Department of Molecular Cytogenetics, Medical Research Institute and School of Biomedical Science, Tokyo Medical and Dental University, Tokyo, Japan.
PLoS One. 2009 Sep 29;4(9):e7099. doi: 10.1371/journal.pone.0007099.
Neuroblastoma (NB) is the most frequently occurring solid tumor in children, and shows heterogeneous clinical behavior. Favorable tumors, which are usually detected by mass screening based on increased levels of catecholamines in urine, regress spontaneously via programmed cell death (PCD) or mature through differentiation into benign ganglioneuroma (GN). In contrast, advanced-type NB tumors often grow aggressively, despite intensive chemotherapy. Understanding the molecular mechanisms of PCD during spontaneous regression in favorable NB tumors, as well as identifying genes with a pro-death role, is a matter of urgency for developing novel approaches to the treatment of advanced-type NB tumors.
We found that the expression of lysosomal associated protein multispanning transmembrane 5 (LAPTM5) was usually down-regulated due to DNA methylation in an NB cell-specific manner, but up-regulated in degenerating NB cells within locally regressing areas of favorable tumors detected by mass-screening. Experiments in vitro showed that not only a restoration of its expression but also the accumulation of LAPTM5 protein, was required to induce non-apoptotic cell death with autophagic vacuoles and lysosomal destabilization with lysosomal-membrane permeabilization (LMP) in a caspase-independent manner. While autophagy is a membrane-trafficking pathway to degrade the proteins in lysosomes, the LAPTM5-mediated lysosomal destabilization with LMP leads to an interruption of autophagic flux, resulting in the accumulation of immature autophagic vacuoles, p62/SQSTM1, and ubiqitinated proteins as substrates of autophagic degradation. In addition, ubiquitin-positive inclusion bodies appeared in degenerating NB cells.
We propose a novel molecular mechanism for PCD with the accumulation of autophagic vacuoles due to LAPTM5-mediated lysosomal destabilization. LAPTM5-induced cell death is lysosomal cell death with impaired autophagy, not cell death by autophagy, so-called autophagic cell death. Thus LAPTM5-mediated PCD is closely associated with the spontaneous regression of NBs and opens new avenues for exploring innovative clinical interventions for this tumor.
神经母细胞瘤（NB）是儿童中最常见的实体肿瘤，具有异质性的临床行为。良性肿瘤通常通过尿液中儿茶酚胺水平升高的大规模筛查来发现，它们通过程序性细胞死亡（PCD）自发消退，或者通过分化为良性神经节细胞瘤（GN）成熟。相比之下，晚期 NB 肿瘤即使经过强化化疗，也常常生长迅速。了解良性 NB 肿瘤自发消退过程中 PCD 的分子机制，以及确定具有促死亡作用的基因，对于开发治疗晚期 NB 肿瘤的新方法至关重要。
我们发现，溶酶体相关蛋白多跨膜 5（LAPTM5）的表达通常由于 DNA 甲基化而在 NB 细胞特异性下调，但在通过大规模筛查检测到的局部消退的有利肿瘤内退化的 NB 细胞中上调。体外实验表明，不仅需要恢复其表达，还需要积累 LAPTM5 蛋白，才能以不依赖半胱天冬酶的方式诱导具有自噬空泡和溶酶体膜通透性（LMP）的非凋亡性细胞死亡。虽然自噬是一种将蛋白质在溶酶体中降解的膜转运途径，但 LAPTM5 介导的溶酶体破坏导致自噬流中断，导致不成熟自噬空泡、p62/SQSTM1 和泛素化蛋白作为自噬降解底物的积累。此外，退化的 NB 细胞中出现了泛素阳性包涵体。
我们提出了一种新的分子机制，即 LAPTM5 介导的溶酶体破坏导致自噬泡积累而发生 PCD。LAPTM5 诱导的细胞死亡是由于自噬受损导致的溶酶体细胞死亡，而不是自噬细胞死亡，即所谓的自噬性细胞死亡。因此，LAPTM5 介导的 PCD 与 NB 的自发消退密切相关，为探索这种肿瘤的创新临床干预提供了新的途径。
