Off-targets of BRAF inhibitors disrupt endothelial signaling and vascular barrier function.

Targeted therapies against mutant BRAF are effectively used in combination with MEK inhibitors (MEKi) to treat advanced melanoma. However, treatment success is affected by resistance and adverse events (AEs). Approved BRAF inhibitors (BRAFi) show high levels of target promiscuity, which can contribute to these effects. The blood vessel lining is in direct contact with high plasma concentrations of BRAFi, but effects of the inhibitors in this cell type are unknown. Hence, we aimed to characterize responses to approved BRAFi for melanoma in the vascular endothelium. We showed that clinically approved BRAFi induced a paradoxical activation of endothelial MAPK signaling. Moreover, phosphoproteomics revealed distinct sets of off-targets per inhibitor. Endothelial barrier function and junction integrity were impaired upon treatment with vemurafenib and the next-generation dimerization inhibitor PLX8394, but not with dabrafenib or encorafenib. Together, these findings provide insights into the surprisingly distinct side effects of BRAFi on endothelial signaling and functionality. Better understanding of off-target effects could help to identify molecular mechanisms behind AEs and guide the continued development of therapies for BRAF-mutant melanoma.

Targeting mTOR and survivin concurrently potentiates radiation therapy in renal cell carcinoma by suppressing DNA damage repair and amplifying mitotic catastrophe.

BACKGROUND: Renal cell carcinoma (RCC) was historically considered to be less responsive to radiation therapy (RT) compared to other cancer indications. However, advancements in precision high-dose radiation delivery through single-fraction and multi-fraction stereotactic ablative radiotherapy (SABR) have led to better outcomes and reduced treatment-related toxicities, sparking renewed interest in using RT to treat RCC. Moreover, numerous studies have revealed that certain therapeutic agents including chemotherapies can increase the sensitivity of tumors to RT, leading to a growing interest in combining these treatments. Here, we developed a rational combination of two radiosensitizers in a tumor-targeted liposomal formulation for augmenting RT in RCC. The objective of this study is to assess the efficacy of a tumor-targeted liposomal formulation combining the mTOR inhibitor everolimus (E) with the survivin inhibitor YM155 (Y) in enhancing the sensitivity of RCC tumors to radiation. EXPERIMENTAL DESIGN: We slightly modified our previously published tumor-targeted liposomal formulation to develop a rational combination of E and Y in a single liposomal formulation (EY-L) and assessed its efficacy in RCC cell lines in vitro and in RCC tumors in vivo. We further investigated how well EY-L sensitizes RCC cell lines and tumors toward radiation and explored the underlying mechanism of radiosensitization. RESULTS: EY-L outperformed the corresponding single drug-loaded formulations E-L and Y-L in terms of containing primary tumor growth and improving survival in an immunocompetent syngeneic mouse model of RCC. EY-L also exhibited significantly higher sensitization of RCC cells towards radiation in vitro than E-L and Y-L. Additionally, EY-L sensitized RCC tumors towards radiation therapy in xenograft and murine RCC models. EY-L mediated induction of mitotic catastrophe via downregulation of multiple cell cycle checkpoints and DNA damage repair pathways could be responsible for the augmentation of radiation therapy. CONCLUSION: Taken together, our study demonstrated the efficacy of a strategic combination therapy in sensitizing RCC to radiation therapy via inhibition of DNA damage repair and a substantial increase in mitotic catastrophe. This combination therapy may find its use in the augmentation of radiation therapy during the treatment of RCC patients.

Vorolanib, sunitinib, and axitinib: A comparative study of vascular endothelial growth factor receptor inhibitors and their anti-angiogenic effects.

PURPOSE: Pathological angiogenesis and vascular instability are observed in diabetic retinopathy (DR), diabetic macular edema (DME), and wet age-related macular degeneration (wAMD). Many receptor tyrosine kinases (RTKs) including vascular endothelial growth factor receptors (VEGFRs) contribute to angiogenesis, whereas the RTK TIE2 is important for vascular stability. Pan-VEGFR tyrosine kinase inhibitors (TKIs) such as vorolanib, sunitinib, and axitinib are of therapeutic interest over current antibody treatments that target only one or two ligands. This study compared the anti-angiogenic potential of these TKIs. METHODS: A kinase HotSpot™ assay was conducted to identify TKIs inhibiting RTKs associated with angiogenesis and vascular stability. Half-maximal inhibitory concentration (IC50) for VEGFRs and TIE2 was determined for each TKI. In vitro angiogenesis inhibition was investigated using a human umbilical vein endothelial cell sprouting assay, and in vivo angiogenesis was studied using the chorioallantoic membrane assay. Melanin binding was assessed using a melanin-binding assay. Computer modeling was conducted to understand the TIE2-axitinib complex as well as interactions between vorolanib and VEGFRs. RESULTS: Vorolanib, sunitinib, and axitinib inhibited RTKs of interest in angiogenesis and exhibited pan-VEGFR inhibition. HotSpot™ assay and TIE2 IC50 values showed that only axitinib potently inhibited TIE2 (up to 89%). All three TKIs effectively inhibited angiogenesis in vitro. In vivo, TKIs were more effective at inhibiting VEGF-induced angiogenesis than the anti-VEGF antibody bevacizumab. Of the three TKIs, only sunitinib bound melanin. TKIs differ in their classification and binding to VEGFRs, which is important because type II inhibitors have greater selectivity than type I TKIs. CONCLUSIONS: Vorolanib, sunitinib, and axitinib exhibited pan-VEGFR inhibition and inhibited RTKs associated with pathological angiogenesis. Of the three TKIs, only axitinib potently inhibited TIE2 which is an undesired trait as TIE2 is essential for vascular stability. The findings support the use of vorolanib for therapeutic inhibition of angiogenesis observed in DR, DME, and wAMD.

The Remarkable Anti-Breast Cancer Efficacy and Anti-Metastasis by Multifunctional Nanoparticles Co-Loading Squamocin, R848 and IR 780.

Background: Breast cancer is a heterogeneous disease globally accounting for approximately 1 million new cases annually. Chemotherapy remains the main therapeutic option, but the antitumor efficacy needs to be improved. Methods: Two multifunctional nanoparticles were developed in this paper using oleic acid and mPEG2k-PCL2k as the drug carriers. Squamocin (Squ) was employed as a chemotherapeutic agent. Resiquimod (R848) or ginsenoside Rh2 was co-encapsulated in the nanoparticles to remold the immunosuppressive tumor microenvironment, and IR780 was coloaded as a photosensitizer to realize photothermal therapy. Results: The obtained Squ-R848-IR780 nanoparticles and Squ-Rh2-IR780 nanoparticles were uniformly spherical and approximately (162.200 ± 2.800) nm and (157.300 ± 1.1590) nm, respectively, in average diameter, with good encapsulation efficiency (above 85% for each drug), excellent stability in various physiological media and high photothermal conversion efficiency (24.10% and 22.58%, respectively). After intravenous administration, both nanoparticles quickly accumulated in the tumor and effectively enhanced the local temperature of the tumor to over 45 °C when irradiated by an 808 nm laser. At a low dose of 0.1 mg/kg, Squ nanoparticles treatment alone displayed a tumor inhibition rate of 55.28%, pulmonary metastasis inhibition rate of 59.47% and a mean survival time of 38 days, which were all higher than those of PTX injection (8 mg/kg) (43.64%, 25 days and 37.25%), indicating that Squ was a potent and effective antitumor agent. Both multifunctional nanoparticles, Squ-Rh2-IR780 nanoparticles and Squ-R848-IR780 nanoparticles, demonstrated even better therapeutic efficacy, with tumor inhibition rates of 90.02% and 97.28%, pulmonary metastasis inhibition rates of 95.42% and 98.09, and mean survival times of 46 days and 52 days, respectively. Conclusion: The multifunctional nanoparticles coloaded with squamocin, R848 and IR 780 achieved extraordinary therapeutic efficacy and excellent antimetastasis activity and are thus promising in the future treatment of breast tumors and probably other tumors.

Identification of triciribine as a novel myeloid cell differentiation inducer.

Differentiation therapy using all-trans retinoic acid (ATRA) for acute promyelocytic leukemia (APL) is well established. However, because the narrow application and tolerance development of ATRA need to be improved, we searched for another efficient myeloid differentiation inducer. Kinase activation is involved in leukemia biology and differentiation block. To identify novel myeloid differentiation inducers, we used a Kinase Inhibitor Screening Library. Using a nitroblue tetrazolium dye reduction assay and real-time quantitative PCR using NB4 APL cells, we revealed that, PD169316, SB203580, SB202190 (p38 MAPK inhibitor), and triciribine (TCN) (Akt inhibitor) potently increased the expression of CD11b. We focused on TCN because it was reported to be well tolerated by patients with advanced hematological malignancies. Nuclear/cytoplasmic (N/C) ratio was significantly decreased, and myelomonocytic markers (CD11b and CD11c) were potently induced by TCN in both NB4 and acute myeloid leukemia (AML) M2 derived HL-60 cells. Western blot analysis using NB4 cells demonstrated that TCN promoted ERK1/2 phosphorylation, whereas p38 MAPK phosphorylation was not affected, suggesting that activation of the ERK pathway is involved in TCN-induced differentiation. We further examined that whether ATRA may affect phosphorylation of ERK and p38, and found that there was no obvious effect, suggesting that ATRA induced differentiation is different from TCN effect. To reveal the molecular mechanisms involved in TCN-induced differentiation, we performed microarray analysis. Pathway analysis using DAVID software indicated that "hematopoietic cell lineage" and "cytokine-cytokine receptor interaction" pathways were enriched with high significance. Real-time PCR analysis demonstrated that components of these pathways including IL1ß, CD3D, IL5RA, ITGA6, CD44, ITGA2B, CD37, CD9, CSF2RA, and IL3RA, were upregulated by TCN-induced differentiation. Collectively, we identified TCN as a novel myeloid cell differentiation inducer, and trials of TCN for APL and non-APL leukemia are worthy of exploration in the future.

Simulating BRAFV600E-MEK-ERK signalling dynamics in response to vertical inhibition treatment strategies.

In vertical inhibition treatment strategies, multiple components of an intracellular pathway are simultaneously inhibited. Vertical inhibition of the BRAFV600E-MEK-ERK signalling pathway is a standard of care for treating BRAFV600E-mutated melanoma where two targeted cancer drugs, a BRAFV600E-inhibitor, and a MEK inhibitor, are administered in combination. Targeted therapies have been linked to early onsets of drug resistance, and thus treatment strategies of higher complexities and lower doses have been proposed as alternatives to current clinical strategies. However, finding optimal complex, low-dose treatment strategies is a challenge, as it is possible to design more treatment strategies than are feasibly testable in experimental settings. To quantitatively address this challenge, we develop a mathematical model of BRAFV600E-MEK-ERK signalling dynamics in response to combinations of the BRAFV600E-inhibitor dabrafenib (DBF), the MEK inhibitor trametinib (TMT), and the ERK-inhibitor SCH772984 (SCH). From a model of the BRAFV600E-MEK-ERK pathway, and a set of molecular-level drug-protein interactions, we extract a system of chemical reactions that is parameterised by in vitro data and converted to a system of ordinary differential equations (ODEs) using the law of mass action. The ODEs are solved numerically to produce simulations of how pathway-component concentrations change over time in response to different treatment strategies, i.e., inhibitor combinations and doses. The model can thus be used to limit the search space for effective treatment strategies that target the BRAFV600E-MEK-ERK pathway and warrant further experimental investigation. The results demonstrate that DBF and DBF-TMT-SCH therapies show marked sensitivity to BRAFV600E concentrations in silico, whilst TMT and SCH monotherapies do not.

Imidazole[1,5-a]pyridine derivatives as EGFR tyrosine kinase inhibitors unraveled by umbrella sampling and steered molecular dynamics simulations.

Although the use of the tyrosine kinase inhibitors (TKIs) has been proved that it can save live in a cancer treatment, the currently used drugs bring in many undesirable side-effects. Therefore, the search for new drugs and an evaluation of their efficiency are intensively carried out. Recently, a series of eighteen imidazole[1,5-a]pyridine derivatives were synthetized by us, and preliminary analyses pointed out their potential to be an important platform for pharmaceutical development owing to their promising actions as anticancer agents and enzyme (kinase, HIV-protease,…) inhibitors. In the present theoretical study, we further analyzed their efficiency in using a realistic scenario of computational drug design. Our protocol has been developed to not only observe the atomistic interaction between the EGFR protein and our 18 novel compounds using both umbrella sampling and steered molecular dynamics simulations, but also determine their absolute binding free energies. Calculated properties of the 18 novel compounds were in detail compared with those of two known drugs, erlotinib and osimertinib, currently used in cancer treatment. Inspiringly the simulation results promote three imidazole[1,5-a]pyridine derivatives as promising inhibitors into a further step of clinical trials.

Predictive DNA damage signaling for low­dose ionizing radiation.

Numerous studies have attempted to develop biological markers for the response to radiation for broad and straightforward application in the field of radiation. Based on a public database, the present study selected several molecules involved in the DNA damage repair response, cell cycle regulation and cytokine signaling as promising candidates for low­dose radiation­sensitive markers. The HuT 78 and IM­9 cell lines were irradiated in a concentration­dependent manner, and the expression of these molecules was analyzed using western blot analysis. Notably, the activation of ataxia telangiectasia mutated (ATM), checkpoint kinase 2 (CHK2), p53 and H2A histone family member X (H2AX) significantly increased in a concentration­dependent manner, which was also observed in human peripheral blood mononuclear cells. To determine the radioprotective effects of cinobufagin, as an ATM and CHK2 activator, an in vivo model was employed using sub­lethal and lethal doses in irradiated mice. Treatment with cinobufagin increased the number of bone marrow cells in sub­lethal irradiated mice, and slightly elongated the survival of lethally irradiated mice, although the difference was not statistically significant. Therefore, KU60019, BML­277, pifithrin­α, and nutlin­3a were evaluated for their ability to modulate radiation­induced cell death. The use of BML­277 led to a decrease in radiation­induced p­CHK2 and γH2AX levels and mitigated radiation­induced apoptosis. On the whole, the present study provides a novel approach for developing drug candidates based on the profiling of biological radiation­sensitive markers. These markers hold promise for predicting radiation exposure and assessing the associated human risk.

Self-Assembled Nanoparticles from the Amphiphilic Prodrug of Resiquimod for Improved Cancer Immunotherapy.

Tumor-associated macrophages (TAMs) usually adopt a tumor-promoting M2-like phenotype, which largely impedes the immune response and therapeutic efficacy of solid tumors. Repolarizing TAMs from M2 to the antitumor M1 phenotype is crucial for reshaping the tumor immunosuppressive microenvironment (TIME). Herein, we developed self-assembled nanoparticles from the polymeric prodrug of resiquimod (R848) to reprogram the TIME for robust cancer immunotherapy. The polymeric prodrug was constructed by conjugating the R848 derivative to terminal amino groups of the linear dendritic polymer composed of linear poly(ethylene glycol) and lysine dendrimer. The amphiphilic prodrug self-assembled into nanoparticles (PLRS) of around 35 nm with a spherical morphology. PLRS nanoparticles could be internalized by antigen-presenting cells (APCs) in vitro and thus efficiently repolarized macrophages from M2 to M1 and facilitated the maturation of APCs. In addition, PLRS significantly inhibited tumor growth in the 4T1 orthotopic breast cancer model with much lower systemic side effects. Mechanistic studies suggested that PLRS significantly stimulated the TIME by repolarizing TAMs into the M1 phenotype and increased the infiltration of cytotoxic T cells into the tumor. This study provides an effective polymeric prodrug-based strategy to improve the therapeutic efficacy of R848 in cancer immunotherapy.

Effect of Acridine Orange and Zoledronic Acid on Bone Metastasis in Renal Cell Carcinoma.

BACKGROUND/AIM: The increasing incidence of renal cell carcinoma (RCC) and its associated bone metastasis pose challenges in surgical interventions, warranting the exploration of novel therapeutic approaches. Therefore, this study aimed to assess the impact of hematogenously administering acridine orange (AO) alone and in combination with zoledronic acid (ZA) on bone metastasis in RCC. MATERIALS AND METHODS: RENCA cells (1.0×106 cells/10 µl) were directly injected into the right femur of male BALB/c mice. The mice were categorized into four groups based on the applied therapeutic intervention and were euthanized after five weeks. Micro-computed tomography was performed to quantify the extent of periosteal reaction, indicative of bone metastasis, along the entire length of the femur. Tumor weight and volume were measured at euthanization. Hematoxylin and eosin staining was used to examine the extent of tumor development in the bone. Apoptotic cell, osteoclast, and vascular endothelial growth factor (VEGF)-positive cell counts were assessed using TdT-mediated dUTP-biotin nick end labeling, tartrate-resistant acid phosphatase staining, and VEGF staining, respectively. RESULTS: The periosteal reaction was significantly reduced in the intervention groups compared to the control group (p<0.05). The apoptotic cell numbers in the intervention groups surpassed that in the control group (p<0.05), whereas those of osteoclasts and VEGF-positive cells in the intervention groups were lower than those in the control group (p<0.05). CONCLUSION: AO hinders bone metastasis progression in RCC, and combination therapy with ZA may be more effective than AO administration alone.

Identification of BRAF Inhibitor Resistance-associated lncRNAs Using Genome-scale CRISPR-Cas9 Transcriptional Activation Screening.

BACKGROUND/AIM: Approximately 50% of melanomas harbor the BRAF V600E mutation and targeted therapies using BRAF inhibitors improve patient outcomes. Nonetheless, resistance to BRAF inhibitors develops rapidly and remains a challenge in melanoma treatment. In this study, we attempted to isolate long noncoding RNAs (lncRNAs) involved in BRAF inhibitor resistance using a comprehensive screening method. MATERIALS AND METHODS: We used a CRISPR-Cas9 synergistic activation mediator (SAM) protein complex in a genome-scale transcriptional activation assay to screen for candidate lncRNA genes related to BRAF inhibitor resistance. Correlation analysis was performed between expression levels of isolated lncRNA genes and IC50 of dabrafenib in a BRAF-mutated melanoma cell line. Next, online databases were used to construct the lncRNA-miRNA-mRNA regulatory network. Finally, we evaluated the significance of the expression levels of these lncRNAs and mRNAs as biomarkers using clinical specimens. RESULTS: We isolated three BRAF inhibitor resistance-associated lncRNA genes, namely SNHG16, NDUFV2-AS1, and LINC01502. We constructed a lncRNA-miRNA-mRNA network of 13 nodes consisting of three lncRNAs, six miRNAs, and four mRNAs. The lncRNAs and target mRNAs from each regulatory axis significantly and positively correlated with each other. Finally, Kaplan-Meier analysis showed that higher expression levels of MITF, which was up-regulated by LINC01502, were significantly associated with worse prognosis in BRAF V600E-mutated melanoma. CONCLUSION: The identification of these BRAF inhibitor resistance-associated lncRNA genes at the genomic scale and the establishment of the lncRNA-miRNA-mRNA regulatory network provides new insights into the underlying mechanisms of BRAF inhibitor resistance in melanoma.

Elesclomol-copper synergizes with imidazole ketone erastin by promoting cuproptosis and ferroptosis in myelodysplastic syndromes.

Myelodysplastic syndromes (MDS) encompass a collection of clonal hematopoietic malignancies distinguished by the depletion of peripheral blood cells. The treatment of MDS is hindered by the advanced age of patients, with a restricted repertoire of drugs currently accessible for therapeutic intervention. In this study, we found that ES-Cu strongly inhibited the viability of MDS cell lines and activated cuproptosis in a copper-dependent manner. Importantly, ferroptosis inducer IKE synergistically enhanced ES-Cu-mediated cytotoxicity both in vitro and in vivo. Of note, the combination of IKE and ES-Cu intensively impaired mitochondrial homeostasis with increased mitochondrial ROS, MMP hyperpolarized, down-regulated iron-sulfur proteins and declined oxygen consumption rate. Additionally, ES-Cu/IKE treatment could enhance the lipoylation-dependent oligomerization of the DLAT. To elucidate the specific order of events in the synergistic cell death, inhibitors of ferroptosis and cuproptosis were utilized to further characterize the basis of cell death. Cell viability assays showed that the glutathione and its precursor N-acetylcysteine could significantly rescue the cell death under either mono or combination treatment, demonstrating that GSH acts at the crossing point in the regulation network of cuproptosis and ferroptosis. Significantly, the reconstitution of xCT expression and knockdown of FDX1 cells have been found to contribute to the tolerance of mono treatment but have little recovery impact on the combined treatment. Collectively, these findings suggest that a synergistic interaction leading to the induction of multiple programmed cell death pathways could be a promising approach to enhance the effectiveness of therapy for MDS.

Nature-inspired substituted 3-(imidazol-2-yl) morpholines targeting human topoisomerase IIα: Dynophore-derived discovery.

The molecular nanomachine, human DNA topoisomerase IIα, plays a crucial role in replication, transcription, and recombination by catalyzing topological changes in the DNA, rendering it an optimal target for cancer chemotherapy. Current clinical topoisomerase II poisons often cause secondary tumors as side effects due to the accumulation of double-strand breaks in the DNA, spurring the development of catalytic inhibitors. Here, we used a dynamic pharmacophore approach to develop catalytic inhibitors targeting the ATP binding site of human DNA topoisomerase IIα. Our screening of a library of nature-inspired compounds led to the discovery of a class of 3-(imidazol-2-yl) morpholines as potent catalytic inhibitors that bind to the ATPase domain. Further experimental and computational studies identified hit compound 17, which exhibited selectivity against the human DNA topoisomerase IIα versus human protein kinases, cytotoxicity against several human cancer cells, and did not induce DNA double-strand breaks, making it distinct from clinical topoisomerase II poisons. This study integrates an innovative natural product-inspired chemistry and successful implementation of a molecular design strategy that incorporates a dynamic component of ligand-target molecular recognition, with comprehensive experimental characterization leading to hit compounds with potential impact on the development of more efficient chemotherapies.

Extracellular vesicles promote migration despite BRAF inhibitor treatment in malignant melanoma cells.

Extracellular vesicles (EVs) constitute a vital component of intercellular communication, exerting significant influence on metastasis formation and drug resistance mechanisms. Malignant melanoma (MM) is one of the deadliest forms of skin cancers, because of its high metastatic potential and often acquired resistance to oncotherapies. The prevalence of BRAF mutations in MM underscores the importance of BRAF-targeted therapies, such as vemurafenib and dabrafenib, alone or in combination with the MEK inhibitor, trametinib. This study aimed to elucidate the involvement of EVs in MM progression and ascertain whether EV-mediated metastasis promotion persists during single agent BRAF (vemurafenib, dabrafenib), or MEK (trametinib) and combined BRAF/MEK (dabrafenib/trametinib) inhibition.Using five pairs of syngeneic melanoma cell lines, we assessed the impact of EVs - isolated from their respective supernatants - on melanoma cell proliferation and migration. Cell viability and spheroid growth assays were employed to evaluate proliferation, while migration was analyzed through mean squared displacement (MSD) and total traveled distance (TTD) measurements derived from video microscopy and single-cell tracking.Our results indicate that while EV treatments had remarkable promoting effect on cell migration, they exerted only a modest effect on cell proliferation and spheroid growth. Notably, EVs demonstrated the ability to mitigate the inhibitory effects of BRAF inhibitors, albeit they were ineffective against a MEK inhibitor and the combination of BRAF/MEK inhibitors. In summary, our findings contribute to the understanding of the intricate role played by EVs in tumor progression, metastasis, and drug resistance in MM.

Co-delivery of carboplatin and doxorubicin using ZIF-8 coated chitosan-poly(N-isopropyl acrylamide) nanoparticles through a dual pH/thermo responsive strategy to breast cancer cells.

A dual pH/temperature sensitive core-shell nanoformulation has been developed based on ZIF-8 coated with chitosan-poly(N-isopropyl acrylamide) (CS-PNIPAAm) for co-delivery of doxorubicin (DOX) and carboplatin (CBP) in breast cancer cells. The resulting nanoparticles (NPs) had particle sizes around 200 nm and a zeta potential of about +30 mV. The CBP and DOX loading contents in the final NPs were 11.6 % and 55.54 %, respectively. NPs showed a pH and thermoresponsive drug release profile with a sustained prolonged release under physiological conditions. The in vitro cytotoxicity experiments showed a significant synergism of CBP and DOX to induce the IC50 of 1.96 µg/mL in MCF-7 cells and 4.54 µg/mL in MDA-MB-231 cells. Also, the final NPs were safer than free DOX and CBP on normal cells. The in vitro study confirmed the higher potency of the designed NPs in combination therapy against breast cancer cells with lower side effects than free drugs.

TLR agonists polarize interferon responses in conjunction with dendritic cell vaccination in malignant glioma: a randomized phase II Trial.

In this randomized phase II clinical trial, we evaluated the effectiveness of adding the TLR agonists, poly-ICLC or resiquimod, to autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination in patients with newly-diagnosed or recurrent WHO Grade III-IV malignant gliomas. The primary endpoints were to assess the most effective combination of vaccine and adjuvant in order to enhance the immune potency, along with safety. The combination of ATL-DC vaccination and TLR agonist was safe and found to enhance systemic immune responses, as indicated by increased interferon gene expression and changes in immune cell activation. Specifically, PD-1 expression increases on CD4+ T-cells, while CD38 and CD39 expression are reduced on CD8+ T cells, alongside an increase in monocytes. Poly-ICLC treatment amplifies the induction of interferon-induced genes in monocytes and T lymphocytes. Patients that exhibit higher interferon response gene expression demonstrate prolonged survival and delayed disease progression. These findings suggest that combining ATL-DC with poly-ICLC can induce a polarized interferon response in circulating monocytes and CD8+ T cells, which may represent an important blood biomarker for immunotherapy in this patient population.Trial Registration: ClinicalTrials.gov Identifier: NCT01204684.

Understanding the interactions between repurposed drugs sertindole and temoporfin with receptor for advanced glycation endproducts: Therapeutic implications in cancer and metabolic diseases.

CONTEXT: In the pursuit of novel therapeutic possibilities, repurposing existing drugs has gained prominence as an efficient strategy. The findings from our study highlight the potential of repurposed drugs as promising candidates against receptor for advanced glycation endproducts (RAGE) that offer therapeutic implications in cancer, neurodegenerative conditions and metabolic syndromes. Through careful analyses of binding affinities and interaction patterns, we identified a few promising candidates, ultimately focusing on sertindole and temoporfin. These candidates exhibited exceptional binding affinities, efficacy, and specificity within the RAGE binding pocket. Notably, they displayed a pronounced propensity to interact with the active site of RAGE. Our investigation further revealed that sertindole and temoporfin possess desirable pharmacological properties that highlighted them as attractive candidates for targeted drug development. Overall, our integrated computational approach provides a comprehensive understanding of the interactions between repurposed drugs, sertindole and temoporfin and RAGE that pave the way for future experimental validation and drug development endeavors. METHODS: We present an integrated approach utilizing molecular docking and extensive molecular dynamics (MD) simulations to evaluate the potential of FDA-approved drugs, sourced from DrugBank, against RAGE. To gain deeper insights into the binding mechanisms of the elucidated candidate repurposed drugs, sertindole and temoporfin with RAGE, we conducted extensive all-atom MD simulations, spanning 500 nanoseconds (ns). These simulations elucidated the conformational dynamics and stability of the RAGE-sertindole and RAGE-temoporfin complexes.

Enhanced selectivity towards melanoma cells with zinc(II)-Schiff bases containing imidazole derivatives.

Zinc(II)-complexes with the general formula [Zn(L)2] containing 8-hydroxyquinoline Schiff bases functionalized with 1-(3-aminopropyl)imidazole or 1-(3-aminopropyl)-2-methyl-1H-imidazole on 2-position and their respective ligands (HL1 or HL2) were synthesized and characterized by NMR, UV-Vis, FTIR and CD spectroscopies as well as ESI-MS spectrometry. Single crystals of HL2 and [Zn(L1)2]n were analysed by SC-XRD. [Zn(L1)2]n shows a 1D polymeric chain structure of alternating Zn(II) cations and bridging Schiff base ligands, in contrast to previously reported monomeric structures of analogous complexes. DFT calculations were performed to rationalize the polymeric X-ray structure of Zn(L1)2. Results showed that the ligands can bind as bi- or tridentate to Zn(II) and there is the possibility of a dynamic behavior for the complexes in solution. Both ligands and complexes present limited stability in aqueous media, however, in the presence of bovine serum albumin the complexes are stable. Molecular docking simulations and circular dichroism spectroscopic studies suggest binding to this protein in close proximity to the Trp213 residue. Biological studies on a panel of cancer cells revealed that the Zn(II)-complexes have a lower impact on cell viability than cisplatin, except for triple-negative breast cancer cells in which they were comparable. Notwithstanding, they display much higher selectivity towards cancer cells vs. normal cells, than cisplatin. They induce the generation of ROS and DNA double-strand breaks, primarily through apoptosis as the mode of cell death. Overall, the novel Zn(II)-complexes demonstrate improved induction of apoptosis and higher selectivity, particularly for melanoma cells, compared to previously reported analogues, making them promising candidates for clinical application.

A novel Imidazo[1,2-a]pyridine derivative modulates active KRASG12D through off-like conformational shifts in switch-I and switch-II regions, mimicking inactive KRASG12D.

KRASG12D are the most prevalent oncogenic mutations and a promising target for solid tumor therapies. However, its inhibition exhibits tremendous challenge due to the necessity of high binding affinity to obviate the need for covalent binders. Here we report the evidence of a novel class of Imidazo[1,2-a]pyridine derivative as potentially significant novel inhibitors of KRASG12D, discovered through extensive ligand-based screening against 2-[(2R)-piperidin-2-yl]-1H-indole, an important scaffold for KRASG12D inhibition via switch-I/II (S-I/II) pocket. The proposed compounds exhibited similar binding affinities and overlapped pose configurations to 2-[(2R)-piperidin-2-yl]-1H-indole, serving as a reliable starting point for drug discovery. Comparative free energy profiles demonstrated that C4 [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] effectively shifted the protein to a stable low-energy conformation via a prominent transition state. The conformational changes across the transition revealed the conformational shift of switch-I and II to a previously known off-like conformation of inactive KRASG12D with rmsd of 0.91 Å. These conformations were even more prominent than the privileged scaffold 2-[(2R)-piperidin-2-yl]-1H-indole. The representative structure overlay of C4 and another X-ray crystallography solved BI-2852 bound inactive KRASG12D revealed that Switch-I and II exhibited off-like conformations. The cumulative variance across the first eigenvalue that accounted for 57 % of the collective variance validated this on-to-off transition. In addition, the relative interaction of C4 binding showed consistent patterns with BI-2852. Taken together, our results support the inhibitory activity of [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] by shifting active KRASG12D to an inactive conformation.

Injectable hydrogel with doxorubicin-loaded ZIF-8 nanoparticles for tumor postoperative treatments and wound repair.

The need for tumor postoperative treatments aimed at recurrence prevention and tissue regeneration have raised wide considerations in the context of the design and functionalization of implants. Herein, an injectable hydrogel system encapsulated with anti-tumor, anti-oxidant dual functional nanoparticles has been developed in order to prevent tumor relapse after surgery and promote wound repair. The utilization of biocompatible gelatin methacryloyl (GelMA) was geared towards localized therapeutic intervention. Zeolitic imidazolate framework-8@ceric oxide (ZIF-8@CeO2, ZC) nanoparticles (NPs) were purposefully devised for their proficiency as reactive oxygen species (ROS) scavengers. Furthermore, injectable GelMA hydrogels loaded with ZC NPs carrying doxorubicin (ZC-DOX@GEL) were tailored as multifunctional postoperative implants, ensuring the efficacious eradication of residual tumor cells and alleviation of oxidative stress. In vitro and in vivo experiments were conducted to substantiate the efficacy in cancer cell elimination and the prevention of tumor recurrence through the synergistic chemotherapy approach employed with ZC-DOX@GEL. The acceleration of tissue regeneration and in vitro ROS scavenging attributes of ZC@GEL were corroborated using rat models of wound healing. The results underscore the potential of the multifaceted hydrogels presented herein for their promising application in tumor postoperative treatments.