AIoT enabled resampling filter for temperature extraction of the Brillouin gain spectrum.

The Artificial Intelligence of Things (AIoT) turns passive fiber sensors into learning machines. It can be used to integrate intelligent nodes into a multi-dimensional sensing system. In this study, the temperature measurement system based on Brillouin Gain Spectrum (BGS) test setup is creatively implemented with the AIoT architecture; the training and testing stages of the neural network are divided into different layers of equipment to improve performance and reduce the network traffic. To enable the lightweight and low-power consumption edge computing device to extract accurate temperature from the BGS during testing, we have integrated the post-processing method inspired by curve fitting into the machine learning and proposed the efficient digital resampling filter. The resampling filter approach is achieved by the peak range algorithm with Gauss differential operator and digital band-pass filter. The evaluation results for different methods on the BGS datasets show the superior performance of our approach. Notably, the approach can increase temperature extraction accuracy and compress the sampling data. The RMSEA of the extraction temperature is 0.5635, which can bring an almost 21% accuracy increase over the classic method. Compared with the classic method of processing the original data on the same hardware platform, the amount of data post-processed by the filter is reduced by 75%; the time consumption is reduced by 22%.

miR-219a-5p inhibits the pyroptosis in knee osteoarthritis by inactivating the NLRP3 signaling via targeting FBXO3.

PURPOSE: This work was to identify the function and mechanism of miR-219a-5p in regulating knee osteoarthritis (KOA). METHODS: Rat fibroblast-like synoviocytes (FLSs) were isolated to construct KOA cell model by lipopolysaccharide and adenosine triphosphate treatment. miR-219a-5p and FBXO3 expression in FLSs was modulated by transfection. Flow cytometry was executed to research FLSs apoptosis. Caspase-1 and IL-1ß expression in FLSs was researched by immunofluorescence. The binding between miR-219a-5p and FBXO3 was identified by dual luciferase reporter gene assay. KOA rat model and miR-219a-5p up-modulation KOA rat model were constructed. Step size of rats was analyzed. Knee joints of rats were experienced Safranin O-fast green staining to evaluate the knee joint injury. FBXO3, pyroptosis-associated proteins, and IL-1ß and IL-18 expression in FLSs and articular cartilage tissues of rats were assessed by Western blot, qRT-PCR and Enzyme-linked immunosorbent assay. RESULTS: KOA cell model had higher apoptosis percentage, expression of pyroptosis-associated proteins, and IL-1ß and IL-18 level. miR-219a-5p up-modulation decreased the above indicators, whereas miR-219a-5p down-modulation increased the above indicators. FBXO3 expression was directly repressed by miR-219a-5p. Loss of FBXO3 suppressed the above indicators. FBXO3 counteracted the suppression of miR-219a-5p on the above indicators. miR-219a-5p agomir attenuated knee joint injury, increased step size of KOA rats, and reduced FBXO3, pyroptosis-associated proteins and level of IL-1ß and IL-18 in the articular cartilage tissues of KOA rats. CONCLUSION: miR-219a-5p suppressed the pyroptosis in KOA by inactivating the NLRP3 signaling via targeting FBXO3, which might be a promising target for ameliorating KOA in the clinic.

Sweep frequency method with variance weight probability for temperature extraction of the Brillouin gain spectrum based on an artificial neural network.

The development of optical fiber sensors has led to the possibility of accumulating vast, real-time databases of acoustic and other measurements throughout fiber networks, which brings even more widespread concern on improving the sampling effectiveness. In this paper, we present two kinds of sweep frequency methods based on using a neural network to extract temperature from the Brillouin gain spectrum (BGS). Gauss centralization and variance weight probability methods are proposed to compare with the uniform sweep frequency method. By analyzing formulas of the ideal BGS model, we find the gain near the peak of Brillouin gain spectrum has greater correlation with temperature extraction than other positions. Therefore, the Gaussian centralized sweep method is proposed. We further investigate the variation of the weights in the neural network and Brillouin data distribution in different positions and find that the variance is positively correlated with the weights in hidden layers. So, we propose the sweep frequency method based on variance weight probability and make a complement to interpret the rationality of this method in neural network. In all the aforementioned approaches, 281 points are obtained between the 9.07 GHz to 9.35 GHz range under the same condition. The data of each method is trained ten times and tested through the same neural network structure. All the RMSE of each test stage covers all data collecting the passage. The result shows that the RMSE of variance weight probability sweep frequency method is 0.5277, which is superior to the Gauss centralization sweep frequency method that was 0.6864 and the uniform sweep frequency method that was 0.9140.

KIAA1429 regulates cell proliferation by targeting c-Jun messenger RNA directly in gastric cancer.

N6-methyladenosine (m6A) modification regulatory proteins are involved in the development of many types of cancer. KIAA1429 serves as a scaffold in bridging the catalytic core components of the m6A methyltransferase complex. The role of KIAA1429 in gastric cancer and its related mechanism has not been reported upon. The expression of KIAA1429 was detected in human gastric cancer tissues and cell lines by quantitative real-time polymerase chain reaction and western blot. The effects of KIAA1429 on gastric cancer proliferation were evaluated by cell counting kit assays, colony formation assays, flow cytometry assay, and in vivo experiments with nude mice. And messenger RNA (mRNA) high-throughput sequencing, RNA immunoprecipitation assay (RIP), luciferase assay, and a rescue experiment were used to identify the relationship between KIAA1429 and its specific targeted gene, c-Jun. We found that KIAA1429 was upregulated in gastric cancer tissues, and expressed lower in adjacent tissues. The upregulated KIAA1429 promoted proliferation and downregulated KIAA1429 was proved to inhibit proliferation of gastric cancer in vitro and in vivo. Then, we identified the potential KIAA1429 regulating gene as c-Jun by mRNAs high-throughput sequencing and RIP assay. By luciferase assay, we verified that KIAA1429 regulated the expression of c-Jun in an m6A-independent manner. Finally, the overexpression of c-Jun rescued the inhibition of proliferation caused by KIAA1429 knockdown in gastric cancer cells. KIAA1429 could act as an oncogene in gastric cancer by stabilizing c-Jun mRNA in an m6A-independent manner. This highlights the functional role for KIAA1429 as a potential prognostic biomarker and therapeutic target in gastric cancer.

Preclinical Efficacy of Anti-RON Antibody-Drug Conjugate Zt/g4-MMAE for Targeted Therapy of Pancreatic Cancer Overexpressing RON Receptor Tyrosine Kinase.

Aberrant expression of the RON receptor tyrosine kinase, a cell surface protein, is a pathogenic feature in pancreatic cancer, which renders it a drug target for targeted therapy. Nevertheless, development of therapeutics targeting RON for pancreatic cancer therapy is hampered due to the lack of full addiction by pancreatic cancer cells to RON signaling for growth and survival. Here we describe a novel strategy using anti-RON antibody-directed drug delivery in the form of an antibody-drug conjugate for inhibition and/or eradication of pancreatic cancers. Monoclonal antibody Zt/g4 specific to the RON Sema domain was selected as the drug carrier based on its ability to induce robust RON internalization. Conjugation of Zt/g4 with monomethyl auristatin E, designated as Zt/g4-MMAE, was achieved through a protease-sensitive dipeptide linker to reach a drug to antibody ratio of 3.29:1. Zt/g4-MMAE was stable in human plasma with a dissociation rate less than 4% within a 10 day period. In vitro, Zt/g4-MMAE rapidly induced RON internalization, resulting in cell cycle arrest followed by massive cell death. The maximal effect was seen in pancreatic cancer cells with more than 10 000 receptor molecules per cell. Zt/g4-MMAE also synergized in vitro with chemotherapeutics including gemcitabine, 5-fluorouracil, and oxaliplatin to further reduce PDAC cell viability. In vivo, Zt/g4-MMAE exerts a long-lasting activity, which not only inhibited but also eradicated pancreatic xenograft tumors. These finding indicate that Zt/g4-directed drug delivery is highly effective for eradicating pancreatic tumors. Thus, Zt/g4-MMAE is a novel biotherapeutic with potential for therapy of RON-expressing pancreatic malignancies.

Ribosomal proteins and human diseases: pathogenesis, molecular mechanisms, and therapeutic implications.

Ribosomes are essential components of the protein synthesis machinery. The process of ribosome biogenesis is well organized and tightly regulated. Recent studies have shown that ribosomal proteins (RPs) have extraribosomal functions that are involved in cell proliferation, differentiation, apoptosis, DNA repair, and other cellular processes. The dysfunction of RPs has been linked to the development and progression of hematological, metabolic, and cardiovascular diseases and cancer. Perturbation of ribosome biogenesis results in ribosomal stress, which triggers activation of the p53 signaling pathway through RPs-MDM2 interactions, resulting in p53-dependent cell cycle arrest and apoptosis. RPs also regulate cellular functions through p53-independent mechanisms. We herein review the recent advances in several forefronts of RP research, including the understanding of their biological features and roles in regulating cellular functions, maintaining cell homeostasis, and their involvement in the pathogenesis of human diseases. We also highlight the translational potential of this research for the identification of molecular biomarkers, and in the discovery and development of novel treatments for human diseases.

Identification of a new class of MDM2 inhibitor that inhibits growth of orthotopic pancreatic tumors in mice.

BACKGROUND & AIMS: The oncogene MDM2, which encodes an E3 ubiquitin ligase, is overexpressed in pancreatic cancers and is therefore a therapeutic target. Current inhibitors of MDM2 target the interaction between MDM2 and P53; these would have no effect on cancer cells that do not express full-length P53, including many pancreatic cancer cells. We searched for a compound that specifically inhibits MDM2 itself. METHODS: We performed a virtual screen and structure-based design to identify specific inhibitors of MDM2. We tested the activities of compounds identified on viability, proliferation, and protein levels of HPAC, Panc-1, AsPC-1, and Mia-Paca-2 pancreatic cancer cell lines. We tested whether intraperitoneal injections of one of the compounds identified affected growth of xenograft tumors from Panc-1 cells, or orthotopic tumors from Panc-1 and AsPC-1 cells (injected into pancreata), in nude mice. RESULTS: We identified a compound, called SP141, which bound directly to MDM2, promoting its auto-ubiquitination and degradation by the proteasome. The compound reduced levels of MDM2 in pancreatic cancer cell lines, as well as their proliferation, with 50% inhibitory concentrations <0.5 µM (0.38-0.50 µM). Increasing concentrations of SP141 induced increasing levels of apoptosis and G2-M-phase arrest of pancreatic cancer cell lines, whether or not they expressed functional P53. Injection of nude mice with SP141 (40 mg/kg/d) inhibited growth of xenograft tumors (by 75% compared with control mice), and led to regression of orthotopic tumors. CONCLUSIONS: In a screen for specific inhibitors of MDM2, we identified a compound called SP141 that reduces levels of MDM2 in pancreatic cancer cell lines, as well as their proliferation and ability to form tumors in nude mice. SP141 is a new class of MDM2 inhibitor that promotes MDM2 auto-ubiquitination and degradation. It might be further developed as a therapeutic agent for pancreatic cancer.

Development and validation of a rapid HPLC method for quantitation of SP-141, a novel pyrido[b]indole anticancer agent, and an initial pharmacokinetic study in mice.

There is an increasing interest in targeting the MDM2 oncogene for cancer therapy. SP-141, a novel designed small molecule MDM2 inhibitor, exerts excellent in vitro and in vivo anticancer activity. To facilitate the preclinical development of this candidate anticancer agent, we have developed an HPLC method for the quantitative analysis of SP-141. The method was validated to be precise, accurate, and specific, with a linear range of 16.2-32,400 ng/mL in plasma, 16.2-6480 ng/mL in homogenates of brain, heart, liver, kidneys, lungs, muscle and tumor, and 32.4-6480 ng/mL in spleen homogenates. The lower limit of quantification was 16.2 ng/mL in plasma and all the tissue homogenates, except for spleen homogenates, where it was 32.4 ng/mL. The intra- and inter-assay precisions (coefficient of variation) were between 0.86 and 13.39%, and accuracies (relative errors) ranged from -8.50 to 13.92%. The relative recoveries were 85.6-113.38%. SP-141 was stable in mouse plasma, modestly plasma bound and metabolized by S9 microsomal enzymes. We performed an initial pharmacokinetic study in tumor-bearing nude mice, demonstrating that SP-141 has a short half-life in plasma and wide tissue distribution. In summary, this HPLC method can be used in future preclinical and clinical investigations of SP-141.

Classifications of clinical and bowel morphological changes and their relationship with characteristics of patients with incarcerated groin hernias.

BACKGROUND: There is currently no grading standard for the degree of clinical and bowel morphological changes. The objective of this study was to define clinical and bowel morphological classifications and investigate the possible relationship with the characteristics of patients with incarcerated groin hernias. MATERIAL AND METHODS: We retrospectively studied 195 patients who underwent emergency hernia repair with simultaneous bowel resection between January 1992 and January 2012. We classified the degree of clinical and bowel morphological changes into 3 grades based on the incarceration time, intestinal morphology after damage, hernia sac integrity, degree of inflammation, and the presence/absence of bacterial growth, peritonitis signs, mechanical obstruction, cellulitis, and systemic shock. We also recorded patient characteristics and analyzed their relationships with these degrees according to our grading system. RESULTS: We identified 134, 42, and 19 cases of Grades I, II, and III of clinical and bowel morphological changes, respectively. Pearson's chi-squared tests revealed that advanced age (P=0.001), presence of comorbid disease (P=0.002), and high American Society of Anesthesiologists (ASA) score (P=0.017) were related to the degree. Morbidity and mortality also showed significant relationships with the degree (P<0.001, P=0.005, respectively), especially with regard to post-operative infection. CONCLUSIONS: The proposed 3-stage classifications of clinical and bowel morphological changes can be used to objectively reflect the degree of bowel damage. Greater levels of the changes were associated with higher incidences of complications and increased mortality, especially for older patients with comorbid diseases and poor ASA scores. Urgent surgery should be performed to avoid bowel damage exacerbation.

Pharmaceutical innovation and advanced biotechnology in the biotech-pharmaceutical industry for antibody-drug conjugate development.

Antibody-drug conjugates (ADCs), from prototypes in the 1980s to first- and second-generation products in the 2000s, and now in their multiformats, have progressed tremendously to meet oncological challenges. Currently, 13 ADCs have been approved for medical practice, with over 200 candidates in clinical trials. Moreover, ADCs have evolved into different formats, including bispecific ADCs, probody-drug conjugates, pH-responsive ADCs, target-degrading ADCs, and immunostimulating ADCs. Technologies from biopharmaceutical industries have a crucial role in the clinical transition of these novel biotherapeutics. In this review, we highlight several features contributing to the prosperity of bioindustrial ADC development. Various proprietary technologies from biopharmaceutical companies are discussed. Such advances in biopharmaceutical industries are the backbone for the success of ADCs in development and clinical application.

Combination of Albumin/Fibrinogen Ratio and Admission Hunt-Hess Scale Score as an Independent Predictor of Clinical Outcome in Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND: The albumin/fibrinogen ratio (AFR) is an independent predictor of clinical outcomes of some diseases; however, the prognostic value of AFR and the admission Hunt-Hess (HH) score is still unclear for patients with an aneurysmal subarachnoid hemorrhage (aSAH). This study aimed to assess the relationship between the AFR-HH score and 6-month outcomes of aSAH patients. METHODS: The clinical characteristics of aSAH patients admitted to our department between December 2017 and December 2021 were retrospectively analyzed. The candidate risk factors were screened using univariate regression analysis, and the independence of the resultant risk factors was evaluated by binary logistic regression analysis. The predictive value of the combined AFR and HH score for unfavorable outcomes was assessed using receiver operating characteristic curve analysis. RESULTS: A total of 112 aSAH patients were included. Binary logistic regression analysis showed the perioperative period AFR, Glasgow coma scale score, and admission HH score were independent risk factors for unfavorable outcomes for aSAH patients. The receiver operating characteristic curve analysis showed the predictive capacity of AFR plus the admission HH score outperformed the AFR, Glasgow coma scale score, and admission HH scale alone and the combination of the AFR and Glasgow coma scale score. CONCLUSIONS: A low AFR during the perioperative period is associated with unfavorable outcomes for aSAH patients at 6 months. The combination of the AFR and admission HH scale score provides superior predictive capacity to either the AFR or HH scale score alone.

RON Receptor Tyrosine Kinase in Tumorigenic Stemness as a Therapeutic Target of Antibody-drug Conjugates for Eradication of Triple-negative Breast Cancer Stem Cells.

BACKGROUND: Cancer stem-like cells in triple-negative breast cancer (TNBC-SLCs) are the tumorigenic core for malignancy. Aberrant expression of the RON receptor tyrosine kinase has implications in TNBC tumorigenesis and malignancy. OBJECTIVE: In this study, we identified the RON receptor as a pathogenic factor contributing to TNBC cell stemness and validated anti-RON antibody-drug conjugate Zt/g4-MMAE for eradication of RONexpressing TNBC-SLCs. METHODS: Immunofluorescence and Western blotting were used for analyzing cellular marker expression. TNBC-SLCs were isolated by magnetic-immunofluorescence cell-sorting techniques. Spheroids were generated using the ultralow adhesion culture methods. Levels of TNBC-SLC chemosensitivity were determined by MTS assays. TNBC-SLC mediated tumor growth was determined in athymic nude mice. The effectiveness of Zt/g4-induced RON internalization was measured by immunofluorescence analysis. Efficacies of Zt/g4-MMAE in killing TNBC-SLCs in vitro and in eradicating TNBC-SLCmediated tumors were determined in mouse models. All data were statistically analyzed using the GraphPad Prism 7 software. RESULTS: Increased RON expression existed in TNBC-SLCs with CD44+/CD24- phenotypes and ALDH activities and facilitated epithelial to mesenchymal transition. RON-positive TNBC-SLCs enhanced spheroid-formatting capability compared to RON-negative TNBC-SLCs, which were sensitive to small molecule kinase inhibitor BMS-777607. Increased RON expression also promoted TNBC-SLC chemoresistance and facilitated tumor growth at an accelerated rate. In vitro, Zt/g4-MMAE caused massive TNBC-SLC death with an average IC50 value of ~1.56 µg per/ml and impaired TNBC cell spheroid formation. In mice, Zt/g4-MMAE effectively inhibited and/or eradicated TNBC-SLC mediated tumors in a single agent regimen. CONCLUSION: Sustained RON expression contributes to TNBC-SLC tumorigenesis. Zt/g4-MMAE is found to be effective in vivo in killing TNBC-SLC-mediated xenograft tumors. Our findings highlight the feasibility of Zt/g4-MMAE for the eradication of TNBC-SLCs in the future.

Preclinical evaluation of anticancer efficacy and pharmacological properties of FBA-TPQ, a novel synthetic makaluvamine analog.

We have recently designed and synthesized a novel iminoquinone anticancer agent, 7-(4-fluorobenzylamino)-1,3,4,8-tetrahydropyrrolo[4,3,2-de]quinolin-8(1H)-one (FBA-TPQ) and initiated its preclinical development. Herein we investigated its efficacy, safety, and pharmacokinetics in in vitro and in vivo models of human pancreatic cancer. Our results demonstrated that FBA-TPQ inhibited pancreatic cancer cell growth, induced apoptosis, and caused cell cycle arrest in vitro. It inhibited the growth of xenograft tumors with minimal host toxicity. To facilitate future preclinical and clinical development of the agent, we also developed and validated a Rapid Resolution Liquid Chromatography (RRLC) method for quantitative analysis of FBA-TPQ in plasma and tissue samples. The method was found to be precise, accurate, and specific. Using this method, we carried out in vitro and in vivo evaluations of the pharmacological properties of FBA-TPQ, including stability in plasma, plasma protein binding, metabolism by S9 enzymes, plasma pharmacokinetics, and tissue distribution. Our results indicate that FBA-TPQ is a potential therapeutic agent for pancreatic cancer, providing a basis for future preclinical and clinical development.

[Experimental study of constructing vertebral canal on rabbit model with bone marrow mesenchymal stem cells].

OBJECTIVE: To construct the artificial lamina of vertebral arch with bone marrow mesenchymal stem cells transplanted in collagen sponge on a rabbit model and observe the growth of new bone. METHODS: To draw out the bone marrow blood from the femur of 2 weeks old rabbit and get the bone marrow mesenchymal stem cells by centrifugal and adhesive effect. To induce the MSCs to osteoblasts and transplant the induced cells in collagen sponge to construct the tissue engineering bone. To divide 48 rabbits into 3 groups randomly, namely group A, group B and group C. All of the rabbits are taken laminectomy in L6, and to group B and C, collagen sponge and tissue engineering bone are implanted in the operation area respectively. The artificial lamina of vertebral arch is determined qualitatively and quantitatively by methods including imageology and histomorphometry. RESULTS: The artificial lamina of vertebral arch is successfully constructed 4 weeks after operation in group C, CT examination at 4 weeks shows that new lamina of vertebral arch is formed, and the vertebral canal is intact. CONCLUSIONS: The artificial lamina of vertebral arch can be constructed successfully with the usage of tissue engineering bone transplanted bone marrow mesenchymal stem cells.

Antibody-drug Conjugate PCMC1D3-Duocarmycin SA as a Novel Therapeutic Entity for Targeted Treatment of Cancers Aberrantly Expressing MET Receptor Tyrosine Kinase.

BACKGROUND: Aberrant expression of the MET receptor tyrosine kinase is an oncogenic determinant and a drug target for cancer therapy. Currently, antibody-based biotherapeutics targeting MET are under clinical trials. OBJECTIVE: Here, we report the preclinical and therapeutic evaluation of a novel anti-MET antibody- drug conjugate PCMC1D3-duocarmycin SA (PCMC1D3-DCM) for targeted cancer therapy. METHODS: The monoclonal antibody PCMC1D3 (IgG1a/κ), generated by a hybridoma technique and specific to one of the MET extracellular domains, was selected based on its high specificity to human MET with a binding affinity of 1.60 nM. PCMC1D3 was conjugated to DCM via a cleavable valine-citrulline dipeptide linker to form an antibody-drug conjugate with a drug-to-antibody ratio of 3.6:1. PCMC1D3-DCM in vitro rapidly induced MET internalization with an internalization efficacy ranging from 6.5 to 17.2h dependent on individual cell lines. RESULTS: Studies using different types of cancer cell lines showed that PCMC1D3-DCM disrupted the cell cycle, reduced cell viability, and caused massive cell death within 96h after treatment initiation. The calculated IC50 values for cell viability reduction were 1.5 to 15.3 nM. Results from mouse xenograft tumor models demonstrated that PCMC1D3-DCM in a single dose injection at 10 mg/kg body weight effectively delayed xenograft tumor growth up to two weeks without signs of tumor regrowth. The calculated tumoristatic concentration, a minimal dose required to balance tumor growth and inhibition, was around 2 mg/kg body weight. Taken together, PCMC1D3-DCM was effective in targeting the inhibition of tumor growth in xenograft models. CONCLUSION: This work provides the basis for the development of humanized PCMC1D3-DCM for MET-targeted cancer therapy in the future.

Ribosomal protein S6 kinase (RSK)-2 as a central effector molecule in RON receptor tyrosine kinase mediated epithelial to mesenchymal transition induced by macrophage-stimulating protein.

BACKGROUND: Epithelial to mesenchymal transition (EMT) occurs during cancer cell invasion and malignant metastasis. Features of EMT include spindle-like cell morphology, loss of epithelial cellular markers and gain of mesenchymal phenotype. Activation of the RON receptor tyrosine kinase by macrophage-stimulating protein (MSP) has been implicated in cellular EMT program; however, the major signaling determinant(s) responsible for MSP-induced EMT is unknown. RESULTS: The study presented here demonstrates that RSK2, a downstream signaling protein of the Ras-Erk1/2 pathway, is the principal molecule that links MSP-activated RON signaling to complete EMT. Using MDCK cells expressing RON as a model, a spindle-shape based screen was conducted, which identifies RSK2 among various intracellular proteins as a potential signaling molecule responsible for MSP-induced EMT. MSP stimulation dissociated RSK2 with Erk1/2 and promoted RSK2 nuclear translocation. MSP strongly induced RSK2 phosphorylation in a dose-dependent manner. These effects relied on RON and Erk1/2 phosphorylation, which is significantly potentiated by transforming growth factor (TGF)-ß1, an EMT-inducing cytokine. Specific RSK inhibitor SL0101 completely prevented MSP-induced RSK phosphorylation, which results in inhibition of MSP-induced spindle-like morphology and suppression of cell migration associated with EMT. In HT-29 cancer cells that barely express RSK2, forced RSK2 expression results in EMT-like phenotype upon MSP stimulation. Moreover, specific siRNA-mediated silencing of RSK2 but not RSK1 in L3.6pl pancreatic cancer cells significantly inhibited MSP-induced EMT-like phenotype and cell migration. CONCLUSIONS: MSP-induced RSK2 activation is a critical determinant linking RON signaling to cellular EMT program. Inhibition of RSK2 activity may provide a therapeutic opportunity for blocking RON-mediated cancer cell migration and subsequent invasion.

The monoclonal antibody Zt/f2 targeting RON receptor tyrosine kinase as potential therapeutics against tumor growth-mediated by colon cancer cells.

BACKGROUND: Overexpression of the RON receptor tyrosine kinase contributes to epithelial cell transformation, malignant progression, and acquired drug resistance. RON also has been considered as a potential target for therapeutic intervention. This study determines biochemical features and inhibitory activity of a mouse monoclonal antibody (mAb) Zt/f2 in experimental cancer therapy. RESULTS: Zt/f2 is a mouse IgG2a mAb that is highly specific and sensitive to human RON and its oncogenic variants such as RON160 (ED(50) = 2.3 nmol/L). Receptor binding studies revealed that Zt/f2 interacts with an epitope(s) located in a 49 amino acid sequence coded by exon 11 in the RON ß-chain extracellular sequences. This sequence is critical in regulating RON maturation and phosphorylation. Zt/f2 did not compete with ligand macrophage-stimulating protein for binding to RON; however, its engagement effectively induced RON internalization, which diminishes RON expression and impairs downstream signaling activation. These biochemical features provide the cellular basis for the use of Zt/f2 to inhibit tumor growth in animal model. Repeated administration of Zt/f2 as a single agent into Balb/c mice results in partial inhibition of tumor growth caused by transformed NIH-3T3 cells expressing oncogenic RON160. Colon cancer HT-29 cell-mediated tumor growth in athymic nude mice also was attenuated following Zt/f2 treatment. In both cases, ~50% inhibition of tumor growth as measured by tumor volume was achieved. Moreover, Zt/f2 in combination with 5-fluorouracil showed an enhanced inhibition effect of ~80% on HT-29 cell-mediated tumor growth in vivo. CONCLUSIONS: Zt/f2 is a potential therapeutic mAb capable of inhibiting RON-mediated oncogenesis by colon cancer cells in animal models. The inhibitory effect of Zt/f2 in vivo in combination with chemoagent 5-fluorouracil could represent a novel strategy for future colon cancer therapy.

Sustained expression of the RON receptor tyrosine kinase by pancreatic cancer stem cells as a potential targeting moiety for antibody-directed chemotherapeutics.

Cancer stem cells (CSCs) contribute to pancreatic cancer tumorigenesis through tumor initiation, drug resistance, and metastasis. Currently, therapeutics targeting pancreatic CSCs are under intensive investigation. This study tested a novel strategy that utilizes the RON receptor as a drug delivery moiety for increased therapeutic activity against pancreatic CSCs. CD24(+)CD44(+)ESA(+) triple-positive pancreatic CSCs (CSCs(+24/44/ESA)) were obtained from spheroids of pancreatic L3.6pl cancer cells by sequential magnetic cell sorting methods. These cells displayed a spherical growth pattern, expressed the unique self-renewal marker Bmi-1, redifferentiated into an epithelial phenotype, acquired an epithelial to mesenchymal phenotype, and caused tumor formation in animal models. Among several receptor tyrosine kinases examined, RON was highly expressed and sustained by CSCs(+24/44/ESA). This feature provided the cellular basis for validating the therapeutic effectiveness of anti-RON antibody Zt/c9-directing doxorubicin-immunoliposomes (Zt/c9-Dox-IL). Zt/c9-Dox-IL specifically interacted with CSCs(+24/44/ESA) and rapidly caused RON internalization, which led to the uptake of liposome-coated Dox. Moreover, Zt/c9-Dox-IL was effective in reducing viability of L3.6pl cells and CSCs(+24/44/ESA). The IC(50) values between free Dox (62.0 ± 3.1 µM) and Zt/c9-Dox-IL (95.0 ± 6.1 µM) treated CSCs(+24/44/ESA) were at relatively comparable levels. In addition, Zt/c9-Dox-IL in combination with small molecule inhibitors lapatinib, sunitinib, or dasatinib further reduced the viability of CSCs(+24/44/ESA). In conclusion, RON expression by CSCs(+24/44/ESA) is a suitable molecule for the targeted delivery of chemoagents. The anti-RON antibody-directed delivery of chemotherapeutics is effective in reducing viability of pancreatic CSCs.

Oncogenic mechanism-based pharmaceutical validation of therapeutics targeting MET receptor tyrosine kinase.

Aberrant expression and/or activation of the MET receptor tyrosine kinase is characterized by genomic recombination, gene amplification, activating mutation, alternative exon-splicing, increased transcription, and their different combinations. These dysregulations serve as oncogenic determinants contributing to cancerous initiation, progression, malignancy, and stemness. Moreover, integration of the MET pathway into the cellular signaling network as an addiction mechanism for survival has made this receptor an attractive pharmaceutical target for oncological intervention. For the last 20 years, MET-targeting small-molecule kinase inhibitors (SMKIs), conventional therapeutic monoclonal antibodies (TMABs), and antibody-based biotherapeutics such as bispecific antibodies, antibody-drug conjugates (ADC), and dual-targeting ADCs have been under intensive investigation. Outcomes from preclinical studies and clinical trials are mixed with certain successes but also various setbacks. Due to the complex nature of MET dysregulation with multiple facets and underlying mechanisms, mechanism-based validation of MET-targeting therapeutics is crucial for the selection and validation of lead candidates for clinical trials. In this review, we discuss the importance of various types of mechanism-based pharmaceutical models in evaluation of different types of MET-targeting therapeutics. The advantages and disadvantages of these mechanism-based strategies for SMKIs, conventional TMABs, and antibody-based biotherapeutics are analyzed. The demand for establishing new strategies suitable for validating novel biotherapeutics is also discussed. The information summarized should provide a pharmaceutical guideline for selection and validation of MET-targeting therapeutics for clinical application in the future.

Pharmaceutical strategies in the emerging era of antibody-based biotherapeutics for the treatment of cancers overexpressing MET receptor tyrosine kinase.

Pharmaceutical innovation in the development of novel antibody-based biotherapeutics with increased therapeutic indexes makes MET-targeted cancer therapy a clinical reality.