and MDS/ AML. We identified a family with a novel c1339A>C, pS447R germline GATA2 mutation. Unlike most other disease-causing GATA2 mutations, this mutation occurred in the non-zinc finger domain. Methods: After due IRB approval, clinical and laboratory data were retrospectively abstracted. Results: Seven family members with the mutation are being followed at our institution. (Table 1), of whom two have MDS, one MDS/AML and one with isolated neutropenia. At the time of diagnosis 5/7(71%) pt had decreased monocyte count. 4/7(57%) patients had decreased NK cell count and/or function. Only 1/7(14%) patient had decreased CD4 count. 3/7(42%) patients had reduced B cell numbers and 5/7(71%) patients had persistent HPV warts. Other infections include invasive Absidia sinusitis, infectious mononucleosis, pertussis, and blood stream infections with S. maltophilia and S.aureus. One patient had the Emberger's syndrome presenting with lymphedema and congenital pulmonary stenosis. Pulmonary manifestation included a patient each with decreased DCLO and hyperreactive airway disease. Genitourinary manifestations included pelvic kidney/dysplastic kidney, posterior urethral valve, dermoid cyst and hydrocele. Two female patients developed primary infertility. One patient had a history of appendicular carcinoid. 4/7(57%) patients have successfully undergone allogenic stem-cell transplantation. Conclusion: Germline GATA2 mutations are associated with a high frequency of MDS/AML. We describe a family with a non-zinc finger GATA2 mutation causing MDS/AML, with a spectrum of hematological and non-hematological manifestations, once again, highlighting the heterogeneity of this syndrome and expanding the genotype-phenotype correlation in this disease. Abstract 5614. Table#Age/sexHemDAbso . MonoCount Abso. CD4 TCellNK cyto-toxicityAbso. NK CountAbso.B cell CountInfectiousDermCoag/VascularCVS/LungOthersNo.137/MAMLSeverely DecreasedNormalAbsentSeverely DecreasedAbsentAbsidia S.maltop- hilia S.aureusHPV WartsITP -Hydrocele 10 hyperparathyroidismNo.231/FMDSDecreasedNormalAbsentDecreasedNormalInfectious MonoHPV Warts-Hyper- reactive airwayInfertilityNo.317/FMDSSeverely DecreasedNormalUnknownDecreasedDecreasedHerpes stomatitisHPV Warts-Dec-DCLOPosterior urethral valveNo.49/MNeutropeniaSeverely DecreasedNormalNormalNormalNormalHSV-1HPV Warts Embergers syndrome Pul. Stenosis Congenital dysplastic Rt. Kidney Pelvic Lt. KidneyNo.531/F-DecreasedNormalDecreasedNormalNormal- -Qualitative platelet defectNoneInfertilityNo.635/F-NormalNormalDecreasedDecreasedDecreasedPertussisHPV Warts--Dermoid appendicular carcinoid No.756/F-NormalNormalNormalNormalNormalFrequent PNA---- Disclosures No relevant conflicts of interest to declare.

Abstract BACKGROUND: Complement-mediated thrombotic microangiopathies (TMAs) are a subset of TMAs in which the thrombocytopenia, microangiopathic hemolytic anemia, and end organ damage are caused by mutations in genes encoding the alternative complement pathway. Numerous complement genes have been implicated in complement-mediated TMAs including complement factor H (CFH), CD46 (MCP), complement factor I (CFI), complement component 3 (C3), complement factor B (CFB), factor H related 5 (CFHR5), and thrombomodulin (THBD) among others. Genetic analysis typically yields a mutation in 60% of patients whose TMA is presumed to be mediated by complement dysregulation. However, the description of novel disease-associated variants may increase this proportion. METHODS: A retrospective study of patients with TMAs diagnosed between 2000 and 2014 was performed. TMA diagnosis was made based on thrombocytopenia and evidence of microangiopathic hemolytic anemia. Analysis was performed with Alamut¨ software with additional in silico prediction tools (SIFT, MutationTaster, and Polyphen) for classification of gene variants. Variants of unknown significance (VUS) and likely pathogenic variants were further assessed using several mutation databases, including HGMD, ClinVar, and Factor H database RESULTS: Of patients diagnosed with a TMA, genetic analysis was performed in only a 10% of patients. Of the 29 patients with genetic studies performed, mutations were identified in 18 patients (62%). The majority of the mutations had been described previously in the literature, but four novel variants were identified: three missense and one splice-site. The table below summarizes these variants as well as laboratory findings on presentation. These were two variants of CFH, one variant of CFHR5, and one variant of CFI. In silico modeling of these variants revealed two polymorphisms likely to be pathogenic, one polymorphism likely benign given the lack of predicted splicing changes, and one VUS. Table 1. Protein Mutation Classification Age Sex Hemoglobin (g/dL) Platelets (thousands) Creatinine (mg/dL) CFH c.245-10_245-9dup Likely benign 61 F 11.6 51 6.7 CFH c.476G>A, p.Ser159Asn Likely pathogenic 43 F 9.1 101 9.1 CFHR5 c.1412G>A p.Gly471Glu Likely pathogenic 30 F 9.8 129 4.9 CFI c.1190T>A p.Val397Glu Unknown significance 51 M 9.4 125 6.1 DISCUSSION: With therapy available to target the alternative complement pathway, genetic analysis to identify genetic variants capable of causing complement mediated TMAs is an essential part of the evaluation. This genetic data must be interpreted and correlated with functional analysis and clinical phenotype. The reporting of novel variants in clinical databases, with inclusion of relevant clinical findings, is necessary to accurately classify and verify variants as pathologic mutations or benign polymorphisms. The full understanding of this diverse disease requires a more complete understanding of its genetics. While complement pathway-directed therapies are available, their rational use requires thorough interpretation of laboratory data, including genetic analysis. Disclosures Murray: Mayo Clinic: Patents & Royalties: Patent Application Filed.

Abstract Abstract 2928 Multiple myeloma (MM) is caused by the clonal expansion of malignant plasma cells (PCs) producing high levels of monoclonal immunoglobulin (Ig) or M-protein. However, ∼2% of all patients with monoclonal gammopathies (MG) have more than 1 M-protein, a condition termed biclonal gammopathy or double monoclonal gammopathy (DMG). These patients can express M-proteins with different heavy chain (HC) isotypes and different light chains (LCs) or different HCs that are LC-matched. Reports suggest that ∼87% of DMGs with different HCs have matched LCs, but without molecular analysis at the nucleotide level, clonal relationships are unknown. Thus, DMG patients may have either two independent malignant PC clones that coincidentally express the same type of LC or the two clones are related and reflect previous or ongoing class switch recombination (CSR). This latter variant would offer novel insight into the origin and differentiation of the malignant clone. Of note, prior reports of HC-dissimilar/LC-matched DMGs are limited to the analysis of single patients and with few exceptions; do not include molecular analysis of the variable regions in Ig HC or LC genes at the nucleotide level. In this study, we interrogate a cohort of MM patients with known HC-dissimilar/LC-matched DMGs for the frequency of clonal relatedness. The cohort was identified by searching our MG database for patients with: 1) HC-dissimilar/LC-matched Igs as determined by immunofixation; 2) >10% bone marrow (BM) PCs; and 3) no transplant prior to BM collection. Eight patients (7 MM and 1 smoldering MM/primary amyloid) were identified for further molecular evaluation. All 8 cases expressed IgG and IgA HCs. Three patients expressed kappa LC (K-LC) and 5 expressed lambda LC (L-LC). To determine if the IgG and IgA M-proteins were clonally related in each patient, we performed Ig HC variable gene (IGHV) PCR with sense primers to all IGHV genes and an antisense primer specific to IgG or IgA. In all 8 patients, a single productive IGHV gene was determined with IgG and IgA. In 2/8 patients, the IgG and IgA HC variable regions were clearly distinct and therefore genuinely biclonal. Of note, PCs from 6/8 patients used the same IGH V-D-J genes in both IgG and IgA transcripts. In 5 of these 6 patients, the entire IgG and IgA variable region and HCDR3 were identical at nucleotide and amino acid (AA) levels. Of interest, one patient had common, as well as distinguishing IGHV region somatic mutations (SHM) between the IgG and IgA transcripts. The HCDR3 regions (including N nucleotides) were highly homologous but differed by 7 nucleotides which resulted in 3 AA differences between the 2 clonally related sequences. While these are compelling data, it remained possible that the malignant PCs in these patients simultaneously expressed IgG and IgA as a result of long nuclear transcripts. To test this possibility, we used immunofluorescence (IF) and verified the presence of both IgG and IgA expressing PCs in all cases. Parallel analysis of both K-LC and L-LC variable gene by PCR was done on the 6 patients with the same IGHV gene in both IgG and IgA transcripts. In 3/6 patients, there was a single somatically mutated LC identified corresponding to the LC detected by immunofixation. These results were also confirmed by IF. The remaining 3 patients appeared to express more than 1 LC rearrangement. Taken together these results confirm that HC-dissimilar/LC-matched DMG can present with various phenotypes at the molecular level. We describe the largest cohort to date documenting the frequency of DMG patients that can be classified as expressing 2 identical HCs with either a single LC or >1 LC based on molecular analysis. The former case may reflect an original progenitor cell that divided with one daughter cell subsequently experiencing further CSR yet the LC remained untouched. In the case where the HCs were slightly different, there may have been CSR with additional SHM of a daughter cell, or the molecular differences may reflect random somatic mutations occurring independent of germinal center reactions. These results suggest that HC-dissimilar/LC-matched DMG may occur more often than thought. Larger studies tracking M-protein levels over time accompanied by cytogenetic analyses are needed to determine if the two clonally-related class switch variants in DMG patients exhibit similar biological properties. This analysis would offer new insight into the genesis of this unique subtype of MM. Disclosures: No relevant conflicts of interest to declare.

Abstract Background: In addition to NBEAL2, a single dominant-negative mutation in the GFIB1 gene has been associated with grey platelet syndrome but no association has been mentioned with dense granule deficiency. Here we describe a child with thrombocytopenia, alpha-delta granule deficiency, and a homozygous missense mutation in GFIB1. Case Report:The patient is an 8y/o Hispanic male born to non-consanguineous parents. Prenatal and birth history were unremarkable. No family history of blood disorders or pediatric malignancies. The boy has 2 healthy older siblings, no dysmorphic features, and normal skin pigmentation and eye findings. Renal function and hearing are normal. At 3mo he was noted to have multiple spontaneous petechiae along with an isolated thrombocytopenia of 46K/uL. At 10 months, a bone marrow evaluation showed increased megakaryocytes suggestive of ITP. By 3years of age, he received three treatments of IVIG without an adequate response. His platelet counts have generally ranged between 30-50K/uL. With acute illnesses, they drop to 15-20K/uL. His bleeding symptoms have primarily been spontaneous bruising and petechiae as well as prolonged epistaxis. His symptoms have been generally controlled with anti-fibrinolytic agents alone. Platelet transfusions have been reserved for surgical procedures or significant bleeding symptoms. Further evaluations over the past 5 years have included peripheral smears, showing atypical large hypo-granular platelets, and two additional bone marrow aspirates, showing megakaryocytic hyperplasia with numerous osteoclast-like forms and occasional small mono-lobated megakaryocytes and evident emperipolesis. Anti-nuclear antibody testing and platelet direct and indirect antibody testing were negative. Platelet electron microscopy showed that platelets virtually contained no dense granules (0.05 dense granules/platelet, 200 platelets) and about 30-40% of the platelets had markedly decreased alpha granules. Some platelets had complex canalicular networks and membrane vacuoles. These features are consistent with an alpha-delta platelet storage-pool deficiency. A myeloid malignancy mutation panel, performed using next generation sequencing, detected no variants of known significance. Gene sequencing for MHY9-related disorders as well as DNA breakage analysis for Fanconi's anemia were negative. However, DNA sequencing of the patient's sample revealed a homozygous missense mutation (c923 T>C; p.Leu308Pro) in the GFIB1gene; each parent carried one copy of this change. Conclusion: GFIB1, mapped to chromosome 9q34.13, encodes a transcriptional repressor that is important for megakaryopoiesis. It has been reported in patients with gray platelet syndrome; an inherited platelet disorder associated with thrombocytopenia and decreased alpha granules. This case points to an association of biallelic GFIB1 mutations with alpha-delta granule deficiency. This case underscores the importance of platelet esoteric testing and molecular analysis in the diagnosis of hereditary platelet disorders. As a targetable mutation in the future, this could revolutionize the early diagnosis and treatment of this rare platelet disorder. Disclosures No relevant conflicts of interest to declare.

Abstract Abstract 1362 The role of anti-tumor immunity is not well understood in CLL but does have the potential to be manipulated to improve treatment responses. We studied the predictive values of 43 immune phenotypes in patients enrolled in a clinical trial of early treatment of high risk CLL. The study enrolled 33 patients with CLL diagnosed using standard NCI-WG96 criteria who had previously untreated early - intermediate stage disease and did not meet NCI-WG96 guidelines for conventional treatment. High risk disease was defined as at least one of the following: 17p13 deletion; 11q22 deletion or unmutated IGHV together with either expression of ZAP-70 (>20%) or CD38 (>30%). Patients were treated with GM-CSF 250 mcg sc 3 × week × 5 weeks, alemtuzumab sc with dose escalation over 3 days then 30 mg/d 3 × week × 4 weeks, and rituximab 375 mg/m2 weekly × 4 weeks for a total duration of therapy of 5 weeks. We measured immune phenotypes (representing Tregs, naïve, effector memory, central memory T cells, B cells, NK cells, monocytes, among others) at the time of enrollment, through treatment and at 1 year after enrollment in 28 patients. We identified pre-treatment immune phenotypes that correlated with disease burden (as measured by the number of circulating B cells). Increased disease burden was correlated with increased absolute numbers of CD3+, CD8+, CD4+ T cells, NK (CD56+CD16+), Dr+CD8+ T cells, and Tregs (CD4+CD25+CD127+); p<0.001. The disease burden was inversely related to the number of circulating monocytes (p<0.01) and unrelated to the circulating number of naïve T cells. We repeated the pre-treatment phenotype correlation studies using the CD19+ cell counts 12 months after treatment for 13 patients who have completed the final analysis. Interestingly, the number of naïve CD4 and CD8 cells (CD4+RA+62L+CD27+ and CD8+RA+62L+CD27+) of the patient prior to treatment correlated with CD19+ cells 12 months after treatment (p=0.024 and p=0.002 respectively; n=13). Finally, we correlated pre-treatment immune phenotypes with time to progression. Again, an elevated T cell phenotype (CD4+RA+62L+CD27- as a percent of CD4+ cells) correlated to increased time to progression (p=0.04; n=28). It is noteworthy that the naïve T cell phenotype did not correlate with tumor burden prior to therapy. Thus, while pre-treatment lymphocyte expansion is commensurate with tumor burden in CLL, the frequency and numbers of naïve T cells prior to treatment has some predictive value to response to treatment. These data suggest that CLL induces the expansion of lymphocytes, suppresses monocytes with an induction in naïve T cells acting as an indicator of higher B cell counts at 12 months in patients with durable remissions. Disclosures: Zent: Genzyme: Research Funding; Genentech: Research Funding; Novartis: Research Funding; G.S.K.: Research Funding.

Abstract Background A cluster of severe lung disease occurred at a manufacturing facility making industrial machines. We aimed to describe disease features and workplace exposures. Methods Clinical, functional, radiologic, and histopathologic features were characterized. Airborne concentrations of thoracic aerosol, metalworking fluid, endotoxin, metals, and volatile organic compounds were measured. Facility airflow was assessed using tracer gas. Process fluids were examined using culture, polymerase chain reaction, and 16S ribosomal RNA sequencing. Results Five previously healthy male never‐smokers, ages 27 to 50, developed chest symptoms from 1995 to 2012 while working in the facility's production areas. Patients had an insidious onset of cough, wheeze, and exertional dyspnea; airflow obstruction (mean FEV 1 = 44% predicted) and reduced diffusing capacity (mean = 53% predicted); and radiologic centrilobular emphysema. Lung tissue demonstrated a unique pattern of bronchiolitis and alveolar ductitis with B‐cell follicles lacking germinal centers, and significant emphysema for never‐smokers. All had chronic dyspnea, three had a progressive functional decline, and one underwent lung transplantation. Patients reported no unusual nonoccupational exposures. No cases were identified among nonproduction workers or in the community. Endotoxin concentrations were elevated in two air samples; otherwise, exposures were below occupational limits. Air flowed from areas where machining occurred to other production areas. Metalworking fluid primarily grew Pseudomonas pseudoalcaligenes and lacked mycobacterial DNA, but 16S analysis revealed more complex bacterial communities. Conclusion This cluster indicates a previously unrecognized occupational lung disease of yet uncertain etiology that should be considered in manufacturing workers (particularly never‐smokers) with airflow obstruction and centrilobular emphysema. Investigation of additional cases in other settings could clarify the cause and guide prevention.