Clinicopathological, cytogenetic, and molecular profiles of primary cutaneous diffuse large B-cell lymphomas.

We analyzed the clinicopathological, cytogenetic, and molecular features of 18 primary cutaneous diffuse large B-cell lymphomas (PCDLBCLs) and 15 DLBCLs secondarily localized to the skin (SCDLBCLs), highlighting biological similarities and differences between the 2 groups. PCDLBCLs were subclassified after histopathological review as PCDLBCL-leg type (PCDLBCL-LT, 10 cases) and the PCDLBCL-not otherwise specified (PCDLBCL-NOS, 8 cases). Immunohistochemistry for Hans' algorithm markers, BCL2, and MYC was performed. The molecular study included the determination of the cell of origin (COO) by Lymph2Cx assay on NanoString platform, FISH analysis of IgH, BCL2, BCL6, and MYC genes, as well as the mutation analysis of MYD88 gene. In immunohistochemistry analysis, BCL2 and MYC hyperexpression was more frequent in LT than in NOS cases and, according to Hans' algorithm, PCDLBCL-LTs were mostly of the non-GC type (8/10), whereas in PCDLBCL-NOS, the GC type prevailed (6/8). The determination of COO using Lymph2Cx supported and further confirmed these results. In FISH analysis, all but one LT cases versus 5 of 8 PCDLBCL-NOS showed at least one gene rearrangement among IgH, BCL2, MYC, or BCL6. In addition, MYD88 mutations were more frequently present in LT than in NOS subtypes. Interestingly, MYD88-mutated patients were older, with a non-GC phenotype and had worse OS, compared to MYD88 WT cases. Overall, SCDLBCL did not show, at the genetic and expression level, different profiles than PCDLBCL, even if they bear a significantly worse prognosis. At survival analysis, the most important prognostic factors in patients with PCDLBCL were age and MYD88 mutation, whereas relapse and high Ki-67 expression were relevant in patients with SCDLBCL. Our study comprehensively analyzed the clinicopathological and molecular features of PCDLBCL-LT, PCDLBCL-NOS, and SCDLBCL, underlining the differences among them and the importance of properly identifying these entities at the time of diagnosis.

New occurrence of Meloidogyne graminicola (Nematoda: Meloidogyninae) from rice fields in Italy: Variability and phylogenetic relationships.

Since the first detection of Meloidogyne graminicola in Piedmont, North Italy, in 2016, further inspections for the presence of the rice root-knot nematode were carried out in rice fields of neighboring regions, in accordance with the Italian NPPO (National Plant Protection Organization) to support the official phytosanitary measures, to enable the early detection of the rice pest, and to prevent its spread within the national territory. In 2018, surveys of rice fields in Lombardy region revealed a new occurrence of M. graminicola. In the present study, we confirmed the identification of the rice nematode in Lombardy using the ribosomal ITS region and the mitochondrial COI and COII genes. The sequences and phylogenetic analyses revealed that Lombardy M. graminicola population grouped in all trees in the main cluster containing Meloidogyne species belonging to graminis group, but always in a different subgroup compared to the Piedmont population of M. graminicola. These results clearly suggest that the two Italian populations have been recently and independently introduced and confirm that the geographic origin is not the main factor leading to M. graminicola population variability.

Cytokine Release Syndrome Associated with T-Cell-Based Therapies for Hematological Malignancies: Pathophysiology, Clinical Presentation, and Treatment.

Cytokines are a broad group of small regulatory proteins with many biological functions involved in regulating the hematopoietic and immune systems. However, in pathological conditions, hyperactivation of the cytokine network constitutes the fundamental event in cytokine release syndrome (CRS). During the last few decades, the development of therapeutic monoclonal antibodies and T-cell therapies has rapidly evolved, and CRS can be a serious adverse event related to these treatments. CRS is a set of toxic adverse events that can be observed during infection or following the administration of antibodies for therapeutic purposes and, more recently, during T-cell-engaging therapies. CRS is triggered by on-target effects induced by binding of chimeric antigen receptor (CAR) T cells or bispecific antibody to its antigen and by subsequent activation of bystander immune and non-immune cells. CRS is associated with high circulating concentrations of several pro-inflammatory cytokines, including interleukins, interferons, tumor necrosis factors, colony-stimulating factors, and transforming growth factors. Recently, considerable developments have been achieved with regard to preventing and controlling CRS, but it remains an unmet clinical need. This review comprehensively summarizes the pathophysiology, clinical presentation, and treatment of CRS caused by T-cell-engaging therapies utilized in the treatment of hematological malignancies.

Improving the international prognostic index score using peripheral blood counts: Results of a large multicenter study involving 520 patients with diffuse large B cell lymphoma.

The main purpose of this study was to assess whether it is possible to improve the prognostic impact of international prognostic index (IPI) score by combining it with peripheral blood counts. Thus, we evaluated the prognostic power of lymphocyte, neutrophil, and monocyte counts in 520 patients with diffuse large B cell lymphoma treated with R-CHOP, confirming that these parameters have a strong impact on overall survival (OS). Using revised IPI (R-IPI), 44% of patients were categorized as poor-risk and showed an OS at 5 years of 46%. As OS at 5 years of the 520 patients is 67%, it is clearly evident that R-IPI tends to overestimate the proportion of patients with poor prognosis. Accordingly, in an attempt to improve the discriminating power of R-IPI, we evaluated and compared three different scores by combining the neutrophil lymphocyte ratio (NLR) and absolute monocyte count (AMC) with the following values: (a) IPI score 3-5, (b) age > 60 years and performance status, (c) age ≥ 65 years and LDH > ULN. The three indexes studied, had a similar 5 years OS for the high-risk group (46%-52%), but the proportion of patients classified as poor-risk were 37%, 20%, and 32%, respectively, which are lower than 44% identified with R-IPI. Thus, while R-IPI overestimates the number of high-risk patients, after applying our models, it is possible to recognize patients who are truly at high-risk. Of the three scores, the most accurate appears to be that based on NLR, AMC, LDH > ULN and age ≥ 65 years, which identifies 32% of high-risk patients, correlating well with what is seen in clinical practice.

Citarinostat and Momelotinib co-target HDAC6 and JAK2/STAT3 in lymphoid malignant cell lines: a potential new therapeutic combination.

Histone deacetylase (HDAC) inhibitors represent an encouraging class of antitumor drugs. HDAC inhibitors induce a series of molecular and biological responses and minimal toxicity to normal cells. Citarinostat (Acy-241) is a second generation, orally administered, HDAC6-selective inhibitor. Momelotinib (CYT387) is an orally administered inhibitor of Janus kinase/signal transducer of transcription-3 (JAK/STAT3) signaling. Momelotinib showed efficacy in patients with myelofibrosis. We hypothesized that both HDAC and JAK/STAT pathways were important in lymphoproliferative disorders, and that inhibiting JAK/STAT3 and HDAC simultaneously might enhance the efficacy of momelotinib and citarinostat without increasing toxicity. Accordingly, we tested the citarinostat + momelotinib combination in lymphoid cell lines. Citarinostat + momelotinib showed strong cytotoxicity; it significantly reduced mitochondrial membrane potential, down-regulated Bcl-2 and Bcl-xL, and activated caspases 9 and 3. Caspase-8 was upregulated in only two lymphoid cell lines, which indicated activation of the extrinsic apoptotic pathway. We identified a lymphoid cell line that was only slightly sensitive to the combination treatment. We knocked down thioredoxin expression by transfecting with small interfering RNA that targeted thioredoxin. This knockdown increased cell sensitivity to the combination-induced cell death. The combination treatment reduced Bcl-2 expression, activated caspase 3, and significantly inhibited cell viability and clonogenic survival.

Nail StrainStress Meter NM 100: A novel in vivo method to characterize biomechanical properties of nails.

BACKGROUND: Nowadays, nail care products are extremely important both in medical and cosmetic fields. Actually, there are only a very few "in vivo" methods to evaluate the safety and the efficacy of nail products. METHODS: The new apparatus, based on a recently patented technology, is developed for the "in vivo" evaluation of nails in terms of thickness, structural firmness, flattening, and bending properties. The device analyzes nails by an "in vivo" non-invasive methodology in a timely way and with high accuracy. The assessment of the resistance to compression measures the cohesion of the nail matrix (nail firmness), while the evaluation of the resistance to transversal deformation detects the elasticity of the nail plate. Furthermore, the apparatus is able to assess the nail thickness and the flexibility of their distal edge. RESULTS: The instrument provides nail thickness and several parameters reflecting mechanical properties of nail plate: Viscoelasticity expressed as viscoelasticity index (VI), structural strength/ firmness expressed as Firmness Index (FI), and viscoelasticity of the distal edge expressed as Bending Index (BI). CONCLUSIONS: The instruments described in this work represent an innovative apparatus for the safety and efficacy evaluation of nail products in several fields: cosmetics, pharmaceuticals, and medical devices.

The classic prognostic factors in advanced Hodgkin's lymphoma patients are losing their meaning at the time of Pet-guided treatments.

The International Prognostic Score (IPS) is the most commonly used risk stratification tool for patients with advanced Hodgkin lymphoma (HL). It incorporates seven clinical parameters independently associated with a poorer outcome: male sex, age, stage IV, hemoglobin level, white blood cell and lymphocyte counts, and albumin level. Since the development of the IPS, there have been significant advances in therapy and supportive care. Recent studies suggest that the IPS is less discriminating due to improved outcomes with ABVD therapy. The aim of the present study was to asses if classic prognostic factors maintain their prognostic meaning at the time of response-adapted treatment based on interim PET scans. We evaluated the prognostic significance of IPS in the 520 advanced stage HL patients enrolled in the PET-guided, HD0801 trial in which PET2-positive patients underwent a more intense treatment with an early stem-cell transplantation after 2 cycles of ABVD. We observed that in these patients, the IPS completely loses its prognostic value together with all the single parameters that contribute to the IPS. Furthermore, neutrophils, monocytes, lymphocytes, and the ratio among them also no longer had any predictive value. We believe that the substantial improvement in survival outcomes in PET2-positive patients treated with early autologous transplantation could explain the complete disappearance of the residual prognostic significance of the IPS.

The prognostic role of end of treatment FDG-PET-CT in patients with diffuse large B cell lymphoma can be improved by considering it with absolute monocyte count at diagnosis.

It is well established that some patients with diffuse large B-cell lymphoma (DLBCL) and the negative end of treatment PET-CT (EOT-PET-CT) will relapse, while a proportion with positive uptake can still obtain long-term EFS. We reviewed data of 200 consecutive, previously untreated patients with DLBCL recorded in Italy and Israel between 2007 and 2015. We found that patients with negative EOT-PET-CT with AMC > 630/mmc have a 3-years EFS of 72%, compared to those with AMC ≤ 630/mmc that have an EFS of 84%. Furthermore, considering patients with positive EOT-PET-CT, those with AMC > 630/mmc have a 3-years EFS of 8%, while those with AMC ≤ 630/mmc have an EFS of 38%. Thus, it appears that combining the gold standard for response evaluation EOT-PET-CT with a simple and inexpensive parameter like AMC at diagnosis, further improves prognostication in DLBCL. Applying this simple method can be useful for all doctors working in lymphoma clinical practice.

Cell-Penetrating CaCO3 Nanocrystals for Improved Transport of NVP-BEZ235 across Membrane Barrier in T-Cell Lymphoma.

Owing to their nano-sized porous structure, CaCO3 nanocrystals (CaCO3NCs) hold the promise to be utilized as desired materials for encapsulating molecules which demonstrate wide promise in drug delivery. We evaluate the possibility to encapsulate and release NVP-BEZ235, a novel and potent dual PI3K/mTOR inhibitor that is currently in phase I/II clinical trials for advanced solid tumors, from the CaCO3NCs. Its chemical nature shows some intrinsic limitations which induce to administer high doses leading to toxicity; to overcome these problems, here we proposed a strategy to enhance its intracellular penetration and its biological activity. Pristine CaCO3 NCs biocompatibility, cell interactions and internalization in in vitro experiments on T-cell lymphoma line, were studied. Confocal microscopy was used to monitor NCs-cell interactions and cellular uptake. We have further investigated the interaction nature and release mechanism of drug loaded/released within/from the NCs using an alternative approach based on liquid chromatography coupled to mass spectrometry. Our approach provides a good loading efficiency, therefore this drug delivery system was validated for biological activity in T-cell lymphoma: the anti-proliferative test and western blot results are very interesting because the proposed nano-formulation has an efficiency higher than free drug at the same nominal concentration.

Ruxolitinib combined with vorinostat suppresses tumor growth and alters metabolic phenotype in hematological diseases.

JAK-2 dysregulation plays an important role as an oncogenic driver, and is thus a promising therapeutic target in hematological malignancies. Ruxolitinib is a pyrrolo[2.3-d]pyrimidine derivative with inhibitory activity against JAK1 and JAK2, moderate activity against TYK2, and minor activity against JAK3. Vorinostat is an HDAC inhibitor that reduces JAK-2 expression, thus affecting JAK-2 mRNA expression and increasing JAK-2 proteasomal deterioration. Here we hypothesized that the combination of ruxolitinib and vorinostat could have synergistic effects against hematological disease. We tested combinations of low doses of ruxolitinib and vorinostat in 12 cell lines, and observed highly synergistic cytotoxic action in six cell lines, which was maintained for up to 120 h in the presence of stromal cells. The sensitivity of the six cell lines may be explained by the broad effects of the drug combination, which can affect various targets. Treatment with the combination of ruxolitinib and vorinostat appeared to induce a possible reversal of the Warburg effect, with associated ROS production, apoptotic events, and growth inhibition. Decreased glucose metabolism may have markedly sensitized the six more susceptible cell lines to combined treatment. Therapeutic inhibition of the JAK/STAT pathway seems to offer substantial anti-tumor benefit, and combined therapy with ruxolitinib and vorinostat may represent a promising novel therapeutic modality for hematological neoplasms.

Hexamermis popilliae n. sp. (Nematoda: Mermithidae) parasitizing the Japanese beetle Popillia japonica Newman (Coleoptera: Scarabaeidae) in Italy.

A new species of mermithid nematode, Hexamermis popilliae n. sp. (Nematoda: Mermithidae) is described from the Japanese beetle Popillia japonica Newman in Italy, an area of new introduction for this invasive pest. The combination of the following characters separates H. popilliae from other members of the genus Hexamermis Steiner, 1924: adult head obtuse; amphidial pouches slightly posterior to lateral head papillae in female but adjacent to lateral head papillae in males; amphidial openings large, well developed; amphidial pouches elliptical in females and oblong in males; cuticular vulvar cone well developed, vulvar lips greatly reduced or lacking, vagina curved at tip where meeting uteri, without reverse bend (not S-shaped), spicules slightly curved, with a slight bend in the basal portion, approximately equal to body width at cloaca. This is the first record of a species of Hexamermis parasitizing the Japanese beetle Popillia japonica. The only previous mention of mermithid nematodes from P. japonica was an undescribed species of Psammomermis in North America. Hexamermis popilliae will be evaluated as a potential biological control agent in an integrated control program of the Japanese beetle in Italy.

Ricolinostat, a selective HDAC6 inhibitor, shows anti-lymphoma cell activity alone and in combination with bendamustine.

Histone deacetylase inhibitors (HDACis) have emerged as a new class of anticancer agents, targeting the biological process including cell cycle and apoptosis. We investigated and explained the anticancer effects of an HDAC6 inhibitor, ricolinostat alone and in combination with bendamustine in lymphoma cell lines. Cell viability was measured by MTT assay. Apoptosis, reactive oxygen species (ROS) generation, Bcl-2 protein expression, cell cycle progression and tubuline expression were determined by flow cytometry. The effects of ricolinostat alone and in combination on the caspases, PI3K/Akt, Bcl-2 pathways, ER stress and UPR were assessed by immunoblotting. Ricolinostat shows anti lymphoma activity when used as single agent and its capability to induce apoptosis is synergistically potentiated by the bendamustine in lymphoma cell lines. Drug combination reduced the proportion of cells in the G0/G1 and S phases and caused an increase of "sub-G0/G1" peak. The synergistic effect accompanied with the increased ROS, activation of caspase-8, -9, and -3, the cleavage of PARP and modulated by Bcl-2 proteins family. In addition, the exposure of ricolinostat induced the acetylation level of α-tubulin, the extend of which was not further modified by bendamustine. Finally, the apoptosis effect of ricolinostat/bendamustine may be mediated by a corresponding effect on microtubule stabilization. Our data suggest that ricolinostat in combination with bendamustine may be a novel combination with potential for use as an antitumor agent in lymphoma.

Neutrophil-lymphocyte ratio at diagnosis is an independent prognostic factor in patients with nodular sclerosis Hodgkin lymphoma: results of a large multicenter study involving 990 patients.

Several studies have demonstrated the prognostic value of neutrophil-lymphocyte ratio (NLR) in patients with solid tumors and non-Hodgkin lymphoma. In contrast, there is only sparse data on its prognostic role in patients with classical Hodgkin lymphoma (cHL). The aim of our study was to establish whether NLR could serve as an independent prognostic factor in a cohort of 990 patients with nodular sclerosis (NS)-cHL. After analysis of the log hazard ratio (HR) as a function of NLR, we chose the value 6 as cutoff. Patients with NLR >6 had a worse progression-free survival and overall survival compared to those with NLR ≤6; 84% vs 75% and 92% vs 88%, at 5 years, with an HR of 1.65 and 1.82, respectively. Multivariate analysis showed that the risk remained high with HR 1.44 and HR 1.54 in progression-free survival and overall survival, respectively. In summary, our study shows that NLR is a robust and independent prognostic parameter in NS-cHL, both in early and advanced disease. It is inexpensive and simple to apply. Thus, we conclude that NLR, possibly in combination with the international prognostic score and absolute monocyte count, is a useful guide for physicians treating NS-cHL patients.

A Multicenter Phase II Study of Twice-Weekly Bortezomib plus Rituximab in Patients with Relapsed Follicular Lymphoma: Long-Term Follow-Up.

Single-agent bortezomib (B) has shown activity in heavily pretreated patients with relapsed/refractory indolent lymphoma. On the basis of these findings, we performed a phase II study of B combined with rituximab (R) in patients with relapsed follicular lymphoma (FL). Forty-five patients with fairly good prognostic profiles were enrolled from 2007 to 2011 and received a total of 6 cycles of the B+R combination. The endpoints were the overall response rate (ORR), progression-free survival (PFS), duration of remission (DoR), overall survival (OS), and toxicity evaluation. When considering all the enrolled patients the ORR was 64%. At 5 years, the estimated PFS, DoR, and OS were 34, 49, and 70%, respectively. After excluding the 7 R-naïve patients, the ORR was 58%, with a PFS of 19 months. The most common grade >2 toxicities were thrombocytopenia (18%), peripheral neuropathy (13%), and neutropenia (2%). Our study shows the feasibility, long-term efficacy, and excellent tolerability of the B+R combination. We are aware that our study has specific limitations, such as the small sample size consisting of patients with a relatively good prognostic profile. However, because FL patients will be treated with subsequent chemotherapy regimens, a well-tolerated and effective chemotherapy-free therapy could be considered an additional tool for long-term disease control.

Bendamustine, Low-dose dexamethasone, and lenalidomide (BdL) for the treatment of patients with relapsed/refractory multiple myeloma confirms very promising results in a phase I/II study.

Lenalidomide and dexamethasone are an effective treatment for naïve and relapsed multiple myeloma (MM) patients. Bendamustine is a good option for B-cell malignancies showing only partial cross resistance with alkylating agents used in MM patients. Based on these considerations, we proposed a phase I/II study testing escalating doses of bendamustine and lenalidomide and fixed low doses of dexamethasone (BdL). Fifteen patients were enrolled in phase I study. Maximum tolerated dose was established at dose "level 0": bendamustine 40 mg/m2 days 1,2; lenalidomide 10 mg days 1-21; d 40 mg days 1,8,15,22 every 28-day cycle, for six cycles. We enrolled 23 patients in the phase II study. BdL combination showed mainly hematological toxicities, fever and infections. Overall response rate was 47%. After median follow up of 22 months, median PFS was 10 months. Two-years OS rate was 65%. BdL combination confirmed to be a promising treatment for patients with relapsed/refractory MM.

The histone deacetylase inhibitor romidepsin synergizes with lenalidomide and enhances tumor cell death in T-cell lymphoma cell lines.

We investigated the cytotoxic interactions of romidepsin, a histone deacetylase inhibitor, and lenalidomide, an immunomodulatory agent, in a T-cell lymphoma preclinical model. Hut-78 and Karpas-299 cells were treated with romidepsin and lenalidomide alone and in combination. The interaction between romidepsin and lenalidomide was evaluated by the Chou-Talalay method, and cell viability and clonogenicity were also evaluated. Apoptosis, reactive oxygen species (ROS) levels, and cell cycle distribution were determined by flow cytometry. ER stress, caspase activation, and the AKT, MAPK/ERK, and STAT-3 pathways were analyzed by Western blot. Combination treatment with romidepsin and lenalidomide had a synergistic effect in Hut-78 cells and an additive effect in Karpas-299 cells at 24 hours and did not decrease the viability of normal peripheral blood mononuclear cells. This drug combination induced apoptosis, increased ROS production, and activated caspase-8, -9, -3 and PARP. Apoptosis was associated with increased hallmarks of ER stress and activation of UPR sensors and was mediated by dephosphorylation of the AKT, MAPK/ERK, and STAT3 pathways.The combination of romidepsin and lenalidomide shows promise as a possible treatment for T-cell lymphoma. This work provides a basis for further studies.