Calpain is a major cell death effector in selective striatal degeneration induced in vivoin\ vivo by 3-nitropropionate: implications for Huntington's disease.

International audienceStriatal cell death in Huntington's Disease (HD) may involve mitochondrial defects, NMDA-mediated excitotoxicity, and activation of death effector proteases such as caspases and calpain. However, the precise contribution of mitochondrial defects in the activation of these proteases in HD is unknown. Here, we addressed this question by studying the mechanism of striatal cell death in rat models of HD using the mitochondrial complex II inhibitor 3-nitropropionic acid (3-NP). The neurotoxin was either given by intraperitoneal injections (acute model) or over 5 d by constant systemic infusion using osmotic pumps (chronic model) to produce either transient or sustained mitochondrial deficits. Caspase-9 activation preceded neurodegeneration in both cases. However, caspase-8 and caspase-3 were activated in the acute model, but not in the chronic model, showing that 3-NP does not require activation of these caspases to produce striatal degeneration. In contrast, activation of calpain was specifically detected in the striatum in both models and this was associated with a calpain-dependent cleavage of huntingtin. Finally, in the chronic model, which mimics a steady blockade of complex II activity reminiscent of HD, selective calpain inhibition prevented the abnormal calpain-dependent processing of huntingtin, reduced the size of the striatal lesions, and almost completely abolished the 3-NP-induced DNA fragmentation in striatal cells. The present results demonstrate that calpain is a predominant effector of striatal cell death associated with mitochondrial defects $in\ vivo$. This suggests that calpain may play an important role in HD pathogenesis and could be a potential therapeutic target to slow disease progression

Corticostriatopallidal Neuroprotection by Adenovirus-Mediated Ciliary Neurotrophic Factor Gene Transfer in a Rat Model of Progressive Striatal Degeneration

International audienceCiliary neurotrophic factor (CNTF) is a potent protective factor for striatal neurons in animal models of Huntington's disease (HD). Clinical application of this potential therapeutic still requires the design and optimization of delivery systems. In the case of HD, spatial spread in the vast volume occupied by the striatum and long-term delivery of the factor are particular challenges for these systems. We explored the potential of adenovirus-mediated gene transfer to fulfill these requirements by studying the functional and anatomical effects of single-site striatal delivery of CNTF recombinant vectors in a rat model of HD. In an initial series of experiments, unilateral injections of CNTF adenovirus were performed in rats 10, 30, or 90 d before a 5 d neurotoxic treatment with systemic 3-nitropropionic acid (3NP). Preservation of striatal neurons was observed at all time points, demonstrating temporally extended neuroprotective effects of the CNTF adenovirus. In a second series of experiments , bilateral injections of CNTF adenovirus were performed in the medial aspect of the striatum 10 d before starting 3NP intoxication. Despite placement of the CNTF-producing vector outside the lateral striatal area susceptible to lesion, massive protection of corticostriatopallidal circuits was observed, associated with significant behavioral benefits. This spatial spread of neuroprotection is discussed with reference to the retrograde transport of the adenovirus vector and the anterograde transport of the transgenic CNTF. Overall, adenovirus-mediated CNTF gene transfer appears to be a potentially useful delivery system for widespread, long-term circuit neuroprotection in HD patients

Therapeutic Vaccination with TNF-Kinoid in TNF Antagonist-Resistant Rheumatoid Arthritis: A Phase II Randomized, Controlled Clinical Trial.

Active immunization, or vaccination, with tumor necrosis factor (TNF)-Kinoid (TNF-K) is a novel approach to induce polyclonal anti-TNF antibodies in immune-mediated inflammatory diseases. This study was performed to transfer the proof of concept obtained in mice model of rheumatoid arthritis (RA) into human. We designed a pilot study to demonstrate the feasibility of therapeutic vaccination in RA

Study design.

<p>In the first stage, 8 patients were randomized 3∶1 to receive 90 µg TNF-K or placebo, in the second, 16 patients were randomized 3∶1 to receive 180 µg TNF-K or placebo, and in the third, 17 patients were randomized 3∶1 to receive 360 µg TNF-K or placebo. In each stage, patients were also randomized 1∶1 to receive 2 doses (day 0, 28) or 3 doses (day 0, 7, 28) (arrows). For stages 1 and 2, after 3 patients had been enrolled and no safety issues had been reported for at least 7 days, enrolment in the subsequent stage started in parallel. One patient randomized to receive 3 doses of 360 µg TNF-K withdrew consent prior to treatment. The principal analysis portion of the study continued up to day 84, and the follow-up portion continued up to month 12.</p

Humoral immune response to TNF.

<p>Patients were treated with 2 doses (days 0 and 28) or 3 doses (days 0, 7, and 28) of placebo or 90, 180, or 360 µg TNF-K. Anti-TNF antibody titers were determined by enzyme-linked immunosorbent assay. (A) GMTs. (B) Percent of patients in each treatment group with detectable anti-TNF antibodies (titer ≥200) up to month 3 (at study day 38, 56, or 84), at month 6, at month 12, or at any time up to month 12 (i.e., antibody responders).</p

Demographics of treated patients at baseline (day 0).

<p>Abbreviations: SD, standard deviation.</p><p>Demographics of treated patients at baseline (day 0).</p

T cell response to TNF.

<p>Peripheral blood mononuclear cells were collected on days 0 and 56 and treated in vitro with medium (control) or with 10 µg/mL TNF-K, TNF, or KLH. Lymphoproliferation was assessed after 72 h by ³H-thymidine incorporation. Shown is the stimulation index (fold-increase in lymphoproliferation vs. control) for cells from patients treated with placebo (n = 6) or TNF-K (n = 10).</p

Difference from baseline for clinical assessments in anti-TNF antibody responders and non-responders.

<p>Post-hoc analyses: Mean changes in clinical assessments from baseline are shown at months 3, 6, and 12 for antibody responders (detectable anti-TNF antibodies at any time; gray bars) and for non-responders (white bars). (A) DAS28-CRP. (B) CRP. (C) Tender joints count. (D) Swollen joints count. (E) Patient’s global activity score. (F) Physician’s global activity score. (G) Patient assessment of pain. (H) Change in HAQ disability/function score. Error bars indicate standard error of the mean. P-values were determined by Wilcoxon rank-sum test. NS, not significant (p≥0.05). In responders, n = 19 at month 3, 16 at months 6 and 12 except at month 3 for Physician GAS n = 18 and for CRP n = 20. In non-responders, n = 17 at month 3 and 15 at months 6 and 12 except at month 6 n = 14 for CRP and DAS 28 score.</p