Cell plasticity modulation by flavonoids in resistant breast carcinoma targeting the nuclear factor kappa B signaling.

Cancer cell plasticity plays a crucial role in tumor initiation, progression, and metastasis and is implicated in the multiple cancer defense mechanisms associated with therapy resistance and therapy evasion. Cancer resistance represents one of the significant obstacles in the clinical management of cancer. Some reversal chemosensitizing agents have been developed to resolve this serious clinical problem, but they have not yet been proven applicable in oncological practice. Activated nuclear factor kappa B (NF-κB) is a frequently observed biomarker in chemoresistant breast cancer (BC). Therefore, it denotes an attractive cellular target to mitigate cancer resistance. We summarize that flavonoids represent an essential class of phytochemicals that act as significant regulators of NF-κB signaling and negatively affect the fundamental cellular processes contributing to acquired cell plasticity and drug resistance. In this regard, flavokawain A, icariin, alpinetin, genistein, wogonin, apigenin, oroxylin A, xanthohumol, EGCG, hesperidin, naringenin, orientin, luteolin, delphinidin, fisetin, norwogonin, curcumin, cardamonin, methyl gallate and catechin-3-O-gallate, ampelopsin, puerarin, hyperoside, baicalein, paratocarpin E, and kaempferol and also synthetic flavonoids such as LFG-500 and 5,3'-dihydroxy-3,6,7,8,4'-pentamethoxyflavone have been reported to specifically interfere with the NF-κB pathway with complex signaling consequences in BC cells and could be potentially crucial in re-sensitizing unresponsive BC cases. The targeting NF-κB by above-mentioned flavonoids includes the modification of tumor microenvironment and epithelial-mesenchymal transition, growth factor receptor regulations, and modulations of specific pathways such as PI3K/AKT, MAP kinase/ERK, and Janus kinase/signal transduction in BC cells. Besides that, NF-κB signaling in BC cells modulated by flavonoids has also involved the regulation of ATP-binding cassette transporters, apoptosis, autophagy, cell cycle, and changes in the activity of cancer stem cells, oncogenes, or controlling of gene repair. The evaluation of conventional therapies in combination with plasticity-regulating/sensitizing agents offers new opportunities to make significant progress towards a complete cure for cancer.

Therapeutic antibodies for the prevention and treatment of cancer.

The developments of antibodies for cancer therapeutics have made remarkable success in recent years. There are multiple factors contributing to the success of the biological molecule including origin of the antibody, isotype, affinity, avidity and mechanism of action. With better understanding of mechanism of cancer progression and immune manipulation, recombinant formats of antibodies are used to develop therapeutic modalities for manipulating the immune cells of patients by targeting specific molecules to control the disease. These molecules have been successful in minimizing the side effects instead caused by small molecules or systemic chemotherapy but because of the developing therapeutic resistance against these antibodies, combination therapy is thought to be the best bet for patient care. Here, in this review, we have discussed different aspects of antibodies in cancer therapy affecting their efficacy and mechanism of resistance with some relevant examples of the most studied molecules approved by the US FDA.

Molecular pathways and therapeutic targets linked to triple-negative breast cancer (TNBC).

Cancer is a group of diseases characterized by uncontrolled cellular growth, abnormal morphology, and altered proliferation. Cancerous cells lose their ability to act as anchors, allowing them to spread throughout the body and infiltrate nearby cells, tissues, and organs. If these cells are not identified and treated promptly, they will likely spread. Around 70% of female breast cancers are caused by a mutation in the BRCA gene, specifically BRCA1. The absence of progesterone, oestrogen and HER2 receptors (human epidermal growth factor) distinguishes the TNBC subtype of breast cancer. There were approximately 6,85,000 deaths worldwide and 2.3 million new breast cancer cases in women in 2020. Breast cancer is the most common cancer globally, affecting 7.8 million people at the end of 2020. Compared to other cancer types, breast cancer causes more women to lose disability-adjusted life years (DALYs). Worldwide, women can develop breast cancer at any age after puberty, but rates increase with age. The maintenance of mammary stem cell stemness is disrupted in TNBC, governed by signalling cascades controlling healthy mammary gland growth and development. Interpreting these essential cascades may facilitate an in-depth understanding of TNBC cancer and the search for an appropriate therapeutic target. Its treatment remains challenging because it lacks specific receptors, which renders hormone therapy and medications ineffective. In addition to radiotherapy, numerous recognized chemotherapeutic medicines are available as inhibitors of signalling pathways, while others are currently undergoing clinical trials. This article summarizes the vital druggable targets, therapeutic approaches, and strategies associated with TNBC.

YAP/TEAD involvement in resistance to paclitaxel chemotherapy in lung cancer.

The Yes-associated protein (YAP) oncoprotein has been linked to both metastases and resistance to targeted therapy of lung cancer cells. We aimed to investigate the effect of YAP pharmacological inhibition, using YAP/TEA domain (TEAD) transcription factor interaction inhibitors in chemo-resistant lung cancer cells. YAP subcellular localization, as a readout for YAP activation, cell migration, and TEAD transcription factor functional transcriptional activity were investigated in cancer cell lines with up-regulated YAP, with and without YAP/TEAD interaction inhibitors. Parental (A549) and paclitaxel-resistant (A549R) cell transcriptomes were analyzed. The half-maximal inhibitory concentration (IC50) of paclitaxel or trametinib, which are Mitogen-Activated protein kinase and Erk Kinase (MEK) inhibitors, combined with a YAP/TEAD inhibitor (IV#6), was determined. A three-dimensional (3D) microfluidic culture device enabled us to study the effect of IV#6/paclitaxel combination on cancer cells isolated from fresh resected lung cancer samples. YAP activity was significantly higher in paclitaxel-resistant cell lines. The YAP/TEAD inhibitor induced a decreased YAP activity in A549, PC9, and H2052 cells, with reduced YAP nuclear staining. Wound healing assays upon YAP inhibition revealed impaired cell motility of lung cancer A549 and mesothelioma H2052 cells. Combining YAP pharmacological inhibition with trametinib in K-Ras mutated A549 cells recapitulated synthetic lethality, thereby sensitizing these cells to MEK inhibition. The YAP/TEAD inhibitor lowered the IC50 of paclitaxel in A549R cells. Differential transcriptomic analysis of parental and A549R cells revealed an increased YAP/TEAD transcriptomic signature in resistant cells, downregulated upon YAP inhibition. The YAP/TEAD inhibitor restored paclitaxel sensitivity of A549R cells cultured in a 3D microfluidic system, with lung cancer cells from a fresh tumor efficiently killed by YAP/TEAD inhibitor/paclitaxel doublet. Evidence of the YAP/TEAD transcriptional program's role in chemotherapy resistance paves the way for YAP therapeutic targeting.

N6-methyladenosine demethylase FTO enhances chemo-resistance in colorectal cancer through SIVA1-mediated apoptosis.

N6-methyladenosine (m6A) is the most pervasive RNA modification and is recognized as a novel epigenetic regulation in RNA metabolism. Although the m6A modification involves various physiological processes, its roles in drug resistance in colorectal cancer (CRC) still remain unknown. We analyzed the RNA expression profile of m6A/A (%) with MRM mass spectrometry in human 5-fluorouracil (5-FU)-resistant CRC tissues, and used the m6A RNA immunoprecipitation assay to validate the m6A-regulated target. Our results have shown that the m6A demethylase FTO was up-regulated in human primary and 5-FU-resistant CRC. Depletion of FTO decreased cell growth, colony formation and metastasis in 5-FU-resistant CRC cells in vitro and in vivo. Mechanistically, we identified SIVA1, a critical apoptotic gene, as a key downstream target of the FTO-mediated m6A demethylation. The m6A demethylation of SIVA1 at the CDS region induced its mRNA degradation via a YTHDF2-dependent mechanism. The SIVA1 levels were negatively correlated with the FTO levels in clinical CRC tissues. Notably, inhibition of FTO significantly reduced the tolerance of 5-FU in 5-FU-resistant CRC cells via the FTO-SIVA1 axis, whereas SIVA1-depletion could restore the m6A-dependent 5-FU sensitivity in CRC cells. In summary, our findings demonstrate a critical role of FTO as an m6A demethylase enhancing chemo-resistance in CRC cells, and suggest that FTO inhibition may restore the sensitivity of chemo-resistant CRC cells to 5-FU.

Unravelling the role of Silibinin in targeting CD44+ cancer stem cells: Therapeutic implications, effective strategies and approaches.

CD44+ cancer stem cells (CSCs) are believed to account for drug resistance and tumour recurrence due to their potential to self-renew and differentiate into heterogeneous lineages. Therefore, efficient treatment strategies targeting and eliminating these CSCs are required. The flavonolignan, Silibinin, has gained immense attention in targeting CD44+ CSCs as it alters functional properties like cell cycle arrest, apoptosis, inhibition of invasion and metastasis and also inhibits a range of molecular pathways. However, its limited bioavailability is a major hurdle in asserting Silibinin as a translational therapeutic agent. Combinatorial therapy of Silibinin with conventional chemotherapeutic drugs is an alternative approach in targeting CD44+ CSCs as it increases the efficacy and reduces the cytotoxicity of chemotherapeutic drugs, thus preventing drug resistance. Certain Silibinin-conjugated nano-formulations have also been successfully developed, through which there is improved absorptivity/bioavailability of Silibinin and a decrease in the concentration of therapeutic drugs leading to reduced cytotoxicity. In this review, we summarise the effectiveness of the synergistic therapeutic approach for Silibinin in targeting the molecular mechanisms of CD44+ CSCs and emphasise the potential role of Silibinin as a novel therapeutic agent.

Cannabigerol Induces Autophagic Cell Death by Inhibiting EGFR-RAS Pathways in Human Pancreatic Ductal Adenocarcinoma Cell Lines.

Pancreatic ductal adenocarcinoma (PDAC) is the most frequent infiltrating type of pancreatic cancer. The poor prognosis associated with this cancer is due to the absence of specific biomarkers, aggressiveness, and treatment resistance. PDAC is a deadly malignancy bearing distinct genetic alterations, the most common being those that result in cancer-causing versions of the KRAS gene. Cannabigerol (CBG) is a non-psychomimetic cannabinoid with anti-inflammatory properties. Regarding the anticancer effect of CBG, up to now, there is only limited evidence in human cancers. To fill this gap, we investigated the effects of CBG on the PDAC cell lines, PANC-1 and MIAPaCa-2. The effect of CBG activity on cell viability, cell death, and EGFR-RAS-associated signaling was investigated. Moreover, the potential synergistic effect of CBG in combination with gemcitabine (GEM) and paclitaxel (PTX) was investigated. MTT was applied to investigate the effect of CBG on PDAC cell line viabilities. Annexin-V and Acridine orange staining, followed by cytofluorimetric analysis and Western blotting, were used to evaluate CBG's effect on cell death. The modulation of EGFR-RAS-associated pathways was determined by Western blot analysis and a Milliplex multiplex assay. Moreover, by employing the MTT data and SynergyFinder Plus software analysis, the effect of the combination of CBG and chemotherapeutic drugs was determined.

Polyploid giant cancer cell characterization: New frontiers in predicting response to chemotherapy in breast cancer.

Although polyploid cells were first described nearly two centuries ago, their ability to proliferate has only recently been demonstrated. It also becomes increasingly evident that a subset of tumor cells, polyploid giant cancer cells (PGCCs), play a critical role in the pathophysiology of breast cancer (BC), among other cancer types. In BC, PGCCs can arise in response to therapy-induced stress. Their progeny possess cancer stem cell (CSC) properties and can repopulate the tumor. By modulating the tumor microenvironment (TME), PGCCs promote BC progression, chemoresistance, metastasis, and relapse and ultimately impact the survival of BC patients. Given their pro- tumorigenic roles, PGCCs have been proposed to possess the ability to predict treatment response and patient prognosis in BC. Traditionally, DNA cytometry has been used to detect PGCCs.. The field will further derive benefit from the development of approaches to accurately detect PGCCs and their progeny using robust PGCC biomarkers. In this review, we present the current state of knowledge about the clinical relevance of PGCCs in BC. We also propose to use an artificial intelligence-assisted image analysis pipeline to identify PGCC and map their interactions with other TME components, thereby facilitating the clinical implementation of PGCCs as biomarkers to predict treatment response and survival outcomes in BC patients. Finally, we summarize efforts to therapeutically target PGCCs to prevent chemoresistance and improve clinical outcomes in patients with BC.

Bladder cancer: therapeutic challenges and role of 3D cell culture systems in the screening of novel cancer therapeutics.

Bladder cancer (BC) is the sixth most common worldwide urologic malignancy associated with elevated morbidity and mortality rates if not well treated. The muscle-invasive form of BC develops in about 25% of patients. Moreover, according to estimates, 50% of patients with invasive BC experience fatal metastatic relapses. Currently, resistance to drug-based therapy is the major tumble to BC treatment. The three-dimensional (3D) cell cultures are clearly more relevant not only as a novel evolving gadget in drug screening but also as a bearable therapeutic for different diseases. In this review, various subtypes of BC and mechanisms of drug resistance to the commonly used anticancer therapies are discussed. We also summarize the key lineaments of the latest cell-based assays utilizing 3D cell culture systems and their impact on understanding the pathophysiology of BC. Such knowledge could ultimately help to address the most efficient BC treatment.

Role of the long non-coding RNAs in regulation of Gemcitabine response in tumor cells.

Chemotherapy is widely used as one of the first line therapeutic methods in cancer patients. However, chemotherapeutic resistance is one of the most common problems in cancer patients, which leads to the therapeutic failure and tumor relapse. Considering the side effects of chemotherapy drugs in normal tissues, it is required to investigate the molecular mechanisms involved in drug resistance to improve the therapeutic strategies in cancer patients. Long non-coding RNAs (lncRNAs) have pivotal roles in regulation of cellular processes associated with drug resistance. LncRNAs deregulations have been frequently reported in a wide range of chemo-resistant tumors. Gemcitabine (GEM) as a nucleoside analog has a wide therapeutic application in different cancers. However, GEM resistance is considered as a therapeutic challenge. Considering the role of lncRNAs in the occurrence of GEM resistance, in the present review we discussed the molecular mechanisms of lncRNAs in regulation of GEM response among cancer patients. It has been reported that lncRNAs have mainly an oncogenic role as the inducers of GEM resistance through direct or indirect regulation of transcription factors, autophagy, polycomb complex, and signaling pathways such as PI3K/AKT, MAPK, WNT, JAK/STAT, and TGF-ß. This review paves the way to present the lncRNAs as non-invasive markers to predict GEM response in cancer patients. Therefore, lncRNAs can be introduced as the efficient markers to reduce the possible chemotherapeutic side effects in GEM resistant cancer patients and define a suitable therapeutic strategy among these patients.

FAP, CD10, and GPR77-labeled CAFs cause neoadjuvant chemotherapy resistance by inducing EMT and CSC in gastric cancer.

OBJECTIVE: A significant proportion of patients can not benefit from neoadjuvant chemotherapy (NCT) due to drug resistance. Cancer-associated fibroblasts (CAFs) influence many biological behaviours of tumors, including chemo-resistance. This study aims to explore whether CAFs expressing FAP, CD10, and GPR77 affect the efficacy of NCT and the prognosis of patients with gastric cancer, and its mechanism. METHODS: One hundred seventy-one patients with locally progressive gastric adenocarcinoma who had undergone NCT and radical surgery were collected. Immunohistochemistry was used to detect the expression of FAP, CD10, and GPR77 in CAFs; the EMT markers (N-cadherin, Snail1, and Twist1) and the CSC markers (ALDH1, CD44, and LGR5) in gastric cancer cells. The χ2 test was used to analyze the relationship between the expression of CAF, EMT, and CSC markers and the clinicopathological factors, as well as the relationship between CAF markers and EMT, and CSC markers. Logistic regression and Cox risk regression were used to analyze the relationship between the expression of CAF, EMT, and CSC markers and TRG grading and OS; Kaplan-Meier analysis was used for survival analysis and plotting the curves. RESULTS: The expression of CAF markers FAP, CD10, and GPR77 was closely associated with that of EMT markers; FAP and CD10 were closely related to CSC markers. In the univariate analysis of pathological response, CAF markers (FAP, CD10, GPR77), EMT markers (N-cadherin, Snail1, Twist1), and CSC markers (ALDH1, LGR5, CD44), were all closely associated with pathological response (all p < 0.05). Only Twist1 was an independent factor affecting pathological response in multifactorial analysis (p = 0.001). In a univariate analysis of OS, expression of FAP and CD10 in CAF, as well as expression of EMT biomarkers (N-cadherin, Snail1), were significant factors influencing patient prognosis (all p < 0.05). Multifactorial analysis revealed N-cadherin (p = 0.032) and Snail1 (p = 0.028), as independent prognostic factors affecting OS. CONCLUSION: FAP, CD10, and GPR77 labeled CAF subgroup may lead to NCT resistance and poor prognosis by inducing EMT and CSC of gastric cancer cells in locally advanced gastric cancer patients.

Targeted Silencing of NRF2 by rituximab-conjugated nanoparticles increases the sensitivity of chronic lymphoblastic leukemia cells to Cyclophosphamide.

BACKGROUND: Targeting influential factors in resistance to chemotherapy is one way to increase the effectiveness of chemotherapeutics. The nuclear factor erythroid 2-related factor 2 (Nrf2) pathway overexpresses in chronic lymphocytic leukemia (CLL) cells and appears to have a significant part in their survival and chemotherapy resistance. Here we produced novel nanoparticles (NPs) specific for CD20-expressing CLL cells with simultaneous anti-Nrf2 and cytotoxic properties. METHODS: Chitosan lactate (CL) was used to produce the primary NPs which were then respectively loaded with rituximab (RTX), anti-Nrf2 Small interfering RNA (siRNAs) and Cyclophosphamide (CP) to prepare the final version of the NPs (NP-Nrf2_siRNA-CP). All interventions were done on both peripheral blood mononuclear cells (PBMCs) and bone marrow mononuclear cells (BMNCs). RESULTS: NP-Nrf2_siRNA-CP had satisfying physicochemical properties, showed controlled anti-Nrf2 siRNA/CP release, and were efficiently transfected into CLL primary cells (both PBMCs and BMNCs). NP-Nrf2_siRNA-CP were significantly capable of cell apoptosis induction and proliferation prevention marked by respectively decreased and increased anti-apoptotic and pro-apoptotic factors. Furthermore, use of anti-Nrf2 siRNA was corresponding to elevated sensitivity of CLL cells to CP. CONCLUSION: Our findings imply that the combination therapy of malignant CLL cells with RTX, CP and anti-Nrf2 siRNA is a novel and efficient therapeutic strategy that was capable of destroying malignant cells. Furthermore, the use of NPs as a multiple drug delivery method showed fulfilling properties; however, the need for further future studies is undeniable. Video Abstract.

RAD51D Secondary Mutation-Mediated Resistance to PARP-Inhibitor-Based Therapy in HGSOC.

Ovarian cancer is the leading cause of gynecologic cancer-related death, and PARP inhibitors (PARPis) are becoming a promising treatment option, as demonstrated by recent clinical trials. After PARPi exposure, somatic reversion mutations in the homologous recombination genes may be a mechanism of PARPi resistance in ovarian carcinoma. We present an ovarian cancer case of a 61-year-old woman, who underwent routine tumor reduction surgery followed by platinum and PARPis. She demonstrated a good response to PARPis for 15 months before recurrence and secondary tumor reduction surgery. However, post-surgery platinum and PARPi treatment only kept the disease stable for 5 months. A potential molecular mechanism for PARPi resistance was investigated using next-generation sequencing, immunohistochemical (IHC) staining, and other functional assays. A germline RAD51D loss-of-function mutation was found in the reported case (LRG_516t1:c.270_271dup p1:p.(Lys91fs*13)). Subsequently, a secondary mutation (LRG_516t1:c.271_282 del) was identified in the same locus of the germline duplication in the post-progression biopsies and ctDNA. The IHC staining supported low expression of RAD51D in the initial tumor tissue, but the expression was restored after the correction of the open reading frame by the secondary mutation. The in vitro results supported that the loss-of-function mutation of RAD51D was the basis for the initial response to the platinum and PARPi therapy, while the newly acquired reversion mutation could be attributed to the observed PARPi resistance. An acquired mutation can reverse a loss-of-function change in RAD51D and can result in PARPi resistance in a hereditary ovarian cancer patient. Liquid biopsy could be considered for longitudinal monitoring in ovarian patients under PARPi-based therapy, which can identify acquired resistant mutations earlier and facilitate precision management.

Estrogen Receptor ß4 Regulates Chemotherapy Resistance and Induces Cancer Stem Cells in Triple Negative Breast Cancer.

Triple Negative Breast Cancer (TNBC) has the worst prognosis among all breast cancers, and survival in patients with recurrence is rarely beyond 12 months due to acquired resistance to chemotherapy, which is the standard of care for these patients. Our hypothesis is that Estrogen Receptor ß1 (ERß1) increases response to chemotherapy but is opposed by ERß4, which it preferentially dimerizes with. The role of ERß1 and ERß4 in influencing chemotherapy sensitivity has never been studied before. CRISPR/CAS9 was used to truncate ERß1 Ligand Binding Domain (LBD) and knock down the exon unique to ERß4. We show that the truncated ERß1 LBD in a variety of mutant p53 TNBC cell lines, where ERß1 ligand dependent function was inactivated, had increased resistance to Paclitaxel, whereas the ERß4 knockdown cell line was sensitized to Paclitaxel. We further show that ERß1 LBD truncation, as well as treatment with ERß1 antagonist 2-phenyl-3-(4-hydroxyphenyl)-5,7-bis(trifluoromethyl)-pyrazolo[1,5-a] pyrimidine (PHTPP), leads to increase in the drug efflux transporters. Hypoxia Inducible Factors (HIFs) activate factors involved in pluripotency and regulate the stem cell phenotype, both in normal and cancer cells. Here we show that the ERß1 and ERß4 regulate these stem cell markers like SOX2, OCT4, and Nanog in an opposing manner; and we further show that this regulation is mediated by HIFs. We show the increase of cancer cell stemness due to ERß1 LBD truncation is attenuated when HIF1/2α is knocked down by siRNA. Finally, we show an increase in the breast cancer stem cell population due to ERß1 antagonist using both ALDEFLUORTM and SOX2/OCT4 response element (SORE6) reporters in SUM159 and MDA-MB-231 cell lines. Since most TNBC cancers are ERß4 positive, while only a small proportion of TNBC patients are ERß1 positive, we believe that simultaneous activation of ERß1 with agonists and inactivation of ERß4, in combination with paclitaxel, can be more efficacious and yield better outcome for chemotherapy resistant TNBC patients.

Striatins and STRIPAK complex partners in clinical outcomes of patients with breast cancer and responses to drug treatment.

Objective: Striatins (STRNs) family, which contains three multi-domain scaffolding proteins, are cornerstones of the striatins interacting phosphatase and kinase (STRIPAK) complex. Although the role of the STRIPAK complex in cancer has become recognized in recent years, its clinical significance in breast cancer has not been fully established. Methods: Using a freshly frozen breast cancer tissue cohort containing both cancerous and adjacent normal mammary tissues, we quantitatively evaluated the transcript-level expression of all members within the STRIPAK complex along with some key interacting and regulatory proteins of STRNs. The expression profile of each molecule and the integrated pattern of the complex members were assessed against the clinical-pathological factors of the patients. The Cancer Genome Atlas (TCGA) dataset was used to evaluate the breast cancer patients' response to chemotherapies. Four human breast cancer cell lines, MDA-MB-231, MDA-MB-361, MCF-7, and SK-BR-3, were subsequently adopted for in vitro work. Results: Here we found that high-level expressions of STRIP2, calmodulin, CCM3, MINK1 and SLMAP were respectively associated with shorter overall survival (OS) of patients. Although the similar pattern observed for STRN3, STRN4 and a contrary pattern observed for PPP2CA, PPP2CB and PPPR1A were not significant, the integrated expression profile of STRNs group and PPP2 group members constitutes a highly significant prognostic indicator for OS [P<0.001, hazard ratio (HR)=2.04, 95% confidence interval (95% CI), 1.36-3.07] and disease-free survival (DFS) (P=0.003, HR=1.40, 95% CI, 1.12-1.75). Reduced expression of STRN3 has an influence on the biological functions including adhesiveness and migration. In line with our clinical findings, the breast cancer cells responded to STRN3 knockdown with changes in their chemo-sensitivity, of which the response is also breast cancer subtype dependent. Conclusions: Our results suggest a possible role of the STRIPAK complex in breast cancer development and prognosis. Among the members, the expression profile of STRN3 presents a valuable factor for assessing patients' responses to drug treatment.

AP4 suppresses DNA damage, chromosomal instability and senescence via inducing MDC1/Mediator of DNA damage Checkpoint 1 and repressing MIR22HG/miR-22-3p.

BACKGROUND: AP4 (TFAP4) encodes a basic helix-loop-helix leucine zipper (bHLH-LZ) transcription factor and is a direct target gene of the oncogenic transcription factor c-MYC. Here, we set out to determine the relevance of AP4 in human colorectal cancer (CRC) cells. METHODS: A CRISPR/Cas9 approach was employed to generate AP4-deficient CRC cell lines with inducible expression of c-MYC. Colony formation, ß-gal staining, immunofluorescence, comet and homologous recombination (HR) assays and RNA-Seq analysis were used to determine the effects of AP4 inactivation. qPCR and qChIP analyses was performed to validate differentially expressed AP4 targets. Expression data from CRC cohorts was subjected to bioinformatics analyses. Immunohistochemistry was used to evaluate AP4 targets in vivo. Ap4-deficient APCmin/+ mice were analyzed to determine conservation. Immunofluorescence, chromosome and micronuclei enumeration, MTT and colony formation assays were used to determine the effects of AP4 inactivation and target gene regulation on chromosomal instability (CIN) and drug sensitivity. RESULTS: Inactivation of AP4 in CRC cell lines resulted in increased spontaneous and c-MYC-induced DNA damage, chromosomal instability (CIN) and cellular senescence. AP4-deficient cells displayed increased expression of the long non-coding RNA MIR22HG, which encodes miR-22-3p and was directly repressed by AP4. Furthermore, Mediator of DNA damage Checkpoint 1 (MDC1), a central component of the DNA damage response and a known target of miR-22-3p, displayed decreased expression in AP4-deficient cells. Accordingly, MDC1 was directly induced by AP4 and indirectly by AP4-mediated repression of miR-22-3p. Adenomas and organoids from Ap4-deficient APCmin/+ mice displayed conservation of these regulations. Inhibition of miR-22-3p or ectopic MDC1 expression reversed the increased senescence, DNA damage, CIN and defective HR observed in AP4-deficient CRC cells. AP4-deficiency also sensitized CRC cells to 5-FU treatment, whereas ectopic AP4 conferred resistance to 5-FU in a miR-22-3p and MDC1-dependent manner. CONCLUSIONS: In summary, AP4, miR-22-3p and MDC1 form a conserved and coherent, regulatory feed-forward loop to promote DNA repair, which suppresses DNA damage, senescence and CIN, and contributes to 5-FU resistance. These findings explain how elevated AP4 expression contributes to development and chemo-resistance of colorectal cancer after c-MYC activation.

Potential role of autophagy induced by FLT3-ITD and acid ceramidase in acute myeloid leukemia chemo-resistance: new insights.

Acute myeloid leukemia (AML) is a type of leukemia with a poor prognosis and survival characterized by abnormal cell proliferation and differentiation. Despite advances in treatment, AML still has a low complete remission rate, particularly in elderly patients, and recurrences are frequently seen even after complete remissions. The major challenge in treating AML is the resistance of leukemia cells to chemotherapy drugs. Thus, to overcome this issue, it can be crucial to conduct new investigations to explore the mechanisms of chemo-resistance in AML and target them. In this review, the potential role of autophagy induced by FLT3-ITD and acid ceramidase in chemo-resistance in AML patients are analyzed. With regard to the high prevalence of FLT3-ITD mutation (about 25% of AML cases) and high level of acid ceramidase in these patients, we hypothesized that both of these factors could lead to chemo-resistance by inducing autophagy. Therefore, pharmacological targeting of autophagy, FLT3-ITD, and acid ceramidase production could be a promising therapeutic approach for such AML patients to overcome chemo-resistance. Video abstract.

Overexpressed or hyperactivated Rac1 as a target to treat hepatocellular carcinoma.

Despite novel targeted and immunotherapies, the prognosis remains bleak for patients with hepatocellular carcinoma (HCC), especially for advanced and/or metastatic forms. The rapid emergence of drug resistance is a major obstacle in the success of chemo-, targeted-, immuno-therapies of HCC. Novel targets are needed. The prominent roles of the small GTPase Rac1 in the development and progression of HCC are discussed here, together with its multiple protein partners, and the targeting of Rac1 with RNA-based regulators and small molecules. We discuss the oncogenic functions of Rac1 in HCC, including the contribution of Rac1 mutants and isoform Rac1b. Rac1 is a ubiquitous target, but the protein is frequently overexpressed and hyperactivated in HCC. It contributes to the aggressivity of the disease, with key roles in cancer cell proliferation, tumor metastasis and resistance to treatment. Small molecule targeting Rac1, indirectly or directly, have shown anticancer effects in HCC experimental models. Rac1-binding agents such as EHT 1864 and analogues offer novel opportunities to combat HCC. We discuss the different modalities to repress Rac1 overactivation in HCC with small molecules and the combination with reference drugs to promote cancer cell death and to repress cell invasion. We highlight the necessity to combine Rac1-targeted approach with appropriate biomarkers to select Rac1 activated tumors. Our analysis underlines the prominent oncogenic functions of Rac1 in HCC and discuss the modalities to target this small GTPase. Rac1 shall be considered as a valid target to limit the acquired and intrinsic resistance of HCC tumors and their metastatic potential.

LncRNA H19 Impairs Chemo and Radiotherapy in Tumorigenesis.

Various treatments based on drug administration and radiotherapy have been devoted to preventing, palliating, and defeating cancer, showing high efficiency against the progression of this disease. Recently, in this process, malignant cells have been found which are capable of triggering specific molecular mechanisms against current treatments, with negative consequences in the prognosis of the disease. It is therefore fundamental to understand the underlying mechanisms, including the genes-and their signaling pathway regulators-involved in the process, in order to fight tumor cells. Long non-coding RNAs, H19 in particular, have been revealed as powerful protective factors in various types of cancer. However, they have also evidenced their oncogenic role in multiple carcinomas, enhancing tumor cell proliferation, migration, and invasion. In this review, we analyze the role of lncRNA H19 impairing chemo and radiotherapy in tumorigenesis, including breast cancer, lung adenocarcinoma, glioma, and colorectal carcinoma.

Secreted HSP90α-LRP1 Signaling Promotes Tumor Metastasis and Chemoresistance in Pancreatic Cancer.

The extracellular heat shock protein 90α (eHSP90α) has been reported to promote cancer cell motility. However, whether pancreatic cancer (PC) cells expressed membrane-bound or secreted HSP90α, as well as its underlying mechanism for PC progression, were still unclear. Our study demonstrated that the amounts of secreted HSP90α proteins were discrepant in multiple PC cells. In addition, highly invasive Capan-2 cells have a higher level of secreted HSP90α compared with those of less invasive PL45 cells. The conditioned medium of Capan-2 cells or recombinant HSP90α treatment stimulated the migration and invasion of PC cells, which could be prevented with a neutralizing anti-HSP90α antibody. Furthermore, secreted HSP90α promoted elements of epithelial-mesenchymal transition in PL45 cells, including increases in vimentin and Snail expressions, decreases in E-cadherin expression, and changes in cell shape towards a mesenchymal phenotype, but these phenomena were reversed by the anti-HSP90α antibody in Capan-2 cells. In addition, high levels of low-density lipoprotein receptor-related protein 1 (LRP1) were associated with worsened patient survival in pancreatic adenocarcinoma. We demonstrated LRP1 as a receptor of eHSP90α for its stimulatory role in metastasis, by activating the AKT pathway. In addition, silencing LRP1 enhanced the chemosensitivity to gemcitabine and doxorubicin in Capan-2 cells. Therefore, our study indicated that blocking secreted HSP90α underlies an aspect of metastasis and chemoresistance in PC.