Amit U. Joshi

TGF-beta signaling, tumor microenvironment and tumor progression: the butterfly effect.

Transforming growth factor-beta (TGF-beta) signals through receptor serine/threonine kinases and intracellular Smad effectors, regulating numerous epithelial cell processes. TGF-beta plays a crucial role in the cancer initiation and progression through tumor cell autonomous signaling and interactions with tumor microenvironment, but is featured with a butterfly effect upon the stages of tumorigenesis. TGF-beta signaling acts as a suppressor of epithelial cell tumorigenesis at early stages, but promotes tumor progression by enhancing migration, invasion, and survival of the tumor cells during the later stages. TGF-beta signaling also cross-talks with other cell survival signaling pathways. Tumor microenvironment contains many distinct cell types, which substantially influences the tumor cell growth and survival, and the invasion and metastasis. TGF-beta in the microenvironment, produced by cancer and/or stromal cells, is high and negatively correlates with disease progression and patient prognosis. Therefore, TGF-beta may affect tumor progression by multiple mechanisms in addition to its direct action on tumor cells, and the diversities of TGF-beta signaling in tumors imply a need for caution to TGF-beta-targeted strategies of tumor prevention and/or therapeutics.

The entangled ER-mitochondrial axis as a potential therapeutic strategy in neurodegeneration: A tangled duo unchained.

Endoplasmic reticulum (ER) and mitochondrial function have both been shown to be critical events in neurodegenerative diseases. The ER mediates protein folding, maturation, sorting as well acts as calcium storage. The unfolded protein response (UPR) is a stress response of the ER that is activated by the accumulation of misfolded proteins within the ER lumen. Although the molecular mechanisms underlying ER stress-induced apoptosis are not completely understood, increasing evidence suggests that ER and mitochondria cooperate to signal cell death. Similarly, calcium-mediated mitochondrial function and dynamics not only contribute to ATP generation and calcium buffering but are also a linchpin in mediating cell fate. Mitochondria and ER form structural and functional networks (mitochondria-associated ER membranes [MAMs]) essential to maintaining cellular homeostasis and determining cell fate under various pathophysiological conditions. Regulated Ca(2+) transfer from the ER to the mitochondria is important in maintaining control of pro-survival/pro-death pathways. In this review, we summarize the latest therapeutic strategies that target these essential organelles in the context of neurodegenerative diseases.

VCP recruitment to mitochondria causes mitophagy impairment and neurodegeneration in models of Huntington’s disease

Mutant Huntingtin (mtHtt) causes neurodegeneration in Huntingtons disease (HD) by evoking defects in the mitochondria, but the underlying mechanisms remains elusive. Our proteomic analysis identifies valosin-containing protein (VCP) as an mtHtt-binding protein on the mitochondria. Here we show that VCP is selectively translocated to the mitochondria, where it is bound to mtHtt in various HD models. Mitochondria-accumulated VCP elicits excessive mitophagy, causing neuronal cell death. Blocking mtHtt/VCP mitochondrial interaction with a peptide, HV-3, abolishes VCP translocation to the mitochondria, corrects excessive mitophagy and reduces cell death in HD mouse- and patient-derived cells and HD transgenic mouse brains. Treatment with HV-3 reduces behavioural and neuropathological phenotypes of HD in both fragment- and full-length mtHtt transgenic mice. Our findings demonstrate a causal role of mtHtt-induced VCP mitochondrial accumulation in HD pathogenesis and suggest that the peptide HV-3 might be a useful tool for developing new therapeutics to treat HD.

Plasma factor VII-activating protease antigen levels and activity are increased in ischemic stroke

Summary.  Background:  Factor VII‐activating protease (FSAP) is a recently discovered plasma protease with a role in the regulation of hemostasis and vascular remodeling processes. Higher levels and activity of FSAP have been reported in patients with deep vein thrombosis, but there are no data on plasma FSAP in ischemic stroke (IS).

Potential biomarkers to follow the progression and treatment response of Huntington's disease.

Disatnik et al. identify mitochondrial DNA levels, 8-OHdG, and inflammation factors as potential peripheral biomarkers to follow progression and treatment response of Huntington’s disease.

Murine aldo-keto reductase family 1 subfamily B: identification of AKR1B8 as an ortholog of human AKR1B10.

Abstract Aldo-keto reductase family 1 member B10 (AKR1B10), over-expressed in multiple human cancers, might be implicated in cancer development and progression via detoxifying cytotoxic carbonyls and regulating fatty acid synthesis. In the present study, we investigated the ortholog of AKR1B10 in mice, an ideal modeling organism greatly contributing to human disease investigations. In the mouse, there are three aldo-keto reductase family 1 subfamily B (AKR1B) members, i.e., AKR1B3, AKR1B7, and AKR1B8. Among them, AKR1B8 has the highest similarity to human AKR1B10 in terms of amino acid sequence, computer-modeled structures, substrate spectra and specificity, and tissue distribution. More importantly, similar to human AKR1B10, mouse AKR1B8 associates with murine acetyl-CoA carboxylase-α and mediates fatty acid synthesis in colon cancer cells. Taken together, our data suggest that murine AKR1B8 is the ortholog of human AKR1B10.

Deficiency of Factor VII activating protease alters the outcome of ischemic stroke in mice

Factor VII activating protease (FSAP) is a circulating protease with a putative role in hemostasis, remodeling and inflammation. A polymorphism giving rise to low proteolytic activity has been associated with an increased risk of stroke and carotid stenosis. To date, no in vivo studies or mechanistic information is available to explain these results. Based on the polymorphism data we hypothesize that a lack of endogenous FSAP will increase the severity of stroke. Stroke was induced by applying thrombin in the middle cerebral artery in wild‐type (WT) and FSAP−/− mice. Increased stroke volume and worsened neurological deficit were observed in FSAP−/− mice. Raised levels of FSAP protein were detected in the infarcted area of WT mice together with enhanced leukocyte infiltration and apoptosis in FSAP−/− mice. There was a concomitant increase in the activation of the NFκB pathway and decrease in expression of the PI3K/AKT pathway proteins. At a cellular level, FSAP increased cell survival and decreased apoptosis in primary cortical neurons and astrocytes exposed to tPA/NMDA excitotoxicity or oxygen glucose deprivation (OGD)/reoxygenation, respectively. This was mediated via the PI3K/AKT pathway with involvement of the protease activated receptor‐1. To corroborate the human epidemiological data, which link FSAP with stroke, we now show that the lack of FSAP in mice worsens the outcome of stroke. In the absence of FSAP there was a stronger inflammatory response and lower cell survival due to insufficient activation of the PI3K/AKT pathway.

Inhibition of Drp1/Fis1 interaction slows progression of amyotrophic lateral sclerosis

Bioenergetic failure and oxidative stress are common pathological hallmarks of amyotrophic lateral sclerosis (ALS), but whether these could be targeted effectively for novel therapeutic intervention needs to be determined. One of the reported contributors to ALS pathology is mitochondrial dysfunction associated with excessive mitochondrial fission and fragmentation, which is predominantly mediated by Drp1 hyperactivation. Here, we determined whether inhibition of excessive fission by inhibiting Drp1/Fis1 interaction affects disease progression. We observed mitochondrial excessive fragmentation and dysfunction in several familial forms of ALS patient‐derived fibroblasts as well as in cultured motor neurons expressing SOD1 mutant. In both cell models, inhibition of Drp1/Fis1 interaction by a selective peptide inhibitor, P110, led to a significant reduction in reactive oxygen species levels, and to improvement in mitochondrial structure and functions. Sustained treatment of mice expressing G93A SOD1 mutation with P110, beginning at the onset of disease symptoms at day 90, produced an improvement in motor performance and survival, suggesting that Drp1 hyperactivation may be an attractive target in the treatment of ALS patients.

Drp1/Fis1 interaction mediates mitochondrial dysfunction, bioenergetic failure and cognitive decline in Alzheimer’s disease

Mitochondrial dynamics, involving a balance between fusion and fission, regulates mitochondrial quality and number. Increasing evidence suggests that dysfunctional mitochondria play a role in Alzheimers disease (AD). We observed that Drp1 interaction with one of the adaptors, Fis1, is significantly increased in Aβ-treated neurons and AD patient-derived fibroblasts. P110, a seven-amino acid peptide, which specifically inhibits Drp1/Fis1 interaction without affecting the interaction of Drp1 with its other adaptors, attenuated Aβ42-induced mitochondrial recruitment of Drp1 and prevented mitochondrial structural and functional dysfunction in cultured neurons, in cells expressing mutant amyloid precursor protein (KM670/671NL), and in five different AD patient-derived fibroblasts. Importantly, sustained P110 treatment significantly improved behavioral deficits, and reduced Aβ accumulation, energetic failure and oxidative stress in the brain of the AD mouse model, 5XFAD. This suggests that Drp1/Fis1 interaction and excessive mitochondrial fission greatly contribute to Aβ-mediated and AD-related neuropathology and cognitive decline. Therefore, inhibiting excessive Drp1/Fis1-mediated mitochondrial fission may benefit AD patients.

Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) Protein-Protein Interaction Inhibitor Reveals a Non-catalytic Role for GAPDH Oligomerization in Cell Death

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), an important glycolytic enzyme, has a non-catalytic (thus a non-canonical) role in inducing mitochondrial elimination under oxidative stress. We recently demonstrated that phosphorylation of GAPDH by δ protein kinase C (δPKC) inhibits this GAPDH-dependent mitochondrial elimination. δPKC phosphorylation of GAPDH correlates with increased cell injury following oxidative stress, suggesting that inhibiting GAPDH phosphorylation should decrease cell injury. Using rational design, we identified pseudo-GAPDH (ψGAPDH) peptide, an inhibitor of δPKC-mediated GAPDH phosphorylation that does not inhibit the phosphorylation of other δPKC substrates. Unexpectedly, ψGAPDH decreased mitochondrial elimination and increased cardiac damage in an animal model of heart attack. Either treatment with ψGAPDH or direct phosphorylation of GAPDH by δPKC decreased GAPDH tetramerization, which corresponded to reduced GAPDH glycolytic activity in vitro and ex vivo. Taken together, our study identified the potential mechanism by which oxidative stress inhibits the protective GAPDH-mediated elimination of damaged mitochondria. Our study also identified a pharmacological tool, ψGAPDH peptide, with interesting properties. ψGAPDH peptide is an inhibitor of the interaction between δPKC and GAPDH and of the resulting phosphorylation of GAPDH by δPKC. ψGAPDH peptide is also an inhibitor of GAPDH oligomerization and thus an inhibitor of GAPDH glycolytic activity. Finally, we found that ψGAPDH peptide is an inhibitor of the elimination of damaged mitochondria. We discuss how this unique property of increasing cell damage following oxidative stress suggests a potential use for ψGAPDH peptide-based therapy.