key: cord-0864607-zr7066rp authors: Elfiky, Abdo A.; Baghdady, Ahmed M.; Ali, Shehab A.; Ahmed, Marwan I. title: GRP78 targeting: Hitting two birds with a stone date: 2020-08-22 journal: Life Sci DOI: 10.1016/j.lfs.2020.118317 sha: 66d98b8e4622431a651b9966aeba2b24d50bd95a doc_id: 864607 cord_uid: zr7066rp BACKGROUND: Glucose regulating protein 78 (GRP78) is one member of the Heat Shock Protein family of chaperone proteins (HSPA5) found in eukaryotes. It acts as the master of the Unfolded Protein Response (UPR) process in the lumen of the Endoplasmic Reticulum (ER). SCOPE: Under the stress of unfolded proteins, GRP78 binds to the unfolded proteins to prevent misfolding, while under the load of the unfolded protein, it drives the cell to autophagy or apoptosis. Several attempts reported the overexpression of GRP78 on the cell membrane of cancer cells and cells infected with viruses or fungi. MAJOR CONCLUSIONS: Cell-surface GRP78 is used as a cancer cell target in previous studies. Additionally, GRP78 is used as a drug target to stop the progression of cancer cells by different compounds, including peptides, antibodies, and some natural compounds. Additionally, it can be used as a protein target to reduce the infectivity of different viruses, including the pandemic SARS-CoV-2. Besides, GRP78 targeting is used in diagnosis and imaging modalities using radionuclides. GENERAL SIGNIFICANCE: This review summarizes the various attempts that used GRP78 both in therapy (fighting cancer, viral and fungal infections) and diagnosis (imaging). In eukaryotes, different mechanisms regulate the cell proteostasis, which can be understood as a cell's response to a signal (Ellgaard and Helenius, 2003 , Gething and Sambrook, 1992 , Hu et al. , 2019 , Ibrahim et al. , 2019 , Little et al. , 1994 . Proteins represent the cell machines and tools used to performs specific functions or biochemical reactions harmonically (Brocchieri et al. , 2008) . With time, the biomacromolecules are affected by the cellular environment and may undergo partial unfolding or misfolding and need to be revisited by the cellular refolding or degradation mechanisms (Ellgaard and Helenius, 2003 , Ismail et al. , 2020 , Liao et al. , 2008 , Little et al., 1994 . Glucose regulating protein 78 (GRP78) or Heat Shock Protein A5 (HSPA5) is the master protein responsible for directing the misfolded proteins in the ER for refolding or degradation mechanisms to keep the unfolded protein concentration at a minimum (Gething and Sambrook, 1992 , Haas, 1991 , Ibrahim et al., 2019 . Cell degrade proteins either by the ubiquitin-proteasome system (UPS) or autophagy-lysosome pathway where, in both cases, chaperones play a crucial role (Li et al. , 2008) . and aid in improving resistance against anticancer drugs. Inhibitors of GRP78 as an anticancer agent is used as a cancer-fighting strategy (Srivastava et al. , 2018) . The review discusses the three main approaches that are utilized to target GRP78; phytochemicals inhibitors, peptide inhibitors, and monoclonal antibodies (see the graphical abstract). Inhibiting such target protein reduces the virulence of pathogens and reduces the therapeutic resistance in the case of cancer. We first come to the basic understanding of the role of GRP78 in healthy and diseased cells. J o u r n a l P r e -p r o o f Journal Pre-proof On the other hand, under the stress of unfolded proteins, GRP78 can escape the ER retention and translocates to the cytoplasm and on the cell membrane and become membrane-exposed, termed Cell-Surface CS-GRP78 ( Figure 1 ) (Ibrahim et al., 2019, Tsai and Lee, 2018a) . This CS-GRP78 characterizes many aggressive types of cancers such as breast, ovarian, pancreatic, and colon cancers (Chang et al. , 2012 , Chen and Xu, 2017 , Ibrahim et al., 2019 , Kim et al. , 2006b , Li et al. , 2013 , Niu et al. , 2015 , Tian et al. , 2015 , Tsai and Lee, 2018a , Wang et al., 2009 , Xie et al. , 2016 , Yuan et al., 2015 , Zhang et al. , 2006 . Additionally, CS-GRP78 was reported to facilitate pathogenic entry, J o u r n a l P r e -p r o o f reported GRP78 association with viral proteins and GRP78 upregulation in infected cells , Choukhi et al. , 1998 , Chu et al. , 2018 , Das et al. , 2009 , Honda et al. , 2009 , Jindadamrongwech et al. , 2004 , Liberman et al. , 1999 , Nain et al. , 2017 , Pujhari et al. , 2017 , Reyes-del Valle et al. , 2005 , Shurtleff et al. , 2014 . Additionally, recent studies hypothesized the association of CS-GRP78 with the spike protein of SARS-CoV-2 to help in virus attachment and host cell entry (Ibrahim et al. , 2020b) . The primary binding viral protein to GRP78 are spike proteins in coronaviruses and envelope proteins for the other viruses (Elfiky, 2020a , d, Elfiky and Ibrahim, 2020 , Ibrahim et al. , 2020a . Besides, the spore coat protein homolog (CotH3) of Rhizopus oryzae, the causative fungus for Mucormycosis, is reported to bind to CS-GRP78 on endothelial cells and the binding is responsible for adherence and invasion of the fungus (Gebremariam et al. , 2014) . Since the association of the viral or pathogen infection and elevated levels of CS- Chemoresistance is the resistance of a tumor to chemotherapy. It was an old observation, while the mechanism of GRP78-induced chemo-resistance in cancer cells was not fully understood (Wang et al. , 1999) . Two mechanisms may be responsible for the chemo-resistance; the UPR pro-survival branch and the receptor-mediated activation of the Akt/PI3K (Phosphoinositide 3-kinase) pathway (Roller and Maddalo, 2013b) . Alternatively, the proapoptotic action of the UPR could be compensated by the activation of the Akt/PI3K pathway, resulting in cell survival. The extracellular loop of Cleft Lip and Palate Transmembrane 1-Like (CLPTM1L) is essential for gemcitabine resistance and interaction with GRP78 (Clarke et al. , 2019) . Additionally, natural products such as isoliquiritigenin, a chalcone-type flavonoid, was able to reduce the chemoresistance and colony-forming ability of oral squamous cell carcinomas (Hu et al. , 2017) . It is reported that the prior treatment of acidic stress protects the human dermal microvascular endothelial cells from apoptosis by reduced the cleavage of caspase 7, which was supposed to be due to the presence of GRP78 on the membrane of ER that suppress caspase 7 activation (Visioli et al. , 2014) . Non-small-cell lung cancer (NSCLC) and glioblastoma multiforme (GBM) have a low survival rate. The overexpressed GRP78 on the cell surface is the primary reason for the radio-resistance in NSCLC and GBM (Dadey et al. , 2017) . Targeting cell-surface GRP78 enhances the apoptosis and reduces cell proliferation, colony formation, and downregulates the crucial intracellular phosphatidylinositol-3-kinase / protein kinase B / J o u r n a l P r e -p r o o f Journal Pre-proof mammalian target of rapamycin (PI3K/Akt/mTOR) signaling essential in the cell cycle, growth and survival (Chang et al., 2012 , Tian et al., 2015 , Xie et al., 2016 . Besides, tumor growth is delayed with enhanced efficacy of the radiation treatment upon anti-GRP78 antibody administration in mice (Dadey et al., 2017) . In breast cancer, GRP78 is overexpressed while the amount of cell-surface GRP78 is increased upon the treatment with the anti-angiogenic factor Combretastatin A4P (Gazit et al. , 1999) . Additionally, cancer cells treated with doxorubicin showed less resistance when treated with GRP78 neutralizing antibodies (Gazit et al., 1999 , Hannun, 1997 , Roller and Maddalo, 2013a , Shen et al. , 1987 . Generally, elevated levels of GRP78 is indicative of cancer aggressivity. Targeting cancer cell-surface GRP78 is a successful strategy to reduce the radio-resistance and chemo-resistance of tumors (Kuroda et al. , 2011 , Li et al., 2013 , Niu et al., 2015 , Pfaffenbach and Lee, 2011 . In this review, the focus is not only on some of the previous trials to use anti-GRP78 to treat cancer but also in the diagnosis (see the peptide inhibitors section below). Different strategies are used to reduce the burden of overexpressed CS-GRP78 (Bailly and Waring, 2019) . Different compounds show binding affinity to CS-GRP78. Once bound to a substrate, the CS-GRP78 will be internalized to the cell; hence the concentration of the membrane-bound GRP78 will be reduced. Once the level of GRP78 over the cell surface is dropped, the pathogens will not be able to enter the host cell through GRP78; hence the virulence will be reduced. The inhibitory molecules that can J o u r n a l P r e -p r o o f target CS-GRP78 include phytochemicals, peptides, and antibodies and will be discussed in detail in the next sections. The inhibitors compete with the pathogen recognizing proteins (such as spike, Envelope, or Coat proteins) for the CS-GRP78 substrate-binding domain β. This domain of the GRP78 is reported to be responsible for the binding of GRP78 to unfolded proteins inside the lumen of the ER through its hydrophobic batches (Gonzalez-Gronow et al. , 2009 , Quinones et al. , 2008 . Phytochemicals are compounds found in plants and have a variety of effects on protein function (Dillard and German, 2000) . They are derived from fruits, vegetables, beans, grains, and some other plants (Dillard and German, 2000) . Phytochemicals have a protective role because their antioxidant characteristics which play a vital role in the protection of cells against oxidative damage and decreasing the probability of cancer propagation via the reactive oxygen species (ROS), which can induce stress in ER. Apoptosis initiated by the ER If there is uncontrolled damage in cells (Hetz, 2012) . We summarize some phytochemicals crucial in cell stress relief through inhibiting the master of UPR, GRP78. Galangin is a flavonol produced from rhizomes of Alphina officinarum, which belongs to the ginger family and grows in Southeast Asia. Galangin work as a suppressor for cell proliferation in hepatocellular carcinoma (Su et al. , 2013) . It raises ER stress through the upregulation of the UPR target genes C/EBP Homologous Protein (CHOP), GRP78, Glucose regulating protein 94 (GRP94), and cytosolic Ca² + (Hu et al., 2019) . ER is the primary site for intercellular calcium ions, hence rising cytosolic Ca² + disrupts the J o u r n a l P r e -p r o o f function of ER chaperones, which induce ER stress leading to the activation of UPR and subsequent upregulation of GRP78 (Hotamisligil, 2010) . Galangin upregulates ER stress, which inhibits tumor progression through inducing apoptosis (Su et al., 2013) . 6-Shogaol is produced by dehydration of 6-gingerol and generated from rhizomes of ginger. When treating hepatocellular carcinoma (HCC) cell-line with 6-Shogaol, cancer cells develop apoptotic phenotypes signs such as nuclear shrinkage and condensation in chromatin (Srivastava et al., 2018) . Activation of CHOP expression and PERK de-phosphorylation initiates reactions of caspase cascade, which induce apoptosis in HCC. Significant stimulation was observed in ER stress-related proteins, which induce apoptosis by 6-Shogaol through rising in the UPR expression (GRP94, GRP78, and HSP70) (Hu et al. , 2012b) . Studies proved that exposing cancer cells to 6-Shogaol and the activator of the PERK/eIF2α pathway, salubrinal, together for a specific time induce ER stress, which leads to cell apoptosis (Srivastava et al., 2018) . Salubrinal alone enhances the phosphorylation of eIF2α in the human hepatocarcinoma cell line SMMC-7721 with negligible toxicity (Hu et al. , 2012a) . This reveals the significant therapeutic effect of anti-GRP78 against malignancies. Sulphureuine B is produced from Lattiporus sulphureus and tested by glioma cells to detect anti-proliferative properties. Studies revealed that Sulphureuine B provides ER stress by raising the level of expression of CHOP, caspase-12, and GRP78, which prevents separation of GRP78 from PERK, ATF6, and IRE1 which initiates UPR (Biswas et al. , 2015 , Ren et al. , 2015 . Additionally, Mushrooms contain p-Coumaric acid and Caffeic acid that proved its binding affinity against GRP78 SBDβ in silico, hence J o u r n a l P r e -p r o o f Journal Pre-proof suggested to be a possible inhibitor for overexpressed GRP78 in cancer cells or cell infected with viruses including SARS-CoV-2 (Elfiky, 2020c). Proanthocyanidins and resveratrol extracted from Grapeseeds. Exposing colorectal cancer cell (CRC) to Grape seed extract which has a high amount of proanthocyanidins and resveratrol leads to a modification in GRP78 and protein disulfide isomerase (PDI) which have a significant role in cell apoptosis which leads CRC to undergo apoptotic pathway leading to inhibition of the targeted cell to proliferation (Srivastava et al., 2018) . On the other hand, Caffeic acid and p-Coumaric acid polyphenols, found in the grape skin, have a protective role against photooxidative damage . Additionally, it has a pre-exposure protective role for the human retinal pigment epithelial cells (ARPE-19) against blue light-associated apoptosis in a dose-dependent manner by promoting GRP78 expression. In contrast, GRP78 knockdown inhibited this protective role . As mentioned before, Caffeic acid and p-Coumaric, are suitable binders to GRP78 SBDβ in silico (Elfiky, 2020c) . Estrogen receptor-positive breast cancer cells are responsive to hormonal therapy by blocking the estrogen synthesis leading to estrogen-starvation (Fu et al. , 2007) . It was reported that GRP78 plays a vital role in resist estrogen-starvation induced apoptosis in breast cancer cells. Hence, it was suggested to dual-target the GRP78 during treating estrogen-positive breast cancer, if the expression level of the GRP78 is high, to improve J o u r n a l P r e -p r o o f the efficacy and reduce the resistance (Fu et al., 2007) . It was reported that GRP78 interacts with estrogen due to the critical role of GRP78 in folding the hormone-binding domain of estrogen receptors (Avila et al. , 2013 , Fu et al., 2007 . Additionally, GRP78 targeting was suggested as a therapeutic strategy to sensitize cancer cells to chemotherapy in endometrial cancer (estrogen induced GRP78 expression) (Luvsandagva et al. , 2012) . Phytoestrogens are found in Cicer arietinum and include daidzein, genistein, formononetin, and biochanin A (Sayed and Elfiky, 2018) . Both Estrogens (estriol and βestradiol) and the four phytoestrogens are found to be recognized by GRP78 SBDβ and hence are suggested as possible GRP78 inhibitors in silico (Elfiky, 2020c) . It was concluded that estrogens and phytoestrogens are the best binders to the GRP78, while the binding affinities range from -7.0 down to -8.5 kcal/mol. This indicates an excellent binding affinity to GRP78 SBDβ, even better than a selective cyclic peptide, Pep42 (Elfiky, 2020c) . Despite its phytoestrogen activity, genistein activates the apoptosis process through UPR by upregulation of GRP78 and C/EBP homologous protein (CHOP), also termed growth arrest and DNA damage 153 (GADD153), and nuclear translation of GADD153 in HCC cells (Yeh et al. , 2007) . Epigallocatechin-3-gallate (EGCG) is a polyphenol found in green tea and has an anti-proliferative effect on breast cancer and melanoma (Nihal et al. , 2005) . Besides, it has an inhibition effect against GRP78 function through direct interaction with the ATP binding site of GRP78 competing against ATP binding (Ermakova et al. , 2006) . EGCG increases the therapeutic efficacy of tomozolomid when exposed to glioblastoma cells in vivo by inhibition of GRP78 (Chen et al. , 2011) . The olive leaf extracts active ingredient, hydroxytyrosol, show good binding affinity to the GRP78 SBDβ in silico (Elfiky, 2020c) . Hydroxytyrosol proved its role as a prophylactic agent against myocardial infarction-mediated apoptosis (Wu et al. , 2018) . Caffeic acid phenethyl ester (CAPE) is found in the hive propolis of the honeybee. CAPE shows in silico binding affinity against GRP78 SBDβ that is comparable to that of the cyclic Pep42, a selective GRP78 peptide (Elfiky, 2020c) . Besides, CAPE induces ER stress in human SH-SY5Y neuroblastoma in an autophagy-dependent manner (Tanida et al. , 2008 , Tomiyama et al. , 2018 . Different peptides are used to target the cell-surface GRP78 specifically (Bailly and Waring, 2019 , Goldenberg-Cohen et al. , 2012 , Palmeira et al. , 2020 . Peptides as anticancer drugs have two main types (i) short naked peptides to induce apoptosis (ii) conjugated peptides to deliver an anticancer drug into cells. For the first use, GRP78 serves as a receptor for the peptide and facilitates the internalization of the peptide, which can then modulate various pathways. Multidrug Resistance (MDR) is drug resistance that happens when cancer cells treated with one anticancer drug develop resistance to different drugs that are different from the used drug in structure and function (Gottesman, 2002, Zhang and Fan, 2010 ). An example of binding peptides that use GRP78 as a receptor is GMBP1, which is used J o u r n a l P r e -p r o o f in reversing gastric cancer MDR. GRP78 facilitates GMBP1 internalization into cells through the transferrin-related pathway . Gonadotropin-releasing hormone (GnRHa) is a hypothalamus secreted hormone that affects sex hormones, testosterone, and estrogen. Modified GnRHa is more efficient than the natural form; hence it is used as a drug depending on the analog (Bethesda, 2012) . GnRHa used as a drug against Endometriosis (a case in which cells like that lining the inside of the uterus grow outside it in other parts of the body) (Bulun, 2009 ). GnRHa inhibits proliferation and induces apoptosis of defected cells by inhibiting GRP78, thus leading to apoptosis (Weng et al. , 2014) . Cell targeting is the solution for the non-specific toxicity of anticancer drugs that affect cancer and healthy cells altogether and thus resulting in severe side effects (Liu et al. , 2007) . Peptides can target cancer cells and deliver anticancer drugs into the cell. In cancer cells, the peptide can bind to the surface, a membrane-bound form of the overexpressed chaperone, GRP78 (Kim et al. , 2006a) . To choose the peptide for a particular cancer cell, in vitro trials are required, such as phage display. Phage display is a technique for studying molecular interactions such as protein-DNA, protein-protein, and protein-peptide utilizing the bacteriophages to encode peptides to genetic information (Smith, 1985) . A pool of cyclic peptides tested against the cancer cells, and then a peptide is chosen to be used for drug delivery (Kim et al., 2006a) . Pep42 is a cyclic peptide (CTVALPGGYVRVC) identified by the phage display technique against human melanoma cell line Me6652/4 (Kim et al., 2006a) . CS-GRP78 is the receptor for Pep42 and facilitates it's internalization to the cell (Tsai and Lee, 2018b) . Pep42-taxol and Pep42-doxorubicin conjugates bind to GRP78 in highly metastatic human melanoma cells leading to its death in vitro, leading to cancer cell death (Ibrahim et al., 2019) . Pep42 selectively bind to GRP78 and enter the cell and thus make it a powerful tool to deliver anticancer drugs to various cancer cells (Kim et al., 2006a , Liu et al., 2007 , Yoneda et al. , 2008 . Pep42 was used as a profiler for in silico predicting the CS-GRP78 and viral proteins of the Zika virus, Human papillomavirus, SARS-CoV-2, and Ebola virus (Elfiky, 2020a , Elfiky, 2020b , Elfiky and Ibrahim, 2020 , Ibrahim et al., 2020a . WIFPWIQL peptide binds to GRP78 expressed in breast cancer cells surface in the breast and metastatic cells. Subtilase cytotoxin is a toxin from the AB5 toxins family. Subtilase cytotoxin composed of two subunits; SubA, which is responsible for the toxicity, and SubB, which is responsible for Subtilase cytotoxin internalization to the cell (Beddoe et al. , 2010) . SubA toxic effect is that it induces cell apoptosis by cleaving GRP78 between the amino acid residues Leu416 and Leu417 (Paton et al. , 2006) . As indicated, WIFPWIQL peptide binds to GRP78 over cancer cells (CS-GRP78). WIFPWIQL-SubA fusion resulting in an efficient anticancer agent. WIFPWIQL-SubA works simultaneously, WIFPWIQL is responsible for GRP78 recognition and J o u r n a l P r e -p r o o f internalization to the cancer cells, while SubA is responsible for the toxic effect on the cell by cleaving GRP78 inside the cell and thus leading to apoptosis . WIFPWIQL liposomes loaded with doxorubicin are used to target CS-GRP78 overexpressed over vascular endothelial growth factor (VEGF)-activated human umbilical vein endothelial cells (Katanasaka et al. , 2010) . WIFPWIQL bound N-(2hydroxypropyl)methacrylamide (HPMA) copolymer aminohexyl-geldanamycin conjugates were able to target CS-GRP78 and hence inhibit human prostate cancer cells (Larson et al. , 2010) . Additionally, the genetic engineered mung bean trypsin inhibitor (GBP-TI) that includes the WIFPWIQL peptide was able to induce apoptosis in colorectal cancer cells . BMTP78 composed of a peptide (WIFPWIQL) conjugated with proapoptotic moiety D(KLAKLAK)2 (Dobroff et al. , 2015) . GRP78 facilitates the internalization of BMTP78 into the cytoplasm. In vitro trials showed that BMTP78 induces apoptosis in human and mouse mammary cell lines. D(KLAKLAK)2 after internalization disrupts mitochondrial membrane permeability and thus kills the cell (Miao et al. , 2013) . BMTP78 induced dose-dependent cytotoxicity in human leukemia and lymphoma cell lines and acute myeloid leukemia patients (Araujo et al. , 2018 , Staquicini et al. , 2018 . Additionally, the GRP78 receptor/BMTP78 system was used to image breast tumors accurately. The adeno-associated virus-M13-derived phage (AAVP) can be used clinically to detect (imaging) and eradicate (targeted therapy) of Inflammatory breast cancer utilizing CS-GRP78 as a target (Dobroff et al., 2015) . GIRLRG is a peptide identified using phage display and binds GRP78. GIRLRG conjugated to paclitaxel-encapsulated nanoparticles, specifically targeted breast cancer and glioblastoma (Passarella et al. , 2010) . It was predicted (in silico) that GIRLRG binds to the ATPase domain of GRP78. GIRLRG conjugated with Poly Ethylene Glycol (PEG) can efficiently target different tumor cell lines including, heterotopic cervical (HT3), esophageal (OE33), pancreatic (BXPC3), lung (A549), and glioma (D54) (Kapoor et al. , 2016b) . Additionally, the radio-labeled 111 In-PEG-GIRLRG show specificity toward cervical, esophageal, pancreatic, lung, and brain tumors using SPECT imaging (Kapoor et al. , 2015) . e. VAP peptide SNTRVAP (VAP) is a peptide identified using the phage display technique, and it binds to GRP78 specifically (Mandelin et al. , 2015) . SNTRVAP coupling with a siRNA for GRP78, effectively downregulated its expression (Stone, 2017) . VAP modified micelles (RI-VAP (retro Inverso isomer of L-VAP) and D-VAP (retro isomer of L-VAP)) could effectively achieve glioma-targeted drug delivery, through GRP78. At the same time, it improved the therapeutic efficacy of paclitaxel for glioma (Ran et al. , 2017) . As we mentioned before, peptides could be used for drug delivery; it can be used as a carrier for radiolabels for imaging purposes, such as in the Positron Emission Tomography (PET), utilizing the same concept of targeting GRP78 over cancer cells. Radiolabeled Polyethylene glycol (PEG)-GIRLRG is used in targeting many cancers as heterotopic cervical, esophageal, pancreatic, lung, and glioma tumors (Kapoor et al. , 2016a Antibody (Ab), also called immunoglobulin (Ig), is a huge, Y-shaped protein produced mainly by plasma cells that are used by the immune system to neutralize pathogens such as viruses and bacteria. The pathogenic molecule that is recognized by the antibody is called an antigen (Janeway et al. , 1999) . The antibody bind with the antigen with a key-lock mechanism. Once the interaction established, the cell bearing the antigen triggers a response such as metabolic inhibition (Tesniere et al. , 2008) . MAb159 is a highly specific monoclonal antibody against the human GRP78 (Kd = 1.7 nM) (Gill et al. , 2020) . When administered, MAb159 found localized on the membranes of cancer cells but not normal cell-lines. Upon glucose starvation stress, MAb159 is found more abundant on the cell membrane (Wang and Kaufman, 2014) . As the CS-GRP78 is PI3K/AKT signaling upstream regulator through its interaction with Crypto and alpha2-macroglobulin over the cell membrane, it is required for these factors to activate the PI3K/AKT signaling (Liu et al. , 2013a) . Once bound to CS-GRP78, J o u r n a l P r e -p r o o f MAb159 endocytosed and modulate the PI3K pathway leading to inhibition for cell proliferation, tumor growth, and metastasis. At the same time, it enhances tumor cell death both in vitro and in vivo (Liu et al. , 2013b) . The efficacy of MAb159 was examined in various tumor xenograft models, including HT29 (colon cancer), H249 (small cell lung carcinoma), and A549 (lung adenocarcinoma). These cells have relatively higher (4.6%-9.4%) surface GRP78 expression compared to healthy cells . MAb159 treatment led to 50%, 58%, and 78% tumor growth inhibition in these models, respectively (Liu et al., 2013b) . The fully human monoclonal IgM antibody, SAM-6, was isolated from a gastric cancer patient, and it binds to an O-glycosylated form of GRP78. SAM-6 is internalized via endocytosis and is finally responsible for a lethal accumulation of oxidized lipoproteins followed by apoptosis in cancer cells (Vercauteren et al. , 2010) . SAM-6 not only bind to GRP78 on the cancer cell membrane but also it reduces the drug resistance and kills the cancer cell (Roller and Maddalo, 2013b) . PAT-SM6 specifically binds to primary multiple myelomas cells. Staining the cells by immunohistochemistry reveals binding to GRP78 of the PAT-SM6. This binding induces apoptosis and complement-dependent cytotoxicity (Lee, 2014) . α2M is associated with the N-terminal region of cell-surface GRP78. The binding activates Akt to suppress apoptotic pathways and promotes cell proliferation (Misra et al. , 2009a, Misra and Pizzo, 2004b) . J o u r n a l P r e -p r o o f The mouse monoclonal antibody C38 recognizes the C-terminal domain of the murine GRP78 exposed on the cell membrane. The binding induces inhibition of the Akt/PI3K proliferative pathway in melanoma cells (de Ridder et al. , 2012) . A comparable experiment done on melanoma mouse model show that the antibody C107 also bind to GRP78. In both experiments, the binging with the antibody decreases the tumor growth (de Ridder et al., 2012) . Anti GRP78-C-terminal domain (CTD) antibodies are tested against human prostate cancer cells. It significantly reduces tumor growth, inhibits cell proliferation, while promotes apoptosis. Besides, in the prostate cancer patients, the anti-CTD GRP78 antibody binds the cell-expressed, GRP78 in human prostate cancer cells Pizzo, 2004a, Misra et al. , 2009b) . GRP78, a master chaperone protein of the unfolded protein response, plays an essential role in cancer chemo-resistance and virulence of the pathogenic infections. Targeting GRP78 was utilized to defeat aggressive types of cancer like triple-negative breast cancer. Additionally, inhibiting GRP78 overexpressed in viral infections is suggested as a promising strategy to reduce the virulence of many viruses and fungal infections. The present review article summarizes the up to date targeting strategies used to inhibit cell-surface GRP78 illuminating the potential use of these strategies to defeat both cancer chemo-resistance and viral and fungal infections. J o u r n a l P r e -p r o o f Cell Surface GRP78: A Novel Regulator of Tissue Factor Procoagulant Activity. 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The international journal of biochemistry & cell biology Reconstructed mung bean trypsin inhibitor targeting cell surface GRP78 induces apoptosis and inhibits tumor growth in colorectal cancer Involvement of endoplasmic reticulum in paclitaxel-induced apoptosis Activation of the grp78 andgrp94 Promoters by Hepatitis C Virus E2 Envelope Protein The glucose-regulated proteins (GRP78 and GRP94): functions, gene regulation, and applications. Critical reviews in eukaryotic gene expression The endothelial cell receptor GRP78 is required for mucormycosis pathogenesis in diabetic mice Monoclonal Antibody against Cell Surface GRP78 as a Novel Agent in Suppressing PI3K/AKT Signaling, Tumor Growth, and Metastasis Monoclonal antibody against cell surface GRP78 as a novel agent in suppressing PI3K/AKT signaling, tumor growth, and metastasis Mechanistic studies of a peptidic GRP78 ligand for cancer cell-specific drug delivery. Molecular pharmaceutics GRP78/BiP is required for cell proliferation and protecting the inner cell mass from apoptosis during early mouse embryonic development GRP78 induced by estrogen plays a role in the chemosensitivity of endometrial cancer Selection and identification of ligand peptides targeting a model of castrate-resistant osteogenic prostate cancer and their receptors The roles of apoptosis, autophagy and unfolded protein response in arbovirus, influenza virus, and HIV infections Inhibition of Established Micrometastases by Targeted Drug Delivery via Cell Surface-Associated GRP78 Potentiation of signal transduction mitogenesis and cellular proliferation upon binding of receptor-recognized forms of α2-macroglobulin to 1-LN prostate cancer cells Ligation of cancer cell surface GRP78 with antibodies directed against its COOH-terminal domain up-regulates p53 activity and promotes apoptosis Ligation of cancer cell surface GRP78 with antibodies directed against its COOH-terminal domain up-regulates p53 activity and promotes apoptosis Potentiation of signal transduction mitogenesis and cellular proliferation upon binding of receptor-recognized forms of α2-macroglobulin to 1-LN prostate cancer cells GRP78 is an important host-factor for Japanese encephalitis virus entry and replication in mammalian cells Anti-proliferative and proapoptotic effects of (-)-epigallocatechin-3-gallate on human melanoma: possible implications for the chemoprevention of melanoma Elevated GRP78 expression is associated with poor prognosis in patients with pancreatic cancer Preliminary Virtual Screening Studies to Identify GRP78 Inhibitors Which May Interfere with SARS-CoV-2 Infection Targeted nanoparticles that deliver a sustained, specific release of Paclitaxel to irradiated tumors Pfaffenbach KT, Lee AS. The critical role of GRP78 in physiologic and pathologic stress. Current opinion in cell biology Heat shock protein 70 (Hsp70) is involved in the Zika virus cellular infection process GRP78, a chaperone with diverse roles beyond the endoplasmic reticulum GRP78 enabled micelle-based glioma targeted drug delivery Sulphureuine B, a drimane type sesquiterpenoid isolated from Laetiporus sulphureus induces apoptosis in glioma cells Heat shock protein 90 and heat shock protein 70 are components of dengue virus receptor complex in human cells Roller C, Maddalo D. The Molecular Chaperone GRP78/BiP in the Development of Chemoresistance: Mechanism and Possible Treatment In silico estrogen-like activity and in vivo osteoclastogenesis inhibitory effect of Cicer arietinum extract Coinduction of glucose-regulated proteins and doxorubicin resistance in Chinese hamster cells HSPA5 is an essential host factor for Ebola virus infection Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface Phytochemicals Targeting Endoplasmic Reticulum Stress to Inhibit Cancer Cell Proliferation Therapeutic targeting of membrane-associated GRP78 in leukemia and lymphoma: preclinical efficacy in vitro and formal toxicity study of BMTP-78 in rodents and primates On target -theranostic imaging for aggressive disease Galangin inhibits proliferation of hepatocellular carcinoma cells by inducing endoplasmic reticulum stress Immunogenic cancer cell death: a key-lock paradigm The interplay between GRP78 expression and Akt activation in human colon cancer cells under celecoxib treatment 3,4-dihydroxybenzalacetone and caffeic acid phenethyl ester induce preconditioning ER stress and autophagy in SH-SY5Y cells Cell Surface GRP78: Anchoring and Translocation Mechanisms and Therapeutic Potential in Cancer. Cell Surface GRP78, a New Paradigm Chapter 3 -Cell Surface GRP78: Anchoring and Translocation Mechanisms and Therapeutic Potential in Cancer The use of inhibitors to study endocytic pathways of gene carriers: optimization and pitfalls. Molecular therapy : the journal of the American Society of Gene Therapy Glucose-Regulated Protein 78 (Grp78) Confers Chemoresistance to Tumor Endothelial Cells under Acidic Stress Control of inducible chemoresistance: enhanced anti-tumor therapy through increased apoptosis by inhibition of NF-kappaB The impact of the endoplasmic reticulum protein-folding environment on cancer development Role of the unfolded protein response regulator GRP78/BiP in development, cancer, and neurological disorders Mechanism study of peptide GMBP1 and its receptor GRP78 in modulating gastric cancer MDR by iTRAQ-based proteomic analysis GnRH agonists induce endometrial epithelial cell apoptosis via GRP78 down-regulation Hydroxytyrosol and olive leaf extract exert cardioprotective effects by inhibiting GRP78 and CHOP expression Glucose regulated protein 78 (GRP78) inhibits apoptosis and attentinutes chemosensitivity of gemcitabine in breast cancer cell via AKT/mitochondrial apoptotic pathway Genistein induces apoptosis in human hepatocellular carcinomas via interaction of endoplasmic reticulum stress and mitochondrial insult A cell-penetrating peptidic GRP78 ligand for tumor cell-specific prodrug therapy GRP78 promotes the invasion of pancreatic cancer cells by FAK and JNK. Molecular and cellular biochemistry New insights into the mechanisms of gastric cancer multidrug resistance and future perspectives Association of elevated GRP78 expression with increased lymph node metastasis and poor prognosis in patients with gastric cancer Involvement of GRP78 in the resistance of ovarian carcinoma cells to paclitaxel Design, purification and assessment of GRP78 binding peptidelinked Subunit A of Subtilase cytotoxic for targeting cancer cells Cell surface relocalization of the endoplasmic reticulum chaperone and unfolded protein response regulator GRP78/BiP Noninvasive Classification of Human Triple Negative Breast Cancer by PET Imaging with GRP78-Targeted Molecular Probe [68 Ga] DOTA-VAP Photooxidative damage in retinal pigment epithelial cells via GRP78 and the protective role of grape skin polyphenols All the authors declare no conflict of interest for this work.