ALBERT

Description: Abstract 2638 Although patients with early-stage Hodgkin's lymphoma (HL) overall have a high rate of cure, they cannot be considered as a homogeneous group. In fact, a portion of these patients are resistant to or may relapse after standard treatment. Current prognostic criteria based on clinical and laboratory parameters at diagnosis do not allow to accurately identify the subset of patients with less favorable clinical outcome. In a study aimed at defining new biomarkers for risk stratification, an increased number of tumor-associated macrophages was found to be strongly associated with shortened survival in patients with classic Hodgkin's lymphoma [N Engl J Med. 2010 Mar 11;362(10):875-85]. The aim of this study was to evaluate the clinical significance of the proportion of CD68-positive infiltrating macrophages in patients with early-stage Hodgkin's lymphoma. By using an immunohistochemistry method, we analyzed diagnostic biopsies of 63 patients followed at our institution between 2006 and 2010, and uniformly treated with ABVD (adriamycin, bleomycin, vinblastine, dacarbazine) chemotherapy. Thirty-nine (62%) patients were males and 24 (38%) were females; median age at diagnosis was 30 years (range 17–85). Histological subtype was nodular sclerosis HL in 55 cases, mixed cellularity HL in 3 cases, lymphocyte-rich HL in 3 cases, and not classified in 2 cases. Five patients had subdiaphragmatic disease while 58 had supradiaphragmatic localizations. Forty-four patients with supradiaphragmatic disease were classified in the EORTC unfavorable subset: in detail, 25 patients had B symptoms and ESR ≥30, or ESR ≥50 despite the absence of B symptoms, 29 patients had bulky disease, 7 patients were older than 50 years, and 6 patients had more than 3 nodal areas involved. Thirty-six out of 63 (59%) patients received radiotherapy as a consolidation treatment after chemotherapy. After completion of the therapeutic program, 54 out of 63 (86%) patients obtained complete remission, while 9 (14%) had refractory disease; 15 out of 54 (28%) patients in complete remission relapsed during follow up. Diagnostic biopsies were reviewed by expert hematopathologists and classified into 3 groups according to the intensity of CD68 expression [N Engl J Med. 2010 Mar 11;362(10):875-85]. CD68-positive infiltrate was lower than 5% in 14 patients (group A), between 5 and 25% in 43 patients (group B), and greater than 25% in 6 patients (group C). Overall survival (OS) and progression-free survival (PFS) were estimated using the Kaplan-Meier method with observation time calculated from diagnosis. Comparison between survival curves was performed by means of the Gehan-Wilcoxon test. The 2-year OS and PFS in the entire cohort were 98% and 79%, respectively. There was a difference in the 2-year PFS between patients with favorable and those with unfavorable prognosis according to the EORTC risk criteria (100% vs 72%, P

Description: Abstract 2667 Waldenström Macroglobulinemia (WM) is a B-cell lymphoproliferative disorder characterized by bone marrow infiltration by lymphoplasmacytic lymphoma associated with a monoclonal component of IgM type in the serum. WM is often preceded by an IgM monoclonal gammopathy of undetermined significance (IgM-MGUS). The cumulative probability of progression of IgM-MGUS to WM or to other lymphoproliferative disorders is approximately 1.5% per year. Other mature B-cell neoplasms such as splenic marginal zone lymphoma (SMZL) and B-cell chronic lymphoproliferative disorders (B-CLPD) can carry an IgM monoclonal component and should therefore be considered in differential diagnosis with WM. In a study based on parallel sequencing of the whole genome of lymphoplasmacytic cells and paired normal tissue from WM patients, Treon et al (Blood. 2011;118:Abstract 300) have identified a highly recurrent somatic mutation with oncogenic activity in the myeloid differentiation primary response (MYD88) gene, leading to a change from leucine to proline at position 265 of the aminoacid sequence [MYD88 (L265P)]. Targeted Sanger resequencing showed MYD88 (L265P) in 90% of WM patients, but only in a minority of patients with IgM-MGUS or other mature B-cell neoplasms such as SMZL. We developed an allele-specific PCR for the MYD88 (L265P) mutation, and studied 58 patients with WM, 77 with IgM-MGUS, 84 with splenic marginal zone lymphoma (SMZL) and 52 with B-cell chronic lymphoproliferative disorders (B-CLPD). DNA was obtained from bone marrow cells (n=204) and peripheral blood (n=67). The aims of this study were: i) to assess the prevalence of the mutation in WM, IgM-MGUS, SMZL, and B-CLPD; ii) to analyze the relationship between MYD88 (L265P) mutation and clinical phenotype; iii) to evaluate the impact of the mutation on the risk of progression from IgM-MGUS WM or other lymphoproliferative disorders. The MYD88 (L265P) mutation was detected in 58/58 (100%) patients with WM, either asymptomatic (n=39) or symptomatic (n=18), and in 36/77 (47%) patients with IgM-MGUS. In addition, it was detected in 5/84 (6%) patients with SMZL and in 3/52 (6%) with B-CLPD; of these MYD88 (L265P)-positive subjects, 4 SMZL and 2 B-CLPD patients carried a serum IgM monoclonal component, while the remaining B-CLPD patient carried a double (IgM and IgG) monoclonal component. Compared with IgM-MGUS patients with wild-type MYD88, those carrying MYD88 (L265P) had significantly higher levels of IgM (P

Description: Abstract 3037 Fotemustine (Muphoran), a nitrosourea alkylating agent approved for use in the treatment of metastatic melanoma, has proven to be effective as single agent in relapsed/refractory multiple mieloma (MM). We report preliminary data of a phase II single centre study exploring the feasibility and the efficacy of the combination bortezomib (B) + fotemustine (Mu) + dexamethasone (D) (B-MuD) in relapsed/refractory MM patients. This study has been approved by local ethical committee; all patients (pts) signed written informed consent before the enrolment. MM pts relapsed or refractory after at least one therapy were eligible for the study. Pts who received prior bortezomib-containing regimen were included only if not considered bortezomib-refractory. Fotemustine at the escalating doses of 80 and 100 mg/m2 i.v. on day 1 was associated to Bortezomib 1,3 mg/m2 i.v. on days 1,4,8,11 + Dexamethasone 20 mg orally on days 1–2, 4–5, 8–9, 11–12 of 21-day cycle for a total of 6 cycles. Protocol was amended after the enrolment of the first five pts due to a considerable toxicity. We observed 3 grade 3–4 peripheral neuropathy, 1 grade 3 pneumonia, 4 grade 4 thrombocytopenia and two pts dropped-out (one for grade 3 pneumonia at 2° cycle, and one for grade 4 peripheral neuropathy at 3° cycles). Thus the schedule was modified as following: Fotemustine at escalating doses of 80 and 100 mg/m2 i.v. on day 1, Bortezomib 1,3 mg/m2 i.v. once weekly on days 1, 8, 15, 22, Dexamethasone 20 mg i.v. on days 1, 8, 15, 22 for six 35-day cycles. An interim analysis of feasibility and efficacy was planned after the inclusion of the first two cohort of 6 pts each, treated with escalating dose of Fotemustine according to the amended schedule. Up to now, 18 pts have been enrolled (5 pts before and 13 after the amendment): M/F 10/8, median age 69 years (44-82), median number of previous therapies 2 (1-5). Previous treatments included autologous transplant in 10 pts (59%), bortezomib in 8 pts (44%), oral melphalan in 7 pts (41%) and thalidomide in 12 (71%). After the inclusion of 12 pts the MTD for Fotemustine was established to be 100 mg/m2. No drop-outs were registered after the amendment. Preliminary data on response are available in 10 pts. Nine pts (90%) obtained at least a PR, 8 pts (80%) registered ≥VGPR (CR 10%). At time of this analysis 79 cycles were delivered: 14 before, 65 after the amendment. Eighty-nine AE of any grade were observed, 43 hematological and 46 non-hematological. Thrombocytopenia was the most common AE either before and after the amendment. Need for dose reduction was significantly lower after the amendment. In detail fotemustine was reduced in 14% of cycles before and never after the amendment (p=0.0001), bortezomib dose reduction were performed in 36% of cycles before and 15% after the amendment (p=0.08), dexamethasone dose reduction occurred in 64% of cycles before and 13% after the amendment (p=0.0001). In conclusion, this interim analysis shows that fotemustine in combination with bortezomib and dexamethasone is safe and gives encouraging results in relapsed/refractory myeloma patients with 80% of ≥VGPR. Updated results will be presented at the meeting. Disclosures: No relevant conflicts of interest to declare.

Description: Introduction: Accordingto National Comprehensive Cancer Network (NCCN) and ESMO guidelines on Waldenstrom’s Macroglobulinemia (WM) bendamustine may be considered as a therapeutic option in first line treatment or in relapsed refractory disease. Even though there are only two clinical trials including a limited number of patients addressing the role of bendamustine and rituximab (BR) treatment in WM. Patients and Methods: To define the efficacy and tolerability of BR combination as salvage regimen in WM patients, we retrospectively analyzed the outcome of symptomatic refractory relapsed patients treated with BR in 14 Italian centres. All patients receiving at least one day of treatment were included in the study. Treatment consisted of: R 375 mg/sqm iv day 1 and B iv days 1, 2. Therapy was administered every 4 weeks up to 6 courses. Results: Seventy-one patients are included in the study. As regards B dosage; 45 patients (63%) received the highest dose of 90 mg/sqm while 22 (31%) were treated with 70 mg/sqm. The 4 patients (6%) with a cumulative illness rating scale ≥ 6, received the lowest dose of 50 mg/sqm. At treatment, median age was 72 years (49-88), sex ratio M/F 46/25. Mediannumber of prior regimens was 2 (range 1-6). Twenty-four patients (34%) presented with refractory disease. The majority (90%) of patients had been previously treated with alkylating agents, 30% had also received purine analogues based treatments. Previous R was administered in the 77% of cases. The main reason (62%) for starting treatment was anemia followed by adenopathy and/or splenomegaly (35%). Median IgM level at treatment was 3815 mg/dL.Overall 361 courses of BR treatment were administered, median number 6 (range 1-6) with 47 (66%) of patients completing the 6 planned courses. Toxicity was discontinuation cause in 10 patients (14%): 4 infection, 1 fatal, 6 myelosuppression. In the remaining 14 treatment was discontinued for clinical clinical decision after disease reassessment. No difference in terms of treatment discontinuation was observed according to B dosage and age. Overall response rate (ORR) was 80.3% including: 7% complete remissions (CR), 15.5 % very good partial remissions (VGPR), 52.2% partial remissions (PR) and 5.6% of minor responses. A stable disease was observed in 16.9% of patients. One (1.4%) disease progression and one death were recorded. A progressive decrease of IgM level was observed during follow-up leading to an amelioration of response in 4 cases leading to a final ORR of 84.5%. None of the clinical and biological characteristics considered (age, sex, disease status, previous lines of treatment, previous fludarabine, bulky disease, Hb and IgM level, beta 2 microglobulin, B dosage) had an impact on ORR achievement. A better quality of response (CR plus VGPR) was observed in patients with an IgM level 〈 3000 mg/dL and in those treated with the higher dosage of B (90 mg/sqm). After a median follow-up of 19 months (3-54) 11 of the 57 responding patients met the criteria for disease progression. No difference was observed when patients were stratified according to the quality of response. B dosage did not impact disease progression. Considering that most of the patients received prophylactic growth factors, grade 3-4 neutropenia developed in only 13% of courses, 36% of patients. Dose modification or delayed treatment administration was necessary in 4 and 10% of courses respectively. During treatment we recorded 14 episodes of FUO and 5 major infections, leading death in one case. After a median follow up of 19 months none of the patients developed secondary myelodisplastic syndrome, acute leukemia or diffuse large B-cell lymphoma. In 3 cases a solid cancer was observed. Conclusion: BR combination showed to be as effective as more intensive salvage regimens in pretreated WM patients. Treatment showed to be well tolerated even in elderly patients with limited episodes of myelosuppression and infections when compared to purine analogues including regimens. Disclosures No relevant conflicts of interest to declare.

Description: Abstract 1585 Splenic marginal zone lymphoma (SMZL) is frequently associated with HCV infection and autoimmune disorders. Previous studies demonstrated a biased usage of immunoglobulin heavy variable genes (IGHV) and, in some cases, stereotyped B-cell receptors (BCRs). This characterization, however, is mainly based on the heavy chain alone, even if strong evidences are emerging on the role of light chain (Bikos et al. Leukemia 2012). The aim of this study was to analyze IG light variable genes (IGLV) of SMZL BCRs, VL-VH pairing and structural information and to investigate the sequence-structure-antigen (AG) relationship. To this end, we analyzed the VL-VH paired sequences of BCR from 52 SMZL pts (38 BM and 14 PB) diagnosed according to Matutes criteria (Leukemia, 2008). Sequences were analyzed using the IMGT/DBs and the IMGT/V-QUEST tool. The PIGS web server was used to build 3-D models of all antibodies (Abs). The Ab structures were compared using LGA and clustered together according to a score accounting for structure and sequence similarity. Using the DIGIT DB and tools, all the clusters were analyzed and compared to other IGs. Based on the IGHV nucleotide sequence identity to the germline, 7 sequences (13%) were considered ‘truly unmutated’ (100% sequence identity), 20 (39%) were ‘minimally or borderline mutated’ (97–99.9%) whereas 25 (48%) were ‘significantly mutated’ (3%), whereas samples using the IGHV1-02 gene (n=10) but a VL gene other than from IGLV1-47 displayed a low number of mutations, suggesting a significant role for the light chain. In order to analyze the possible functional role of light chain, we analyzed the structural similarity of AG binding sites (ABSs), performing hierarchical clustering on the similarity obtained by an all-against-all structural superposition of each ABS. Twenty structural clusters were identified (8 with ≥3 samples) (Fig. 1). Considering IGs in the same major groups, they showed a similar mutation rate, pointing out a likely common AG selection at least in a fraction of pts (Fig. 1). In most cases, IGs in the same clusters display ABSs with similar physicochemical characteristics: positively charged binding sites (2 clusters), hydrophobic patches (3 clusters) or small pockets in the middle of the ABS (3 clusters) might be clue for different AGs specific for each cluster. HCV infection was found in 1 major and 2 minor clusters (Fig. 1), mainly associated with unmutated clones, indicating a likely common antigenic stimulation. In the other major clusters, the role for an AG-driven selection different from HCV in SMZL lymphomagenesis can be postulated. In particular, 3 clusters, containing both mutated and unmutated samples, displayed a statistically significant similarity to CLL clones (p

Description: Abstract 262 Hairy cell leukemia (HCL) is an indolent neoplasm of small mature B-lymphoid cells, which are found in peripheral blood and bone marrow (BM), and are characterized by hairy projections of their abundant cytoplasm. In clinical practice, HCL needs to be differentiated from similar indolent lymphoid neoplasms. In a study based on massively parallel sequencing of the whole exome of leukemic and matched normal cells from a HCL patient and subsequent targeted resequencing in additional patients, Tiacci et al (N Engl J Med. 2011 Jun 16;364:2305–15) have recently identified the BRAF V600E mutation as a genetic alteration associated with this disease. This somatic mutation was previously detected in diverse human cancers, with a particularly high frequency in melanoma (Nature. 2002 Jun 27;417:949–54; N Engl J Med. 2005 Nov 17;353:2135–47). In order to develop a reliable molecular diagnostic tool and verify its sensitivity and specificity in the diagnosis of HCL, we developed an allele-specific PCR for the BRAF V600E mutation, and searched for this molecular lesion in a series of 239 patients with mature B-cell lymphoid neoplasms. The study population included 62 patients with HCL, 91 with splenic marginal zone lymphoma (SMZL), 29 with Waldenström macroglobulinemia (WM), and 57 with B-cell chronic lymphoproliferative disorders (B-CLPD). Genomic DNA was extracted from bone marrow (BM) biopsies in 61 cases of HCL, from BM in 90 patients with diverse lymphoid neoplasms (33 SMZL, 29 WM, 28 B-CLPD), and from peripheral blood (PB) in the remaining 88 patients (1 HCL, 58 SMZL, 29 B-CLPD). The BRAF V600E mutation was detected in all patients with HCL (62/62) and in none of those with SMZL or WM. Two of the 57 patients with B-CLPD carried the mutation, and their clinical features are as follows. Case #1. This 41 year-old woman presented in November 2008 with asymptomatic lymphocytosis, without any evidence of lymphadenopathy, splenomegaly or hepatomegaly. Laboratory data showed: Hb 12.9 g/dL, WBC count 16 × 109/L (62% lymphoid cells), and PLT count 283 × 109/L. On BM biopsy, an interstitial lymphoid infiltrate (60% of the whole cellularity) composed by small, lymphocyte/centrocyte-like cells was found. By immunohistochemistry, neoplastic cells showed expression of CD20, CD79a and cyclin-D1, but were uniformly negative for CD5, CD10, CD23, CD25 and DBA44, and annexin A1. At flow cytometry analysis, they were CD20 and FMC7 positive and CD10, CD38, CD5, CD23, CD11c, CD25, DBA44 and CD103 negative. FISH for t(11;14), performed for cyclin D1 expression, was negative. Immunoglobulin rearrangement was IGHV3-48*02, IGHD7-27*01 IGHJ4*02. So far, lymphocytosis has remained stable and the patient is regularly followed without any need for treatment. Case #2. This 62 year-old male presented in 2006 with thrombocytopenia and splenomegaly, and was diagnosed with HCL was established in another hospital (no additional data are available). He was treated with cladribine with a partial response. In May 2008, we evaluated this patient in Pavia. The spleen was palpable 3 cm under the costal margin, and laboratory data showed: Hb 15.4 g/dL, WBC count 3.9 × 109/L, and PLT vount 95 × 109/L. BM biopsy showed a 20% lymphoid infiltrate with interstitial and sinusoidal pattern, composed by small to medium sized cells with evident nucleoli, resembling pro-lymphocytes. By immunohistochemistry, cells were positive for CD20 and negative for CD5, CD23, cyclin-D1, CD25, DBA44, and annexin A1. Flow cytometry demonstrated the expression of CD20, FMC7 and CD11c, partial expression (25%) of CD103, and negativity for CD5, CD10, CD38, CD23, DBA44, CD11c and CD25. This patient was asymptomatic and a watch-and wait-policy was adopted. These findings indicate that the allele-specific PCR we developed is able to detect the mutation in the bone marrow of all patients with HCL, and confirm that the BRAF V600E mutation is highly specific for HCL within mature B-cell neoplasms. Only 2/177 (1.1%) patients with lymphoid neoplasms other than HCL (2/57 or 3.5% of patients with B-CLPD) were positive for BRAF V600E. This is in agreement with a previous study that found BRAF mutations in 2.4% of patients with non-Hodgkin's lymphoma (Br J Cancer. 2003 Nov 17;89:1958–60). The detection of the BRAF V600E mutation in the clone (or, at least, in a subclone) of mature B-cell lymphoid neoplasms without typical HCL features might help to define their biology. Disclosures: No relevant conflicts of interest to declare.

Description: Key Points MYD88 L265P is expressed in WM and IgM MGUS patients using AS-PCR assays with potential use in diagnostic discrimination and response assessment.

Description: BACKGROUND AND AIMS: Hairy cell leukemia (HCL) is very sensitive to purine analogs (PAs), but ~40% of patients relapse and become progressively less responsive to these myelotoxic and immune-suppressive drugs. Having discovered the BRAF-V600E kinase-activating mutation as the genetic lesion underlying HCL (Tiacci et al, NEJM 2011;364:2305), we performed the first clinical trial of a BRAF inhibitor (vemurafenib) in refractory/relapsed HCL. In particular, this is a phase-2, academic, single-arm, Italian, multi-center (n=8) study (HCL-PG01; EudraCT 2011-005487-13). METHODS: In 11 months we enrolled 28 BRAF-V600E+ HCL patients, needing therapy due to cytopenias and including: i) 6 patients primary refractory to a PA; ii) 21 patients who relapsed early and/or repeatedly after PAs and had received a median of 4 previous therapies; and iii) a 81-year old patient showing severe myelotoxicicity after a PA (discouraging its further use). Previous treatments other than PAs included interferon, rituximab and splenectomy in 12, 14 and 8 patients, respectively. Complete remission (CR) required resolution of cytopenias (N≥1500/mmc, PLT≥100000/mmc, Hb≥11 g/dl), no morphological evidence of HCL cells in the bone marrow biopsy and blood smear, and no splenomegaly. Partial remission (PR) required resolution of cytopenias, and a ≥50% reduction of splenomegaly and of marrow and blood HCL involvement by immunophenotyping. Two patients were not evaluable as they went off-study after ≤1 week of treatment (due to drug-unrelated acute myocardial infarction and consent withdrawal after grade-3 drug-related reversible pancreatitis). RESULTS: Vemurafenib, given orally at the dose of 960 mg twice daily on an outpatient basis for a median of 16 weeks, was generally well tolerated. Drug-related adverse events (mainly arthralgias, skin toxicities, pancreatitis; no myelosuppression) were frequent, but reversible in all patients, and were typically grade 1-2. Only 7 patients developed grade 3 events, and none grade 4 events. Although we did not observe any cutaneous squamous cell carcinomas/keratoachantomas (as reported in BRAF-V600E+ melanoma patients treated with vemurafenib), 3 patients developed 2 basaliomas and 1 superficial melanoma, all treated with a simple excision. Notably, overall response rate was 96% (25/26 patients): 9/26 (34.6%) CRs and 16/26 (61.4%) PRs, obtained after a median of 8 and 9 weeks respectively. CR and PR patients included 1 and 5 primary refractory ones, respectively, as well as 4 and 10 not responding to the last prior treatment, respectively. In all CR patients immunohistochemistry showed minimal residual disease (≤10%) at the end of treatment. Six of 9 (67%) CR patients enjoyed normal blood counts at a median of 13 (range 12-15) months from the end of treatment (see Figure): 3 of these 6 patients showed no morphological evidence of HCL in the bone marrow biopsy (complying with a continuous CR) at 12, 13 and 15 months, respectively, whereas the other 3 lost the bone marrow CR status, all at 12 months. The remaining 3/9 CR patients (33%) developed a mild cytopenia (N ~1000/mmc or PLT ~80000/mmc) 5, 9 and 12 months post-treatment, respectively: in the 2nd patient the cytopenia remained stable until the last follow-up at 15 months, whereas in the other two cases it worsened requiring therapy 9 and 18 months post-treatment, respectively (see Figure). These two latter patients were recently retreated with vemurafenib for 12 and 4 weeks, and obtained a PR and a second CR. Among the 16 PR patients, 5 (31%) mantain normal blood counts at a median of 12 (range 8-17) months post-treatment (see Figure). The other 11 PR patients developed cytopenia(s) after 3 months of median follow-up (range 5-10): in 6 patients (38%) no anti-leukemic therapy was started at a median of 9 (range 6-12) months post-treatment, whereas in the remaining 5 cases (31%) cytopenia(s) worsened requiring therapy at a median of 8 (range 5-11) months of follow-up (see Figure). Four of these latter 5 patients were retreated with vemurafenib for 12 weeks: 3 cases had a minor response and the last one witnessed a second PR that lasted less than the first PR (3 versus 9 months). CONCLUSIONS: In heavily pre-treated HCL patients, a short oral course of vemurafenib was safe, and proved quickly and highly active. Retreatment with vemurafenib was able to reinduce remissions in patients relapsing after a CR, but was less effective in patients relapsing after a PR. Figure 1 Figure 1. Disclosures Off Label Use: Off-label use of vemurafenib in hairy cell leukemia will be discussed as part of a clinical research protocol..

Description: The somatically acquired V600E mutation of the BRAF gene has been recently described as a molecular marker of hairy cell leukemia (HCL). We developed an allele-specific PCR for this mutation and studied 62 patients with HCL, 1 with HCL variant, 91 with splenic marginal zone lymphoma, 29 with Waldenström macroglobulinemia, and 57 with B-cell chronic lymphoproliferative disorders. The BRAF V600E mutation was detected in all HCL cases and in only 2 of the remaining 178 patients. These 2 subjects had B-cell chronic lymphoproliferative disorders that did not fulfill the diagnostic criteria for HCL. Despite the positive PCR finding, the mutation could not be detected by Sanger sequencing in these 2 cases, suggesting that it was associated with a small subclone. We conclude that the BRAF V600E mutation is present in all patients with HCL and that, in combination with clinical and morphologic features, represents a reliable molecular marker for this condition.

Description: Abstract 3495 WM is a rare malignant B-cell disorder characterized by lymphoplasmacytic infiltration of the bone marrow (BM) and hypersecretion of monoclonal IgM. IgMMGUS is an asymptomatic condition characterized by the presence of a serum monoclonal IgM protein and bone marrow infiltration 〈 10%. WM (symptomatic and indolent) and IgMMGUS can be identified based on two main features, the bone marrow infiltration and the existence of signs and symptoms. The biological and genetic characteristics of both conditions need to be explored. Our study aims to highlight the different expression profiles between WM and IgMMGUS comparing CD19+ as well as CD138+ cells. We have investigated patients with WM (n =21) and patients affected by IgMMGUS (n=10). BM CD19+ and BM CD138+ cells were isolated from 21 WM patients, while BM CD19+ and BM CD138+ were isolated from 10 and 4 IgMMGUS patients, respectively. Microarray analysis was performed using Affymetrix GeneChip Human Genome U133 Plus 2.0 Array. Data was preprocessed using Robust Multi-Array Average (RMA) software. Differential expression analysis was performed using Significant Analysis of Microarrays (SAM). Genes showing a q value lower than 5% and an absolute fold change greater than 2 were selected for further clustering analysis which was performed applying complete linkage hierarchical agglomerative clustering on Euclidean pairwise distances between genes. Microarray of WM vs IgMMGUS CD19+ cells has highlighted 151 differently expressed genes (Fig. 1). Among them we have found 33 genes involved in the regulation of transcription which were significantly overexpressed in WM vs. IgMMGUS. BM WM CD19+ cells overexpressed the following 14 Zinc Finger Protein (ZNF) genes: ZBTB40, ZNF83, ZNF137P, ZNF177, ZNF224, ZNF264, ZNF320, ZNF395, ZNF514, ZNF532, ZNF623, ZNF767, ZNF785, ZNF850. Other genes acting as regulators of transcription overexpressed in WM B cells were: HIF1AN, BHLHE41, EZH1, CCNL2, TCFL5, BLZF1, CIITA, PER2, MECP2, PRDM2, and ELP2. In particular, HIF1AN belongs to the PI3K/Akt/mTOR pathway, ELP2 is involved in the JAK/STAT process, and BHLHE41 and PER2 are included in the cicardian clock showing that these different biological mechanisms differently develop in WM with respect to IgMMGUS. TNFRSF10A, MAP4K4, TNFRSF10B, WNK1, DUSP22, ITPKB genes were overexpressed in WM B cells showing the involvement of Akt and MAPK signaling pathways which can play an important role in WM cells as they regulate several biological processes including cell growth, differentiation, survival, migration and metabolism. Gene expression profiling across CD138+ cells have demonstrated 43 differently expressed genes between WM vs. IgMMGUS (Fig. 2). MS4A1, BANK1 genes were overexpressed with high fold changes (FC) of 11.6 and 9.4, respectively, as well as GPR183, SWAP70 which were significantly overexpressed in WM vs. IGMMGUS, both genes showing a FC of 5. These results suggest that B cell activation and immune response are biological processes which act differently in WM compared to IgMMGUS. RALGPS2(FC=4), PLEKHG1 (FC=10) and ARHGAP24 (FC=4) genes mediating GTPase regulator activity of signal transduction were overexpressed in WM. ARHGAP24 could also be involved in the modulation of angiogenesis. FCRL2 and FCRLA genes involved in cell-cell signaling and cell differentiation, were significantly overexpressed in WM vs. IgMMGUS CD138+ with a fold change of 4.6 and 9, respectively. Again, the immune response seems to be a biological mechanism involved in WM CD138+ cells as CD79B (FC=5) and HLA-DOA (FC=4) genes were overexpressed in WM in respect to IgMMGUS. These differences may reflect varied immune mechanisms in the two disorders. In conclusion, the regulation of transcription, PI3K/Akt/mTOR and MAPK signaling pathways are the most relevant gene ontology biological processes occurring in CD19+ cells, while immune response, cell activation and signaling processes developing in CD138+ cells mainly distinguish WM and IgMMGUS. Future studies of the biological role of the genes differently expressed in WM vs IgMMGUS could clarify the pathogenetic processes underlying IgMMGUS and WM. The understanding of the molecular mechanisms leading to the progression of IgMMGUS to WM could help the identification of IgMMGUS patients who are at high risk for progression to WM. Fig. 1 GEP of BM WM CD19+ cells vs. IgMMGUS CD19+ cells. Fig. 1. GEP of BM WM CD19+ cells vs. IgMMGUS CD19+ cells. Fig. 2 GEP of BM WM CD138+ cells vs. IgMMGUS CD138+ cells. Fig. 2. GEP of BM WM CD138+ cells vs. IgMMGUS CD138+ cells. Disclosures: No relevant conflicts of interest to declare.