Normal hemostasis but defective hematopoietic response to growth factors in mice deficient in phospholipid scramblase 1

Phospholipid scramblase 1 (PLSCR1) is an endofacial plasma membrane protein proposed to participate in transbilayer movement of phosphatidylserine and other phospholipids. In addition to its putative role in the reorganization of plasma membrane phospholipids, PLSCR1 is a substrate of intracellular kinases that imply its possible participation in diverse signaling pathways underlying proliferation, differentiation, or apoptosis. Because PLSCR1 is prominently expressed in a variety of blood cells, we evaluated PLSCR activity in platelets and erythrocytes, and cytokine-dependent growth of hematopoietic precursor cells, of PLSCR1 knock-out mice. Adult PLSCR1(-/-) mice showed no obvious hematologic or hemostatic abnormality, and blood cells from these animals normally mobilized phosphatidylserine to the cell surface upon stimulation. Whereas blood cell counts in adult PLSCR1(-/-) mice were normal, in both fetus and newborn animals neutrophil counts were significantly depressed relative to age-matched wild type (WT). Furthermore, when compared with WT, hematopoietic precursor cells from PLSCR1(-/-) mice showed defective colony formation and impaired differentiation to mature granulocytes as stimulated by stem cell factor and granulocyte colony-stimulating factor (G-CSF). By contrast, PLSCR1(-/-) cells showed normal colony formation stimulated by interleukin-3 or granulocyte-macrophage CSF, and expansion of megakaryocytic and erythroid progenitors by thrombopoietin or erythropoietin was unaffected. Stem cell factor and G-CSF were also found to induce marked increases in PLSCR1 levels in WT cells. Consistent with in vitro assays, PLSCR1(-/-) mice treated with G-CSF showed less than 50% of the granulocytosis observed in identically treated WT mice. These data provide direct evidence that PLSCR1 functionally contributes to cytokine-regulated cell proliferation and differentiation and suggest it is required for normal myelopoiesis.     

Liver-directed gene therapy corrects cardiovascular lesions in feline mucopolysaccharidosis type I

Patients with mucopolysaccharidosis type I (MPS I), a genetic deficiency of the lysosomal enzyme α-l-iduronidase (IDUA), exhibit accumulation of glycosaminoglycans in tissues, with resulting diverse clinical manifestations including neurological, ocular, skeletal, and cardiac disease. MPS I is currently treated with hematopoietic stem cell transplantation or weekly enzyme infusions, but these therapies have significant drawbacks for patient safety and quality of life and do not effectively address some of the most critical clinical sequelae, such as life-threatening cardiac valve involvement. Using the naturally occurring feline model of MPS I, we tested liver-directed gene therapy as a means of achieving long-term systemic IDUA reconstitution. We treated four MPS I cats at 3–5 mo of age with an adeno-associated virus serotype 8 vector expressing feline IDUA from a liver-specific promoter. We observed sustained serum enzyme activity for 6 mo at ∼30% of normal levels in one animal, and in excess of normal levels in three animals. Remarkably, treated animals not only demonstrated reductions in glycosaminoglycan storage in most tissues, but most also exhibited complete resolution of aortic valve lesions, an effect that has not been previously observed in this animal model or in MPS I patients treated with current therapies. These data point to clinically meaningful benefits of the robust enzyme expression achieved with hepatic gene transfer that extend beyond the economic and quality of life advantages over lifelong enzyme infusions.Mucopolysaccharidosis type I (MPS I) is a recessive genetic disorder caused by deficiency of the lysosomal enzyme α-l-iduronidase (IDUA). In the absence of IDUA, cells are unable to catabolize the ubiquitous glycosaminoglycans (GAGs) heparan and dermatan sulfates. The resulting lysosomal GAG storage causes multisystem organ pathology and diverse clinical manifestations, including bone and joint deformity, upper airway obstruction, hepatosplenomegaly, corneal clouding, and cognitive impairment (1). Most patients also develop cardiac disease, which arises from the combined effects of GAG deposition in the myocardium, coronary arteries, and left-sided heart valves (2). Without treatment, median survival in patients with the severe form of the disease is less than 7 y (3).Care of MPS I patients has been vastly improved by the introduction of two disease-modifying therapies—hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy (ERT). Both treatments are based on the principle of cross-correction: that cells can efficiently endocytose extracellular lysosomal enzymes bearing a mannose-6-phosphate residue, allowing IDUA secreted from donor-derived cells after HSCT or recombinant enzyme delivered i.v. to correct the metabolic defect in many tissues (4, 5). The introduction of HSCT has increased the survival of MPS I patients and has demonstrated improvements in growth, mobility, hepatosplenomegaly, and some aspects of cardiac disease such as left ventricular hypertrophy (2, 6–9). ERT has shown a similar capacity to improve many of the clinical features of MPS I (7, 10, 11). ERT is favored in patients with an attenuated disease phenotype because of the high mortality associated with HSCT, although HSCT remains the first-line intervention for patients less than 2 y of age owing to the beneficial effect of early transplantation on cognitive outcomes (7).Despite the enormous advances that have been made in the treatment of MPS I, significant shortcomings remain. Neurological symptoms do not improve with ERT and are highly variable after HSCT (7, 9). Skeletal disease is incompletely treated by both therapies. ERT and HSCT may improve heart disease but do not reverse valvular GAG deposition, often leaving treated patients with persistent aortic and mitral valve insufficiency or stenosis (2, 12). Additionally, the current treatment options are fundamentally limited by the morbidity and mortality associated with HSCT and the need for lifelong, expensive, weekly enzyme infusions in ERT.For diseases such as MPS I that require lifelong systemic enzyme replacement, liver-directed gene therapy has emerged as a potential therapeutic option. The high synthetic capacity of the liver, coupled with the discovery of adeno-associated virus (AAV) vectors capable of safe and efficient hepatic targeting, make this a feasible alternative to exogenous enzyme infusion (13, 14). The first clinical success of AAV-mediated liver gene therapy was recently demonstrated in a trial for hemophilia B, in which some patients were able to discontinue prophylactic factor IX injections (15). Apart from the potential safety and quality of life benefits over HSCT and ERT, respectively, we hypothesized that liver-directed gene therapy could have three potential benefits specific to MPS I. First, the liver is a therapeutic target in MPS I, making the high local concentrations of enzyme potentially useful for efficiently treating hepatomegaly due to GAG storage. Second, liver-mediated expression could theoretically result in circulating concentrations of IDUA higher than those achieved with HSCT, and more stable than those achieved with i.v. infusion of the enzyme, which has a serum half-life of less than 4 h (11). Maintaining high levels of serum IDUA could drive greater enzyme uptake and improve efficacy in difficult-to-treat tissues. Finally, antibody responses to IDUA, which develop in the vast majority of patients receiving ERT, seem to limit treatment efficacy (16, 17). Evidence from mouse models suggests that AAV-mediated hepatic expression of an enzyme is less immunogenic than i.v. delivery of the recombinant protein, indicating that this approach could exhibit improved efficacy by eliciting less-robust immune responses to IDUA (18).In the present study we tested liver-directed gene therapy in the naturally occurring feline model of MPS I, which recapitulates many of the clinical and pathological features of the disease, including progressive cardiac valve involvement (19–23). Four animals were treated at 3–5 mo of age with an i.v. injection of an AAV serotype 8 vector expressing feline IDUA from a liver-specific promoter. Three of the animals exhibited sustained supraphysiologic IDUA expression, with subsequent GAG clearance from all tissues examined. Remarkably, aortic valve lesions were reversed in these three animals, indicating the potential utility of this approach for targeting treatment refractory tissues in MPS I.     

Outcome and determinants of failure to complete primary R-CHOP treatment for reasons other than non-response among patients with diffuse large B-cell lymphoma

Patients with diffuse large B-cell lymphoma (DLBCL) who fail to complete planned treatment with R-CHOP due to toxicity are sparsely described. We investigated the extent of failure to complete treatment (six cycles or more, or three cycles + RT for patients with stage I disease) with R-CHOP for reasons unrelated to non-response, the determinants of such failure and the outcome among these patients. Three thousand one hundred and forty nine adult DLBCL patients who started primary treatment with R-CHOP were identified through the Swedish lymphoma register 2007-2014. Of these, 147 (5%) stopped prematurely after 1-3 cycles of R-CHOP for reasons unrelated to non-response, 168 (5%) after 4-5 cycles and 2639 patients (84%) completed planned treatment. Additionally, 195 (6%) patients did not complete treatment due to non-response or death before treatment end. In a multivariable logistic regression model, age > 75 years, poor performance status, extranodal disease and Charlson Comorbidity Index ≥1 were significantly associated with failure to complete planned R-CHOP treatment for other reasons than non-response. Non-completion of treatment strongly correlated with survival. Five-year overall survival for patients who received 1-3 cycles was 26% (95% CI: 19%-33%), 49% (95% CI: 41%-57%) for 4-5 cycles and 76% (74%-77%) for patients who completed treatment. Failure to complete planned R-CHOP treatment is an important clinical issue associated with inferior survival. Old age and poor performance status most strongly predict such failure. These results indicate a need for improved treatment tailoring for patients with certain baseline demographics to improve tolerability and chance for treatment completion.     

Eculizumab treatment for rescue of renal function in IgA nephropathy

Background

Immunoglobulin A (IgA) nephropathy is a chronic glomerulonephritis with excessive glomerular deposition of IgA1, C3 and C5b-9, which may lead to renal failure.

Case Diagnosis/Treatment

We describe the clinical course of an adolescent with rapidly progressive disease leading to renal failure in spite of immunosuppressive treatment. Due to refractory disease the patient was treated with eculizumab (anti-C5) for 3 months in an attempt to rescue renal function. Treatment led to clinical improvement with stabilization of the glomerular filtration rate and reduced proteinuria. Discontinuation of treatment led to a rapid deterioration of renal function. This was followed by a single dose of eculizumab, which again reduced creatinine levels temporarily.

Conclusions

Early initiation of eculizumab therapy in patients with progressive IgA nephropathy may have a beneficial effect by blocking complement-mediated renal inflammation.     

An empirical typology of seriously and persistently mentally ill patients using symptom and social functioning factors

This study generated an empirical, statistically based typology that used both symptom data and social functioning dimensions to define subtypes of seriously mentally ill patients. The intent of using social functioning information was to explore the degree to which it improves clinicians' understanding of and treatment planning for diverse subgroups of patients. Social functioning dimensions and symptom data collected on a group of SPMI (seriously and persistently mentally ill) patients were factor analyzed and then submitted to a cluster analysis that yielded five meaningful patient subtypes. Demonstrations of the usefulness of this classification included significant subgroup differences on clinically important external variables, including needs for specific types of treatment and service consumption. For comparison purposes, the failure of the DSM-III diagnostic classification to distinguish patients on the clinically relevant criteria also was demonstrated.