Phase I Study of mTORC1/2 Inhibitor Sapanisertib (CB-228/TAK-228) in Combination with Metformin in Patients with mTOR/AKT/PI3K Pathway Alterations and Advanced Solid Malignancies.

BACKGROUND: Sapanisertib (CB-228/TAK-228) is a potent, selective ATP-competitive, dual inhibitor of mTORC1/2. Metformin is thought to inhibit the mTOR pathway through upstream activation of 5'-AMP-activated protein kinase (AMPK) suggesting combination therapy may enhance antitumor activity of sapanisertib. We report preliminary safety, tolerability, and efficacy from the dose-escalation study of sapanisertib in combination with metformin in patients with advanced solid tumors. METHODS: Patients with advanced metastatic solid tumors resistant or refractory to standard treatment, with and without mTOR/AKT/PI3K pathway alterations, received sapanisertib 3 or 4 mg daily together with metformin once to three times daily (500-1,500 mg). All patients underwent 14-day titration period for metformin in cycle 1. Tumor measurements were performed following cycle 2 and subsequently every 8 weeks. RESULTS: A total of 30 patients were enrolled across four cohorts (3 mg/500 mg; 3 mg/1,000 mg, 4 mg/1,000 mg; 4 mg/1,500 mg). 19 were female (63%), median age was 57 (range: 30-77), all were Eastern Cooperative Oncology Group performance status 1. Tumor types included sarcoma (6), breast (4), ovarian (4), head and neck (3), colorectal (2), lung (2), renal cell (2), endometrial (2), gastroesophageal junction (1), prostate (1), stomach (1), urachus (1), and cervical cancer (1). Median number of prior lines of therapy was 4. Most common genomic alterations included PIK3CA (27%), PTEN (17%), AKT1/2 (10%), mTOR (10%). Of 30 patients evaluable for response, 4 patients achieved partial response (PR); 15 patients achieved stable disease (SD) as best response. Disease control rate (PR+SD) was 63%. Of the responders in PR, 3 of 4 patients had documented PTEN mutations (3/5 patients enrolled with PTEN mutations had PR); 2 of 4 of patients in PR had comutations (patient with leiomyosarcoma had both PTEN and TSC; patient with breast cancer had both PTEN and STK11); 1 of 4 patients in PR had AKT and mTOR mutation; tumor types included leiomyosarcoma (n = 2), breast (n = 1), and endometrial cancer (n = 1). Most common treatment-emergent adverse events included nausea, anorexia, diarrhea, and rash. Grade (G) 3-5 treatment-related adverse events included hyperglycemia (4/30; 13%), fatigue (2/30; 7%), hypertriglyceridemia (1/30; 3%), rash (2/20; 7%), diarrhea (2/30; 7%), creatinine increase (1/30; 3%), acidosis (1/30; 3%). No dose-limiting toxicities (DLT) were reported in the 3 mg/500 mg cohort. One of 6 patient had DLT in the 3 mg/1,000 mg cohort (G3 diarrhea) and 2 of 11 patients had DLTs in the 4 mg/1,500 mg cohort (G3 fatigue, G3 rash). 4 mg/1,000 mg was defined as the MTD. CONCLUSIONS: The safety profile of mTORC1/2 inhibitor sapanisertib in combination with metformin was generally tolerable, with antitumor activity observed in patients with advanced malignancies harboring PTEN mutations and AKT/mTOR pathway alterations. SIGNIFICANCE: Sapanisertib (CB-228/TAK-228) is a potent, selective ATP-competitive, next-generation dual inhibitor of mTORC1/2. Metformin is thought to inhibit the mTOR pathway through upstream activation of AMPK suggesting combination therapy may enhance antitumor activity of sapanisertib. This dose-escalation study of sapanisertib and metformin in advanced solid tumors and mTOR/AKT/PI3K pathway alterations, demonstrates safety, tolerability, and early clinical activity in advanced malignancies harboring PTEN mutations and AKT/mTOR pathway alterations.Clinical trial information: NCT03017833.

Phase I study of sapanisertib with carboplatin and paclitaxel in mTOR pathway altered solid malignancies.

Pre-clinically, the mTORC1/2 inhibitor sapanisertib restored sensitivity to platinums and enhanced paclitaxel-induced cancer cell killing. NCT03430882 enrolled patients with mTOR pathway aberrant tumors to receive sapanisertib, carboplatin and paclitaxel. Primary objective was safety and secondary objectives were clinical response and survival. One patient had a dose-limiting toxicity at dose level 4. There were no unanticipated toxicities. Grade 3-4 treatment-related adverse events included anemia (21%), neutropenia (21%), thrombocytopenia (10.5%), and transaminitis (5%). Of 17 patients evaluable for response, 2 and 11 patients achieved partial response and stable disease, respectively. Responders included a patient with unclassified renal cell carcinoma harboring EWSR1-POU5F1 fusion and a patient with castrate resistant prostate cancer harboring PTEN loss. Median progression free survival was 3.84 months. Sapanisertib in combination with carboplatin plus paclitaxel demonstrated a manageable safety profile, with preliminary antitumor activity observed in advanced malignancies harboring mTOR pathway alterations.

Zoning Optimization Method of a Riverfront Greenspace Service Function Oriented to the Cooling Effect: A Case Study in Shanghai.

Blue-green space commonly provides multiple ecological service functions, especially thermal environment comfort for citizens. The greenspace of the riparian buffers along 22 river channels in Shanghai was selected as the study object, and remote sensing and GIS technologies were used to obtain the quantitative composition and morphological indices of riverfront greenspace and the spatial distribution data of the land surface temperature in the study area. Through BRT modelling and statistical analyses, the interactive correlations among the three aspects, namely, the spatial patterns of riverfront greenspace, their specific functional zoning, and cooling island differentiation characteristics, were explored. The results showed that different river types served for different functional zones of the city, namely, high-density built-up zoning, new urban-growth zoning in built-up areas, suburban areas, and rural areas, and had specific regular patterns of morphosis and service function of riverfront greenspace. These also led to a significant spatial differentiation pattern of cooling intensity levels, which generally appeared in the approximate circle differentiation structure of the cooling island in the city riverfront area. The study further proposed the key factors and corresponding strategies for optimizing the greenspace pattern to strengthen the cooling intensity levels of different river types. This study summarizes the landscape composition paradigm of riverfront greenspaces at the urban mesoscale and provides adaptive planning methods for better local microclimate conditions.

Interaction of Urban Rivers and Green Space Morphology to Mitigate the Urban Heat Island Effect: Case-Based Comparative Analysis.

The spatial morphology of waterfront green spaces helps generate cooling effects to mitigate the urban heat island effect (UHI) in metropolis cities. To explore the contribution and influence of multi-dimensional spatial indices on the mitigation of UHIs, the green space of the riparian buffer along 18 river channels in Shanghai was considered as a case study. The spatial distribution data of the land surface temperature (LST) in the study area were obtained by using remote sensing images. By selecting the related spatial structure morphological factors of the waterfront green space as the quantitative description index, the growth regression tree model (BRT) was adapted to analyze the contribution of various indexes of the waterfront green space on the distribution of the LST and the marginal effect of blue-green synergistic cooling. In addition, mathematical statistical analysis and spatial analysis methods were used to study the influence of the morphological group (MG) types of riparian green spaces with different morphological characteristics on the LST. The results showed that in terms of the spatial structure variables between blue and green spaces, the contribution of river widths larger than 30 m was more notable in decreasing the LST. In the case of a larger river width, the marginal effect of synergistic cooling could be observed in farther regions. The green space that had the highest connectivity degree and was located in the leeward direction of the river exhibited the lowest LST. In terms of the spatial morphology, the fractional cover values of the vegetation (Fv) and area (A) of the green space were the main factors affecting the cooling effect of the green space. For all MG types, a large green patch that had a high green coverage and connectivity degree, as well as was distributed in the leeward direction of the river, corresponded to the lowest LST. The research presented herein can provide methods and development suggestions for optimizing spatial thermal comfort in climate adaptive cities.

Cooling Island Effect of Blue-Green Corridors: Quantitative Comparison of Morphological Impacts.

The patterns of green corridors in urban riverfront districts provide different synergistic cooling effects of blue-green space in urban areas. The purpose of this study is to quantify the spatial morphological impact of green corridors in riverfront block-scale area on the cooling effect. Three representative patterns (radiate, grid and dendritic) were selected in the study. The comprehensive influences analysis between multi-dimensional factors of spatial structure and morphology of green corridors and Ta (air temperature) distribution are processed by Envi-met4.4.5 simulation data and statistical analysis methods, such as regression tree model (BRT), were combined. The results showed that the D (distance from riverbank) has the greatest impact on the cooling effect of each belt green space. The D in the range of 600-750 m was affected by the cooling effect of blue-green space; The orientation with parallel to (southeast-northwest) or roughly the same as the prevailing wind direction (north-south) green corridors had relatively better cooling effect. When the width of green corridor was 20-25 m, the ME (marginal effect) of cooling was the largest; at 30-35 m (corridor width), the overall ME of cooling was the best; When the dPC (decreased probability connectivity, here the index was adapted to describe the connectivity degree) of green corridors was in the range of 0.5-1.5, the cooling effect of green corridor could be significantly improved. When dPC is 1.5, its marginal effect on temperature reached the maximum. The study provided a quantitative correlation technology for the morphological influence of blue-green space on the distribution of UCI (urban cooling island), which can guide the spatial layout control of green corridors in the planning and design of urban riverfront district.

Comparative Study on the Cooling Effects of Green Space Patterns in Waterfront Build-Up Blocks: An Experience from Shanghai.

Different structural patterns of waterfront green space networks in built-up areas have different synergistic cooling characteristics in cities. This study's aim is to determine what kinds of spatial structures and morphologies of waterfront green spaces offer a good cooling effect, combined with three different typical patterns in Shanghai. A multidimensional spatial influence variable system based on the cooling effect was constructed to describe the spatial structural and morphological factors of the green space network. The ENVI-met 4.3 software, developed by Michael Bruse at Bochum, German, was used to simulate the microclimate distribution data, combined with the boosted regression tree (BRT) model and the correlation analysis method. The results showed that at the network level, the distance from the water body and the connectivity of green space had a stronger cooling correlation. The orientation of green corridors consistent with a summer monsoon had larger cooling effect ranges. In terms of spatial morphology, the vegetation sky view factor (SVF) and Vegetation Surface Albedo (VSAlbedo) had an important correlation with air temperature (T), and the green corridor with a 20-25 m width had the largest marginal effect on cooling. These results will provide useful guidance for urban climate adaptive planning and design.

Dual EGFR blockade with cetuximab and erlotinib combined with anti-VEGF antibody bevacizumab in advanced solid tumors: a phase 1 dose escalation triplet combination trial.

BACKGROUND: Angiogenesis and activation of the epidermal growth factor (EGFR) pathway play an essential role in tumor proliferation and metastasis. Targeting angiogenesis or EGFR alone does not yield adequate tumor control in most solid tumors. Overcoming intrinsic and/or acquired resistance may need a doublet or triplet therapy strategy. Herein, we report the safety and feasibility of dual EGFR blockade with EGFR monoclonal antibody and EGFR tyrosine kinase inhibitor combined with anti-VEGF antibody in advanced solid tumors. METHODS: We conducted a phase I study combining erlotinib, cetuximab, and bevacizumab. Patients with advanced or metastatic solid tumors (excluding colorectal and non-small cell lung cancers) were analyzed for safety, toxicity profile, and response. Anti-tumor activity was evaluated per response evaluation criteria in solid tumors (RECIST 1.0). RESULTS: Thirty-six patients received treatment on a range of dose-levels. The most frequent tumor types enrolled were cervical (n = 10), head and neck squamous cell (n = 10), and follicular thyroid (n = 4) cancers. The most common treatment-related grade ≥ 2 adverse events were rash (56%), hypomagnesemia (17%), pruritus (11%), diarrhea (8%), and tumor-related bleeding (8%). Seventeen of 19 patients (89%) treated at the maximum tolerated dose did not present treatment-related dose-limiting toxicity. Fifteen (63%) of the 24 evaluable patients achieved a disease control (stable disease ≥ 4 months (n = 14) and partial response (n = 1). The median number of prior lines of therapies was 3 (range 1-10). CONCLUSIONS: The triplet combination of erlotinib, cetuximab, and bevacizumab was well tolerated, conferring clinical benefit in heavily pretreated patients. Future studies are warranted with second or third-generation EGFR tyrosine kinase triplet combinations in the EGFR pathway aberrant patients.Trial Registration: ClinicalTrials.gov Identifier: NCT00543504. Sponsor(s): National Cancer Institute (NCI), MD Anderson Cancer Center.

Genomic consequences of population decline in critically endangered pangolins and their demographic histories.

Pangolins are among the most critically endangered animals due to heavy poaching and worldwide trafficking. However, their demographic histories and the genomic consequences of their recent population declines remain unknown. We generated high-quality de novo reference genomes for critically endangered Malayan (Manis javanica, MJ) and Chinese (M. pentadactyla, MP) pangolins and re-sequencing population genomic data from 74 MJs and 23 MPs. We recovered the population identities of illegally traded pangolins and previously unrecognized genetic populations that should be protected as evolutionarily distinct conservation units. Demographic reconstruction suggested environmental changes have resulted in a population size fluctuation of pangolins. Additionally, recent population size declines due to human activities have resulted in an increase in inbreeding and genetic load. Deleterious mutations were enriched in genes related to cancer/diseases and cholesterol homeostasis, which may have increased their susceptibility to diseases and decreased their survival potential to adapt to environmental changes and high-cholesterol diets. This comprehensive study provides not only high-quality pangolin reference genomes, but also valuable information concerning the driving factors of long-term population size fluctuations and the genomic impact of recent population size declines due to human activities, which is essential for pangolin conservation management and global action planning.

[Effect of moxibustion at "Shenque" (CV8) and "Zusanli" (ST36) on expression of adiponectin and adiponectin receptor in Alzheimer's disease rats].

OBJECTIVE: To observe the effect of moxibustion on serum adiponectin content, and expression of adiponectin and adiponectin receptor in adipose tissue in Alzheimer's disease (AD) rats, so as to explore its mechanism underlying improvement of AD. METHODS: Fifty male SD rats were randomly divided into control, model, Shenque (CV8), Zusanli (ST36) and CV8＋ST36 groups (n＝10 in each group). The AD model was established by intraperitoneal injection of D-galactose (400 mg•kg－1•d－1) for 5 weeks and scopolamine hydrobromide (3 mg •kg－1•d－1) for 2 weeks. Moxibustion was applied to CV8, ST36 and CV8＋ST36 respectively for 3 moxa-cones every time, once daily for 5 weeks. Morris water maze tests were used to assess the rats' learning-memory ability. The contents of serum adiponectin were assayed using ELISA, and the expression of adiponectin and adiponectin receptor in the adipose tissue was detected by quantitative real-time PCR and Western blot, separately. RESULTS: Following modeling, the average escape latency of Morris water maze tests was significantly prolonged (P<0.05), the content of serum adiponectin and the expression level of adiponectin mRNA in adipose tissue were significantly decreased (P<0.01), the expression of adiponectin receptor protein significantly decreased in the model group relevant to the control group (P<0.01). After the intervention, the average escape latency was significantly shortened (P<0.05), the decreased serum adiponectin content and adiponectin mRNA expression, and the decreased adiponectin receptor protein expression in adipose tissue were all reversed in the 3 treatment groups (P<0.01，P<0.05). No significant differences were found among the three moxibustion groups in the above indexes (P>0.05). CONCLUSION: Moxibustion at CV8, ST36 and CV8＋ST36 is effective in up-regulating serum adiponectin content，adiponectin mRNA expression and adiponectin receptor protein expression in adipose tissue, which may provide evidence for clinical election of acupoints.

Microclimatic Impact Analysis of Multi-Dimensional Indicators of Streetscape Fabric in the Medium Spatial Zone.

Different historical backgrounds and planning ideas have created different urban streetscape fabrics. The patterns of the streetscape fabric have affected urban microclimate factors and formed a unique local microclimate. This paper simulated the microclimatic effects in four study areas with different streetscape fabrics in Shanghai to compare the microclimatic conditions with a system of multi-dimensional street morphological indices using ENVI-met 4.3 software. At the street network fabric level, the results showed that streets with a south⁻north orientation, a small junction spacing, and a street network with better connectivity were conducive to mitigation of the air temperature heating intensity in the street space and improving the ventilation effect; at the street-site level: The indices of Build-to-line ratio (BL), Height-width ratio (H/W), and Sky view factors (SVF) played different roles that affected the distribution characteristics of the microclimate factors. The BL value of the streets between 0.5 and 0.8 generally had a positive relationship with the air temperature. The SVF value of the streets was positively correlated with the microclimate index, while the H/W values were negatively correlated with them. The morphological indicators of different levels also had a synergistic effect on the microclimatic impact of the street space fabric. This comparative analysis of microclimatic characteristics at the medium spatial scale will provide useful suggestions for urban climate adaptability in urban spatial morphology optimization in future urbanization development.

Hyaluronic Acid Stabilized Iodine-Containing Nanoparticles with Au Nanoshell Coating for X-ray CT Imaging and Photothermal Therapy of Tumors.

In recent years, considerable efforts have been made for the development of multifunctional nanoparticles with diagnosis and therapy functions. To achieve enhanced CT imaging and photothermal therapy on the tumor, we employed iodinated nanoparticles as template to construct Au nanoshell structure and demonstrated a facile but effective approach to synthesize biocompatible and well-dispersed multifunctional nanoparticles by coating iodinated nanoparticles with Au nanoshell and subsequent surface modification by hyaluronic acid. The resultant poly(2-methacryl(3-amide-2,4,6-triiodobenzoic acid))/polyethylenimine/Au nanoshell/hyaluronic acid (PMATIB/PEI/Au nanoshell/HA) nanoparticles had relatively high X-ray attenuation coefficient and photothermal efficiency. After intravenous injection into MCF-7 tumor-bearing mice, PMATIB/PEI/Au nanoshell/HA nanoparticles were efficiently accumulated in the tumor, remarkably enhanced the tumor CT imaging, and selectively ablated the tumor through the thermal treatment of lesions under the NIR irradiation. Thus, PMATIB/PEI/Au nanoshell/HA nanoparticles displayed a great potential for CT diagnosis and CT-guided, focused photothermal tumor therapy.

Enhanced Electrochemical Oxidation of p-Nitrophenol Using Single-Walled Carbon Nanotubes/Silver Nanowires Hybrids Modified Electrodes.

The electrochemical oxidation of p-nitrophenol (p-Np) has been studied on glassy carbon electrode modified with the single-walled carbon nanotubes/silver nanowires hybrids (SWNTs-Ag) by using cyclic and differential pulse voltammetry. p-Np is irreversibly oxidized at +0.88 V (vs. the Ag/AgCl) in PBS solutions of pH 7.4. The modified electrodes display the detection sensitivity of 0.0212 µA/µM with an unusually wide linear response of 5-1700 µM (R2 = 0.998) and the detection limit of 1 µM. The current response of SWNTs-Ag modified electrode to p-Np is better than that of SWNTs or Ag nanowires modified electrode under the same concentration. Combining the adsorption ability of SWNTs and the conductivity of SWNTs and Ag nanowires, the detection performance of SWNTs-Ag modified electrode to p-Np was greatly improved.

A phase I trial of combination trastuzumab, lapatinib, and bevacizumab in patients with advanced cancer.

PURPOSE: Preclinical data indicate that combination HER2-directed and anti-VEGF therapy may bypass resistance to trastuzumab. A phase I trial was performed to assess safety, activity, and correlates. EXPERIMENTAL DESIGN: Patients with advanced, refractory malignancy were enrolled (modified 3 + 3 design with expansions for responding tumor types). Patients received lapatinib daily for 21 days, and bevacizumab and trastuzumab every 3 weeks. Correlates included HER2 extracellular domain levels (ECD) and single nucleotide polymorphisms (SNPs). RESULTS: Ninety-four patients were treated (median = four prior systemic therapies). The most common related adverse events ≥ grade 2 were diarrhea (n = 33, 35 %) and hypertension (n = 10, 11 %). The recommended phase 2 dose was trastuzumab 6 mg/m(2) (loading = 8 mg/m(2)) and bevacizumab 15 mg/kg every 3 weeks, with lapatinib 1,250 mg daily (full FDA-approved dose of each drug). One patient (1 %) achieved a complete response (CR); eight (9 %), a partial response (PR) (includes breast (n = 7, one of which was HER2 2+ by IHC) and salivary ductal carcinoma (n = 1); and 14 (15 %), stable disease (SD) ≥6 months (total SD ≥ 6 months/PR/CR =23 (25 %). All patients with PR/CR received prior trastuzumab +/- lapatinib. SD ≥ 6 months/PR/CR rate and time to treatment failure (TTF) correlated with elevated baseline HER2 ECD (N = 75 patients tested) but not with HER2 SNPs. CONCLUSIONS: Combination trastuzumab, lapatinib, and bevacizumab was well-tolerated and demonstrated antitumor activity in heavily pretreated patients with advanced malignancy.

Dual antiangiogenic inhibition: a phase I dose escalation and expansion trial targeting VEGF-A and VEGFR in patients with advanced solid tumors.

PURPOSE: Angiogenesis plays a pivotal role in tumor growth and metastasis. Sorafenib, a tyrosine kinase inhibitor of vascular endothelial growth factor receptor (VEGFR), combined with bevacizumab, a monoclonal antibody to vascular endothelial growth factor (VEGF-A), would vertically inhibit VEGF/VEGFR signaling. A phase I trial was performed to assess safety, maximum tolerated dose (MTD), and clinical correlates. EXPERIMENTAL DESIGN: Patients with advanced solid tumors refractory to standard therapy were eligible. In cohorts of escalating doses, patients received sorafenib daily for 28 days and bevacizumab every two weeks. Clinical correlates included VEGF polymorphisms. Expansion cohorts of responding tumor types were enrolled. RESULTS: One hundred fifteen patients were treated, and the MTD was identified as 200 mg twice daily sorafenib and 5 mg/kg bevacizumab every two weeks. Median number of prior therapies was four. Twenty-nine patients (25 %) achieved stable disease ≥6 months; six patients (5 %) achieved a partial response (total SD ≥ 6 months/PR=35 (30 %)). 76 patients (66 %) experienced adverse events of grade 2 or higher, most commonly hand and foot syndrome (n = 27, 24 %) and hypertension (n = 24, 21 %). Dose-limiting toxicity occurred in eight patients (7 %), and 45 patients (39 %) required dose reduction for toxicity. Grade 3 and 4 hypertension was associated with longer time to treatment failure, overall survival, and higher response rate. CONCLUSIONS: Combination sorafenib and bevacizumab was well-tolerated and demonstrated antitumor activity in heavily pretreated patients with advanced solid tumors.

Dual EGFR inhibition in combination with anti-VEGF treatment: a phase I clinical trial in non-small cell lung cancer.

BACKGROUND: Preclinical data indicate EGFR signals through both kinase-dependent and independent pathways and that combining a small-molecule EGFR inhibitor, EGFR antibody, and/or anti-angiogenic agent is synergistic in animal models. METHODS: We conducted a dose-escalation, phase I study combining erlotinib, cetuximab, and bevacizumab. The subset of patients with non-small cell lung cancer (NSCLC) was analyzed for safety and response. RESULTS: Thirty-four patients with NSCLC (median four prior therapies) received treatment on a range of dose levels. The most common treatment-related grade ≥2 adverse events were rash (n=14, 41%), hypomagnesemia (n=9, 27%), and fatigue (n=5, 15%). Seven patients (21%) achieved stable disease (SD) ≥6 months, two achieved a partial response (PR) (6%), and two achieved an unconfirmed partial response (uPR) (6%) (total=32%). We observed SD≥6 months/PR/uPR in patients who had received prior erlotinib and/or bevacizumab, those with brain metastases, smokers, and patients treated at lower dose levels. Five of 16 patients (31%) with wild-type EGFR experienced SD≥6 months or uPR. Correlation between grade of rash and rate of SD≥6 months/PR was observed (p less than 0.01). CONCLUSION: The combination of erlotinib, cetuximab, and bevacizumab was well-tolerated and demonstrated antitumor activity in heavily pretreated patients with NSCLC.

BiovaxID: a personalized therapeutic cancer vaccine for non-Hodgkin's lymphoma.

The clonal immunoglobulin molecule, idiotype (ID), expressed on the surface of B-cell malignancies can function as a tumor-specific antigen. BiovaxID is a patient-specific therapeutic cancer vaccine composed of the tumor idiotype conjugated to a carrier protein, keyhole limpet hemocyanin (KLH). In a Phase II clinical trial, administration of ID-KLH vaccine together with granulocyte-macrophage colony-stimulating factor to follicular lymphoma patients in complete remission induced tumor-specific cellular and humoral immunity and molecular remissions, and was associated with prolonged disease-free survival. A randomized, double-blind, Phase III clinical trial is ongoing to definitively determine the clinical benefit of BiovaxID plus granulocyte-macrophage colony-stimulating factor vaccination in patients with follicular lymphoma.

Establishment and characterization of a new mantle cell lymphoma cell line M-1.

A new mantle cell lymphoma cell line, M-1, was established from peripheral blood mononuclear cells of a patient with a diagnosis of blastoid variant of mantle cell lymphoma in leukemic phase. This cell line showed cell surface antigens identical to the original tumor and demonstrated the profile of a mature B-cell phenotype typical of mantle cell lymphoma: positive for CD5, CD19, CD20, sIgM and FMC7, and negative for CD3, CD10 and CD23. Cytogenetically, the M-1 cell line showed chromosomal alterations similar to the initial clinical specimen, among which a translocation t(11;14) (q13;q32) resulting in the overexpression of cyclin D1 as well as additional abnormalities involving chromosomes 3, 9 and 10. This cell line was used as a model to investigate the activity of the three drugs doxorubicin, cyclophosphamide and vincristine, commonly used in the treatment of mantle cell lymphoma patients. The effect of the drugs was evaluated by a 24 h cytotoxicity test and a 7-days anti-proliferation test using a microculture tetrazolium-based assay (MTT). Both assays indicated a higher sensitivity of the cell line to vincristine when compared to doxorubicin and cyclophosphamide. The characterization of a new mantle cell lymphoma cell line is a unique tool for studying the biology of this subtype of lymphoma for which only a few cell lines have been established.

Prognostic significance of MUC-1 expression in systemic anaplastic large cell lymphoma.

Systemic anaplastic large cell lymphoma (ALCL) frequently carries the t(2;5)(p23;q35) and overexpresses anaplastic lymphoma kinase (ALK). MUC-1, a highly glycosylated transmembrane protein, is detected in normal and malignant epithelial cells and has been associated with a poorer patient survival in various human malignancies. We have shown previously that MUC-1 is expressed as a consequence of t(1;14)(q21;32) in a subset of diffuse large B-cell lymphomas. ALCLs are known to express MUC-1, but its clinical significance is undefined. For this study, eligible patients with ALCL were HIV negative, received anthracycline-containing regimens, and had pretreatment archival tissue. Expression of MUC-1 and ALK was determined immunohistochemically after heat-induced antigen retrieval. A 10% cutoff for MUC-1 positivity was used. We identified 63 patients with systemic ALCL (22 ALK+, 41 ALK-) with a median age of 47 years, and 41 were male. MUC-1 was detected in 16 of 22 (73%) ALK-positive and 20 of 41 (49%) ALK-negative ALCL (P = 0.06, chi(2) test). MUC-1 expression was not associated with apoptotic rate as detected by terminal deoxynucleotidyl transferase-mediated nick end labeling assay or proliferation index as evaluated by MIB-1 antibody. For 48 patients with ALCL (16 ALK+, 32 ALK-) and complete clinical follow-up, 5-year progression-free survival (PFS) was 39.7% for patients with MUC-1-positive tumors versus 75.2% (P = 0.027 by Log-rank) for patients with MUC-1-negative tumors. For the ALK-negative ALCL group of 32 patients, the 5-year PFS was 26 versus 70.8% for patients with MUC-1-positive versus MUC-1-negative tumors (P = 0.0096 by Log-rank). For the ALK-positive ALCL group of 16 patients, the 5-year PFS was 52 versus 100% for patients with MUC-1-positive versus MUC-1-negative tumors (P, not significant). In summary, MUC-1 is frequently expressed in systemic ALCL, and its expression is associated with significantly inferior outcome in patients untreated previously with ALK-negative tumors. Future studies should explore the underlying molecular mechanisms of MUC-1 expression in these tumors and its role as a target for novel therapeutic strategies.

Quantitative real-time polymerase chain reaction for detection of circulating cells with t(14;18) in volunteer blood donors and patients with follicular lymphoma.

The chromosomal rearrangement t(14;18)(q32;21) involves the major (MBR) or minor (mcr) breakpoint cluster regions and the immunoglobulin heavy chain joining regions (JH) in most follicular lymphomas. As a first step towards determining the clinical significance of circulating cells with t(14;18), we detected and quantitated circulating cells in samples obtained from volunteer blood donors and follicular lymphoma patients. The t(14;18) was co-amplified with beta-actin with real-time quantitative PCR (QRT-PCR) in reactions containing 1 microg of DNA from peripheral blood or bone marrow aspirates. The cell number was quantitated using linear regression and an external standard of serially diluted DNA from cell lines with MBR/JH or mcr/JH rearrangements. At dilutions of 10(5) and 106, sensitivity was 100 and 55% for MBR/JH, and 100 and 10% for mcr/JH rearrangements. Among 102 volunteer blood donors MBR/JH vs. mcr/JH amplicons were detected in 22 vs. 4% with duplicate 1 microg DNA reactions, and in 41 vs. 6% with a total 10 microg DNA analyzed in multiple reactions. Among volunteer blood donors the mean number of circulating cells with MBR/JH vs. mcr/JH rearrangements were 0.8 vs. 0.1/microg DNA, and exceeded the upper normal limit (defined as the mean of all volunteer samples plus two standard deviations) in 3% vs. 2%, respectively. Analysis for MBR/JH rearrangements revealed that follicular lymphoma patients vs. volunteer blood donors were positive in 76% vs. 22% (p = 0.008 by Fisher's exact test); that the mean number of MBR/JH cells per microg of DNA was 91 vs. 0.8 (p = 0.0002 by Mann-Whitney test); and the number of the MBR/JH cells exceeded the upper normal limit in 32% vs. 3% of subjects (p = 0.0001 by Fisher's exact test). Circulating cells with mcr/JH were not detected among any of these 25 lymphoma patients. We conclude that patients with follicular lymphoma are more frequently positive, have higher numbers of circulating cells with t(14;18), which exceed upper normal limit more frequently than in volunteer blood donors.