MYD88 wild-type Waldenstrom Macroglobulinaemia: differential diagnosis, risk of histological transformation, and overall survival.

MYD88 mutations are present in 95% of Waldenstrom Macroglobulinaemia (WM) patients, and support diagnostic discrimination from other IgM-secreting B-cell malignancies. Diagnostic discrimination can be difficult among suspected wild-type MYD88 (MYD88WT ) WM cases. We systematically reviewed the clinical, pathological and laboratory studies for 64 suspected MYD88WT WM patients. World Health Organization and WM consensus guidelines were used to establish clinicopathological diagnosis. Up to 30% of suspected MYD88WT WM cases had an alternative clinicopathological diagnosis, including IgM multiple myeloma. The estimated 10-year survival was 73% (95% confidence interval [CI] 52-86%) for MYD88WT versus 90% (95% CI 82-95%) for mutated (MYD88MUT ) WM patients (Log-rank P < 0·001). Multivariate analysis only showed MYD88 mutation status (P < 0·001) as a significant determinant for overall survival. Diffuse large B-cell lymphoma (DLBCL) was diagnosed in 7 (15·2%) and 2 (0·76%) of MYD88WT and MYD88MUT patients, respectively (Odds ratio 23·3; 95% CI 4·2-233·8; P < 0·001). Overall survival was shorter among MYD88WT patients with an associated DLBCL event (Log-rank P = 0·08). The findings show that among suspected MYD88WT WM cases, an alternative clinicopathological diagnosis is common and can impact clinical care. WM patients with MYD88WT disease have a high incidence of associated DLBCL events and significantly shorter survival versus those with MYD88MUT disease.

Transcriptome sequencing reveals a profile that corresponds to genomic variants in Waldenström macroglobulinemia.

Whole-genome sequencing has identified highly prevalent somatic mutations including MYD88, CXCR4, and ARID1A in Waldenström macroglobulinemia (WM). The impact of these and other somatic mutations on transcriptional regulation in WM remains to be clarified. We performed next-generation transcriptional profiling in 57 WM patients and compared findings to healthy donor B cells. Compared with healthy donors, WM patient samples showed greatly enhanced expression of the VDJ recombination genes DNTT, RAG1, and RAG2, but not AICDA Genes related to CXCR4 signaling were also upregulated and included CXCR4, CXCL12, and VCAM1 regardless of CXCR4 mutation status, indicating a potential role for CXCR4 signaling in all WM patients. The WM transcriptional profile was equally dissimilar to healthy memory B cells and circulating B cells likely due increased differentiation rather than cellular origin. The profile for CXCR4 mutations corresponded to diminished B-cell differentiation and suppression of tumor suppressors upregulated by MYD88 mutations in a manner associated with the suppression of TLR4 signaling relative to those mutated for MYD88 alone. Promoter methylation studies of top findings failed to explain this suppressive effect but identified aberrant methylation patterns in MYD88 wild-type patients. CXCR4 and MYD88 transcription were negatively correlated, demonstrated allele-specific transcription bias, and, along with CXCL13, were associated with bone marrow disease involvement. Distinct gene expression profiles for patients with wild-type MYD88, mutated ARID1A, familial predisposition to WM, chr6q deletions, chr3q amplifications, and trisomy 4 are also described. The findings provide novel insights into the molecular pathogenesis and opportunities for targeted therapeutic strategies for WM.

Self-starting optical-electrical-optical homodyne clock recovery for phase-modulated signals.

We propose a novel self-homodyne optical-electrical-optical clock recovery technique for binary phase-shift keying (BPSK) signals using commercial optical and electrical components. We present the principle of operation as well as a proof-of-concept experiment for a 10.7 Gb/s BPSK signal clock recovery transmitted over a dispersion-compensated link of 20 km of single-mode fiber. Suppression of pattern-related frequency noise at the output of the recovered clock is shown. The timing jitter of the recovered clock at 10.7 GHz was measured to be ∼450 fs (integration range: 100 Hz-10 MHz).

Renal disease related to Waldenström macroglobulinaemia: incidence, pathology and clinical outcomes.

The incidence and prognostic impact of nephropathy related to Waldenström macroglobulinaemia (WM) is currently unknown. We performed a retrospective study to assess biopsy-confirmed WM-related nephropathy in a cohort of 1391 WM patients seen at a single academic institution. A total of 44 cases were identified, the estimated cumulative incidence was 5·1% at 15 years. There was a wide variation in kidney pathology, some directly related to the WM: amyloidosis (n = 11, 25%), monoclonal-IgM deposition disease/cryoglobulinaemia (n = 10, 23%), lymphoplasmacytic lymphoma infiltration (n = 8, 18%), light-chain deposition disease (n = 4, 9%) and light-chain cast nephropathy (n = 4, 9%), and some probably related to the WM: thrombotic microangiopathy (TMA) (n = 3, 7%), minimal change disease (n = 2, 5%), membranous nephropathy (n = 1, 2%) and crystal-storing tubulopathy (n = 1, 2%). The median overall survival in patients with biopsy-confirmed WM-related nephropathy was 11·5 years, shorter than for the rest of the cohort (16 years, P = 0·03). Survival was better in patients with stable or improved renal function after treatment (P = 0·05). Based on these findings, monitoring for renal disease in WM patients should be considered and a kidney biopsy pursued in those presenting with otherwise unexplained renal failure and/or nephrotic syndrome.

The genomic landscape of Waldenstrom macroglobulinemia is characterized by highly recurring MYD88 and WHIM-like CXCR4 mutations, and small somatic deletions associated with B-cell lymphomagenesis.

The genetic basis for Waldenström macroglobulinemia (WM) remains to be clarified. Although 6q losses are commonly present, recurring gene losses in this region remain to be defined. We therefore performed whole genome sequencing (WGS) in 30 WM patients, which included germline/tumor sequencing for 10 patients. Validated somatic mutations occurring in >10% of patients included MYD88, CXCR4, and ARID1A that were present in 90%, 27%, and 17% of patients, respectively, and included the activating mutation L265P in MYD88 and warts, hypogammaglobulinemia, infection, and myelokathexis-syndrome-like mutations in CXCR4 that previously have only been described in the germline. WGS also delineated copy number alterations (CNAs) and structural variants in the 10 paired patients. The CXCR4 and CNA findings were validated in independent expansion cohorts of 147 and 30 WM patients, respectively. Validated gene losses due to CNAs involved PRDM2 (93%), BTG1 (87%), HIVEP2 (77%), MKLN1 (77%), PLEKHG1 (70%), LYN (60%), ARID1B (50%), and FOXP1 (37%). Losses in PLEKHG1, HIVEP2, ARID1B, and BCLAF1 constituted the most common deletions within chromosome 6. Although no recurrent translocations were observed, in 2 patients deletions in 6q corresponded with translocation events. These studies evidence highly recurring somatic events, and provide a genomic basis for understanding the pathogenesis of WM.

MYD88 L265P somatic mutation in Waldenström's macroglobulinemia.

BACKGROUND: Waldenström's macroglobulinemia is an incurable, IgM-secreting lymphoplasmacytic lymphoma (LPL). The underlying mutation in this disorder has not been delineated. METHODS: We performed whole-genome sequencing of bone marrow LPL cells in 30 patients with Waldenström's macroglobulinemia, with paired normal-tissue and tumor-tissue sequencing in 10 patients. Sanger sequencing was used to validate the findings in samples from an expanded cohort of patients with LPL, those with other B-cell disorders that have some of the same features as LPL, and healthy donors. RESULTS: Among the patients with Waldenström's macroglobulinemia, a somatic variant (T→C) in LPL cells was identified at position 38182641 at 3p22.2 in the samples from all 10 patients with paired tissue samples and in 17 of 20 samples from patients with unpaired samples. This variant predicted an amino acid change (L265P) in MYD88, a mutation that triggers IRAK-mediated NF-κB signaling. Sanger sequencing identified MYD88 L265P in tumor samples from 49 of 54 patients with Waldenström's macroglobulinemia and in 3 of 3 patients with non-IgM-secreting LPL (91% of all patients with LPL). MYD88 L265P was absent in paired normal tissue samples from patients with Waldenström's macroglobulinemia or non-IgM LPL and in B cells from healthy donors and was absent or rarely expressed in samples from patients with multiple myeloma, marginal-zone lymphoma, or IgM monoclonal gammopathy of unknown significance. Inhibition of MYD88 signaling reduced IκBα and NF-κB p65 phosphorylation, as well as NF-κB nuclear staining, in Waldenström's macroglobulinemia cells expressing MYD88 L265P. Somatic variants in ARID1A in 5 of 30 patients (17%), leading to a premature stop or frameshift, were also identified and were associated with an increased disease burden. In addition, 2 of 3 patients with Waldenström's macroglobulinemia who had wild-type MYD88 had somatic variants in MLL2. CONCLUSIONS: MYD88 L265P is a commonly recurring mutation in patients with Waldenström's macroglobulinemia that can be useful in differentiating Waldenström's macroglobulinemia and non-IgM LPL from B-cell disorders that have some of the same features. (Funded by the Peter and Helen Bing Foundation and others.).

A mutation in MYD88 (L265P) supports the survival of lymphoplasmacytic cells by activation of Bruton tyrosine kinase in Waldenström macroglobulinemia.

Myeloid differentiation factor 88 (MYD88) L265P somatic mutation is highly prevalent in Waldenström macroglobulinemia (WM) and supports malignant growth through nuclear factor κB (NF-κB). The signaling cascade(s) by which MYD88 L265P promotes NF-κB activation in WM remain unclear. By lentiviral knockdown or use of a MYD88 inhibitor, decreased phosphorylation of the NF-κB gatekeeper IκBα and survival occurred in MYD88 L265P-expressing WM cells. Conversely, WM cells engineered to overexpress MYD88 L265P showed enhanced survival. Coimmunoprecipitation studies identified Bruton tyrosine kinase (BTK) complexed to MYD88 in L265P-expressing WM cells, with preferential binding of MYD88 to phosphorylated BTK (pBTK). Increased pBTK was also observed in WM cells transduced to overexpress L265P vs wild-type MYD88. Importantly, MYD88 binding to BTK was abrogated following treatment of MYD88 L265P-expressing cells with a BTK kinase inhibitor. Inhibition of BTK or interleukin-1 receptor-associated kinase 1 and 4 (IRAK-1 and -4) kinase activity induced apoptosis of WM cells, and their combination resulted in more robust inhibition of NF-κB signaling and synergistic WM cell killing. The results establish BTK as a downstream target of MYD88 L265P signaling, and provide a framework for the study of BTK inhibitors alone, and in combination with IRAK inhibitors for the treatment of WM.

MYD88 L265P in Waldenström macroglobulinemia, immunoglobulin M monoclonal gammopathy, and other B-cell lymphoproliferative disorders using conventional and quantitative allele-specific polymerase chain reaction.

By whole-genome and/or Sanger sequencing, we recently identified a somatic mutation (MYD88 L265P) that stimulates nuclear factor κB activity and is present in >90% of Waldenström macroglobulinemia (WM) patients. MYD88 L265P was absent in 90% of immunoglobulin M (IgM) monoclonal gammopathy of undetermined significance (MGUS) patients. We therefore developed conventional and real-time allele-specific polymerase chain reaction (AS-PCR) assays for more sensitive detection and quantification of MYD88 L265P. Using either assay, MYD88 L265P was detected in 97 of 104 (93%) WM and 13 of 24 (54%) IgM MGUS patients and was either absent or rarely expressed in samples from splenic marginal zone lymphoma (2/20; 10%), CLL (1/26; 4%), multiple myeloma (including IgM cases, 0/14), and immunoglobulin G MGUS (0/9) patients as well as healthy donors (0/40; P < 1.5 × 10(-5) for WM vs other cohorts). Real-time AS-PCR identified IgM MGUS patients progressing to WM and showed a high rate of concordance between MYD88 L265P ΔCT and BM disease involvement (r = 0.89, P = .008) in WM patients undergoing treatment. These studies identify MYD88 L265P as a widely present mutation in WM and IgM MGUS patients using highly sensitive and specific AS-PCR assays with potential use in diagnostic discrimination and/or response assessment. The finding of this mutation in many IgM MGUS patients suggests that MYD88 L265P may be an early oncogenic event in WM pathogenesis.

Practical and cost-effective high-fidelity optical carrier dissemination using coherent communication techniques.

We report a unidirectional frequency dissemination scheme for high-fidelity optical carriers deployable over telecommunication networks. For the first time, a 10 Gb/s Binary Phase Shift Keying (BPSK) signal from an ultra-narrow linewidth laser was transmitted through a field-installed optical fibre with round-trip length of 124 km between Cork City and town of Clonakilty, without inline optical amplification. At the receiver, using coherent communication techniques and optical injection-locking the carrier was recovered with noise suppression. The beat signal between the original carrier at the transmitter and recovered carrier at the receiver shows a linewidth of 2.8 kHz. Long term stability measurements revealed fractional instabilities (True Allan deviation) of 3.3 × 10(-14) for 1 s averaging time, prior to phase noise cancellation.

Four-wave mixing for clock recovery of phase modulated optical OFDM superchannel.

We simulate and experimentally demonstrate a novel all-optical clock recovery technique for a BPSK OFDM superchannel. Four-wave mixing in SOAs is used to strip the modulation from the superchannel sub-carriers, two of which are filtered and beat together in a photodiode to recover the clock.

All-optical phase discrimination using SOA.

We describe the first experimental demonstration of a novel all-optical phase discrimination technique, which can separate the two orthogonal phase components of a signal onto different frequencies. This method exploits nonlinear mixing in a semiconductor optical amplifier (SOA) to separate a 10.65 Gbaud QPSK signal into two 10.65 Gb/s BPSK signals which are then demodulated using a delay interferometer (DI). Eye diagrams and spectral measurements verify correct operation and a conversion efficiency greater than 9 dB is observed on both output BPSK channels when compared with the input QPSK signal.

High speed cross-amplitude modulation in concatenated SOA-EAM-SOA.

We observe a near-ideal high speed amplitude impulse response in an SOA-EAM-SOA configuration under optimum conditions. Full amplitude recovery times as low as 10 ps with modulation depths of 70% were observed in pump-probe measurements. System behavior could be controlled by the choice of signal wavelength, SOA current biases and EAM reverse bias voltages. Experimental data and impulse response modelling indicated that the slow tail in the gain response of first SOA was negated by a combination of cross-absorption modulation between pump and modulated CW probe, and self-gain modulation of the modulated CW probe in both the EAM and second SOA.

Maintenance Rituximab is associated with improved clinical outcome in rituximab naïve patients with Waldenstrom Macroglobulinaemia who respond to a rituximab-containing regimen.

This study examined the outcome of 248 Waldenstrom macroglobulinaemia (WM) rituximab-naïve patients who responded to a rituximab-containing regimen. Eighty-six patients (35%) subsequently received maintenance rituximab (M-Rituximab). No differences in baseline characteristics, and post-induction categorical responses between cohorts were observed. The median rituximab infusions during induction was 6 for both cohorts; and 8 over a 2-year period for patients receiving M-Rituximab. Categorical responses improved in 16/162 (10%) of observed, and 36/86 (41·8%) of M-Rituximab patients respectively, following induction therapy (P < 0·0001). Both progression-free (56·3 vs. 28·6 months; P = 0·0001) and overall survival (Not reached versus 116 months; P = 0·0095) were longer in patients who received M-Rituximab. Improved progression-free survival was evident despite previous treatment status, induction with rituximab alone or in combination therapy (P ≤ 0·0001). Best serum IgM response was lower (P < 0·0001), and haematocrit higher (P = 0·001) for patients receiving M-Rituximab. Among patients receiving M-Rituximab, an increased number of infectious events were observed, but were mainly ≤ grade 2 (P = 0·008). The findings of this observational study suggest improved clinical outcomes following M-Rituximab in WM patients who respond to induction with a rituximab-containing regimen. Prospective studies aimed at clarifying the role of M-Rituximab therapy in WM patients are needed to confirm these findings.

Attainment of complete/very good partial response following rituximab-based therapy is an important determinant to progression-free survival, and is impacted by polymorphisms in FCGR3A in Waldenstrom macroglobulinaemia.

The incorporation of rituximab into various regimens has improved depth of response in Waldenstrom macroglobulinaemia (WM), though the impact of achieving better responses remains to be determined. We examined response depth on progression-free survival (PFS) in 159 rituximab-naïve WM patients who received rituximab-based therapy. The median follow-up was 33·5 months, and categorical responses were as follows: complete response (CR, 8·8%); very good partial response (VGPR, 13·2%); partial response (50%); minor response (18·9%); Non-Responders (8·8%). Sequencing for polymorphic variants of FCGR2A, FCGR2B, and FCGR3A was performed, and impact on response depth determined. Achievement of better categorical responses was incrementally associated with improved PFS (P < 0·0001). No separation was observed between CR and VGPR, and attainment of at least a VGPR was associated with improved time-to-progression. Neither age, serum IgM, haematocrit, platelet count, serum ß(2) microglobulin, WM International Prognostic Scoring System score, and treatment group predicted for CR/VGPR. Polymorphisms at FCGR3A-48 and -158 were associated with improved categorical responses, particularly attainment of CR/VGPR (P ≤ 0·03). The attainment of CR/VGPR was associated with significantly longer PFS in rituximab-naïve WM patients undergoing rituximab-based therapy, and was predicted by polymorphisms in FCGR3A.

Endoplasmic reticulum stress is a target for therapy in Waldenstrom macroglobulinemia.

Waldenstrom macroglobulinemia (WM) is an incurable low-grade lymphoma characterized by bone marrow (BM) involvement of IgM secreting lymphoplasmacytic cells. The induction of unfolded protein response (UPR) genes ("physiologic" UPR) enables cells to differentiate into professional secretory cells capable of production of high amounts of endoplasmic reticulum (ER)-processed proteins, such as immunoglobulins. Ultimately, the initially cytoprotective UPR triggers an apoptotic cascade if ER stress is not corrected, called proapoptotic/terminal UPR. We show that WM cells inherently express the physiologic UPR machinery compared with normal BM cells, and that increased ER stress leads to proapoptotic/terminal UPR in WM cells. We therefore examined tunicamycin, ER stress inducer, for potential antitumor effects in WM. Tunicamycin induced significant cytotoxicity, apoptosis and cell-cycle arrest, and inhibited DNA synthesis in WM cell lines and primary BM CD19(+) cells from patients with WM with an inhibitory concentration (IC(50)) of 0.5 microg/mL to 1 microg/mL, but not in healthy donor cells. Importantly, coculture of WM cells in the context of the BM microenvironment did not inhibit tunicamycin-induced cytotoxicity. Finally, we demonstrate that ER stress inducer synergizes with other agents used in the treatment of WM. These preclinical studies provide a framework for further evaluation of ER stress inducing agents as therapeutic agents in WM.

High-bandwidth generation of duobinary and alternate-mark-inversion modulation formats using SOA-based signal processing.

We report on the novel all-optical generation of duobinary (DB) and alternate-mark-inversion (AMI) modulation formats at 42.6 Gb/s from an input on-off keyed signal. The modulation converter consists of two semiconductor optical amplifier (SOA)-based Mach-Zehnder interferometer gates. A detailed SOA model numerically confirms the operational principles and experimental data shows successful AMI and DB conversion at 42.6 Gb/s. We also predict that the operational bandwidth can be extended beyond 40 Gb/s by utilizing a new pattern-effect suppression scheme, and demonstrate dramatic reductions in patterning up to 160 Gb/s. We show an increasing trade-off between pattern-effect reduction and mean output power with increasing bitrate.

CD27-CD70 interactions in the pathogenesis of Waldenstrom macroglobulinemia.

Waldenström macroglobulinemia (WM) is a B-cell malignancy characterized by an IgM monoclonal gammopathy and bone marrow (BM) infiltration with lymphoplasmacytic cells (LPCs). Excess mast cells (MCs) are commonly present in WM, and provide growth and survival signals to LPCs through several TNF family ligands (CD40L, a proliferation-inducing ligand [APRIL], and B-lymphocyte stimulator factor [BLYS]). As part of these studies, we demonstrated that WM LPCs secrete soluble CD27 (sCD27), which is elevated in patients with WM (P < .001 vs healthy donors), and serves as a faithful marker of disease. Importantly, sCD27 stimulated expression of CD40L on 10 of 10 BM MC samples and APRIL on 4 of 10 BM MC samples obtained from patients with WM as well as on LAD2 MCs. Moreover, the SGN-70 humanized monoclonal antibody, which binds to CD70 (the receptor-ligand partner of CD27), abrogated sCD27 mediated up-regulation of CD40L and APRIL on WM MCs. Last, treatment of severe combined immunodeficiency-human (SCID-hu) mice with established WM using the SGN-70 antibody blocked disease progression in 12 of 12 mice, whereas disease progressed in all 5 untreated mice. The results of these studies demonstrate a functional role for sCD27 in WM pathogenesis, along with its utility as a surrogate marker of disease and a target in the treatment of WM.

Fast gain recovery rates with strong wavelength dependence in a non-linear SOA.

We report remarkably fast and strongly wavelength-dependent gain recovery in a single SOA without the aid of an offset filter. Full gain recovery times as short as 9 ps were observed in pump-probe measurements when pumping to the blue wavelength side of a continuous wave probe, in contrast to times of 25 to 30 ps when pumping to the red wavelength side. Experimental and numerical analysis indicate that the long effective length and high gain led to deep saturation of the second half of the SOA by the probe. The consequent absorption of blue-shifted pump pulses in this region resulted in device dynamics analogous to those of the Turbo-Switch.

IgA and IgG hypogammaglobulinemia in Waldenström's macroglobulinemia.

BACKGROUND: Hypogammaglobulinemia is common in Waldenström's macroglobulinemia. The etiology of this finding remains unclear, but it has been speculated to be based on tumor-induced suppression of the 'uninvolved' immunoglobulin production DESIGN AND METHODS: We evaluated the incidence of IgA and IgG hypogammaglobulinemia in 207 untreated patients with Waldenström's macroglobulinemia and investigated the associated clinicopathological findings and impact of therapy. We also sequenced eight genes (AICDA, BTK, CD40, CD154, NEMO, TACI, SH2D1A, UNG) implicated in immunoglobulin deficiency in 19 Waldenström's macroglobulinemia patients with IgA and/or IgG hypogammaglobulinemia. RESULTS: At baseline 63.3%, 58.0% and 49.3% of the 207 patients had abnormally low serum levels of IgA, IgG, or both. No association between IgA and IgG hypogammaglobulinemia and disease burden, serum IgM levels, beta(2)-microglobulin, International Prognostic Scoring System score, or incidence of recurrent infections was observed, although the presence of adenopathy and/or splenomegaly was associated with a lower incidence of hypogammaglobulinemia. Lower IgA and IgG levels were associated with disease progression in patients managed with a 'watch and wait' strategy. IgA and/or IgG levels remained abnormally low despite response to treatment, including complete remissions. A missense mutation in the highly conserved catalytic site of UNG was observed in a patient with hypogammaglobulinemia, warranting further study of this pathway in Waldenström's macroglobulinemia. CONCLUSIONS: IgA and IgG hypogammaglobulinemia is common in Waldenström's macroglobulinemia and persists despite therapeutic intervention and response. IgA and IgG hypogammaglobulinemia does not predict the risk of recurrent infections in patients with Waldenström's macroglobulinemia, although lower levels of serum IgA and IgG are associated with disease progression in Waldenström's macroglobulinemia patients being managed with a 'watch and wait' strategy.