Herein, we show that an enzymatic reaction can generate peptide assemblies that sequestrate proteins to selectively kill cancer cells. A phosphopeptide bearing the antagonistic motif (AVPI) to the ...inhibitors of apoptotic proteins (IAPs) enters cancer cells and normal cells by caveolin‐dependent endocytosis and macropinocytosis, respectively. The AVPI‐bearing peptide assemblies sequestrates IAPs and releases bortezomib (BTZ), a proteasome inhibitor, in the cytosol of cancer cells, but rescues the normal cells (namely, HS‐5 cells) by trafficking the BTZ into lysosomes. Alkaline phosphatase (ALP) acts as a context‐dependent signal for trafficking the peptide/BTZ assemblies and selectively induces the death of the cancer cells. The assemblies of AVPI exhibit enhanced proteolytic resistance. This work, which utilizes the difference in endocytic uptake of enzymatically formed peptide assemblies to selectively kill cancer cells, promises a new way to develop selective cancer therapeutics.
The enzymatic self‐assembly: A proapoptotic‐peptide–bortezomib (BTZ) assembly enters cancer cells by caveolin‐dependent endocytosis and is dephosphorylated by alkaline phosphatase (ALP), releasing BTZ. The peptides sequestrate the inhibitors of apoptotic proteins (IAPs), promoting cell death. In contrast, this assembly enters normal cells by macropinocytosis and is transported to the lysosome, reducing side effects of cancer therapy for normal cells.
Presently, little is known of how the inter‐organelle crosstalk impacts cancer cells owing to the lack of approaches that can manipulate inter‐organelle communication in cancer cells. We found that a ...negatively charged, enzyme cleavable peptide (MitoFlag) enables the trafficking of histone protein H2B, a nuclear protein, to the mitochondria in cancer cells. MitoFlag interacts with the nuclear location sequence of H2B to block it from entering the nucleus. A protease on the mitochondria cleaves the Flag from the MitoFlag/H2B complex to form assemblies that retain H2B on the mitochondria and facilitate H2B entering the mitochondria. Adding NLS, replacing aspartic acid by glutamic acid residues, or changing the l‐ to d‐aspartic acid residue on MitoFlag abolishes the trafficking of H2B into mitochondria of HeLa cells. As the first example of the enzyme‐instructed self‐assembly of a synthetic peptide for trafficking endogenous proteins, this work provides insights for understanding and manipulating inter‐organelle communication in cells.
Cellular pep talk: Carrying a negative charge, the synthetic peptideMitoFlag(l‐D) forms complexes with the nuclear protein, H2B, and traffics H2B to mitochondria through enzyme‐instructed self‐assembly. This result provides a supramolecular approach for understanding and molecular insights into the inter‐organelle communication.
Emerging evidence indicates that mitochondria contribute to drug resistance in cancer, but how to selectively target the mitochondria of cancer cells remains less explored. Here, we show ...perimitochondrial enzymatic self-assembly for selectively targeting the mitochondria of liver cancer cells. Nanoparticles of a peptide–lipid conjugate, being a substrate of enterokinase (ENTK), encapsulate chloramphenicol (CLRP), a clinically used antibiotic that is deactivated by glucuronidases in cytosol but not in mitochondria. Perimitochondrial ENTK cleaves the Flag-tag on the conjugate to deliver CLRP selectively into the mitochondria of cancer cells, thus inhibiting the mitochondrial protein synthesis, inducing the release of cytochrome c into the cytosol and resulting in cancer cell death. This strategy selectively targets liver cancer cells over normal liver cells. Moreover, blocking the mitochondrial protein synthesis sensitizes the cancer cells, relying on glycolysis and/or OXPHOS, to cisplatin. This work illustrates a facile approach, selectively targeting mitochondria of cancer cells and repurposing clinically approved ribosome inhibitors, to interrupt the metabolism of cancer cells for cancer treatment.
Most of the reported mitochondria-targeting molecules are lipophilic and cationic, and thus they may become cytotoxic with accumulation. Here we show enzymatic cleavage of branched peptides that ...carry negative charges for targeting mitochondria. Conjugating a well-established protein tag (i.e., FLAG-tag) to self-assembling motifs affords the precursors that form micelles. Enzymatic cleavage of the hydrophilic FLAG motif (DDDDK) by enterokinase (ENTK) turns the micelles to nanofibers. After being taken up by cells, the micelles, upon the action of intracellular ENTK, turn into nanofibers to locate mainly at mitochondria. The micelles of the precursors are able to deliver cargos (either small molecules or proteins) into cells, largely to mitochondria and within 2 h. Preventing ENTK proteolysis diminishes mitochondria targeting. As the first report of using enzymatic self-assembly for targeting mitochondria and delivery cargos to mitochondria, this work illustrates a fundamentally new way to target subcellular organelles for biomedicine.
Purpose
To develop a fast, sub‐millimeter 3D magnetic resonance fingerprinting (MRF) technique for whole‐brain quantitative scans.
Methods
An acquisition trajectory based on multi‐axis spiral ...projection imaging (maSPI) was implemented for 3D MRF with steady‐state precession and slab excitation. By appropriately assigning the in‐plane and through‐plane rotations of spiral interleaves in a novel acquisition scheme, an maSPI‐based acquisition was implemented, and the total acquisition time was reduced by up to a factor of 8 compared to stack‐of‐spiral (SOS)‐based acquisition. A sliding‐window method was also used to further reduce the required number of time points for a faster acquisition. The experiments were conducted both on a phantom and in vivo.
Results
The results from the phantom measurements with the proposed and gold standard methods were consistent with a good linear correlation and an R2 value approaching 0.99. The in vivo experiments achieved whole‐brain parametric maps with isotropic resolutions of 1 mm and 0.8 mm in 5.0 and 6.0 min, respectively, with potential for further acceleration. An in vivo experiment with intentionally moving subjects demonstrated that the maSPI scheme largely outperforms the SOS scheme in terms of robustness to head motion.
Conclusion
3D MRF with an maSPI acquisition scheme enables fast and robust scans for high‐resolution parametric mapping.
Neurite orientation dispersion and density imaging (NODDI) has become a popular diffusion MRI technique for investigating microstructural alternations during brain development, maturation and aging ...in health and disease. However, the NODDI model of diffusion does not explicitly account for compartment-specific T2 relaxation and its model parameters are usually estimated from data acquired with a single echo time (TE). Thus, the NODDI-derived measures, such as the intra-neurite signal fraction, also known as the neurite density index, could be T2-weighted and TE-dependent. This may confound the interpretation of studies as one cannot disentangle differences in diffusion from those in T2 relaxation. To address this challenge, we propose a multi-TE NODDI (MTE-NODDI) technique, inspired by recent studies exploiting the synergy between diffusion and T2 relaxation. MTE-NODDI could give robust estimates of the non-T2-weighted signal fractions and compartment-specific T2 values, as demonstrated by both simulation and in vivo data experiments. Results showed that the estimated non-T2 weighted intra-neurite fraction and compartment-specific T2 values in white matter were consistent with previous studies. The T2-weighted intra-neurite fractions from the original NODDI were found to be overestimated compared to their non-T2-weighted estimates; the overestimation increases with TE, consistent with the reported intra-neurite T2 being larger than extra-neurite T2. Finally, the inclusion of the free water compartment reduces the estimation error in intra-neurite T2 in the presence of cerebrospinal fluid contamination. With the ability to disentangle non-T2-weighted signal fractions from compartment-specific T2 relaxation, MTE-NODDI could help improve the interpretability of future neuroimaging studies, especially those in brain development, maturation and aging.
•Conventional NODDI-derived compartment fractions are T2-weighted and TE-dependent.•MTE-NODDI disentangles non-T2-weighted signal fractions from T2 relaxation.•Robust intra-neurite T2 estimation in WM even with CSF contamination.•Non-T2-weighted fractions may improve the interpretability of neurodevelopmental studies.
Purpose
In diffusion‐weighted magnetic resonance imaging (DW‐MRI), the fiber orientation distribution function (fODF) is of great importance for solving complex fiber configurations to achieve ...reliable tractography throughout the brain, which ultimately facilitates the understanding of brain connectivity and exploration of neurological dysfunction. Recently, multi‐shell multi‐tissue constrained spherical deconvolution (MSMT‐CSD) method has been explored for reconstructing full fODFs. To achieve a reliable fitting, similar to other model‐based approaches, a large number of diffusion measurements is typically required for MSMT‐CSD method. The prolonged acquisition is, however, not feasible in practical clinical routine and is prone to motion artifacts. To accelerate the acquisition, we proposed a method to reconstruct the fODF from downsampled diffusion‐weighted images (DWIs) by leveraging the strong inference ability of the deep convolutional neural network (CNN).
Methods
The method treats spherical harmonics (SH)‐represented DWI signals and fODF coefficients as inputs and outputs, respectively. To compensate for the reduced gradient directions with reduced number of DWIs in acquisition in each voxel, its surrounding voxels are incorporated by the network for exploiting their spatial continuity. The resulting fODF coefficients are fitted with applying the CNN in a multi‐target regression model. The network is composed of two convolutional layers and three fully connected layers. To obtain an initial evaluation of the method, we quantitatively measured its performance on a simulated dataset. Then, for in vivo tests, we employed data from 24 subjects from the Human Connectome Project (HCP) as training set and six subjects as test set. The performance of the proposed method was primarily compared to the super‐resolved MSMT‐CSD with the decreasing number of DWIs. The fODFs reconstructed by MSMT‐CSD from all available 288 DWIs were used as training labels and the reference standard. The performance was quantitatively measured by the angular correlation coefficient (ACC) and the mean angular error (MAE).
Results
For the simulated dataset, the proposed method exhibited the potential advantage over the model reconstruction. For the in vivo dataset, it achieved superior results over the MSMT‐CSD in all the investigated cases, with its advantage more obvious when a limited number of DWIs were used. As the number of DWIs was reduced from 95 to 25, the median ACC ranged from 0.96 to 0.91 for the CNN, but 0.93 to 0.77 for the MSMT‐CSD (with perfect score of 1). The angular error in the typical regions of interest (ROIs) was also much lower, especially in multi‐fiber regions. The average MAE for the CNN method in regions containing one, two, three fibers was, respectively, 1.09°, 2.75°, and 8.35° smaller than the MSMT‐CSD method. The visual inception of the fODF further confirmed this superiority. Moreover, the tractography results validated the effectiveness of the learned fODF, in preserving known major branching fibers with only 25 DWIs.
Conclusion
Experiments on HCP datasets demonstrated the feasibility of the proposed method in recovering fODFs from up to 11‐fold reduced number of DWIs. The proposed method offers a new streamlined reconstruction procedure and exhibits promising potential in acquisition acceleration for the reconstruction of fODFs with good accuracy.
Purpose To improve diagnosis of hippocampal sclerosis (HS) in patients with mesial temporal lobe epilepsy (MTLE) by using MR fingerprinting and compare with visual assessment of T1- and T2-weighted ...MR images. Materials and Methods For this prospective study performed between April and November 2016, T1 and T2 maps were obtained and tissue segmentation performed in consecutive patients with drug-resistant MTLE with unilateral or bilateral HS. T1 and T2 maps were compared between 33 patients with MTLE (23 women and 10 men; mean age, 32.6 years; age range, 16-60 years) and 30 healthy participants (20 women and 10 men; mean age, 28.8 years; age range, 18-40 years). Differences in individual bilateral hippocampi were compared by using a Wilcoxon signed rank test, whereas the Wilcoxon rank-sum test was used for difference analysis between healthy control participants and patients with MTLE. Results The diagnosis rate (ie, ratio of HS diagnosed on the basis of a 2.5-minute MR fingerprinting examination compared with standard methods: MRI, electroencephalography, and PET) was 32 of 33 (96.9%; 95% confidence interval: 84.9%, 100%), reflecting improved accuracy of diagnosis (P = 1.92 × 10
) over routine MR examinations that had a diagnostic rate of 23 of 33 (69.7%; 95% confidence interval: 51.5%, 81.6%). The comparison between atrophic and normal-appearing hippocampus in 33 patients with MTLE and healthy control participants demonstrated that both T1 and T2 values in HS lesions were higher than those of normal hippocampal tissue of healthy participants (T1: 1361 msec ± 85 vs 1249 msec ± 59, respectively; T2: 135 msec ± 15 vs 104 msec ± 9, respectively; P < .0001). Conclusion MR fingerprinting allowed for multiparametric mapping of temporal lobe within 2.5 minutes and helped to identify lesions suspicious for HS in patients with MTLE with improved accuracy.
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