Degree Of Bone Marrow Infiltration Research Articles

Correlation between whole skeleton dual energy CT calcium-subtracted attenuation and bone marrow infiltration in multiple myeloma

ObjectivesObjective evaluation of the extent of skeletal marrow involvement in multiple myeloma remains a clinical gap for CT. We aimed to develop a quantitative segmentation pipeline for dual energy CT and to assess whether quantified whole skeleton calcium-subtracted attenuation values correlate with biopsy-derived bone marrow infiltration in multiple myeloma. MethodsConsecutive prospective patients with suspected/established myeloma underwent dual source CT from the skull vertex to proximal tibia. Whole skeleton segmentation was performed for 120 kVp-equivalent images as follows: following Hounsfield unit (HU) thresholding, a Chan-Vese morphological operation was implemented to generate a whole skeleton segmentation mask. This mask was then applied to corresponding whole skeleton material decomposition calcium-subtracted maps, generating whole skeleton HU values. Associations with biopsy-derived bone marrow plasma cell infiltration percentage were assessed with Spearman’s rank correlation; significance was at 5%. Results21 patients (12 females; median (IQR) 67 (61, 73) years) were included; 16 patients had osteolytic bone lesions; 15 patients underwent bone marrow biopsy. Segmentation and quantification were feasible in all patients. Median (IQR) of the average skeletal calcium-subtracted attenuation was −59.9 HU (-66.3, −51.8HU). There was a positive correlation with bone marrow plasma cell infiltration percentage (Spearman’s rho: + 0.79, p < 0.001). ConclusionWhole skeleton calcium-subtracted attenuation is associated with the degree of bone marrow infiltration by plasma cells, providing an objective measure of marrow involvement with the potential to allow earlier detection of disease.

Role of whole-body diffusion-weighted imaging in evaluation of multiple myeloma.

The evaluation of bone disease in multiple myeloma (MM) is an important topic in imaging. This study retrospectively investigated whole-body diffusion-weighted imaging (WB-DWI) in the evaluation of bone marrow infiltration and treatment response in MM.A total of 126 patients with MM who underwent WB-DWI between January 2016 and December 2020 were enrolled. All the patients received 4-course induction chemotherapy. WB-DWI was performed before and after chemotherapy to measure the apparent diffusion coefficient (ADC) values. According to gender and Revised International Staging System (RISS) staging groups, the relationship between ADC value and bone marrow plasma cell infiltration ratio before treatment were explored using Spearman and Pearson correlation coefficients. Comparison of ADC values before and after treatment according to different chemotherapy regimens and treatment response was performed by 2-independent samples non-parametric tests and t test.There was a negative correlation between the ADC value and the degree of bone marrow infiltration and this was statistically significant (r = –0.843, P < .001). In different gender and RISS groups, ADC value before treatment was negatively correlated with the proportion of plasma cell infiltration (male, r = –0.849; female, r = –0.836; Stage I, r = –0.659; Stage II, r = –0.870; Stage III, r = –0.745; all P < .001). The ADC values of all subjects increased to varying degrees after 4-course induction chemotherapy, including different chemotherapy regimens and treatment responses (all P < .05 except for progressive disease group).The ADC value was negatively correlated with the degree of bone marrow infiltration in different gender and RISS stages. The ADC value increased after treatment, but it was not consistent with progressive disease group. The increase of ADC value may indicate the disease burden and outcome of MM induced chemotherapy.

Correlation between the Degree of Bone Marrow Involvement and the Results of NGS in Plasma Cell Neoplasms

Practical guidance on diagnostic and prognostic use of next generation sequencing (NGS) in multiple myeloma (MM) and other plasma cell neoplasms (PCN) has yet to be developed. To this end, we analyzed the correlation of NGS data with the degree of bone marrow (BM) plasma cell (PC) involvement obtained by cytomorphology (CM), histopathology (HP), and multiparameter flow cytometry (MFC) from patients with PCN/MM. To perform this correlation, we analyzed 90 PCN cases, that included MM (n=77), MGUS (n=7), AL-amyloidosis (ALA) (n=4) and solitary plasmocytoma (SP) (n=2). The degree of BM infiltration (percent of PC) was defined by MFC as grade I &amp;lt;1%, grade II 1-3%, grade III &amp;gt;3%; by CM as grade I &amp;lt;10%; grade II, 10-30%; grade III &amp;gt;30% and by HP as grade I &amp;lt;10%, grade II 10-30%, grade III &amp;gt;30%. The newly designed NGS panel consisted of 15 genes including splice sites or hotspots: BRAF (exons 11, 15), CCND1, DIS3, EGR1, TENT5C (FAM46C), FGFR3, IDH1 (exon 4), IDH2 (exon 4), IRF4 (exon 3), KRAS (exons 2, 3), MYD88 (L265P), NRAS (exons 2, 3) PRDM1, TP53 and TRAF3. Genes and hotspots were chosen according to the frequency in the literature, prognostic impact, and possible function as therapeutic targets (Chapman et al., Nature 2011; Walker et al., JCO 2015). PC enrichment based on CD138+ magnetic cell sorting of marrow samples was successfully performed before the NGS procedure in all cases. In total, 102 mutations were detected by NGS in 64/90 (71%) cases analysed, 26 cases (29%) showed no mutations. In 41 cases (45%) one mutation/sample was detected and in 23 (26%) two to five mutations/sample. The proportion of cases affected by mutations was 59/77 patients (77%) in MM, 2/7 (29%) in MGUS and 1/4 (25%) in ALA. The most frequent mutations across all PCNs types were KRAS 22/90 (24%) and NRAS 14/90 (16%), followed by DIS3 and TENT5C in 11/90 cases each or 12% (Table 1). Mutational load correlated with degree of bone marrow infiltration: BM samples with no mutation had the lowest plasma cell infiltration: mean 1.6% by MFC (range, 0.003 - 11.7%), 20% by CM (1 - 90%), and 24% by HP (1 - 80%). Presence of at least one mutation/sample corresponded to a higher degree of BM involvement with a mean of 11% pathologic PC by MFC (range, 0.002 - 62%), and ~50% (3 - 100%) as defined by both CM and HP (Figure 1). The majority of samples without mutation (26 cases, 29%) were obtained from patients with initial BM evaluation for PCN suspicion (19/26 cases or 73% of this subgroup), whereas BM samples with &amp;gt;1 mutation (23 cases, 26%) were mostly acquired from patients with relapsed/refractory disease (15/23 cases, or 65% of this subgroup). TP53 mutated cases had the highest percentage of PC infiltration by all methods and were mostly correlated to relapsed/refractory MM (rrMM). In conclusion, the probability to detect a mutation by NGS in the BM was highest in samples with &amp;gt;10% clonal PC by flow cytometry, or &amp;gt;20% PC by CM/HP. We propose further evaluation of these thresholds as a practical cut-off for processing of samples by NGS at initial PCN/MM diagnosis, whereas disease progression is commonly associated to higher rates of mutations. TP53 mutation was detected in all but one samples from rrMM and was associated with the highest degree of BM involvement. Further large-scale studies are warranted. Disclosures No relevant conflicts of interest to declare.

Association between Clinical Presentation of Multiple Myeloma, Immunoglobulin Isotypes, and Cytogenetic Risk Groups

Introduction: In multiple myeloma (MM), chromosomal abnormalities (CAs) are valuable in risk stratifying patients and predicting disease free survival. While immunoglobulin isotypes have historically contributed to MM staging, more recently established classifications of CAs have allowed for a revised staging system that better predicts disease behavior. In this retrospective study, we hypothesized that CAs correlate with disease characteristics including immunoglobulin heavy chain isotype, degree of bone marrow infiltration (PC%), and end-organ damage. Methods: MM patients diagnosed between 2013 to 2019 were included in this retrospective chart review using electronic records from two distinct sources: (1) 442 patients from an independent pathology database and (2) a validation cohort composed of 110 patients from our institution. CAs were stratified by Mayo mSMART 2.0 criteria into standard, intermediate, and high-risk groups (Mikhael et al. Mayo Clin Proc 2013). End-organ damage was defined as the presence of lytic bone lesions, anemia, hypercalcemia, or renal failure on clinical presentation. Within each cohort, associations between categorical variables were made using the chi-squared test or Fisher's exact test, as deemed appropriate. The Mann-Whitney test was used to compare between groups for continuous variables. A result was considered statistically significant at the p&amp;lt;0.05 level of significance. All analyses were performed using SAS version 9.4 (SAS Institute Inc., Cary, NC). Results: 552 MM patients were included in the study. Multi-variate analysis revealed that del(13q14), dup(1q21), t(4;14), trisomy (11q13), del(16q23), and hypodiploidy were associated with a significantly higher PC%, whereas kappa light chain only (LCO) disease was associated with a lower PC%. Higher median PC% was found when comparing the intermediate to standard CA risk group. IgA isotype was associated with intermediate risk CAs including del(13q) and standard risk CAs including t(11;14), while IgG isotype was associated with dup(1q21), and kappa LCO disease correlated with a higher rate of higher risk CAs including deletion of p53 at 17p13 and dup(1q21). With regard to clinical presentation, lytic lesions were more frequent in patients with normal cytogenetics and trisomy 11 and less frequent in IgA isotype, whereas the presence of anemia on presentation correlated with IgA isotype. Renal failure was associated with MAF translocations including t(14;16), a high-risk CA. Conclusions: We demonstrate that a relationship exists between specific CAs, immunoglobulin isotypes, and clinical presentations in MM. Our data indicate that IgA isotype is significantly associated with intermediate-risk cytogenetics including del(13q) and anemia on presentation, and that light chain disease and renal failure correlate with high risk CAs including del(17p13). These associations between biological and clinical features further support the concept of divergent cytogenetic evolution in MM as being an underlying factor leading to distinctive disease presentations. Disclosures Braunstein: Takeda: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Epizyme: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees; Verastem: Membership on an entity's Board of Directors or advisory committees; Morphosys: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding.

Role of computed tomography texture analysis using dual-energy-based bone marrow imaging for multiple myeloma characterization: comparison with histology and established serologic parameters

ObjectiveTo identify textural features on dual-energy CT (DECT)–based bone marrow images in myeloma which correlate with serum markers of myeloma activity and the degree of medullary involvement.MethodsA total of 110 patients (63.0 ± 11.0 years, 51 female) who underwent unenhanced whole-body DECT between September 2015 and February 2019 were retrospectively included, which was approved by our institutional ethics committee with a waiver of the informed consent requirement. All patients had current hematologic laboratory tests. Using DECT post-processing, non-calcium bone marrow images were reconstructed. The vertebral bodies T10–L5 were segmented for quantification of textural features, which were compared with serologic parameters and myeloma stages by the Mann-Whitney U test. In a subgroup of 56/110 patients with current bone marrow biopsies, textural features were correlated with the degree of bone marrow infiltration.ResultsFirst-order features were higher in patients with advanced stage of myeloma (p < .02), whereas the 2nd-order “gray-level co-occurrence matrix (GLCM) cluster prominence” was lower (p < .04). In patients with elevated serum-free light chains (SFLC) or kappa/lambda SFLC ratio above 1.56, the “entropy” and 2nd-order GLCM features were lower (p < .03). The degree of bone marrow infiltration correlated with 1st-order features (e.g., “uniformity”; rP = 0.49; p < .0001), whereas “entropy” and 2nd-order GLCM features were negatively correlated (e.g., “difference entropy”; rP = − 0.54; p < .0001).ConclusionsCT textural features applied on non-calcium bone marrow images correlate well with myeloma-related serologic parameters and histology showing a more uniform tissue structure and higher attenuation with increasing medullary infiltration and could therefore be used as additional imaging biomarkers for non-invasive assessment of medullary involvement.Key Points• Texture analysis applied on dual-energy reconstructed non-calcium bone marrow images provides information about marrow structure and attenuation.• Myeloma-related serologic parameters and the degree of myeloma cell infiltration correlate with 1st- and 2nd-order features which could be useful as additional imaging biomarkers for non-invasive assessment of medullary involvement.

Extended Texture Analysis of Non-Enhanced Whole-Body MRI Image Data for Response Assessment in Multiple Myeloma Patients Undergoing Systemic Therapy.

Identifying MRI-based radiomics features capable to assess response to systemic treatment in multiple myeloma (MM) patients. Retrospective analysis of whole-body MR-image data in 67 consecutive stage III MM patients (40 men; mean age, 60.4 years). Bone marrow involvement was evaluated using a standardized MR-imaging protocol consisting of T1w-, short-tau inversion recovery- (STIR-) and diffusion-weighted-imaging (DWI) sequences. Ninety-two radiomics features were evaluated, both in focally and diffusely involved bone marrow. Volumes of interest (VOI) were used. Response to treatment was classified according to International Myeloma Working Group (IMWG) criteria in complete response (CR), very-good and/or partial response (VGPR + PR), and non-response (stable disease (SD) and progressive disease (PD)). According to the IMWG-criteria, response categories were CR (n = 35), VGPR + PR (n = 19), and non-responders (n = 13). On apparent diffusion coefficient (ADC)-maps, gray-level small size matrix small area emphasis (Gray Level Size Zone (GLSZM) small area emphasis (SAE)) significantly correlated with CR (p < 0.001), whereas GLSZM non-uniformity normalized (NUN) significantly (p < 0.008) with VGPR/PR in focal medullary lesions (FL), whereas in diffuse involvement, 1st order root mean squared significantly (p < 0.001) correlated with CR, whereas for VGPR/PR Log (gray-level run-length matrix (GLRLM) Short Run High Gray Level Emphasis) proved significant (p < 0.003). On T1w, GLRLM NUN significantly (p < 0.002) correlated with CR in FL, whereas gray-level co-occurrence matric (GLCM) informational measure of correlation (Imc1) significantly (p < 0.04) correlated with VGPR/PR. For diffuse myeloma involvement, neighboring gray-tone difference matrix (NGTDM) contrast and 1st order skewness were significantly associated with CR and VGPR/PR (p < 0.001 for both). On STIR-images, CR correlated with gray-level co-occurrence matrix (GLCM) Informational Measure of Correlation (IMC) 1 (p < 0.001) in FL and 1st order mean absolute deviation in diffusely involved bone marrow (p < 0.001). VGPR/PR correlated at best in FL with GSZLM size zone NUN (p < 0.019) and in all other involved medullary areas with GLSZM large area low gray level emphasis (p < 0.001). GLSZM large area low gray level emphasis also significantly correlated with the degree of bone marrow infiltration assessed histologically (p = 0.006). GLCM IMC 1 proved significant throughout T1w/STIR sequences, whereas GLSZM NUN in STIR and ADC. MRI-based texture features proved significant to assess clinical and hematological response (CR, VPGR, and PR) in multiple myeloma patients undergoing systemic treatment.

Rebound Thrombocytosis after Induction Chemotherapy Is a Strong Biomarker for Favorable Outcome in AML Patients

Background: Whereas the molecular events underlying acute myeloid leukemia (AML) are increasingly identified, dynamics of hematologic suppression and recovery following induction chemotherapy remain enigmatic. In addition, thrombocytopenia at diagnosis is related to deregulated cytokine expression in specific leukemia subtypes rather than on the degree of bone marrow infiltration per se. Similarly, recovery of platelets after the first induction cycle among individual patients can vary from incomplete to excess recovery even if morphologic remission is achieved. In particular, excess platelet rebound (EPR) with values exceeding 500 G/L can be observed in some patients. However, this clinical observation of EPR is poorly understood so far, and its prognostic relevance is unknown. Methods: We analyzed platelet recovery after the first induction cycle in 291 consecutive AML patients receiving at least one cycle of intensive "3+7" based induction treatment. We defined excess platelet rebound (EPR) as a platelet increase above 500 G/L during hematologic recovery after the first induction cycle. Results: We observed EPR in 120 (41.2%) patients. EPR+ patients had lower platelets at diagnosis, higher marrow infiltration, more frequently NPM1 mutations, and were associated with ELN favorable risk. Absence of EPR correlated with complex karyotypes, ELN intermediate-I and adverse risk, and therapy-related AML. Overall survival was better in EPR+ patients than EPR- (median 125 versus 41 months; P=0.04), as was disease-free survival (Figure 1). By multivariate analysis, achievement of EPR+ was an independent parameter associated with favorable survival. Plasma thrombopoietin (TPO) levels at diagnosis indicated EPR+ (p&lt;0.0001), while GATA-1, GATA-2 and MPL mRNA expression did not differ between EPR+ and EPR- patients. Finally, transcription factors blocking early megakaryopoiesis were upregulated in EPR- patients, while NFE2 involved in late megakaryocyte differentiation was increased in EPR+ patients. Conclusion: This study suggests that excess platelet rebound (EPR+) defined as platelets exceeding 500 G/L is a common event identified in AML patients after the first induction cycle. EPR+ was predominantly observed in ELN favorable risk AML, and it was associated with significantly better overall and disease-free survival as compared to EPR- patients. In addition, achievement of EPR+ represented an independent prognostic parameter for improved survival in a multivariate analysis, and may contribute to prognostic assessment in AML patients undergoing intensive treatment with intention to cure. Disclosures No relevant conflicts of interest to declare.

Utility of FDG-PET/CT in Lymphoblastic Lymphoma

Aim:To explore the utility of [18F] Fluoro-2-deoxyglucose (18-FDG) positron emission tomography computed tomography (PET/CT) in lymphoblastic lymphoma (LBL). Background: LBL is a rare, malignant disorder of precursor T- or B- cells, which forms the lymphoma variant of acute lymphoblastic leukaemia (ALL). Whilst morphologically and immunophenotypically similar to ALL, distinction is made on degree of bone marrow infiltration (typically less than 25% in LBL). Due to the rarity of the condition, investigations at diagnosis, treatment approach and methods to evaluate disease response are not standardised. Bone marrow examination with MRD determination is the gold standard modality for disease monitoring in ALL yet the utility of this investigation is limited in LBL due to the low levels of marrow involvement at presentation. Computed tomography (CT) scanning is widely used in the diagnosis and assessment of lymphomas, however the criteria for disease are based purely on size of a lesion. Difficulties in distinguishing inflammatory from malignant lesions on CT have been widely reported (Elstrom et al. Blood. 2003;101(10):3875-6). 18-FDG PET/CT has been shown to be sensitive for staging high grade non-Hodgkin's lymphoma and is better at differentiating inflammatory from malignant lesions compared to CT alone (Ngeow et al, Ann Oncol. 2009;20(9):1543-7). Normalisation of the SUV to a suitable reference region such as the tumor-to-liver ratio (TLR), has reduced issues with inter-scanner variability. Retrospective data on the use of PET/CT in LBL is limited. In most series, few patients underwent PET/CT at diagnosis making interpretation of subsequent scans difficult thus PET/CT is currently only used at physician discretion in LBL. Methods: Patients with a diagnosis of T- or B-cell LBL managed at University College London Hospital, a large tertiary haematology unit in the UK since 2011 were identified. Data on timing of PET/CT scans in patients with a diagnosis of LBL were collected retrospectively and correlated with clinical outcome data. SUVmax values within the lesion(s) and within the liver, spleen and mediastinal (aortic) blood pool were collected and the TLR calculated. Results:20 patients were identified (14 T-LBL, 6 B-LBL), median age 24.5 years (range 13-66). Ten patients aged 13-24 years were treated using the UKALL 2011 protocol and 10 patients aged &gt;24, years were treated using the UKALL 14 protocol. Sixteen of 20 (80%) of patients had a PET/CT scan at diagnosis before commencing treatment. The remaining 4 patients presented in extremis, requiring intensive care support and immediate empirical treatment. Of the 16 patients who had a PET/CT at diagnosis, all but one had significant FDG avid disease (mean SUVmax 8.6, range 2.7-17.1, TLR &gt;1). Overall survival (OS) for the whole cohort was 70% (median follow up 31 months, range 6-91 months). Consistent with previous published data, OS was superior in the younger patients (90% in the 13-24 year age group vs 50% in those aged &gt;24 years) although due to different patient characteristics and therapy these groups are not comparable. Regardless of age, all patients with a negative PET/CT scan after cycle 1 of chemotherapy survived, disease free (see table 1). Patients with a positive PET/CT after course 2 of treatment or at the end of treatment had a much poorer OS (60% and 0% respectively). In keeping with data from other high-grade lymphomas, SUVmax&gt;10 at presentation may predict an aggressive clinical course, with average SUVmax of 11.9 (TLR4.23) in the patients who died of their disease compared to 6.225 (TLR2.33) in patients who survived. PET/CT had 100% sensitivity and specificity for detecting relapsed disease in this small series. In 8 patients who presented with symptoms suggestive of disease progression or relapse PET/CT detected all 6 patients with true positive disease and excluded relapse in 2 patients with inflammatory lesions. Conclusions: Our data suggests that PET/CT is useful in the diagnosis and response assessment of LBL particularly if the clinical scenario permits a scan at diagnosis. Although patient numbers are small, these data suggest that a high lesion SUVmax at diagnosis correlates with an aggressive subtype and patients with a negative scan after their first cycle of chemotherapy appear to have an excellent prognosis. Further investigation of the utility of PET/CT in risk stratification in LBL is warranted. Disclosures Fielding: Incyte: Consultancy; Pfizer: Consultancy; Amgen: Consultancy; Novartis: Consultancy.

PB2009 PERIPHERAL BLOOD INVOLVEMENT BY FLOW CYTOMETRY IS NOT AN UNUSUAL FEATURE IN IGM MGUS/WALDENSTRÖM MACROGLOBULINEMIA WITH MYD88 L265P MUTATION.

Background:The IgM monoclonal gammopathy of undetermined significance (MGUS) is defined as a serum IgM paraprotein concentration &lt;30 g/L, bone marrow (BM) lymphoplasmacytic infiltration by clonal population &lt;10% and no symptoms or end‐organ damage. Waldenström Macroglobulinemia (WM) is a lymphoplasmacytic lymphoma (LPL) with BM involvement and an IgM monoclonal gammopathy of any concentration. Patients fullfilling criteria for WM and not requiring inmediate treatment are classified as smoldering WM. More than 90% of LPL/WM cases and 60% of IgM MGUS display the MYD88 L265P mutation (MYD88mut), suggesting that IgM MGUS is closer to mature B‐cell neoplasms than to plasma cell myeloma according to 2016 WHO update. From the immunophenotypic point of view, clonal B cells are reported in 75% of IgM MGUS in BM on multiparameter flow cytometry (FC). However, data about peripheral blood (PB) involvement in IgM MGUS/WM are scarce.Aims:To analyze the incidence of PB involvement detected by FC in a series of IgM MGUS/WM with MYD88mut in BM consecutively diagnosed in a single institution.Methods:69 patients (47 male/22 female; median age 77 years, range 38‐92) diagnosed with IgM MGUS/WM MYD88mut between 2000 and 2019 with PB FC availability were selected. The diagnosis was made in BM in all cases. For FC analysis, 100,000 events were acquired in a FACSCanto II (BD) cytometer and were analyzed with the Infinicyt software (Cytognos). The study of the MYD88 L265P mutation was performed in PB by allele‐specific qPCR and the analysis of the IGH gene rearrangement by PCR.Results:Monoclonal gammopathy was detected in routine blood tests in asymptomatic patients in 52/69 (75%) cases. Median lymphocyte count was 2.04x109/L (0.77‐8.81). Only 3/69 (4.3%) patients presented with ≥4x109/L lymphocytes. Leukemic involvement by FC was detected in 36/69 (52%) patients (see Table 1). The phenotype of clonal lymphocytes was the usually described in WM (CD19dimCD22dimCD25dim) in most cases. In those patients, the median percentage of B‐cells from the total lymphocytes was 8% (0.6‐40%) with only 5 (7%) patients presenting ≥20% B‐lymphocytes. The median percentage of clonal B‐cells in respect of the total number of B‐cells was 60% (3‐100%). However, in 9 patients the clonal B‐lymphocytes could not be quantified because of the minor representation of this abnormal population with non‐specific phenotype. In 5 (7%) patients an additional abnormal B‐cell population was detected, 4/5 being phenotypically consistent with chronic lymphocytic leukemia. PB involvement was most frequently observed in patients displaying ≥30% lymphocytes in BM (15/20, 75% vs. 21/48, 44% in patients with &lt;30% lymphocytes, P = 0.032). No differences were observed with regard to the amount of serum monoclonal component (PB involvement: 12/24 cases with ≥15 g/L vs. 24/45 patients with &lt;15 g/L, P = NS). MYD88mut was detected in 31/34 (91%) cases with leukemic expression. In 13/31 (42%) patients without PB involvement by FC, molecular data were consistent with circulating disease (only MYD88mut 2/31, only clonal IGH rearrangement 5/29, both 6/29).Summary/Conclusion:In our series, PB involvement by FC was not unusual in patients with IgM MGUS/MW MYD88mut. However, nearly all patients had a normal lymphocyte count and a percentage of B‐cells within the normal ranges. Leukemic expression by FC correlated with the degree of BM infiltration. Finally, PB involvement could be detected by molecular techniques (MYD88 L265P mutation and/or clonal IGH rearrangement) in almost half of the patients with negative FC.image

Abnormal repression of SHP-1, SHP-2 and SOCS-1 transcription sustains the activation of the JAK/STAT3 pathway and the progression of the disease in multiple myeloma.

Sustained activation of JAK/STAT3 signaling pathway is classically described in Multiple Myeloma (MM). One explanation could be the silencing of the JAK/STAT suppressor genes, through the hypermethylation of SHP-1 and SOCS-1, previously demonstrated in MM cell lines or in whole bone marrow aspirates. The link between such suppressor gene silencing and the degree of bone marrow invasion or the treatment response has not been evaluated in depth. Using real-time RT-PCR, we studied the expression profile of three JAK/STAT suppressor genes: SHP-1, SHP-2 and SOCS-1 in plasma cells freshly isolated from the bone marrows of MM patients and healthy controls. Our data demonstrated an abnormal repression of such genes in malignant plasma cells and revealed a significant correlation between such defects and the sustained activation of the JAK/STAT3 pathway during MM. The repressed expression of SHP-1 and SHP-2 correlated significantly with a high initial degree of bone marrow infiltration but was, unexpectedly, associated with a better response to the induction therapy. Collectively, our data provide new evidences that substantiate the contribution of JAK/STAT suppressor genes in the pathogenesis of MM. They also highlight the possibility that the decreased gene expression of SHP-1 and SHP-2 could be of interest as a new predictive factor of a favorable treatment response, and suggest new potential mechanisms of action of the therapeutic molecules. Whether such defect helps the progression of the disease from monoclonal gammopathy of unknown significance to MM remains, however, to be determined.

Circulating Adamts-13 Is Reduced in Patients with Waldenstrom’s Macroglobulinemia and Is Associated with Increased IgM Levels and Features of the Disease but Not with the Increased Levels of Von Willebrand Factor

ADAMTS-13 is a protease, member of the ADAMTS family (A Disintegrin and Metalloproteinase with ThromboSpondin type 1 repeats-13), which cleaves the cell bound large ultrapolymeric von Willebrand factor (vWF) strings. Circulating ADAMTS-13 is primarily synthesized and released from hepatic stellate and endothelial cells. Acquired functional deficiency of ADAMTS-13, usually due to inhibitory IgG autoantibodies, results in excessive platelet aggregation and disseminated vWF/platelet-rich thrombus formation. A possible association of low levels of ADAMTS-13 Ag with arterial thrombosis and endothelial dysfunction has also been reported. Hivert and colleagues (Blood 2012;120:3214-21) and our group have shown that increased levels of vWF, the only known substrate of ADAMTS-13, are associated with poorer prognosis in patients with symptomatic Waldenstrom’s Macroglobulinemia (WM). Thus, our aim was to investigate the possible association of ADAMTS-13 antigen (Ag) levels with features of WM and possible biologic implications of the ADAMTS-13 / vWF interaction in WM patients’ prognosis.Our study included 42 patients with symptomatic WM who were treated and followed in the Department of Clinical Therapeutics of the University of Athens (Greece), from 1999 to 2012, 22 patients with asymptomatic WM and 19 healthy controls of matched gender and age. For the purpose of this study we used stored serum, which had been collected before initiation of any therapy. ADAMTS-13 antigen levels were measured using a commercially available kit (R&D Systems, Minneapolis, MN, USA), which has a detection limit for ADAMTS-13 13 (ng/mL) with intra- and inter-Assay Precisions of <3.7% and <5.7%, respectively.The median age of patients with symptomatic WM was 65 years (range: 37-83 years) and 54% were males. Anemia (Hb <11.5 g/dL) was present in 78% of patients, low platelet counts (<100x109/L) in 17%, beta2-microglobulin >3 mg/dl in 56%, while 7.5% had serum LDH ≥250 U/L and 58% had serum albumin <3.5 g/dL. Median serum IgM was 3340 mg/L (range 246-9563 mg/dL). According to IPSSWM, 22% had low, 43% intermediate and 35% high risk WM, respectively. Main reasons to initiate therapy included cytopenias in 42% of patients, B-symptoms in 15%, hyperviscosity in 12%, neuropathy in 10% and other reasons in 21%. Primary therapy based on rituximab was given in 93% of the patients and 54% achieved at least 50% reduction of IgM.The median levels of ADAMTS-13 in patients with symptomatic WM were 848 ng/ml (range 471-1622 ng/ml), while those of patients with asymptomatic WM were 875 ng/ml (range 280.5-1466 ng/nl) and of healthy controls were 1170 ng/ml (range 770-1598 ng/ml). Thus, there was a statistically significant difference in the levels of ADAMTS-13 between patients with either symptomatic or asymptomatic WM and normal individuals (p<0.001 and p=0.002 respectively), while there was no difference between those with symptomatic and asymptomatic WM (p=0.934). We then evaluated the association of ADAMTS-13 antigen levels with features of symptomatic WM. There was a negative correlation between low levels of ADAMTS-13 with high levels of IgM (r=-0.544, p<0.001), beta2-microglobulin (r=-0.376, p=0.01) and with the infiltration of the BM by lymphoplasmatic cells (r=-0.338, p=0.03), while there was a positive correlation with serum albumin levels (r=0.373, p=0.016). Accordingly, in patients who required therapy because of hyperviscosity syndrome, serum ADAMTS-13 was lower (p=0.04). There was no association of the levels of ADAMTS-13 and vWF Ag either in patients with symptomatic or asymptomatic WM (p>0.5 for both), and thus, a direct link between increased vWF Ag levels, which are found in patients with WM, and compensatory ADAMTS-13 Ag levels cannot be supported by our data. The levels of ADAMTS-13 Ag had no prognostic association with overall or progression free survival in patients with symptomatic WM.In conclusion, this is the first evaluation of ADAMTS-13 Ag levels in the serum of patients with WM, in which we found that the levels of ADAMTS-13 Ag are lower compared to healthy controls and these reduced levels inversely correlate with IgM levels, beta2-microglobulin and the degree of bone marrow infiltration. Levels of ADAMTS-13 do not increase in compensation to increased levels of vWF Ag, a marker of endothelial activation and this fact may have biological implications yet to be investigated. DisclosuresNo relevant conflicts of interest to declare.

Clinical significance of interleukin-22 in multiple myeloma

ObjectiveInterleukin-22 (IL-22) is a cytokine participating in many aspects of inflammation. Multiple myeloma (MM) is a malignant disease of plasma cells with characteristic immune deregulation. We estimated serum levels of IL-22 in MM patients, both in activity and remission, in order to apprehend its possible participation in MM biology.MethodsWe measured serum levels of IL-22 along with beta-2 microglobulin (B2M), paraprotein, and interleukin-1beta (IL-1beta), as well as degree of bone marrow infiltration, in 51 patients with active MM and in 22 of them in remission.ResultsWe found that IL-22 was higher in active MM patients, compared to both controls and patients in remission, and also in patients in remission compared to controls. Moreover, IL-22 was increasing in parallel with the disease stage and also correlated with B2M, IL1-beta, and degree of infiltration.DiscussionWe suggest that the elevated levels of IL-22 in active MM patients, in parallel with disease activity, and in positive correlation with IL-1beta, may represent the inflammatory element of the disease. This increased occurrence of IL-22 may enhance myeloma proliferation and growth, and moreover, may participate in the mechanisms of immune deregulation.

Molecular Screening In Blastic Plasmacytoid Dendritic Cell Neoplasms Reveals Mutations In TET2 and ASXL1 In The Majority Of Patients

BackgroundBlastic plasmacytoid dendritic cell neoplasm is a rare and clinically aggressive disease with frequent involvement of skin and bone marrow. It derives from precursors of plasmacytoid dendritic cells and is classified according to WHO 2008 into the group of “Acute myeloid leukemia and related precursor neoplasms”. Cytogenetics frequently reveals complex karyotypes data on molecular mutations is scarce. AimsTo characterize blastic plasmacytoid dendritic cell neoplasm on the molecular level applying a comprehensive next-generation sequencing panel targeting 26 genes commonly mutated in myeloid neoplasms. Patients and MethodsWe studied six patients with blastic plasmacytoid dendritic cell neoplasm (5 bone marrow, 1 peripheral blood; 1 female, 5 male). The median age was 71.7 yrs, (range: 58.0-81.9). Cytomorphologic assessment revealed the findings characteristic for blastic plasmacytoid dendritic cell neoplasm in all cases. Furthermore, by multiparameter flow cytometry all cases had the typical immunophenotype with strong expression of CD56, expression of CD4 and CD123 and lack of expression of most myeloid and lymphoid markers. The degree of bone marrow infiltration as quantified by multiparameter flow cytometry ranged from 3% to 83% (median: 41%). One patient had a complex karyotype (44,XY,der(2)t(1;2)(q12;p22),del(5)(q14q35),del(7)(q11q22),-13,-15), four had other aberrations (46,XX,t(1;9)(q23;q33); 46,XY,t(3;8)(p12;q21),der(17)t(1;17)(q24;p13); 46,XY,i(7)(q10); 46,XY,del(13)(q14q31)), and one had a normal karyotype. None of the cases had received anti-neoplastic therapy.Mutational analysis was based on sensitive next-generation sequencing assays comprising in total 26 genes: ASXL1, BCOR, BRAF, CBL, DNMT3A, ETV6, EZH2, FLT3 (TKD), GATA1, GATA2, IDH1, IDH2, JAK2, KIT, KRAS, MPL, NPM1, NRAS, PHF6, RUNX1, SF3B1, SRSF2, TET2, TP53, U2AF1, and WT1. Targets of interest included either complete coding gene regions or hotspots. With the exception of RUNX1, which was sequenced on the 454 Life Sciences NGS platform (Branford, CT), all remainder genes were studied using a combination of a microdroplet-based assay (RainDance, Lexington, MA) and the MiSeq sequencing instrument (Illumina, San Diego, CA). ResultsStrikingly, all six patients had TET2 mutations, two had one and the other four cases had two different TET2 mutations. As described for other hematologic malignancies, there was no hotspot observed and all cases displayed different mutations (p.Leu615Serfs*24 and p.Ser1648Tyrfs*35, p.Asn767Metfs*46, p.Cys1193Tyr and p.Cys1811*, p.Gln1138*, p.Ser657Hisfs*43 and p.Glu798*, p.Ala1344Glyfs*3 and p.His1380Tyr). Of the ten TET2 mutations five were frame-shift mutations, three were nonsense mutations and two were missense mutations. Missense mutations were located in the same conserved region. The mutational load ranged from 12% to 50% (median, 32%). In one case a follow-up analysis after three weeks revealed the same two TET2 mutations with increases of the mutational load from 23% to 44% and from 24% to 39%. Although the sample size of the present series is rather limited, this is the first time that at least almost all cases of a distinct malignant disease entity are reported to carry TET2 mutations. Furthermore, there was also a very high incidence of ASXL1 mutations with 3/6 patients being affected. All cases carried the common mutation p.Gly646Trpfs*12, with mutational loads of 23%, 16% and 29%. Further mutations included SRSF2 (p.Pro95Ala) and ETV6 (p.Ile140Tyrfs*14) in one case and SRSF2 (p.Pro95Arg) with no further mutation in another case, PHF6 (p.Glu293Lys) in one case, and WT1(p.Arg596His) in one case. No mutations in any of the other analyzed genes were found. ConclusionsThe pattern of mutations in genes that have been described in other myeloid malignancies clearly underlines the correct classification of blastic plasmacytoid dendritic cell neoplasm into a myeloid disease category. Based on the present series with TET2 mutations in all (6/6) patients with blastic plasmacytoid dendritic cell neoplasm, TET2 mutations have to be considered to play a central role in the pathogenesis of this malignant disease. Additional mutational analyses on extended patient cohorts including ASXL1 should aim at clarifying the frequencies of these mutations and their potential impact on diagnostic and possible therapeutic interventions. Disclosures:Kern:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kuznia:MLL Munich Leukemia Laboratory: Employment. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Kohlmann:MLL Munich Leukemia Laboratory: Employment.

Assessment of bone marrow infiltration diagnosed by flow cytometry in canine large B cell lymphoma: Prognostic significance and proposal of a cut-off value

The aims of this study were to assess the prognostic significance of bone marrow (BM) infiltration in canine large B cell lymphoma (LBCL) and to establish cut-off values for designating the BM as infiltrated by lymphoid blasts. The degree of BM infiltration by large CD21 positive cells in dogs with LBCL was assessed by flow cytometry (FC) and related to time to progression (TTP) and lymphoma-specific survival (LSS). Forty-six dogs were prospectively enrolled, staged and treated with a dose-intense chemotherapeutic protocol. BM infiltration was directly correlated with peripheral blood infiltration (P=0.001), high lactate dehydrogenase activity (P=0.0024) and substage b disease (P<0.001). In the univariate analysis, there was a significant association between BM infiltration diagnosed by FC and both TTP (P=0.001) and LSS (P<0.001). Substage was the only factor associated with TTP in the multivariate analysis (P=0.002), whereas substage (P<0.001) and anaemia (P=0.008) were associated with LSS. A cut-off of 3% BM infiltration had the strongest prognostic value, since it discriminated between dogs with a poorer prognosis (median TTP 69days; median LSS 155days) and dogs with a better prognosis (median TTP 149days; median LSS 322days). BM analysis is an essential step in the staging of LBCL. The presence of BM infiltration by FC at diagnosis is a negative prognostic indicator in canine LBCL.

Cytogenetic Aberrations In Lymphoplasmacytic Lymphoma (LPL): A Study On 166 Cases

AbstractAbstract 2009Lymphoplasmacytic lymphoma (LPL) belongs to the most frequently occurring mature B-cell neoplasms besides chronic lymphocytic leukemia, splenic marginal zone lymphoma (SMZL), mantle cell lymphoma, follicular lymphoma, and hairy cell leukemia. Data on cytogenetic abnormalities in LPL is limited although these may facilitate the differentiation between LPL and other mature B-cell neoplasms with similar immunophenotypic features like SMZL. Thus, cytogenetic analysis may be a valuable tool to better characterize these cases and may in addition give prognostic information. We therefore analyzed bone marrow samples from 166 patients with LPL in parallel by cytomorphology, multiparameter flow cytometry (MFC) and chromosome banding analysis (CBA). In addition, fluorescence in situ hybridization (FISH) analysis was performed in 98/166 patients applying a standard panel of probes targeting del(6q), del(11q22.3) (ATM), trisomy 12, del(13q14) (D13S25, D13S319), and del(17p13) (TP53). The patients′ ages ranged from 40.9 to 87.2 years (median, 68.5 years), the male/female ratio was 1.18:1.00. The median percentage of LPL cells detected in bone samples by MFC amounted to 8% (range, 0.4% to 79%). In 66 patients (39.8%) an aberrant karyotype was identified by CBA. Specifically, these aberrations were classified into the following categories: complex karyotype (defined as at least three structural or numeric aberrations), 7.8%; loss of Y chromosome, 5.4%; del(5q), 1.2%; trisomies, 7.2%; other deletions, 3.6%; translocations, 1.8%; and other aberrations, 12.6%. FISH analysis identified the following chromosomal aberrations: del(6q), 3/60 (5.0%); del(11q22.3), 2/84 (2.4%); trisomy 12, 7/75 (9.3%); del(13q14), 13/98 (13.3%); and del(17p13), 7/85 (8.2%). Patients with aberrant karyotypes as identified by CBA were older than other patients (mean±SD, 70.0±9.0 vs. 65.7±10.4, p=0.007), and they had a significantly higher percentage of lymphoma cells as identified by MFC (mean±SD, 23.1±21.4% vs. 11.3±12.5%, p<0.001). Cases in which FISH analysis identified a del(13q) had a higher percentage of lymphoma cells as identified by MFC as compared to those without del(13q) (31.2±21.7% vs. 16.0±17.0%, p=0.005). The same was true for cases with del(17p) (40.9±25.3% vs. 14.2±14.6%, p=0.031). There were no significant differences in the percentage of lymphoma cells as identified by MFC for cases with the other chromosomal aberrations detected by FISH analysis. This data suggests that chromosomal aberrations can be detected in bone marrow samples infiltrated by LPL in 40% of the cases. Since in patients with chromosomal aberrations higher degrees of bone marrow infiltration by lymphoma cells were found it is suggested that CBA may miss chromosomal aberrations in cases with a lower degree (<5%) of bone marrow infiltration. For FISH analysis similar observations were made, i.e. a higher degree of bone marrow infiltration in cases with del(13q) or del(17p). This may indicate a more aggressive clinical course of LPL in the presence of these chromosomal aberrations. Furthermore, taking into consideration the level of sensitivity level of FISH analysis (1% to 3%) this data suggests the absence of chromosomal aberrations in cases with a lower degree of bone marrow infiltration rather than the lack of their detection by CBA and FISH analysis. With regard to the potential clinical implications of chromosomal aberrations, i.e. the estimation of the clinical course of the disease and of the patient′s prognosis, CBA and FISH analysis should be considered for routine diagnostic work-up in patients with LPL and bone marrow infiltration as detected by MFC. Disclosures:Kern: MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Employment, Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Employment, Equity Ownership.

Interest of Diffusion Weighted Magnetic Resonance Imaging for the Evaluation of Bone Marrow Infiltration in Multiple Myeloma, Smoldering Myeloma and Monoclonal Gammopathy of Undetermined Significance.

Abstract 4874 IntroductionMonoclonal gammopathy of undetermined significance (MGUS) or smoldering myeloma (SMM) are two asymptomatic early stages of multiple myeloma, and are characterized by the absence of bone involvement. In contrast, bone lytic lesions are common complications of multiple myeloma. Magnetic resonance imaging (MRI) has improved the evaluation of bone involvement in multiple myeloma. Moreover MRI allows visualization of the medullary cavity and a direct assessment of the degree of myeloma cell infiltration before bone destruction becomes visible on plain radiographs. Diffusion weighted magnetic resonance imaging (DWI) probes the diffusion of water in the body. This sequence yields qualitative and quantitative information that reflects the tissue cellularity and cell membrane intregrity and thus complements morphological information obtained by conventional MRI. It shows considerable promise for detecting and characterizing tumors and evaluating treatment response. ObjectiveOur objective was to study the potential interest of diffusion weighted MRI for the evaluation of bone marrow infiltration in multiple myeloma, SMM and MGUS. Diagnostic criteria used were those defined by the International Myeloma Working Group. Patients and methodsThis retrospective study was conducted over a period of 8 months (November 2008 - June 2009) in Internal Medicine and Hematology departments of our institution. All patients who had a whole body MRI for evaluation of multiple myeloma, SMM or MGUS were included. We choose to study simultaneously 3 complementary sequences: T1 weighted, short time inversion recovery (STIR) and diffusion weighted. ResultsSeventy patients were included in this study: 9 MGUS, 19 SMM and 42 symptomatic multiple myeloma (16 at diagnosis and 27 at relapse). They were 30 men and 40 women. The median age was 64 years (36 - 89 years). Thirty one of the 42 multiple myeloma patients have focal bone lesions. Focal lesions were observed in the 3 MRI sequences and diffusion did not detect new lesions. Regarding bone marrow infiltration assessment by MRI, four groups of patients could be identified: - Group 1 (20 patients – 28.6%): normal signal in T1, STIR, and diffusion. - Group 2 (19 patients – 27.1%): abnormal signal involving the whole spine on the STIR or diffusion weighted sequences and with a normal signal of the spine on T1.- Group 3 (25 patients – 35.7%): abnormal signal involving the whole spine on the STIR and diffusion weighted sequences and with a normal signal of the spine on T1.- Group 4 (6 patients – 8.6%): abnormal signal involving the whole spine on the 3 sequences. For the 28 patients with either MGUS or SMM, the distribution in the 4 groups was respectively 13 patients (46.4%), 7 patients (25.0%), 7 patients (25.0%) and 1 patient (3.6%) for group 1,2 3 and 4. Regarding the 42 cases of symptomatic multiple myeloma, 7 patients (16.7%) were classified in group 1, 12 (28.6%) in group 2, 18 (42.9%) in group 3 and 5 (11.9%) in group 4. There was no correlation between number of focal lesions and degree of diffuse infiltration. The degree of bone marrow infiltration evaluated by MRI was not correlated with Durie and Salmon stage, ISS, monoclonal component concentration or bone marrow plasmocytosis. DiscussionDiffusion weighted magnetic resonance imaging combined with conventional sequences (T1 and STIR) can distinguish 4 groups of patients with different aspects of bone marrow infiltration. Fifteen patients (53.6%) with either MGUS or SMM had various degree of bone marrow infiltration without any focal lesions. A prospective study on a large population of MGUS and SMM is needed to assess the impact of these asymptomatic infiltrations on the risk of malignant transformation. The degree of bone marrow infiltration evaluated by MRI is very heterogeneous between patients with multiple myeloma and not correlated with classical prognostic factors. Further prospective studies are necessary to investigate the possible prognostic impact of these different groups on the evolution of myeloma. DisclosuresNo relevant conflicts of interest to declare.

Bone marrow infiltration in infants with disseminated neuroblastoma

10056 Background: The INSS definition of stage 4S neuroblastoma allows a bone marrow infiltration (BMI) no greater than 10%. More extensive involvement is to be consider stage 4. However, the degree of BMI may be difficult to establish, since tumor cells in the marrow commonly aggregate in clumps. So far, no studies have specifically investigated this issue. In this study we have reviewed the bone marrow slides of infants with disseminated neuroblastoma enrolled between January 1998 and July 2008 in the Italian Neuroblastoma Registry. Methods: BMI was arbitrarily considered &lt;10% when it consisted of a maximum of 5 tumor cell aggregates/slide in association to normal cellularity. Marrow was aspirated from iliac crests and slides were stained with Wright-Giemsa method. A minimum of 2 slides for aspirate were examined at low magnification. The same 2 hematologists (CR, MP) examined the slides independently along the study period, blind for clinical data. A third hematologist (FS) joined to perform the reviewing process. Results: Of 142 infants enrolled, we retrieved and reviewed bone marrow slides of 73. Before results of bone marrow study, 31 patients were defined as stage 4, and 42 as stage 4S. Following bone marrow study, 2 patients previously defined as stage 4S who had ≥10% BMI were moved to stage 4. The results of the revision are summarized ( Table ). Conclusions: In this study, only 2/42 infants with stage 4S clinical features had BMI ≥ 10% and were eventually labeled as stage 4. Whether chemotherapy for stage 4 is of advantage for these patients remains to be seen. [Table: see text] No significant financial relationships to disclose.

Clinical Course and Prognosis of Smoldering (Asymptomatic) Waldenstrom's Macroglobulinemia

Smoldering Waldenström's Macroglobulinemia (SWM) is defined as a serum IgM monoclonal protein ≥3 g/dL and/or ≥10% bone marrow lymphoplasmacytic infiltration but no evidence of constitutional symptoms, symptomatic anemia, or hyperviscosity. Forty-eight patients with SWM were identified at Mayo Clinic from 1974 to 1995. Patients with a diagnosis of chronic lymphocytic leukemia or lymphoma or history of any antineoplastic therapy before diagnosis of SWM were excluded. The median age at diagnosis was 63 years (range 39–87 years). Only one patient (2%) was < 40 years of age, but 14 percent were < 50 years old. Thirty-two (67%) were men, and 16 (33%) were women. At diagnosis, hepatomegaly was noted in 10%, splenomegaly in 4%, and lymphadenopathy in 8%. The initial hemoglobin level ranged from 8.7 to 15.3 g/dL (median 11.8). The anemia in all 4 patients with an initial hemoglobin < 10 g/dL was due to other causes such as myelodysplastic syndrome, bronchopleural fistula, Barrett's esophagus, and Coumadin administration. The initial leukocyte level ranged from 3.2 to 13.2 × 109/L (median 5.7), and platelets ranged from 101 to 578 × 109/L (median 285.5). Only one patient had a serum creatinine > 2.0 mg/dL and it was unrelated to SWM. The serum monoclonal protein level at the time of diagnosis ranged from 1.5 to 5.2 g/dL (median 3.3); 12 (25%) were < 3 g/dL, while 10 (21%) were ≥4 g/dL. IgM kappa accounted for 75%, IgM lambda 21%, and 2 (4%) were biclonal. Immunofixation of the urine was positive in 97% (kappa 80%, lambda 17%, indeterminate 3%) of those tested. The size of the urine M protein ranged from unmeasurable to 1.4 g/24h (median 0.04 g/24h). Serum albumin ranged from 2.5 to 4.3 g/dL (median 3.6). Five patients (10%) had an albumin value < 3 g/dL. Ten (26%) had a reduction of 1 uninvolved immunoglobulin, while 8 (21%) had a reduction of both IgG and IgA immunoglobulins. Lymphoplasmacytic infiltration of the bone marrow ranged from 3% to 80%, (median 30%). Three (6%) had less than 10% infiltration, while 13 (27%) had 50% or greater infiltration. The 48 patients were followed for a total of 292 (range 0.5 to 22.6 years; median 3.7) person-years, during which time 33 progressed to symptomatic WM and one to AL amyloidosis. The expected number of WM based on the SEER data was 0.004 cases. Thus, relative risk of progression to WM was increased 7,586-fold (95% CI, 5,083 to 10,373). The median time to progression was 4.6 years. The absolute risk of progression to WM was 6% at 1 year, 39% at 3 years, and 55% at 5 years. The median survival after progression to symptomatic WM was 5.1 years. Seventy-three percent of patients died, indicating a robust follow-up. We evaluated sex, hemoglobin level, size of serum M protein, reduction of uninvolved immunoglobulins, presence of urinary monoclonal light chains, serum albumin level, and the proportion of lymphoplasmacytic infiltration of the bone marrow as risk factors for progression. Significant risk factors for progression with univariate analysis included size of the serum M protein, hemoglobin level, reduction in uninvolved immunoglobulins, and degree of bone marrow lymphoplasmacytic cell infiltration. Multivariate modeling revealed that the size of the M protein and the degree of bone marrow infiltration were the most important risk factors. We conclude that smoldering WM is a distinct clinical entity that needs to be differentiated from IgM MGUS and symptomatic Waldenstrom's Macroglobulinemia.

The CC Genotype of the C825T Polymorphism of the G Protein β3 Gene (GNB3) is Associated with a High Relapse Rate in Patients with Chronic Lymphocytic Leukaemia

A C825T polymorphism has been described in the GNB3 gene which encodes the Gbeta3 subunit of heterotrimeric G proteins.The GNB3 825T allele is predictive of enhanced Gi protein activation. This study was performed to correlate genotypes of the C825T polymorphism with various clinical aspects of chronic lymphocytic leukaemia (B-CLL). The GNB3 genotype distribution in B-CLL patients was similar to that in other Caucasian populations, arguing against a role of the polymorphism in the susceptibility to develop B-CLL. No statistically significant differences were observed at diagnosis between patients with the CC genotype and homozygous or heterozygous T allele carriers with respect to age at disease onset, sex distribution, proportion of patients with CD38+ leukaemia or patients in Binet stage A, blood cell counts, degree of bone marrow infiltration or serum levels of lactate dehydrogenase, thymidine kinase or β2-microglobulin. In a subgroup of 44 patients requiring chemotherapy, the median interval between diagnosis and first treatment and the response to treatment were similar in patients with CC or CT/TT genotypes. A statistically significant difference, however, was found in the proportion of patients relapsing and requiring second line chemotherapy (CC: 95%; CT/TT: 52%; p = 0.0043). The GNB3 genotype (p = 0.024) and age (p = 0.042) were identified as independent prognostic factors for a second therapy. Thus, the long-term success of the treatment appears to be correlated with the GNB3 genotype.

Pitfalls in the diagnosis of childhood leukaemia.

Pitfalls in the diagnosis of childhood leukaemia.