Citations

ABSTRACT Vesicular stomatitis virus (VSV) has long been regarded as a promising recombinant vaccine platform and oncolytic agent but has not yet been tested in humans because it causes encephalomyelitis in rodents and primates. Recent studies have shown that specific tropisms of several viruses could be eliminated by engineering microRNA target sequences into their genomes, thereby inhibiting spread in tissues expressing cognate microRNAs. We therefore sought to determine whether microRNA targets could be engineered into VSV to ameliorate its neuropathogenicity. Using a panel of recombinant VSVs incorporating microRNA target sequences corresponding to neuron-specific or control microRNAs (in forward and reverse orientations), we tested viral replication kinetics in cell lines treated with microRNA mimics, neurotoxicity after direct intracerebral inoculation in mice, and antitumor efficacy. Compared to picornaviruses and adenoviruses, the engineered VSVs were relatively resistant to microRNA-mediated inhibition, but neurotoxicity could nevertheless be ameliorated significantly using this approach, without compromise to antitumor efficacy. Neurotoxicity was most profoundly reduced in a virus carrying four tandem copies of a neuronal mir125 target sequence inserted in the 3′-untranslated region of the viral polymerase (L) gene.

Oncolytic virotherapy makes use of the natural ability of viruses to infect and kill cancer cells. Adenovirus serotype 5 (Ad5) has been approved for use in humans as a therapy for solid cancers. In this study, we have tested whether Ad5 and low-seroprevalence adenoviruses can be used as oncolytics for multiple myeloma (MM). We show that Ad5 productively infects most myeloma cell lines, replicates to various degrees, and mediates oncolytic cell killing in vitro and in vivo. Comparison of Ad5 with low-seroprevalence Ads on primary marrow samples from MM patients revealed striking differences in the abilities of different adenoviral serotypes to kill normal CD138 – cells and CD138 + MM cells. Ad5 and Ad6 from species C and Ad26 and Ad48 from species D all mediated killing of CD138 + cells with low-level killing of CD138 – cells. In contrast, Ad11, Ad35, Ad40, and Ad41 mediated weak oncolytic effects in all of the cells. Comparison of cell binding, cell entry, and replication revealed that Ad11 and Ad35 bound MM cells 10 to 100 times better than other serotypes. However, after this efficient interaction, Ad11 and Ad35 viral DNA was not replicated and cell killing did not occur. In contrast, Ad5, Ad6, Ad26, and Ad48 all replicated 10- to 100-fold in MM cells and this correlated with cell killing. These data suggest that Ad5 and other low-seroprevalence adenoviruses may have utility as oncolytic agents against MM and other hematologic malignancies.

Purpose Thalidomide and lenalidomide are immunomodulatory drugs (IMiDs) that produce high remission rates in the treatment of multiple myeloma. Pomalidomide is a new IMiD with high in vitro potency. We report, to our knowledge, the first phase II trial of pomalidomide administered in combination with low-dose dexamethasone for the treatment of relapsed or refractory multiple myeloma. Patients and Methods Pomalidomide was administered orally at a dose of 2 mg daily on days 1 through 28 of a 28-day cycle. Dexamethasone 40 mg daily was administered orally on days 1, 8, 15, and 22 of each cycle. Responses were recorded using the criteria of the International Myeloma Working Group. Results Sixty patients were enrolled. Thirty-eight patients (63%) achieved confirmed response including complete response in three patients (5%), very good partial response in 17 patients (28%), and partial response in 18 patients (30%). Responses were seen in 40% of lenalidomide-refractory patients, 37% of thalidomide-refractory patients, and 60% of bortezomib-refractory patients. Responses were seen in 74% of patients with high-risk cytogenetic or molecular markers. Toxicity consisted primarily of myelosuppression. Grade 3 or 4 hematologic toxicity consisted of anemia (5%), thrombocytopenia (3%), and neutropenia (32%). One patient (1.6%) had a thromboembolic event. The median progression-free survival time was 11.6 months and was not significantly different in patients with high-risk disease compared with patients with standard-risk disease. Conclusion The combination of pomalidomide and low-dose dexamethasone is extremely active in the treatment of relapsed multiple myeloma, including high response rates in patients refractory to other novel agents.

AbstractUp to now, no lentiviral vector (LV) tool existed to govern efficient and stable gene delivery into quiescent B lymphocytes, which hampers its application in gene therapy and immunotherapy areas. Here, we report that LVs incorporating measles virus (MV) glycoproteins, H and F, on their surface allowed transduction of 50% of quiescent B cells, which are not permissive to VSVG-LV transduction. This high transduction level correlated with B-cell SLAM expression and was not at cost of cell-cycle entry or B-cell activation. Moreover, the naive and memory phenotypes of transduced resting B cells were maintained. Importantly, H/F-LVs represent the first tool permitting stable transduction of leukemic cancer cells, B-cell chronic lymphocytic leukemia cells, blocked in G0/G1 early phase of the cell cycle. Thus, H/F-LV transduction overcomes the limitations of current LVs by making B cell–based gene therapy and immunotherapy applications feasible. These new LVs will facilitate antibody production and the study of gene functions in these healthy and cancer immune cells.

ABSTRACT Mutations that negatively or positively affect the fusion properties of murine leukemia viruses (MLVs) have been found within all subdomains of their SU (surface) and TM (transmembrane) envelope units. Yet, the interrelations between these different regions of the envelope complex during the cell entry process are still elusive. Deletion of the histidine residue of the conserved PHQV motif at the amino terminus of the amphotropic or the ecotropic MLV SU resulted in the AdelH or the MOdelH fusion-defective mutant envelope, respectively. These delH mutant envelopes are incorporated on retroviral particles at normal densities and normally mediate virion binding to cells expressing the retroviral receptors. However, both their cell-cell and virus-cell fusogenicities were fully prevented at an early postbinding stage. We show here that the fusion defect of AdelH or MOdelH envelopes was also almost completely reverted by providing either soluble SU or a polypeptide encompassing the receptor-binding domain (RBD) to the target cells, provided that the integrity of the amino-terminal end of either polypeptide was preserved. Restoration of delH envelope fusogenicity was caused by activation of the target cells via specific interaction of the latter polypeptides with the retrovirus receptor rather than by their association with the delH envelope complexes. Moreover crossactivation of the target cells, leading to fusion activation of AdelH or MOdelH envelopes, was achieved by polypeptides containing various type C mammalian retrovirus RBDs, irrespective of the type of entry-defective glycoprotein that was used for infection. Our results indicate that although they recognize different receptors for binding to the cell surface, type C mammalian retroviruses use a common entry pathway which is activated by a conserved feature of their envelope glycoproteins.

Object The authors describe their experience in a series of cases of intraneural ganglia within the hip and pelvic regions, and explain the mechanism of formation and propagation of this pathological entity. Methods Five patients with 6 intraneural ganglia are presented. Four patients presented with symptomatic intraneural ganglia in the buttock and pelvis affecting the sciatic and lumbosacral plexus elements. An asymptomatic cyst affecting the opposite sciatic nerve was found on MR imaging in 1 patient. The fifth patient, previously reported on by another group, had an obturator intraneural ganglion that the authors reinterpreted. Results All 5 intraneural ganglia affecting the sciatic and lumbosacral plexus elements were found to have a joint connection to the posteromedial aspect of the hip joint; the obturator intraneural cyst had a joint connection to the anteromedial aspect of the hip joint. In all cases, initial review of the MR images led to their misinterpretation. Conclusions To the authors' knowledge, these are the first cases of intraneural ganglia demonstrated to have a connection to the hip joint. This finding at a rare site provides further evidence for the unifying articular (synovial) theory for the formation of intraneural ganglia and reveals a shared mechanism for their propagation. Furthermore, understanding the pathogenesis of these lesions provides insight into their successful treatment and their recurrence.

Abstract The outcome of patients with multiple myeloma is dictated primarily by cytogenetic abnormalities and proliferative capacity of plasma cells. We studied the outcome after initial therapy with lenalidomide-dexamethasone among 100 newly diagnosed patients, risk-stratified by genetic abnormalities and plasma cell labeling index. A total of 16% had high-risk multiple myeloma, defined by the presence of hypodiploidy, del(13q) by metaphase cytogenetics, del(17p), IgH translocations [t(4;14), or t(14;16)] or plasma cell labeling index more than or equal to 3%. Response rates were 81% vs 89% in the high-risk and standard-risk groups, respectively. The median progression-free survival was shorter in the high-risk group (18.5 vs 36.5 months, P < .001), but overall survival was comparable. Because of unavailability of all tests for every patient, we separately analyzed 55 stringently classified patients, and the results were similar. In conclusion, high-risk patients achieve less durable responses with lenalidomide-dexamethasone compared with standard-risk patients; no significant differences in overall survival are apparent so far. These results need confirmation in larger, prospectively designed studies.

The applicability of cytoreductive treatment of malignant diseases using recombinant viruses strongly depends on specific recognition of surface receptors to target exclusively neoplastic cells. A recently generated monoclonal antibody (mAb), Wue-1, specifically detects CD138+ multiple myeloma (MM) cells. In this study, a haemagglutinin (H) protein that was receptor-blinded (i.e. did not bind to CD46 and CD150) was genetically re-engineered by fusing it to a single-chain antibody fragment (scFv) derived from the Wue-1 mAb open reading frame (scFv-Wue), resulting in the recombinant retargeted measles virus (MV)-Wue. MV-Wue efficiently targeted and fully replicated in primary MM cells, reaching titres similar to those seen with non-retargeted viruses. In agreement with its altered receptor specificity, infection of target cells was no longer dependent on CD150 or CD46, but was restricted to cells that had been labelled with Wue-1 mAb. Importantly, infection with MV-Wue rapidly induced apoptosis in CD138+ malignant plasma cell targets. MV-Wue is the first fully retargeted MV using the restricted interaction between Wue-1 mAb and primary MM cells specifically to infect, replicate in and deplete malignant plasma cells.

Abstract Oncolytic measles virus (MV) induces cell fusion and cytotoxicity in a CD46-dependent manner. Development of fully retargeted oncolytic MVs would improve tumor selectivity. The urokinase-type plasminogen activator receptor (uPAR) is a tumor and stromal target overexpressed in multiple malignancies. MV-H glycoproteins fully retargeted to either human or murine uPAR were engineered and their fusogenic activity was determined. Recombinant human (MV-h-uPA) and murine (MV-m-uPA) uPAR–retargeted MVs expressing enhanced green fluorescent protein (eGFP) were rescued and characterized. Viral expression of chimeric MV-H was shown by reverse transcription-PCR and Western blot. In vitro viral replication was comparable to MV-GFP control. The receptor and species specificity of MV-uPAs was shown in human and murine cells with different levels of uPAR expression. Removal of the NH2-terminal fragment ligand from MV-uPA by factor X(a) treatment ablated the MV-uPA functional activity. Cytotoxicity was shown in uPAR-expressing human and murine cells. MV-h-uPA efficiently infected human endothelial cells and capillary tubes in vitro. I.v. administration of MV-h-uPA delayed tumor growth and prolonged survival in the MDA-MB-231 breast cancer xenograft model. Viral tumor targeting was confirmed by immunohistochemistry. MV-m-uPA transduced murine mammary tumors (4T1) in vivo after intratumor administration. MV-m-uPA targeted murine tumor vasculature after systemic administration, as shown by dual (CD31 and MV-N) staining of tumor capillaries in the MDA-MB-231 model. In conclusion, MV-uPA is a novel oncolytic MV associated with potent and specific antitumor effects and tumor vascular targeting. This is the first retargeted oncolytic MV able to replicate in murine cells and target tumor vasculature in a uPAR-dependent manner. [Cancer Res 2009;69(4):1459–68]