PLGA nanocapsules as a delivery system for a recombinant LRP-based therapeutic.

Telomerase activity is directly affected by the laminin receptor precursor (LRP) protein, a highly conserved nonintegrin transmembrane receptor, which has been shown to have therapeutic effects in ageing, and age-related diseases. Recently, it has been found that overexpression of LRP-FLAG, by plasmid transfection, leads to a significant increase in telomerase activity in cell culture models. This may indicate that upregulation of LRP can be used to treat various age-related diseases. However, transfection is not a viable treatment strategy for patients. Therefore, we present a nanoencapsulated protein-based drug synthesised using poly(lactic-co-glycolic acid) (PLGA) nanocapsules for delivery of the 37 kDa LRP protein therapeutic. PLGA nanocapsules were synthesised using the double emulsification-solvent evaporation technique. Different purification methods, including filtration and centrifugation, were tested to ensure that the nanocapsules were within the optimal size range, and the BCA assay was used to determine encapsulation efficiency. The completed drug was tested in a HEK-293 cell culture model, to investigate the effect on cell viability, LRP protein levels and telomerase activity. A significant increase in total LRP protein levels with a concomitant increase in cell viability and telomerase activity was observed. Due to the observed increase in telomerase activity, this approach could represent a safer alternative to plasmid transfection for the treatment of age-related diseases.

Downregulation of LRP/LR with siRNA inhibits several cancer hallmarks in lung cancer cells.

The incidence and mortality rates of cancer are growing rapidly worldwide, with lung cancer being the most commonly occurring cancer in males. Human carcinomas circumvent the inhibitory pathways induced by DNA damage and senescence through the upregulation of telomerase activity. The 37 kDa/67 kDa laminin receptor (LRP/LR) is a cell surface receptor which plays a role in several cancer hallmarks, including metastasis, angiogenesis, cell viability maintenance, apoptotic evasion, and mediating telomerase activity. We have previously shown that the knockdown of LRP/LR with an LRP-specific siRNA significantly impedes adhesion and invasion, induces apoptosis, and inhibits telomerase activity in various cancer cell lines in vitro. Here, we investigated the effect of downregulating LRP/LR with LRP-specific siRNA in A549 lung cancer cells. Downregulation of LRP/LR resulted in a significant decrease in cell viability, migration potential, and telomerase activity, as well as a significant increase in apoptosis. Proteomic analysis further suggested the re-establishment of immune control over the lung cancer cells, a previously unidentified facet of LRP downregulation in cancer. Altogether, we suggest that targeting LRP/LR for downregulation may have therapeutic potential for inhibiting several cancer hallmarks.

Knock-down of LRP/LR influences signalling pathways in late-stage colorectal carcinoma cells.

BACKGROUND: The 37 kDa/67 kDa laminin receptor (LRP/LR) is involved in several tumourigenic-promoting processes including cellular viability maintenance and apoptotic evasion. Thus, the aim of this study was to assess the molecular mechanism of LRP/LR on apoptotic pathways in late stage (DLD-1) colorectal cancer cells upon siRNA-mediated down-regulation of LRP/LR. METHODS: siRNAs were used to down-regulate the expression of LRP/LR in DLD-1 cells which was assessed using western blotting and qPCR. To evaluate the mechanistic role of LRP/LR, proteomic analysis of pathways involved in proliferation and apoptosis were investigated. The data from the study was analysed using a one-way ANOVA, followed by a two-tailed student's t-test with a confidence interval of 95%. RESULTS: Here we show that knock-down of LRP/LR led to significant changes in the proteome of DLD-1 cells, exposing new roles of the protein. Moreover, analysis showed that LRP/LR may alter components of the MAPK, p53-apoptotic and autophagic signalling pathways to aid colorectal cancer cells in continuous growth and survival. Knock-down of LRP/LR also resulted in significant decreases in telomerase activity and telomerase-related proteins in the DLD-1 cells. CONCLUSIONS: These findings show that LRP/LR is critically implicated in apoptosis and cell viability maintenance and suggest that siRNA-mediated knock-down of LRP/LR may be a possible therapeutic strategy for the treatment of colorectal cancer.

LRP::FLAG Reduces Phosphorylated Tau Levels in Alzheimer's Disease Cell Culture Models.

BACKGROUND: Alzheimer's disease (AD) is characterized by amyloid-ß (Aß) plaque and neurofibrillary tangle formation, respectively. Neurofibrillary tangles form as a result of the intracellular accumulation of hyperphosphorylated tau. Telomerase activity and levels of the human reverse transcriptase (hTERT) subunit of telomerase are significantly decreased in AD. Recently, it has been demonstrated that the 37 kDa/67 kDa laminin receptor (LRP/LR) interacts with telomerase and is implicated in Aß pathology. Since both LRP/LR and telomerase are known to play a role in the Aß facet of AD, we hypothesized that they might also play a role in tauopathy. OBJECTIVE: This study aimed to determine if LRP/LR has a relationship with tau and whether overexpression of LRP::FLAG has an effect on tauopathy-related proteins. METHODS: We employed confocal microscopy and FRET to determine whether LRP/LR and tau co-localize and interact. LRP::FLAG overexpression in HEK-293 and SH-SY5Y cells as well as analysis of tauopathy-related proteins was assessed by western blotting. RESULTS: We demonstrate that LRP/LR co-localizes with tau in the perinuclear cell compartment and confirmed a direct interaction between LRP/LR and tau in HEK-293 cells. Overexpression of LRP::FLAG in HEK-293 and SH-SY5Y cells decreased total and phosphorylated tau levels with a concomitant decrease in PrPc levels, a tauopathy-related protein. LRP::FLAG overexpression also resulted in increased hTERT levels. CONCLUSION: This data suggest that LRP/LR extends its role in AD through a direct interaction with tau, and recommend LRP::FLAG as a possible alternative AD therapeutic via decreasing phosphorylated tau levels.

Patented therapeutic approaches targeting LRP/LR for cancer treatment.

Introduction: The ubiquitously expressed 37 kDa/67 kDa high-affinity laminin receptor (laminin receptor precursor/laminin receptor, LRP/LR) is a protein found to play several roles within cells. The receptor is located in the nucleus, cytosol and the cell surface. LRP/LR mediates cell proliferation, cell adhesion and cell differentiation. As a result, it is seen to enhance tumor angiogenesis as well as invasion and adhesion, key steps in the metastatic cascade of cancer. Recent findings have shown that LRP/LR is involved in the maintenance of cell viability through apoptotic evasion, allowing for tumor progression. Thus, several patented therapeutic approaches targeting the receptor for the prevention and treatment of cancer have emerged.Areas covered: The several roles that LRP/LR plays in cancer progression as well as an overview of the current therapeutic patented strategies targeting LRP/LR and cancer to date.Expert opinion: Small molecule inhibitors, monoclonal antibodies and small interfering RNAs might act used as powerful tools in preventing tumor angiogenesis and metastasis through the induction of apoptosis and telomere erosion in several cancers. This review offers an overview of the roles played by LRP/LR in cancer progression, while providing novel patented approaches targeting the receptor as potential therapeutic routes for the treatment of cancer as well as various other diseases.

LRP::FLAG Rescues Cells from Amyloid-ß-Mediated Cytotoxicity Through Increased TERT Levels and Telomerase Activity.

Alzheimer's disease (AD) represents the most common form of neurodegenerative disorders with only palliative treatments currently available. Amyloid plaque formation caused by amyloid-ß (Aß) aggregation and neurofibrillary tangle formation caused by hyperphosphorylated tau are hallmarks for the development of AD. The 37 kDa/67 kDa laminin receptor (LRP/LR) has been implicated in AD and tools blocking or downregulating LRP/LR impede amyloid plaque formation in vitro and in vivo. We have recently shown that LRP::FLAG enhances telomerase activity with a concomitant reduction of senescent markers. Here, we overexpressed LRP::FLAG in HEK293 and SH-SY5Y cells, which resulted in an increase in hTERT levels as well as increased telomerase activity and increased cell viability in the presence of cytotoxic levels of exogenous Aß. LRP::FLAG overexpression decreased Aß shedding and intracellular Aß levels in HEK293 cells. This suggests that LRP::FLAG rescues cells from Aß-induced cytotoxicity through increased telomerase activity. This study recommends LRP::FLAG as a novel alternative therapeutic for AD treatment through activation of telomerase activity.

LRP/LR specific antibody IgG1-iS18 impedes neurodegeneration in Alzheimer's disease mice.

Alzheimer's disease (AD) is a neurodegenerative disease caused by accumulation of amyloid beta (Aß) plaque and neurofibrillary tangle formation. We have shown in vitro, that knock-down and blockade of the 37 kDa/67 kDa Laminin Receptor (LRP/LR) resulted in reduced Aß induced cytotoxicity and Aß accumulation. In order to test the effect of blocking LRP/LR on Aß formation and AD associated symptoms, AD transgenic mice received the anti-LRP/LR specific antibody, IgG1-iS18 through intranasal administration. We show that this treatment resulted in an improvement in memory, and decreased Aß plaque formation. Moreover, a significant decrease in Aß42 protein expression with a concomitant increase in amyloid precursor protein (APP) and telomerase reverse transcriptase (mTERT) levels was observed. These data recommend IgG1-iS18 as a potentially powerful therapeutic antibody for AD treatment.

Knock-down of LRP/LR promotes apoptosis in early and late stage colorectal carcinoma cells via caspase activation.

BACKGROUND: Cancer remains one of the leading causes of death around the world, where incidence and mortality rates are at a constant increase. Tumourigenic cells are characteristically seen to over-express the 37 kDa/67 kDa laminin receptor (LRP/LR) compared to their normal cell counterparts. This receptor has numerous roles in tumourigenesis including metastasis, angiogenic enhancement, telomerase activation, cell viability and apoptotic evasion. This study aimed to expose the role of LRP/LR on the cellular viability of early (SW-480) and late (DLD-1) stage colorectal cancer cells. METHODS: siRNA were used to down-regulate the expression of LRP/LR in SW-480 and DLD-1 cells which was assessed using western blotting. Subsequently, cell survival was evaluated using the MTT cell survival assay and confocal microscopy. Thereafter, Annexin V-FITC/PI staining and caspase activity assays were used to investigate the mechanism underlying the cell death observed upon LRP/LR knockdown. The data was analysed using Student's t-test with a confidence interval of 95%, with p-values of less than 0.05 seen as significant. RESULTS: Here we reveal that siRNA-mediated knock-down of LRP led to notable decreases in cell viability through increased levels of apoptosis, apparent by compromised membrane integrity and significantly high caspase-3 activity. Down-regulated LRP resulted in a significant increase in caspase-8 and -9 activity in both cell lines. CONCLUSIONS: These findings show that the receptor is critically implicated in apoptosis and that LRP/LR down-regulation induces apoptosis in early and late stage colorectal cancer cells through both apoptotic pathways. Thus, this study suggests that siRNA-mediated knock-down of LRP could be a possible therapeutic strategy for the treatment of early and late stage colorectal carcinoma.

siRNA - Mediated LRP/LR knock-down reduces cellular viability of malignant melanoma cells through the activation of apoptotic caspases.

The 37 kDa/67 kDa laminin receptor (LRP/LR) is over-expressed in tumor cells and has been implicated in several tumourigenic processes such as metastasis and telomerase activation, however, more importantly the focus of the present study is on the maintenance of cellular viability and the evasion of apoptosis. The aim of the study was to investigate the role of LRP/LR on the cellular viability of early (A375) and late stage (A375SM) malignant melanoma cells. Flow cytometry and western blot analysis revealed that A375SM cells contain more cell-surface and total LRP/LR levels in comparison to the A375 cells, respectively. In order to determine the effect of LRP/LR on cell viability and apoptosis, LRP was down-regulated via siRNA technology. MTT assays revealed that LRP knock-down led to significant reductions in the viability of A375 and A375SM cells. Confocal microscopy indicated nuclear morphological changes suggestive of apoptotic induction in both cell lines and Annexin-V FITC/PI assays confirmed this observation. Additionally, caspase-3 activity assays revealed that apoptosis was induced in both cell lines after siRNA-mediated down-regulation of LRP. Caspase-8 and -9 activity assays suggested that post LRP knock-down; A375 cells undergo apoptosis solely via the extrinsic pathway, while A375SM cells undergo apoptosis via the intrinsic pathway. IMPLICATIONS: siRNAs mediated LRP knock-down might represent a powerful alternative therapeutic strategy for the treatment of malignant melanoma through the induction of apoptosis.

37 kDa LRP::FLAG enhances telomerase activity and reduces senescent markers in vitro.

One of the core regulators of cellular aging are telomeres, repetitive DNA sequences at the ends of chromosomes that are maintained by the ribonucleoprotein DNA polymerase complex, telomerase. Recently, we demonstrated that knockdown of the 37kDa/ 67kDa laminin receptor (LRP/LR), a protein that promotes cell viability in tumorigenic and normal cells, reduces telomerase activity. We therefore hypothesized that upregulating LRP/LR might increase telomerase activity and impede aging. Here we show that overexpression of LRP::FLAG resulted in significantly elevated hTERT levels, telomerase activity and telomere length, respectively, with concomitantly reduced levels of senescence markers. These data suggest a novel function of LRP/LR hampering the onset of senescence through elevating hTERT levels and telomerase activity, respectively. LRP::FLAG might therefore act as a potential novel anti-aging drug through the impediment of the cellular aging process.

Knockdown of LRP/LR induces apoptosis in pancreatic cancer and neuroblastoma cells through activation of caspases.

The 37kDa/67kDa laminin receptor (LRP/LR) serves various physiological and pathological roles such as enhancing tumour-related processes including metastasis, angiogenesis, cellular viability and telomerase activation in cancerous cell lines. The present study investigates the effect of siRNA mediated downregulation of LRP/LR on pancreatic cancer (AsPC-1) and neuroblastoma (IMR-32) cells. MTT and BrdU assays revealed that siRNA mediated downregulation of LRP resulted in a significant reduction in cell viability and cell proliferation. In addition, knock-down of LRP resulted in phosphatidylserine externalization, diminished nuclear integrity and significantly enhanced caspase-3 activity, which is indicative of apoptosis. LRP downregulation resulted in a significant increase in caspase-8 activity in IMR-32 cells and enhanced caspase-8 and 9 activity in AsPC-1 cells. These data recommend siRNA mediated knock-down of LRP as a potential therapeutic avenue for the treatment of pancreatic cancer and neuroblastoma.

IgG1-iS18 impedes the adhesive and invasive potential of early and late stage malignant melanoma cells.

The 37kDa/67kDa laminin receptor (LRP/LR) is a non-integrin laminin receptor which is overexpressed in tumorigenic cells and supports progression of cancer via promoting metastasis, angiogenesis and telomerase activity and impediment of apoptosis. The present study investigates the role of LRP/LR on the metastatic potential of early (A375) and late (A375SM) stage malignant melanoma cells. Flow cytometry revealed that both early and late stage malignant melanoma cells display high levels of LRP/LR on their cell surface. Flow cytometry and western blot analysis showed that late stage malignant melanoma cells display significantly higher total and cell surface LRP/LR levels in comparison to early stage malignant melanoma cells and the poorly invasive breast cancer (MCF-7) control cell line. Targeting LRP/LR using the LRP/LR specific antibody IgG1-iS18 resulted in a significant reduction of the adhesive potential to laminin-1 and the invasive potential through the 'ECM-simulating' Matrigel™ of both early and late stage malignant melanoma cells. Furthermore, Pearson's correlation coefficient confirmed that increased LRP levels correlate with the increased invasive and adhesive potential in early and late stage melanoma cells. Thus, blocking LRP/LR using the IgG1-iS18 antibody may therefore be a promising therapeutic strategy for early and late stage malignant melanoma treatment.

Anti-LRP/LR-specific antibody IgG1-iS18 impedes adhesion and invasion of pancreatic cancer and neuroblastoma cells.

BACKGROUND: Cancer has become a global burden due to its high incidence and mortality rates, with an estimated 14.1 million cancer cases reported worldwide in 2012 particularly as a result of metastasis. Metastasis involves two crucial steps: adhesion and invasion, and the non-integrin receptor; the 37-kDa/67-kDa laminin receptor precursor/ high affinity laminin receptor (LRP/LR) has been shown to be overexpressed on the surface of tumorigenic cells, thus being implicated in the enhancement of these two crucial steps. The current study investigated the role of LRP/LR on the aggressiveness of pancreatic cancer (AsPC-1) and neuroblastoma (IMR-32) cells with respect to their adhesive and invasive potential. METHODS: AsPC-1 and IMR-32 cells were utilized as the experimental cell lines for the study. Cell surface LRP/LR levels were visualised and quantified on the experimental and control (MCF-7) cell lines via confocal microscopy and flow cytometry, respectively. Total LRP/LR levels in the cell lines were assessed by Western blotting and the adhesive and invasive potential of the above-mentioned cell lines was determined before and after supplementation with the anti-LRP/LR specific antibody IgG1-iS18. Statistical significance of the data was confirmed via the use of the two-tailed student's t-test and Pearson's correlation coefficient. RESULTS: Flow cytometry revealed that AsPC-1 and IMR-32 cells displayed significantly higher cell surface LRP/LR levels in comparison to the MCF-7 control cell line. However, Western blotting and subsequent densitometric analysis revealed that all three tumorigenic cell lines displayed no significant difference in total LRP/LR levels. The treatment of AsPC-1 and IMR-32 cells with IgG1-iS18 caused a significant reduction in the adhesive and invasive potential of the cells to laminin-1 and through the ECM-like Matrigel™, respectively. Pearson's correlation coefficients indicated a high correlation, thus suggesting a directly proportional relationship between cell surface LRP/LR levels and the adhesive and invasive potential of AsPC-1 and IMR-32 cells. CONCLUSION: These findings suggest that through the interference of the LRP/LR-laminin-1 interaction, the anti-LRP/LR specific antibody IgG1-iS18 may act as an alternative therapeutic tool for the treatment of metastatic pancreatic cancer and neuroblastoma.

Anti-LRP/LR specific antibody IgG1-iS18 significantly impedes adhesion and invasion in early and late stage colorectal carcinoma cells.

Cancer is a highly complex disease that has become one of the leading causes of death globally. Metastasis, a major cause of cancer deaths, requires two crucial events known as adhesion and invasion. The 37kDa/67kDa laminin receptor [laminin receptor precursor/high-affinity laminin receptor (LRP/LR)] enhances these two steps, consequently aiding in cancer progression. In this study, the role of LRP/LR in adhesion and invasion of early (SW-480 & HT-29) and late (DLD-1) stage colorectal cancer cells has been investigated. Western blotting revealed that early and late stage colorectal cancer cells contained significantly higher total LRP/LR levels compared to poorly invasive MCF-7 breast cancer control cells. Flow cytometry revealed that all three stages of colorectal cancer displayed significantly higher cell surface LRP/LR levels. Furthermore, upon treatment of the colorectal cancer cells with the anti-LRP/LR specific antibody IgG1-iS18, adhesion to laminin-1 was significantly reduced in all three stages. Each stage's invasive potential was determined using the Matrigel™ invasion assay, which revealed that invasion is significantly impeded in all three colorectal cancer stages when the cells are incubated with IgG1-iS18. In addition, Pearson's correlation coefficients propose that both total and cell surface LRP/LR levels are directly proportional to the adhesive and invasive potential of all three stages of colorectal cancer. Hence, these findings indicate the potential for the use of the IgG1-iS18 antibody as a promising therapeutic tool for colorectal cancer patients of early and late stage.

LRP/LR Antibody Mediated Rescuing of Amyloid-ß-Induced Cytotoxicity is Dependent on PrPc in Alzheimer's Disease.

The neuronal perturbations in Alzheimer's disease are attributed to the formation of extracellular amyloid-ß (Aß) neuritic plaques, composed predominantly of the neurotoxic Aß42 isoform. Although the plaques have demonstrated a role in synaptic dysfunction, neuronal cytotoxicity has been attributed to soluble Aß42 oligomers. The 37kDa/67kDa laminin receptor has been implicated in Aß42 shedding and Aß42-induced neuronal cytotoxicity, as well as internalization of this neurotoxic peptide. As the cellular prion protein binds to both LRP/LR and Aß42, the mechanism underlying this cytotoxicity may be indirectly due to the PrPc-Aß42 interaction with LRP/LR. The effects of this interaction were investigated by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assays. PrPc overexpression significantly enhanced Aß42 cytotoxicity in vitro, while PrP-/-  cells were more resistant to the cytotoxic effects of Aß42 and exhibited significantly less cell death than PrPc expressing N2a cells. Although anti-LRP/LR specific antibody IgG1-iS18 significantly enhanced cell viability in both pSFV1-huPrP1-253 transfected and non-transfected cells treated with exogenous Aß42, it failed to have any cell rescuing effect in PrP-/-  HpL3-4 cells. These results suggest that LRP/LR plays a significant role in Aß42-PrPc mediated cytotoxicity and that anti-LRP/LR specific antibodies may serve as potential therapeutic tools for Alzheimer's disease.

Knock-Down of the 37kDa/67kDa Laminin Receptor LRP/LR Impedes Telomerase Activity.

Cancer has become a major problem worldwide due to its increasing incidence and mortality rates. Both the 37kDa/67kDa laminin receptor (LRP/LR) and telomerase are overexpressed in cancer cells. LRP/LR enhances the invasiveness of cancer cells thereby promoting metastasis, supporting angiogenesis and hampering apoptosis. An essential component of telomerase, hTERT is overexpressed in 85-90% of most cancers. hTERT expression and increased telomerase activity are associated with tumor progression. As LRP/LR and hTERT both play a role in cancer progression, we investigated a possible correlation between LRP/LR and telomerase. LRP/LR and hTERT co-localized in the perinuclear compartment of tumorigenic breast cancer (MDA_MB231) cells and non-tumorigenic human embryonic kidney (HEK293) cells. FLAG® Co-immunoprecipitation assays confirmed an interaction between LRP/LR and hTERT. In addition, flow cytometry revealed that both cell lines displayed high cell surface and intracellular LRP/LR and hTERT levels. Knock-down of LRP/LR by RNAi technology significantly reduced telomerase activity. These results suggest for the first time a novel function of LRP/LR in contributing to telomerase activity. siRNAs targeting LRP/LR may act as a potential alternative therapeutic tool for cancer treatment by (i) blocking metastasis (ii) promoting angiogenesis (iii) inducing apoptosis and (iv) impeding telomerase activity.

Knockdown of LRP/LR Induces Apoptosis in Breast and Oesophageal Cancer Cells.

Cancer is a global burden due to high incidence and mortality rates and is ranked the second most diagnosed disease amongst non-communicable diseases in South Africa. A high expression level of the 37kDa/67kDa laminin receptor (LRP/LR) is one characteristic of cancer cells. This receptor is implicated in the pathogenesis of cancer cells by supporting tumor angiogenesis, metastasis and especially for this study, the evasion of apoptosis. In the current study, the role of LRP/LR on cellular viability of breast MCF-7, MDA-MB 231 and WHCO1 oesophageal cancer cells was investigated. Western blot analysis revealed that total LRP expression levels of MCF-7, MDA-MB 231 and WHCO1 were significantly downregulated by targeting LRP mRNA using siRNA-LAMR1. This knockdown of LRP/LR resulted in a significant decrease of viability in the breast and oesophageal cancer cells as determined by an MTT assay. Transfection of MDA-MB 231 cells with esiRNA-RPSA directed against a different region of the LRP mRNA had similar effects on LRP/LR expression and cell viability compared to siRNA-LAMR1, excluding an off-target effect of siRNA-LAMR1. This reduction in cellular viability is as a consequence of apoptosis induction as indicated by the exposure of the phosphatidylserine protein on the surface of breast MCF-7, MDA-MB 231 and oesophageal WHCO1 cancer cells, respectively, detected by an Annexin-V/FITC assay as well as nuclear morphological changes observed post-staining with Hoechst. These observations indicate that LRP/LR is crucial for the maintenance of cellular viability of breast and oesophageal cancer cells and recommend siRNA technology targeting LRP expression as a possible novel alternative technique for breast and oesophageal cancer treatment.

Novel patented therapeutic approaches targeting the 37/67 kDa laminin receptor for treatment of cancer and Alzheimer's disease.

INTRODUCTION: The 37/67 kDa high-affinity laminin receptor (laminin receptor precursor/laminin receptor, LRP/LR) is a multi-faceted cellular receptor. It plays a vital role in the malignancy of various cancer types where it is seen to contribute to invasion, adhesion, apoptosis evasion and angiogenesis. Furthermore, it has been found to play an important role in facilitating the processes leading to neurotoxicity in Alzheimer's disease (AD). Various therapeutic options targeting this receptor have been patented with the outlook on application for the treatment/prevention of these diseases. AREAS COVERED: The various roles that LRP/LR plays in cancer, AD and infectious diseases caused by viruses and bacteria have been examined in detail and an overview of the current patented therapeutic strategies targeting this receptor is given. EXPERT OPINION: Molecular tools directed against LRP/LR, such as antibodies and small interfering RNA, could prove to be effective in the prevention of metastasis and angiogenesis while inducing apoptosis in cancers. Moreover, these strategies could also be applied to AD where LRP/LR is seen to facilitate the production and internalization of the neurotoxic Aß peptide. This review provides a comprehensive overview of the mechanisms by which LRP/LR is involved in eliciting pathogenic events, while showing how the use of patented approaches targeting this receptor could be used to treat them.

The 37kDa/67kDa laminin receptor acts as a receptor for Aß42 internalization.

Neuronal loss is a major neuropathological hallmark of Alzheimer's disease (AD). The associations between soluble Aß oligomers and cellular components cause this neurotoxicity. The 37 kDa/67 kDa laminin receptor (LRP/LR) has recently been implicated in Aß pathogenesis. In this study the mechanism underlying the pathological role of LRP/LR was elucidated. Försters Resonance Energy Transfer (FRET) revealed that LRP/LR and Aß form a biologically relevant interaction. The ability of LRP/LR to form stable associations with endogenously shed Aß was confirmed by pull down assays and Aß-ELISAs. Antibody blockade of this association significantly lowered Aß42 induced apoptosis. Furthermore, antibody blockade and shRNA mediated downregulation of LRP/LR significantly hampered Aß42 internalization. These results suggest that LRP/LR is a receptor for Aß42 internalization, mediating its endocytosis and contributing to the cytotoxicity of the neuropeptide by facilitating intra-cellular Aß42 accumulation. These findings recommend anti-LRP/LR specific antibodies and shRNAs as potential therapeutic tools for AD treatment.

High resolution imaging study of interactions between the 37 kDa/67 kDa laminin receptor and APP, beta-secretase and gamma-secretase in Alzheimer's disease.

Alzheimer's disease (AD) is the most prevalent form of dementia affecting the elderly. Neurodegeneration is caused by the amyloid beta (Aß) peptide which is generated from the sequential proteolytic cleavage of the Amyloid Precursor Protein (APP) by the ß- and γ- secretases. Previous reports revealed that the 37 kDa/67 kDa laminin receptor (LRP/LR) is involved in APP processing, however, the exact mechanism by which this occurs remains largely unclear. This study sought to assess whether LRP/LR interacted with APP, ß- or γ-secretase. Detailed confocal microscopy revealed that LRP/LR showed a strong co-localisation with APP, ß- and γ-secretase, respectively, at various sub-cellular locations. Superresolution Structured Illumination Microscopy (SR-SIM) showed that interactions were unlikely between LRP/LR and APP and ß-secretase, respectively, while there was strong co-localisation between LRP/LR and γ-secretase at this 80 nm resolution. FRET was further employed to assess the possibility of protein-protein interactions and only an interaction between LRP/LR and γ-secretase was found. FLAG co-immunoprecipitation confirmed these findings as LRP/LR co-immunoprecipitated with γ-secretase, but failed to do so with APP. These findings indicate that LRP/LR exerts its influence on Aß shedding via a direct interaction with the γ-secretase and possibly an indirect interaction with the ß-secretase.