Inhibition Of Cellular Proliferation Research Articles

Clinical Applications of Phototherapy in Treating Skin Disorders

Introduction and Purpose: Phototherapy, involving the therapeutic use of ultraviolet and visible light, has been a cornerstone in dermatology for treating various skin disorders. This review aims to consolidate current knowledge on the mechanisms of action and clinical applications of phototherapy, with a focus on enhancing understanding among clinicians and researchers.State of Knowledge: Phototherapy is particularly effective for chronic conditions such as psoriasis, vitiligo, and atopic dermatitis, where traditional therapies might not suffice. The mechanism involves modulation of immune response and inhibition of cellular proliferation, achieved through various wavelengths, including UVA and UVB. Recent advancements have introduced narrow-band UVB, which offers targeted therapy with fewer side effects compared to broad-band UVB. Moreover, the review explores innovations such as LED-based therapies and photodynamic therapy, expanding the spectrum of phototherapeutic options.Summary: Phototherapy remains a vital and evolving treatment modality in dermatology, offering safe and effective management for multiple skin disorders. Its success hinges on precise application and adherence to treatment protocols. Future research should aim to optimize treatment regimens, reduce side effects, and enhance patient compliance to improve outcomes. By advancing our understanding and application of phototherapy, we can better tailor treatments to meet individual patient needs, thereby improving the quality of life for those afflicted with chronic skin conditions.

Overexpression of Fatty Acid Synthase Upregulates Glutamine-Fructose-6-Phosphate Transaminase 1 and O-Linked N-Acetylglucosamine Transferase to Increase O-GlcNAc Protein Glycosylation and Promote Colorectal Cancer Growth.

Fatty acid synthesis has been extensively investigated as a therapeutic target in cancers, including colorectal cancer (CRC). Fatty acid synthase (FASN), a key enzyme of de novo lipid synthesis, is significantly upregulated in CRC, and therapeutic approaches of targeting this enzyme are currently being tested in multiple clinical trials. However, the mechanisms behind the pro-oncogenic action of FASN are still not completely understood. Here, for the first time, we show that overexpression of FASN increases the expression of glutamine-fructose-6-phosphate transaminase 1 (GFPT1) and O-linked N-acetylglucosamine transferase (OGT), enzymes involved in hexosamine metabolism, and the level of O-GlcNAcylation in vitro and in vivo. Consistently, expression of FASN significantly correlates with expression of GFPT1 and OGT in human CRC tissues. shRNA-mediated downregulation of GFPT1 and OGT inhibits cellular proliferation and the level of protein O-GlcNAcylation in vitro, and knockdown of GFPT1 leads to a significant decrease in tumor growth and metastasis in vivo. Pharmacological inhibition of GFPT1 and OGT leads to significant inhibition of cellular proliferation and colony formation in CRC cells. In summary, our results show that overexpression of FASN increases the expression of GFPT1 and OGT as well as the level of protein O-GlcNAcylation to promote progression of CRC; targeting the hexosamine biosynthesis pathway could be a therapeutic approach for this disease.

USP2 Promotes the Proliferation and Inflammation of Fibroblast-Like Synovial Cells in Rheumatoid Arthritis Through Deubiquitination of TRAF2.

Rheumatoid arthritis (RA) is a prevalent inflammatory disorder affecting about 1% of the global population. The ubiquitin-specific protease 2 (USP2) is known to have a substantial influence on the regulation of several cellular processes. Both in vivo (using rats with collagen-induced arthritis, CIA) and in vitro (using human fibroblast-like synoviocytes, HFLS-RA) models of RA were used to examine the role of USP2 in RA. The proliferation of HFLS-RA cells was assessed using the cell counting kit 8 test and EdU staining. The technique used for the assessment of gene expression was quantitative real-time PCR. Protein expression was quantified using Western blot (WB) analysis, while the quantities of inflammatory factors and matrix metalloproteinases were assessed using an ELISA test. The co-immunoprecipitation and ubiquitination tests investigated the relationships between proteins and the underlying molecular pathways. The results of this study demonstrate an upregulation of USP2 expression in both vivo and vitro models of RA. In addition, our findings indicate that the overexpression of USP2 notably exacerbates both proliferation and inflammation. The consistent downregulation of USP2 resulted in a reduction in the secretion of inflammatory cytokines and a suppression of cellular proliferation. Furthermore, it was shown that USP2 interacts with tumor necrosis factor receptor-associated factor 2 (TRAF2) and facilitates the removal of ubiquitination chains from TRAF2, enhancing its stability. Our findings propose that USP2 functions as a favorable modulator of proliferation and inflammatory reactions in HFLS-RA, thereby indicating its potential as a therapeutic target for the treatment of RA.

Diosgenin inhibits proliferation and migration of ovarian cancer cells and induce apoptosis via upregulation of PTEN.

Diosgenin, a natural steroidal sapogenin, has recently attracted a high amount of attention, as an effective anticancer agent in ovarian cancer. However, diosgenin mediated anticancer impacts are still not completely understood. Thus, the present study evaluated the effect of diosgenin on the proliferation, apoptosis, and metastasis of ovarian cancer cells. OVCAR-3 and SKOV-3 cells were treated with diosgenin, cellular viability was assessed by MTT assay and apoptosis was measured by ELISA and evaluated the protein expression levels of apoptotic markers through western blotting. Cell migration was examined by measuring the mRNA levels of genes involved in the cell invasion. The protein expression levels of main components of PI3K signaling were evaluated via western blotting. Diosgenin led to significant inhibition of cellular proliferation in a dose-dependent manner. It also induced apoptosis through upregulating pro-apoptotic markers and downregulating antiapoptotic mediators. In addition, OVCAR-3 cells exposure to diosgenin decreased cell migration and invasion. More importantly, diosgenin downregulated the expression levels of main proteins in PI3K signaling including PI3K, Akt, mTOR, and GSK3. Diosgenin inhibited the proliferation and migration of OVCAR-3 ovarian cancer cells and induced apoptosis, which may be mediated by targeting PI3K signaling.

Targeting adenosine A2A receptors for early intervention of retinopathy of prematurity.

Retinopathy of prematurity (ROP) continues to pose a significant threat to the vision of numerous children worldwide, primarily owing to the increased survival rates of premature infants. The pathologies of ROP are mainly linked to impaired vascularization as a result of hyperoxia, leading to subsequent neovascularization. Existing treatments, including anti-vascular endothelial growth factor(VEGF) therapies, have thus far been limited to addressing pathological angiogenesis at advanced ROP stages, inevitably leading to adverse side effects. Intervention to promote physiological angiogenesis during the initial stages could hold the potential to prevent ROP. Adenosine A2A receptors (A2AR) have been identified in various ocular cell types, exhibiting distinct densities and functionally intricate connections with oxygen metabolism. In this review, we discuss experimental evidence that strongly underscores the pivotal role of A2AR in ROP. In particular, A2AR blockade may represent an effective treatment strategy, mitigating retinal vascular loss by reversing hyperoxia-mediated cellular proliferation inhibition and curtailing hypoxia-mediated neovascularization in oxygen-induced retinopathy (OIR). These effects stem from the interplay of endothelium, neuronal and glial cells, and novel molecular pathways (notably promoting TGF-β signaling) at the hyperoxia phase. We propose that pharmacological targeting of A2AR signaling may confer an early intervention for ROP with distinct therapeutic benefits and mechanisms than the anti-VEGF therapy.

Gesicles packaging dCas9-VPR ribonucleoprotein complexes can combine with vorinostat and promote HIV proviral transcription

Despite the success of combination antiretroviral therapies (cART) in HIV treatment, a cure for HIV remains elusive. Scientists postulate that HIV latent reservoirs may be a vital target in curative strategies. Vorinostat is a latency-reversing agent which has demonstrated some effectiveness in reactivating latent HIV, but complementary therapies may be essential to enhance its efficacy. One such approach may utilize the CRISPR/Cas9 system which has evolved to include transcriptional activators such as dCas9-VPR. In this study, we explored the effects of combining vorinostat coupled with gesicle-mediated delivery of dCas9-VPR in promoting the transcription of integrated HIV proviruses in HIV-NanoLuc CHME-5 microglia and J-Lat 10.6 lymphocytes. We confirmed that dCas9-VPR ribonucleoprotein complexes can be packaged into gesicles and application to cells successfully induced HIV transcription through interactions with the HIV LTR. Vorinostat also induced significant increases in proviral transcription but generated inhibition of cellular proliferation (microglia) or cell viability (lymphocytes) starting at 1000nM and higher concentrations. Experiments combining dCas9-VPR gesicles and vorinostat confirmed the enhanced transcriptional activation of the HIV provirus in microglia but not lymphocytes. Thus, a combination of dCas9-VPR gesicles with other latency reversing agents may provide a complementary method to activate latent HIV in future studies utilizing patient-derived cells or small animal models.

Efficacy of tyrosine kinase inhibitors examined by a combination of Raman micro-spectroscopy and a deep wavelet scattering-based multivariate analysis framework.

HER2 is a crucial therapeutic target in breast cancer, and the survival rate of breast cancer patients has increased because of this receptor's inhibition. However, tumors have shown resistance to this therapeutic strategy due to oncogenic mutations that decrease the binding of several HER2-targeted drugs, including lapatinib, and confer resistance to this drug. Neratinib can overcome this drug resistance and effectively inhibit HER2 signaling and tumor growth. In the present study, we examined the efficacy of lapatinib and neratinib using breast cancer cells by Raman microscopy combined with a deep wavelet scattering-based multivariate analysis framework. This approach discriminated between control cells and drug-treated cells with high accuracy, compared to classical principal component analysis. Both lapatinib and neratinib induced changes in the cellular biochemical composition. Furthermore, the Raman results were compared with the results of several in vitro assays. For instance, drug-treated cells exhibited (i) inhibition of ERK and AKT phosphorylation, (ii) inhibition of cellular proliferation, (iii) cell-cycle arrest, and (iv) apoptosis as indicated by western blotting, real-time cell analysis (RTCA), cell-cycle analysis, and apoptosis assays. Thus, the observed Raman spectral changes are attributed to cell-cycle arrest and apoptosis. The results also indicated that neratinib is more potent than lapatinib. Moreover, the uptake and distribution of lapatinib in cells were visualized through its label-free marker bands in the fingerprint region using Raman spectral imaging. These results show the prospects of Raman microscopy in drug evaluation and presumably in drug discovery.

The Neuroprotective Effects of Piracetam on Cisplatin-Induced Cognitive Decline

ABSTRACT Aim This work explores the effect of Cisplatin—a chemotherapeutic agent known to cause deterioration in cognitive function in cancer patients, and spatial memory in mice. It also investigates the potential neuroprotective effects of Piracetam, which is a nootropic drug recognized for improving cognitive ability. Materials and Methods The study incorporates four groups of mice receiving varied medication regimens, with memory tested using the Novel Location Recognition (NLR) method. Results The findings from our study revealed that memory decline and a suppression of cellular proliferation were observed in adult male mice subjected to Cisplatin treatment; furthermore, a decline in antioxidant efficacy within the hippocampal dentate gyrus was evident. Moreover, analysis of treatment effects on the animals’ weight revealed that the Cisplatin and Piracetam group exhibited the most significant weight loss during drug administration. Despite the significant weight loss, the simultaneous use of Cisplatin and Piracetam demonstrated a notable improvement in memory and an augmentation of hippocampal proliferation and antioxidant effect. Limitations It is important to note that our study was hampered by budget limits, a lack of additional animals, and mice’s low tolerance for protracted treatment. Conclusions Should the outcomes of Piracetam observed in this investigation be applicable to patients, it might offer a relatively straightforward approach to mitigate the cognitive impacts endured by cancer survivors following exposure to chemotherapy. Future research will be needed to study Piracetam’s effect on mice with brain cancer after Cisplatin treatment in order to extrapolate the results onto cancer patients.

Celastrol suppresses human pancreatic cancer via m6A-YTHDF3-mediated downregulation of Claspin and Bcl-2

BackgroundCelastrol has been revealed to exhibit anticancer pharmacological activity, however, the molecular mechanisms of celastrol involved in pancreatic cancer remain to be further elucidated. The present study was to illustrate whether celastrol suppresses pancreatic cancer through modulating RNA m6A modification.MethodsEffect of celastrol treatment on the malignant phenotypes of pancreatic cancer cells was evaluated by CCK-8 assay, EdU assay, colony formation assay, flow cytometry analysis and subcutaneous xenograft experiments. RNA sequencing (RNA-seq) analysis was carried out to analyze the genes differentially expressed in celastrol-treated pancreatic cancer cells. RT-qPCR, Western blotting and immunohistochemistry were employed to evaluate the expression of the indicated genes. RNA dot blot and quantification of total RNA m6A modification assays, MeRIP-qPCR assay, RIP-qPCR assay, RNA stability and protein stability assays were applied to evaluate the regulatory mechanism of celastrol treatment in pancreatic cancer cells.ResultsWe demonstrated that celastrol suppressed cell proliferation and induced cell cycle arrest and apoptosis of pancreatic cancer cells in vitro, and decreased tumor growth in vivo. Specifically, Bcl-2, Claspin, METTL3 and YTHDF3 were identified as the potential targets of celastrol treatment in pancreatic cancer cells. Moreover, our results indicated that celastrol treatment downregulated METTL3 and decreased m6A levels of Claspin and Bcl-2 mRNA, leading to the degradation of Claspin and Bcl-2 mRNA in pancreatic cancer cells. Furthermore, we revealed that celastrol treatment downregulated Claspin and Bcl-2, at least in part, in an m6A-YTHDF3-mediated manner in pancreatic cancer cells.ConclusionOur study highlighted a novel mechanism underlying celastrol-induced cellular proliferation inhibition and apoptosis in pancreatic cancer cells via m6A-YTHDF3-mediated downregulation of Claspin and Bcl-2.

GPATCH4 contributes to nucleolus morphology and its dysfunction impairs cell viability

The nucleolus serves a multifaceted role encompassing not only rRNA transcription and ribosome synthesis, but also the intricate orchestration of cell cycle regulation and the modulation of cellular senescence. G-patch domain containing 4 (GPATCH4) stands as one among the nucleolar proteins; however, its functional significances remain still unclear. In order to elucidate the functions of GPATCH4, we examined the effects of its dysfunction on cellular proliferation, alterations in nucleolar architecture, apoptotic events, and cellular senescence. Through experimentation conducted on cultured neuroblastoma SH-SY5Y cells, the reduction of GPATCH4 caused inhibition of cellular proliferation, concurrently fostering escalated apoptotic susceptibilities upon exposure to high-dose etoposide. In the realm of nucleolar morphology comparisons, a discernible decline was noted in the count of nucleoli per nucleus, concomitant with a significant expansion in the area occupied by individual nucleoli. Upon induction of senescence prompted by low-dose etoposide, GPATCH4 knockdown resulted in decreased cell viability and increased expression of senescence-associated markers, namely senescence-associated β-galactosidase (SA-β-GAL) and p16. Furthermore, GPATCH4 dysfunction elicited alterations in the gene expression profile of the ribosomal system. In sum, our findings showed that GPATCH4 is a pivotal nucleolar protein that regulates nucleolar morphology and is correlated with cell viability.

Type I interferon signaling induces a delayed antiproliferative response in respiratory epithelial cells during SARS-CoV-2 infection.

Disease progression during SARS-CoV-2 infection is tightly linked to the fate of lung epithelial cells, with severe cases of COVID-19 characterized by direct injury of the alveolar epithelium and an impairment in its regeneration from progenitor cells. The molecular pathways that govern respiratory epithelial cell death and proliferation during SARS-CoV-2 infection, however, remain unclear. We now report a high-throughput CRISPR screen for host genetic modifiers of the survival and proliferation of SARS-CoV-2-infected Calu-3 respiratory epithelial cells. The top four genes identified in our screen encode components of the same type I interferon (IFN-I) signaling complex—IFNAR1, IFNAR2, JAK1, and TYK2. The fifth gene, ACE2, was an expected control encoding the SARS-CoV-2 viral receptor. Surprisingly, despite the antiviral properties of IFN-I signaling, its disruption in our screen was associated with an increase in Calu-3 cell fitness. We validated this effect and found that IFN-I signaling did not sensitize SARS-CoV-2-infected cultures to cell death but rather inhibited the proliferation of surviving cells after the early peak of viral replication and cytopathic effect. We also found that IFN-I signaling alone, in the absence of viral infection, was sufficient to induce this delayed antiproliferative response in both Calu-3 cells and iPSC-derived type 2 alveolar epithelial cells. Together, these findings highlight a cell autonomous antiproliferative response by respiratory epithelial cells to persistent IFN-I signaling during SARS-CoV-2 infection. This response may contribute to the deficient alveolar regeneration that has been associated with COVID-19 lung injury and represents a promising area for host-targeted therapeutic development.

5,6-Dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) induces apoptosis in breast cancer cells through inhibiting of Mcl-1 expression

The effective treatment of breast cancer remains a profound clinical challenge, especially due to drug resistance and metastasis which unfortunately arise in many patients. The transcription inhibitor 5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole (DRB), as a selective inhibitor of cyclin-dependent kinase 9, was shown to be effective in inducing apoptosis in various hematopoietic malignancies. However, the anticancer efficacy of DRB against breast cancer is still unclear. Herein, we demonstrated that administration of DRB to the breast cancer cell line led to the inhibition of cellular proliferation and induction of the typical signs of apoptotic cells, including the increases in Annexin V-positive cells, DNA fragmentation, and activation of caspase-7, caspase-9, and poly (ADP ribose) polymerase (PARP). Treatment of DRB resulted in a rapid decline in the myeloid cell leukemia 1 (Mcl-1) protein, whereas levels of other antiapoptotic proteins did not change. Overexpression of Mcl-1 decreased the DRB-induced PARP cleavage, whereas knockdown of Mcl-1 enhanced the effects of DRB on PARP activation, indicating that loss of Mcl-1 accounts for the DRB-mediated apoptosis in MCF-7 cells, but not in T-47D. Furthermore, we found that co-treatment of MCF-7 cells with an inhibitor of AKT (LY294002) or an inhibitor of the proteasome (MG-132) significantly augmented the DRB-induced apoptosis. These data suggested that DRB in combination with LY294002 or MG-132 may have a greater therapeutic potency against breast cancer cells.

Transcriptome and proteome analysis reveals the anti-cancer properties of Hypnea musciformis marine macroalga extract in liver and intestinal cancer cells

BackgroundMarine seaweeds are considered as a rich source of health-promoting compounds by the food and pharmaceutical industry. Hypnea musciformis is a marine red macroalga (seaweed) that is widely distributed throughout the world, including the Mediterranean Sea. It is known to contain various bioactive compounds, including sulfated polysaccharides, flavonoids, and phlorotannins. Recent studies have investigated the potential anticancer effects of extracts from H. musciformis demonstrating their cytotoxic effects on various cancer cell lines. The anticancer effects of these extracts are thought to be due to the presence of bioactive compounds, particularly sulfated polysaccharides, which have been shown to have anticancer and immunomodulatory effects. However, further studies are needed to fully understand the molecular mechanisms that underlie their anticancer effects and to determine their potential as therapeutic agents for cancer treatment.MethodsH. musciformis was collected from the Aegean Sea (Greece) and used for extract preparation. Transcriptome and proteome analysis was performed in liver and colon cancer human cell lines following treatment with H. musciformis seaweed extracts to characterize its anticancer effect in detail at the molecular level and to link transcriptome and proteome responses to the observed phenotypes in cancer cells.ResultsWe have identified that treatment with the seaweed extract triggers a p53-mediated response at the transcriptional and protein level in liver cancer cells, in contrast to colon cancer cells in which the effects are more associated with metabolic changes. Furthermore, we show that in treated HepG2 liver cancer cells, p53 interacts with the chromatin of several target genes and facilitates their upregulation possibly through the recruitment of the p300 co-activator.ConclusionsOverall, the available evidence suggests that extracts from H. musciformis have the potential to serve as a source of anticancer agents in liver cancer cells mainly through activation of a p53-mediated anti-tumor response that is linked to inhibition of cellular proliferation and induction of cell death.

Inhibitors of Aspartate Transcarbamoylase Inhibit Mycobacterium tuberculosis Growth.

Aspartate transcarbamoylase (ATCase) plays a key role in the second step of de novo pyrimidine biosynthesis in eukaryotes and has been proposed to be a target to suppress cell proliferation in E. coli, human cells and the malarial parasite. We hypothesized that a library of ATCase inhibitors developed for malarial ATCase (PfATCase) may also contain inhibitors of the tubercular ATCase and provide a similar inhibition of cellular proliferation. Of the 70 compounds screened, 10 showed single-digit micromolar inhibition in an in vitro activity assay and were tested for their effect on M. tuberculosis cell growth in culture. The most promising compound demonstrated a MIC90 of 4 μM. A model of MtbATCase was generated using the experimental coordinates of PfATCase. In silico docking experiments showed this compound can occupy a similar allosteric pocket on MtbATCase to that seen on PfATCase, explaining the observed species selectivity seen for this compound series.

Abstract 4045: DCC-3084, a RAF dimer inhibitor, broadly inhibits BRAF class I, II, III, BRAF fusions, and RAS-driven solid tumors leading to tumor regression in preclinical models

Abstract Background: Mutations in the RAS/MAPK pathway are a frequent driver of cancer, with oncogenic RAS or RAF mutations occurring in >30% of all cancers. First generation BRAF inhibitors are approved for use for tumors with Class I BRAF mutations (V600X). However, these drugs are not efficacious in RAF dimer mutant and RAS mutant cancers due to paradoxical activation of RAF dimers. Herein, we describe DCC-3084, a potent and selective investigational Switch Control inhibitor of BRAF and CRAF kinase dimers that targets Class I, II and III BRAF mutations, BRAF fusions, and BRAF/CRAF heterodimers. DCC-3084 combines with inhibitors of additional nodes in the MAPK pathway to potentially target a large unmet medical need in RAS and RAF mutant cancers. Methods: Inhibition of RAF kinases, including off-rate analysis, was measured using recombinant enzymes. X-ray crystallography was used for structure-based drug design. Cellular proliferation was measured using resazurin to monitor cell viability. Synergy in cells was measured using BLISS scores and curve shift analysis. Inhibition of ERK or RSK phosphorylation was measured by AlphaLISA or ELISA. Pharmacokinetics (PK) in the plasma, brain and CSF compartments were measured following oral dosing in Wistar rats. RAF and RAS mutant mouse xenograft models were used to assess PK, pharmacodynamics (PD), and efficacy. Results: DCC-3084 is a potent and selective Switch Control inhibitor of RAF dimers that was designed to target Class I, II, III BRAF mutants, BRAF fusions, and BRAF/CRAF heterodimers. DCC-3084 inhibits BRAF and CRAF, exhibiting slow off-rates (t1/2 >20 hr). Potent single-agent inhibition of MAPK pathway signaling and cellular proliferation was observed in a wide range of Class I, II, III BRAF and BRAF fusion altered cell lines. Synergy was observed in combination with inhibitors of other nodes in the RAS/MAPK pathway in RAS mutant cell lines. DCC-3084 was demonstrated to be CNS penetrable and exhibited dose dependent oral exposure with robust inhibition of the RAS/MAPK pathway in PK/PD models. DCC-3084 accumulated in tumor tissue relative to plasma, further demonstrating a favorable pharmaceutical profile. Oral treatment of DCC-3084 as a single agent resulted in tumor regression in BRAF mutant and KRAS Q61K mutant mouse xenograft models and tumor growth inhibition in KRAS G12C/D mutant models. Additionally, DCC-3084 in combination with a MEKi resulted in tumor regression in KRAS mutant models. Conclusions: The Switch Control inhibitor DCC-3084 broadly inhibits Class I, II and III BRAF mutations, BRAF fusions, and BRAF/CRAF heterodimers leading to tumor regression in preclinical models. The overall preclinical profile of DCC-3084 supports IND-enabling activities towards clinical development in a key area of unmet medical need in RAS and RAF mutant cancers. Citation Format: Stacie L. Bulfer, Bertrand Le Bourdonnec, Jeffery D. Zwicker, Yu Mi Ahn, Gada Al-Ani, Hikmat Al-Hashimi, Chase Crawley, Kristin M. Elliott, Saqib Faisal, Andrew M. Harned, Cale L. Heiniger, Molly M. Hood, Salim Javed, Michael Kennedy, Joshua W. Large, Cynthia B. Leary, Wei-Ping Lu, Kylie Luther, Max D. Petty, Hunter R. Picard, Justin T. Proto, Yeni K. Romero, Forrest A. Stanley, Kristen L. Stoltz, Daniel C. Tanner, Hanumaiah Telikepalli, Mary J. Timson, Lakshminarayana Vogeti, Subha Vogeti, Sihyung Yang, Lexy H. Zhong, Bryan D. Smith, Daniel L. Flynn. DCC-3084, a RAF dimer inhibitor, broadly inhibits BRAF class I, II, III, BRAF fusions, and RAS-driven solid tumors leading to tumor regression in preclinical models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4045.

Abstract 3415: Preclinical characterization of a brain penetrant RAF inhibitor, BDTX-4933, targeting oncogenic BRAF Class I/II/III and RAS mutations

Abstract Mutations in BRAF and RAS are often oncogenic and lead to a constitutively active MAPK pathway that promotes aberrant cell proliferation and tumor growth. Currently approved BRAF inhibitors are selective against monomeric BRAF V600 mutants. These drugs are largely inactive against non-V600 dimeric BRAF mutants and have poor brain penetration. Although there is an FDA-approved KRAS G12C mutant-selective inhibitor, there are no approved inhibitors for cancer patients who harbor other (non-G12C) KRAS and NRAS mutations which promote tumor growth likely through constitutively active RAF dimers. There remains a high unmet clinical need for a highly CNS penetrant oral RAF inhibitor that targets a broad spectrum of BRAF mutations and constitutively active RAF dimers without paradoxical activation of the MAPK signaling pathway. BDTX-4933 is a potent, reversible, CNS penetrant RAF MasterKey inhibitor designed to target a large family of oncogenic BRAF mutations including BRAF monomers and RAF dimers. The compound inhibits not only all classes (I, II, and III) of BRAF mutations but also targets constitutively active RAF dimers promoted by upstream oncogenic MAPK pathway alterations, such as RAS mutations. In a panel of cancer cell lines that endogenously express BRAF or RAS mutations, BDTX-4933 demonstrates inhibition of the MAPK pathway signaling without paradoxical activation, resulting in potent inhibition of cellular proliferation. BDTX-4933 shows target engagement, inhibiting ERK phosphorylation, in tumor models in vivo, achieving strong anti-tumor efficacy and tumor regression across tumor models driven by either BRAF or RAS mutations. Furthermore, BDTX-4933 exhibits high CNS exposure leading to dose-dependent tumor growth inhibition, and survival benefit in mice implanted intracranially with xenograft BRAF mutant tumors. BDTX-4933 has a best-in-class profile to treat cancer patients harboring BRAF mutations or RAF dimer-promoting upstream genetic alterations. IND-enabling studies for BDTX-4933 are on-going. Citation Format: Yoon-Chi Han, Pui-Yee Ng, Luisa Shin Ogawa, Shao Ning Yang, Miao Chen, Noboru Ishiyama, Tai-An Lin, Elizabeth Buck. Preclinical characterization of a brain penetrant RAF inhibitor, BDTX-4933, targeting oncogenic BRAF Class I/II/III and RAS mutations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3415.

Integrating network pharmacology approaches for the investigation of multi-target pharmacological mechanism of 6-shogaol against cervical cancer

Traditional treatment of cancer has been plagued by a number of obstacles, such as multiple drug resistance, toxicity and financial constraints. In contrast, phytochemicals that modulate a variety of molecular mechanisms are garnering increasing interest in complementary and alternative medicine. Therefore, an approach based on network pharmacology was used in the present study to explore possible regulatory mechanisms of 6-shogaol as a potential treatment for cervical cancer (CC). A number of public databases were screened to collect information on the target genes of 6-shogaol (SuperPred, Targetnet, Swiss target prediction and PharmMapper), while targets pertaining to CC were taken from disease databases (DisGeNet and Genecards) and gene expression omnibus (GEO) provided expression datasets. With STRING and Cytoscape, protein-protein interactions (PPI) were generated and topology analysis along with CytoNCA were used to identify the Hub genes. The Gene Ontology (GO) database Enrichr was used to annotate the target proteins, while, using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, signaling pathway enrichment analysis was conducted. Molecular docking and survival analysis for the Hub genes revealed four genes (HSP90AA1, HRAS, ESR1 and EGFR) with lowest binding energy and majority of the Hub genes (EGFR, SRC, CASP-3, HSP90AA1, MTOR, MAPK-1, MDM2 and ESR1) were linked with the overall survival of CC patients. In conclusion, the present study provides the scientific evidence which strongly supports the use of 6-shogoal as an inhibitor of cellular proliferation, growth, migration as well as inducer of apoptosis via targeting the hub genes involved in the growth of CC.Communicated by Ramaswamy H. Sarma

Acid/reduction dual-sensitive amphiphilic graft polyurethane with folic acid and detachable poly(ethylene glycol) as anticancer drug delivery carrier

In order to not only improve the stability of nanomicelles in blood circulation but also promote the cellular uptake in tumors and rapidly release the encapsulated drugs in tumor cells, a kind of acid/reduction dual-sensitive amphiphilic graft polyurethane with folic acid and detachable poly(ethylene glycol) (FA-PUSS-gimi-mPEG) was synthesized by grafting folic acid and monomethoxy poly(ethylene glycol) to the polyurethane side chain. FA-PUSS-gimi-mPEG could self-assemble in aqueous solution to form negatively charged nanomicelles, which endowed them good stability under normal physiological condition. Using ultraviolet-visible spectrometer (UV–vis) and dynamic light scattering (DLS), it was found that the hydrophilic poly(ethylene glycol) layer of FA-PUSS-gimi-mPEG micelles could be detached due to the cleavage of benzoic-imine bond under slightly acidic condition, which resulted in reversing the charge of the micellar surface and exposing folic acid to the micellar surface. FA-PUSS-gimi-mPEG micelles could load doxorubicin (DOX), moreover the drug release rate was faster at pH 5.0 and 10 mM glutathione (GSH) than that under normal physiological condition. The results of cell experiments further demonstrated that FA-PUSS-gimi-mPEG micelles had acid/reduction dual-sensitive property. The changes in the structure of FA-PUSS-gimi-mPEG micelles could enhance the cellular uptake under acid condition and the micelles could accelerate the drug release in tumor cells due to the presence of disulfide bonds in the polymer. Therefore, FA-PUSS-gimi-mPEG micelles could efficiently deliver anticancer drug into tumor cells and enhance the inhibition of cellular proliferation through multi-effect synergy.

Differential Gene Expression Profile in DNA Damage Signaling Pathways, in De Novo Acute Myeloid Leukemia Patients before Induction Chemotherapy

Introduction: Acute Myeloid Leukemia (AML) is a life-threatening myeloid malignancy with poor prognosis. Response to induction treatment determines long-term prognosis; however, mechanisms underlying response have not been thoroughly investigated. DNA damage and repair mechanisms influence not only the genetic predisposition to leukemia but are also important for refractoriness to treatment. Aim: The aim of this study is to investigate the possible alterations in the gene expression profile of DNA damage signaling pathways in two leukemic cell lines following their exposure to chemotherapeutic agents, namely, idarubicin and cytarabine, and to verify the findings in AML patient samples at diagnosis before the onset of chemotherapy. Methods: Cell lines Kasumi-1 with the t(8;21) and MV4-11 (biphenotypic B-myelomonocytic leukemia) were exposed either to idarubicin (0.1μΜ) for 6h or cytarabine (1μΜ) for 48h. Dead cells were eliminated from drug-treated cells using an appropriate commercial kit. Gene expression profiling through PCR arrays analysis (RT2 Profiler, Qiagen) was performed in triplicate after RNA extraction from untreated, chemotherapy-treated and live (chemo resistant) cells following chemotherapy exposure. Human DNA damage signaling pathway-related gene expression and statistical analysis was performed through the RT2 Profiler PCR Array data analysis tool. Moreover, mononuclear cells were isolated from bone marrow of 11 lymphoma patients, used as controls, and from 64 AML patients at diagnosis who subsequently received chemotherapy with the 7+3 regimen. Response to first-line chemotherapy was recorded and patients were assigned in two groups: Responders (n=53 patients) and non-Responders (n=11 patients). Total RNA was isolated (RNeasy plus Mini Kit, Qiagen), quantified and cDNA was synthesized. Relative expression of genes of interest was quantified using SYBR® Greenbased real-time RT-PCR (QuantiTect® Primer Assay, Qiagen) based on the ddCt method (Housekeeping gene GADPH). All samples were obtained from patients after informed consent and treated in accordance with the Declaration of Helsinki. Statistical analysis was performed using R Statistical Software (version 4.2.1). Results: Five genes were found significantly [p < 0.05, Students t-test] up-regulated in live cells from either leukemic cell lines after treatment with both agents and had a FoldChange > 2. More specifically PPP1R15A, CDKN1A and GADD45G genes were up-regulated in both live cell lines, GADD45A in live MV4-11 cells and EXO1 in live Kasumi cells (Figure 1A). The aforementioned genes were selected and their expression levels were quantified in human bone marrow samples. CDKN1A and GADD45G were found down-regulated in Responders [p=0.032 and p=0.001 respectively; Welch t-test] while PPP1R15A was significantly up-regulated [over 8-fold change, p= 2.08E-09; Welch t-test] compared to controls. Likewise, GADD45G and PPP1R15A were also significantly down and up regulated respectively, in non-Responders [p=0.08 and p=1.51E-05, Welch t-test]. Interestingly, PPP1R15A presented a 1.7-fold overexpression trend in non-Responders compared to Responders [p=0.18, Welch t-test] (Figure 1B). Conclusion: The results of our study suggest a dysregulation of the DNA damage and repair pathways in AML. Upregulation of GADD45G is known to dramatically induce apoptosis, differentiation, and growth arrest while increasing sensitivity of AML cells to chemotherapeutic drugs (Guo, Dan et al., Blood 2021). CDKN1A is involved in p53/TP53 mediated inhibition of cellular proliferation in response to DNA damage. Both were down-regulated in AML patients and are likely to play an important role in the pathogenesis of AML. PPP1R15A participates in DNA damage response facilitating the recovery of cells from stress. The up-regulation of PPP1R15A both in live cells treated with Ara-C and Cytarabine, as well as the 1.7-fold overexpression trend in non-Responders compared to Responders, supports the notion that PPP1R15A could be associated with resistance to induction treatment, which may have significant prognostic and therapeutic implications. A greater sample of AML patients both responding and not responding to chemotherapy, would be essential to delineate the role of genes involved in DNA damage and repair pathways in resistance to induction treatment. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Novel protective effect of diosmin against cisplatin-induced prostate and seminal vesicle damage: Role of oxidative stress and apoptosis

Multiple organ toxicity has been associated with cisplatin (CIS) treatment, limiting its clinical use. The human prostate and seminal vesicles are accessory sex organs with androgen-dependent morphogenesis, growth, and secretion. The present study aimed to investigate, for the first time, the toxic effect of CIS on normal prostate and seminal vesicles in the presence and absence of diosmin (DS). The animals were randomized into 4 groups as follows: control (received vehicle), CIS group (7.5 mg/kg, i.p. on 5th and 12th day), DS group (100 mg/kg, p.o. for 15 days), and DS+CIS group. Histopathological and biochemical analyses were conducted to elucidate the goal of this study. CIS administration significantly induced prostate and seminal vesicle toxicity as evidenced by alteration of serum testosterone, LH, FSH, PSA, steroidogenic HSD17B6 as well as seminal analysis markers. Remarkably, marked histopathological changes in thin and ultrathin structures were observed. Besides, CIS significantly boosted oxidative stress as evidenced by the up-regulation of MDA and down-regulation of TAC. CIS significantly induced tissue apoptosis concomitant with suppression of cellular proliferation and stem cell expression as indicated by up-regulation of activated caspase-3 and Bax expression along with down-regulation of Bcl-2, Ki67, and CD44 expression. Interestingly, DS fixed all disturbances in the prostate and seminal vesicles induced by CIS. Together, CIS could cause prostate and seminal vesicle toxicity by affecting hormonal, steroidogenic, oxidative stress, apoptosis, and proliferation processes, and this effect was reversed by DS administration.