Brendan Beaton

Human mesenchymal stromal cells deliver systemic oncolytic measles virus to treat acute lymphoblastic leukemia in the presence of humoral immunity

Clinical trials of oncolytic attenuated measles virus (MV) are ongoing, but successful systemic delivery in immune individuals remains a major challenge. We demonstrated high-titer anti-MV antibody in 16 adults with acute lymphoblastic leukemia (ALL) following treatments including numerous immunosuppressive drugs. To resolve this challenge, human bone marrow-derived mesenchymal stromal cells (BM-MSCs) were used to efficiently deliver MV in a systemic xenograft model of precursor B-lineage-ALL. BM-MSCs were successfully loaded with MV ex vivo, and MV was amplified intracellularly, without toxicity. Live cell confocal imaging demonstrated a viral hand-off between BM-MSCs and ALL targets in the presence of antibody. In a murine model of disseminated ALL, successful MV treatment (judged by bioluminescence quantification and survival) was completely abrogated by passive immunization with high-titer human anti-MV antibody. Importantly, no such abrogation was seen in immunized mice receiving MV delivered by BM-MSCs. These data support the use of BM-MSCs as cellular carriers for MV in patients with ALL.

Mouse xenograft modeling of human adult acute lymphoblastic leukemia provides mechanistic insights into adult LIC biology

The distinct nature of acute lymphoblastic leukemia (ALL) in adults, evidenced by inferior treatment outcome and different genetic landscape, mandates specific studies of disease-initiating mechanisms. In this study, we used NOD/LtSz-scid IL2Rγ null(c) (NSG) mouse xenotransplantation approaches to elucidate leukemia-initiating cell (LIC) biology in primary adult precursor B (pre-B) ALL to optimize disease modeling. In contrast with xenografting studies of pediatric ALL, we found that modification of the NSG host environment using preconditioning total body irradiation (TBI) was indispensable for efficient engraftment of adult non-t(4;11) pre-B ALL, whereas t(4;11) pre-B ALL was successfully reconstituted without this adaptation. Furthermore, TBI-based xenotransplantation of non-t(4;11) pre-B ALL enabled detection of a high frequency of LICs (<1:6900) and permitted frank leukemic engraftment from a remission sample containing drug-resistant minimal residual disease. Investigation of TBI-sensitive stromal-derived factor-1/chemokine receptor type 4 signaling revealed greater functional dependence of non-t(4;11) pre-B ALL on this niche-based interaction, providing a possible basis for the differential engraftment behavior. Thus, our studies establish the optimal conditions for experimental modeling of human adult pre-B ALL and demonstrate the critical protumorogenic role of microenvironment-derived SDF-1 in regulating adult pre-B LIC activity that may present a therapeutic opportunity.

Generation of osteoclasts from type 1 Gaucher patients and correlation with clinical and genetic features of disease

Gaucher disease (GD) is a rare autosomal recessive disorder caused by deficient activity of β-glucocerebrosidase resulting in the accumulation of glucosylceramide. Bone disease is a common feature with radiological evidence in up to 93% of patients. Severity of bone involvement ranges from osteoporosis to pathological fractures. The progressive course of type 1 GD is largely mitigated by treatment with enzyme replacement therapy (ERT) or substrate reduction. A number of studies have shown some patients suffer bone events while receiving ERT. Studies of biochemical markers of bone turnover have generated varied results and as a consequence are not generally used to assess bone disease in GD. In vitro osteoclast generation from peripheral blood samples of 74 Gaucher patients followed over a period of up to 10 years was correlated with bone events, reports of bone pain, anaemia, spleen status, bone mineral density, chitotriosidase activity, treatment with Gaucher specific therapies, bisphosphonates, mutation status and severity. Osteoclast generation, enumerated when cultured on glass, was significantly higher when differentiated from the peripheral blood of Gaucher patients which reported bone pain (116.4 ± 18.0 vs 69.0 ± 8.6, p < 0.01), had anaemia (153.7 ± 34.9 vs 78.5 ± 8.8, p < 0.01), had a splenectomy (137.6 ± 41.1 vs 60.8 ± 13.0, p < 0.05), versus those who did not. Osteoclast generation was also indicative of in vivo Gaucher specific therapy response as those naïve to therapy generated significantly more osteoclasts than those on therapy (111.2 ± 35.8 vs 45.1 ± 10.3, p < 0.05), as did patients receiving therapy but still suffering bone events (125.1 ± 31.37 vs 45.1 ± 10.33, p < 0.05). These findings demonstrate that the in vitro osteoclast assay may be a useful method for following bone disease progression in Gaucher patients.

Abundant pseudo-Gaucher cells result in delay in diagnosis of plasma cell myeloma

A 42-year-old woman was referred for management of Gaucher disease (GD). Prior to referral she gave a history of abdominal pain, fatigue and bruising. Examination revealed moderate splenomegaly. A bone marrow aspirate showed enlarged, atypical macrophages. Trephine biopsy sections showed extensive, diffuse infiltration by CD68-positive macrophages with crumpled tissue-paper appearance, reported as highly suggestive of GD. Tests performed after referral showed haemoglobin concentration 100 g/l, platelet count 85 9 10/l, normal white cells, prominent rouleaux on film, an estimated glomerular filtration rate of 42 ml/min, and immunoparesis with presence of two immunoglobulin A kappa paraproteins (49 g/l; 3 g/l). Plasma chitotriosidase was moderately elevated at 1139 nmol/h/ml (normal range [NR] <100). Leucocyte beta-glucosidase activity was elevated at 21 0 nmol/h/mg protein (NR 5 0–15 0) (not reduced as would be expected in GD) and no mutations associated with GD were detected in the GBA gene sequence. A bone marrow biopsy was repeated based on the discovery of the paraproteins. The aspirate showed frequent, large, atypical macrophages (top panels) together with 40% atypical plasma cells, some with cytoplasmic granulation (top right). The trephine biopsy specimen was hypercellular with sheets of plasma cells (up to 60% of cells) that were CD138 positive with kappa light chain restriction. Adjacent to these clustered plasma cells, sheets of atypical macrophages with fine cytoplasmic fibrils were again noted (bottom left); they were tartrate-resistant acid phosphatase (TRAP) positive (bottom right). Period acid–Schiff staining was negative. Fluorescence in situ hybridisation analysis of a CD138-enriched population showed t(4;14)(p16;q32) and loss of 11q22 (ATM gene). High-risk plasma cell myeloma was diagnosed. Pseudo-Gaucher cells in association with myeloma have been recognised in several cases since first reported by Scullin et al (1979). At the time of the first bone marrow biopsy in this patient, the pseudoGaucher cells were such a dominant feature, they obscured the correct diagnosis, resulting in diagnostic and treatment delay. This case demonstrates that chitotriosidase can be elevated in the presence of pseudo-Gaucher cells as well as in GD and emphasises the need for enzyme activity measurement and genetic analysis to make a diagnosis of GD.

Gaucher disease: risk stratification and comorbidities

ABSTRACT Introduction: Gaucher disease (GD) is characterized clinically by anemia, thrombocytopenia, hepatosplenomegaly and bone disease, with central nervous system (CNS) involvement for certain subtypes. In patients with GD type 1, which conventionally excluded those with primary CNS disease, bone complications represent a major source of morbidity. Some patients also have an increased risk of Parkinson disease (PD) and multiple myeloma (MM). There are several biomarkers, e.g., chitotriosidase and glucosylsphingosine, whose activity or concentrations have been correlated with overall disease burden. Areas covered: This paper reviews the authors experience in the diagnosis and management of patients with GD, including a review of pertinent literature. The availability of treatment, including enzyme therapy and substrate inhibition, has transformed the natural history of GD. However, there remains incomplete understanding of determinants of clinical outcome and critically, the basis of the increased propensity for PD and MM in a subset. Although biomarkers have been incorporated in the monitoring of patients, none have enabled prediction of disease course. The overarching goal of therapy is enhanced patient physical and functional well-being, achieved through the resolution of clinical problems and reduction in risks of disease-related complications. Early diagnosis and timely intervention logically enables the best prospects for the patient. The multiorgan involvement encountered in GD necessitates multidisciplinary care, ideally coordinated at centers of expertise. Expert opinion: The availability of therapeutic options has transformed the natural history of Gaucher disease, although the recognized increased risk of multiple myeloma and Parkinson disease in a subset of patients with type 1, and the devastating neurologic complications associated with type 2 and 3 disease, remain as hurdles that need to be overcome.