Characterization of BCL-X L , MCL-1, and BAX Protein Expression in Response to Neoadjuvant Chemotherapy in Breast Cancer.

The use of chemotherapy has improved the overall treatment of breast cancer, which is frequently administered in the form of neoadjuvant chemotherapy (NAC). Apoptosis is an established cell stress response to NAC in preclinical models; however, there is limited understanding of its role in clinical cancer, specifically, its contribution to favorable pathologic responses in breast cancer therapy. Here, we aimed to characterize the change in protein expression of 3 apoptosis-associated biomarkers, namely, BCL-X L , MCL-1, and BAX in breast cancer in response to NAC. For this, we utilized a set of 68 matched invasive breast cancer FFPE samples that were collected before (pre) and after (post) the exposure to NAC therapy that were characterized by incomplete pathologic response. Immunohistochemistry (IHC) analysis suggested that most of the samples show a decrease in the protein expression of all 3 markers following exposure to NAC as 90%, 69%, and 76% of the matched samples exhibited a decrease in expression for BCL-X L , MCL-1, and BAX, respectively. The median H-score of BCL-X L post-NAC was 150/300 compared with 225/300 pre-NAC ( P value <0.0001). The median H-score of MCL-1 declined from 200 pre-NAC to 160 post-NAC ( P value <0.0001). The median H-score of BAX protein expression decreased from 260 pre-NAC to 190 post-NAC ( P value <0.0001). There was no statistically significant association between the expression of these markers and stage, grade, and hormone receptor profiling (luminal status). Collectively, our data indicate that the expression of apoptosis regulatory proteins changes following exposure to NAC in breast cancer tissue, developing a partial pathologic response.

[Effect of venetoclax plus chemotherapy on treatment-naive acute myeloid leukemia patients with moderate to poor cytogenetic profiles and the combination's influence on the expression of proteins of the anti-apoptoic family].

Objective: This was an open-label observational assessment aimed to evaluate whether venetoclax (VEN) plus chemotherapy could enhance the therapeutic benefits for treatment-naive acute myeloid leukemia (AML) patients with adverse cytogenetic profiles. Methods: A total of 38 adult patients (including 11 patients with moderate risk stratification and 27 patients with high risk stratification) who were treated at the Affiliated Hospital of Inner Mongolia Medical University from April 2019 to May 2022 were enrolled in this study. Patients were randomized into two cohorts according to the random number method to receive single intensive chemotherapy (18/38) alone or VEN+intensive chemotherapy (20/38), respectively. The chemotherapy cohort received 2 cycles of induction chemotherapy (idarbicin or daunorubicin plus cytarabine), followed by 6 cycles of consolidation chemotherapy (cytarabine), while the treatment for the VEN + chemotherapy cohort consisted of the same chemotherapy as above plus oral VEN. Heparinized bone marrow samples were obtained from patients at enrollment de novo and post chemotherapy. The expressions of MCL-1 and BCL-2 were detected by Western blot analysis. Results: Patients with VEN+chemotherapy showed an objective response rate (ORR) of 90.0% (18/20), compared with 55.6% (10/18, P=0.012) of the chemotherapy group. Meanwhile, the VEN + chemotherapy cohort gained more benefits in progression-free survival (PFS) and overall survival (OS) than the chemotherapy cohort (mean PFS: 27.1 months versus 17.9 months, P=0.038; mean OS: 32.2 months versus 21.3 months, P=0.004). For patients with moderate risk stratification, there were no differences in the ORR and PFS between the chemotherapy cohort and the VEN + chemotherapy cohort: the ORR was 80.0% (4/5) versus 100% (6/6, P=0.251), and the PFS was 27.9 months versus 32.0 months (P=0.582). Moreover, the ORR was 85.7% (12/14) for the VEN+chemotherapy cohort and 46.2% (6/13) for the chemotherapy cohort in the high risk profile (P=0.029). The PFS of the VEN+chemotherapy cohort was superior to the chemotherapy cohort in the high risk profile (mean PFS: 23.7 months versus 11.1 months, P=0.002). Meanwhile, in the chemotherapy cohort, there were no difference in the PFS between FAB-M5 patients and non-FAB-M5 patients; the mean PFS was 20.0 months versus 15.5 months (P=0.298) for the two groups. Nevertheless, FAB-M5 patients were inferior to non-FAB-M5 patients in PFS in the VEN + chemotherapy arm (mean PFS: 19.6 months versus 30.2 months, P=0.031). The most frequent grade 4 hematological toxicities (therapy related) were leukopenia and thrombopenia. Grade 3/4 hematological adverse events in patients treated with VEN+chemotherapy were not increased compared with those who received chemotherapy. Western blot showed VEN continuously decreased the expression of BCL-2 proteins in both FAB-M5 and non-FAB-M5 patients, but obviously increased the expression of MCL-1 proteins only in FAB-M5 patients. Conclusions: VEN combined with intensive chemotherapy have yielded high ORR and survival advantages for de novo AML patients with poor cytogenetics profiles. The high-expression of MCL-1 may drive resistance to VEN.

C-X-C motif chemokine receptor 4 inhibition promotes the effect of plantamajoside in hepatocellular carcinoma.

BACKGROUND AND STUDY AIM: Hepatocellular carcinoma (HCC) is the fifth leading cause of cancer-related mortality worldwide, and, more than half of these cases are diagnosed in China. However, effective treatment for HCC is still limited. MATERIAL AND METHODS: C-X-C motif chemokine receptor 4 (CXCR4) was first activated and inhibited in HepG2 cells using a pharmacological method. HepG2 cell proliferation was detected using the CCK-8 method. Metastasis and apoptosis of HepG2 cells were detected using wound healing and flow cytometry. The expression of each target molecule related to metastasis and invasion, such as MMPs, E-cadherin and the PI3K/AKT/Mcl-1/PARP signaling pathway was detected by western blotting. The secretion of molecular metastases was detected using competitive ELISA. RESULTS: This study constructed a CXCR4 activation and inhibition model in HepG2 cells. CXCR4 inhibition promoted the inhibitory effect of plantamajoside on the proliferation and metastasis of cells, which led to apoptosis. Furthermore, we found that the expression of apoptosis-related proteins was increased after treatment with plantamajoside combined with CXCR4 inhibition. In addition, the expression and secretion of pro-metastatic proteins, including MMPs and E-cadherin were decreased. We also noticed that this effect might be mediated by the PI3K/AKT/Mcl-1/PARP signaling pathway. CONCLUSION: CXCR4 inhibition may contribute to the treatment of HCC. Inhibition of CXCR4 expression contributes to the therapeutic effect of plantamajoside; the effect of plantamajoside might be mediated by the PI3K/AKT/Mcl-1/PARP signaling pathway; and CXCR4 might be a therapeutic target of HCC.

FBXW7 regulates the sensitivity of imatinib in gastrointestinal stromal tumors by targeting MCL1.

BACKGROUND: Imatinib contributes to improving prognosis of high-risk or unresectable gastrointestinal stromal tumors (GISTs). As therapeutic efficacy is limited by imatinib resistance and toxicity, the exploration of predictive markers of imatinib therapeutic efficacy that enables patients to utilize more effective therapeutic strategies remains urgent. METHODS: The correlation between FBXW7 and imatinib resistance via FBXW7-MCL1 axis was evaluated in vitro and in vivo experiments. The significance of FBXW7 as a predictor of imatinib treatment efficacy was examined in 140 high-risk patients with GISTs. RESULTS: The ability of FBXW7 to predict therapeutic efficacy of adjuvant imatinib in high-risk GIST patients was determined through 5-year recurrence-free survival (RFS) rates analysis and multivariate analysis. FBXW7 affects imatinib sensitivity by regulating apoptosis in GIST-T1 cells. FBXW7 targets MCL1 to regulate apoptosis. MCL1 involves in the regulation of imatinib sensitivity through inhibiting apoptosis in GIST-T1 cells. FBXW7 regulates imatinib sensitivity by down-regulating MCL1 to enhance imatinib-induced apoptosis in vitro. FBXW7 regulates imatinib sensitivity of GIST cells by targeting MCL1 to predict efficacy of imatinib treatment in vivo. CONCLUSIONS: FBXW7 regulates imatinib sensitivity by inhibiting MCL1 to enhance imatinib-induced apoptosis in GIST, and predicts efficacy of imatinib treatment in high-risk GIST patients treated with imatinib.

Zotiraciclib (TG02) for newly diagnosed glioblastoma in the elderly or for recurrent glioblastoma: The EORTC 1608 STEAM trial.

BACKGROUND: Zotiraciclib (TG02) is an oral multi-cyclin dependent kinase (CDK) inhibitor thought to inhibit tumor growth via CDK-9-dependent depletion of survival proteins such as c-MYC and MCL-1 which are frequently overexpressed in glioblastoma. METHODS: EORTC 1608 (NCT03224104) (STEAM) had a three parallel group (A,B,C) phase Ib, open-label, non-randomized, multicenter design in IDH wild-type newly diagnosed glioblastoma or anaplastic astrocytoma. Groups A and B explored the maximum tolerated dose (MTD) of TG02 in elderly patients, in combination with hypofractionated radiotherapy alone (group A) or temozolomide alone (group B), according to O6-methylguanine DNA methyltransferase promoter methylation status determined centrally. Group C explored single agent activity of TG02 at first relapse after temozolomide chemoradiotherapy with a primary endpoint of progression-free survival at 6 months (PFS-6). Tumor expression of CDK-9, c-MYC and MCL-1 was determined by immunohistochemistry. RESULTS: The MTD was 150 mg twice weekly in combination with radiotherapy alone (group A) or temozolomide alone (group B). Two dose-limiting toxicities were observed at 150 mg: one in group A (grade 3 seizure), one in group B (multiple grade 1 events). Main toxicities included neutropenia, gastrointestinal disorders and hepatotoxicity. PFS-6 in group C was 6.7%. CDK-9, c-MYC and MCL-1 were confirmed to be expressed and their expression was moderately cross-correlated. High protein levels of MCL-1 were associated with inferior survival. CONCLUSIONS: TG02 exhibits overlapping toxicity with alkylating agents and low single agent clinical activity in recurrent glioblastoma. The role of CDK-9 and its down-stream effectors as prognostic factors and therapeutic targets in glioblastoma warrants further study.

EGR3 and estrone are involved in the tamoxifen resistance and progression of breast cancer.

BACKGROUND: Tamoxifen (Tam) is an effective treatment for estrogen receptor (ER) positive breast cancer. However, a significant proportion of patients develop resistance under treatment, presenting a therapeutic challenge. The study aims to determine the role of early growth response protein (EGR) 3 in tamoxifen resistance (TamR) and elucidate its molecular mechanism. METHODS: TamR cell models were established and NGS was used to screening signaling alternation. Western blot and qRT-PCR were used to analysis the expression of ERα, EGR3, MCL1 and factors associated with apoptosis. CCK8, colony formation and apoptosis assay were used to analysis resistance to Tam. Immunofluorescence, chromatin immunoprecipitation, and dual luciferase assays were used to investigate mechanism of regulation. RESULTS: We observed that EGR3, a deeply rooted ERα response factor, showed increased upregulation in response to both estrone (E1) and Tam in TamR cells with elevated level of E1 and ERα expression, indicating a potential connection between EGR3 and TamR. Mechanically, manipulating EGR3 expression revealed that it imparted resistance to Tam through increased expression of the downstream molecule MCL1 (apoptosis suppressor gene) that it regulated. Mechanismly, EGR3 directly binds to the promoter of the anti-apoptotic factor MCL1 gene, facilitating its transcription. Furthermore, apoptosis assays revealed that E1 reduces Tam induced apoptosis by upregulating EGR3 expression. Importantly, clinical public database confirmed the high expression of EGR3 in breast cancer tissue and in Tam-treated patients. CONCLUSIONS: These findings shed light on the novel estrogen/EGR3/MCL1 axis and its role in inducing TamR in ER positive breast cancer. EGR3 emerges as a promising target to overcome TamR. The elucidation of this mechanism holds potential for the development of new therapeutic modalities to overcome endocrine therapy resistance in clinical settings.

Apoptotic activity of genipin in human oral squamous cell carcinoma in vitro by regulating STAT3 signaling.

Genipin, a natural compound derived from the fruit of Gardenia jasminoides Ellis, was reported to have activity against various cancer types. In this study, we determined the underlying mechanism for genipin-induced cell death in human oral squamous cell carcinoma (OSCC). The growth-inhibitory effects of genipin in human OSCC cells was examined by the Cell Counting Kit-8 and soft agar assays. The effects of genipin on apoptosis were assessed by nuclear morphological changes by 4',6-diamidino-2-phenylindole staining, measurement of the sub-G1 population, and Annexin V-fluorescein isothiocyanate/propidium iodide double staining. The underlying mechanism of genipin activity was analyzed by western blot analysis, subcellular fractionation of the nucleus and cytoplasm, immunocytochemistry, and quantitative real-time polymerase chain reaction. Genipin inhibited the growth of OSCC cells and induced apoptosis, which was mediated by a caspase-dependent pathway. Genipin reduced the phosphorylation of signal transducer and activator of transcription 3 (STAT3) at Tyr705 and its nuclear localization. Furthermore, inhibition of p-STAT3Tyr705 levels following genipin treatment was required for the reduction of survivin and myeloid cell leukemia-1 (Mcl-1) expression, leading to apoptotic cell death. The genipin-mediated reduction in survivin and Mcl-1 expression was caused by transcriptional and/or posttranslational regulatory mechanisms. The results provide insight into the regulatory mechanism by which genipin induces apoptotic cell death through the abrogation of nuclear STAT3 phosphorylation and suggest that genipin may represent a potential therapeutic option for the treatment of human OSCC.

Cordycepin improves sensitivity to temozolomide in glioblastoma cells by down-regulating MYC.

PURPOSE: Glioblastoma is one of the malignant tumors with poor prognosis and no effective treatment is available at present. METHODS: To study the effect of cordycepin combined with temozolomide on glioblastoma, we explored the effect of the combination based on network pharmacology and biological verification. RESULTS: It was found that the drug combination significantly inhibited the cell growth, proliferation, migration and invasion of LN-229 cells. Drug combination inhibited epithelial-mesenchymal transition (EMT) by up-regulating the expression of E-cadherin and suppressing the expression of N-cadherin, Zeb1 and Twist1. Through network pharmacology, we further explored the molecular mechanism of drug combination against glioblastoma, and 36 drug-disease common targets were screened. The GO biological process analysis included 44 items (P < 0.01), which mainly involved the regulation of apoptosis, cell proliferation, cell migration, etc. The enrichment analysis of KEGG pathways included 28 pathways (P < 0.05), and the first four pathways were "MicroRNA in cancer, Proteoglycans in cancer, Pathways in cancer and PI3K-AKT signaling pathway". We detected the expression of important genes in the pathways and PPI network, and the results showed that the drug combination down-regulated NFKB1, MYC, MMP-9, MCL1, CTNNB1, and up-regulated PDCD4. CONCLUSION: Cordycepin combined with temozolomide may down-regulate MYC through "MicroRNA in cancer, Proteoglycans in cancer, Pathways in cancer and PI3K-AKT signaling pathway", which in turn regulate the expression of MCL1, CTNNB1, MMP9, PDCD4, thus regulating cell proliferation, migration and apoptosis in glioblastoma.

Osimertinib is a dual inhibitor of hepatocellular carcinoma and angiogenesis in an EGFR-independent manner, and synergizes with venetoclax.

BACKGROUND: To investigate the effects of osimertinib on hepatocellular carcinoma (HCC) and angiogenesis, and its combinatory effects with venetoclax in HCC. METHODS: Viability was assessed by flow cytometry of Annexin V in multiple HCC cell lines after drug treatment. In vitro angiogenesis assay was performed using primary human liver tumor associated endothelial cell (HLTEC). HCC-bearing model was generated by subcutaneous implantation of Hep3B cells to investigate the efficacy of osimertinib alone and its combination with venetoclax. RESULTS: Osimertinib significantly induced apoptosis in a panel of HCC cell lines regardless of EGFR expression level. It inhibited capillary network formation and induced apoptosis in HLTEC. Using HCC xenograft mouse model, we further showed that osimertinib at non-toxic dose inhibited tumor growth by ~ 50% and remarkably decreased blood vessel in tumor. Mechanism studies demonstrated that osimertinib acted on HCC cells in an EGFR-independent manner. It decreased level of VEGF and Mcl-1 in HCC cells via suppressed phosphorylation of eIF4E, thus leading to inhibition of eIF4E-mediated translation. Mcl-1 overexpression reversed pro-apoptotic effect of osimertinib, suggesting an important role of Mcl-1 in osimertinib's action in HCC cells. Of note, the combination of osimertinib and venetoclax achieved approximately complete HCC cell death and tumor growth in mice. CONCLUSIONS: We provide pre-clinical evidence that osimertinib is a promising candidate for the treatment of HCC via targeting tumor cells and angiogenesis. The combination of osimertinib and venetoclax is synergistic in inhibiting HCC.

IFN-γ Signaling Sensitizes Melanoma Cells to BH3 Mimetics.

Immunotherapy targeting PD-1 and/or CTLA4 leads to durable responses in a proportion of patients with melanoma. However, many patients will not respond to these immune checkpoint inhibitors, and up to 60% of responding patients will develop treatment resistance. We describe a vulnerability in melanoma driven by immune cell activity that provides a pathway towards additional treatment options. This study evaluated short-term melanoma cell lines (referred to as PD1 PROG cells) derived from melanoma metastases that progressed on PD-1 inhibitor-based therapy. We show that the cytokine IFN-γ primes melanoma cells for apoptosis by promoting changes in the accumulation and interactions of apoptotic regulators MCL-1, NOXA, and BAK. The addition of pro-apoptotic BH3 mimetic drugs sensitized PD1 PROG melanoma cells to apoptosis in response to IFN-γ or autologous immune cell activation. These findings provide translatable strategies for combination therapies in melanoma.

Targeting MCL-1 and BCL-2 with polatuzumab vedotin and venetoclax overcomes treatment resistance in R/R non-Hodgkin lymphoma: Results from preclinical models and a Phase Ib study.

The treatment of patients with relapsed or refractory lymphoid neoplasms represents a significant clinical challenge. Here, we identify the pro-survival BCL-2 protein family member MCL-1 as a resistance factor for the BCL-2 inhibitor venetoclax in non-Hodgkin lymphoma (NHL) cell lines and primary NHL samples. Mechanistically, we show that the antibody-drug conjugate polatuzumab vedotin promotes MCL-1 degradation via the ubiquitin/proteasome system. This targeted MCL-1 antagonism, when combined with venetoclax and the anti-CD20 antibodies obinutuzumab or rituximab, results in tumor regressions in preclinical NHL models, which are sustained even off-treatment. In a Phase Ib clinical trial (NCT02611323) of heavily pre-treated patients with relapsed or refractory NHL, 25/33 (76%) patients with follicular lymphoma and 5/17 (29%) patients with diffuse large B-cell lymphoma achieved complete or partial responses with an acceptable safety profile when treated with the recommended Phase II dose of polatuzumab vedotin in combination with venetoclax and an anti-CD20 antibody.

Natural Dibenzo[b,f]oxepines, Pacharin and Bauhiniastatin-1, Isolated from Bauhinia acuruana Induce Apoptosis on Breast Cancer Cells via MCL-1 Protein Reduction.

Herein, we describe the antiproliferative effects of two natural dibenzo [b,f]oxepines, pacharin and bauhiniastatin-1, isolated from Bauhinia acuruana on a breast cancer cell line and the mode of action underlying the cytotoxicity. Both compounds were cytotoxic in a panel of six tumor lines analyzed by the MTT assay, and IC50 values ranged from 7.8 to 45.1 µM, including human breast adenocarcinoma (MCF-7) cells. In contrast, none of the compounds were cytotoxic on normal human peripheral blood mononuclear cells (IC50 > 100 µM). Human breast adenocarcinoma (MCF-7) cells treated with pacharin or bauhiniastatin-1 20 µM for 24 h presented a reduction in cell volume and intensification of chromatin condensation, DNA fragmentation, and apoptotic cells. These findings became more evident after 48 h of exposure. Antiapoptotic B-cell lymphoma-2 family members, such as myeloid cell leukemia-1 and B-cell lymphoma-extra large, are important targets in cancer cells since their overexpression confers resistance to cancer treatments. A significant reduction of the myeloid cell leukemia-1 protein levels in human breast adenocarcinoma (MCF-7) cells after 24 h of treatment with pacharin or bauhiniastatin-1 at 20 µM was observed, while the B-cell lymphoma-extra large protein content was reduced in bauhiniastatin-1-treated cells at 40 µM only. The cytotoxic effects of pacharin and bauhiniastatin-1 are likely linked to myeloid cell leukemia-1 inhibition, which leads to the apoptosis of breast adenocarcinoma cells.

Small-molecule High-throughput Screening Identifies an MEK Inhibitor PD198306 that Enhances Sorafenib Efficacy via MCL-1 and BIM in Hepatocellular Carcinoma Cells.

BACKGROUND: Sorafenib is the most widely used systematic therapy drug for treating unresectable Hepatocellular Carcinoma (HCC) but showed dissatisfactory efficacy in clinical applications. OBJECTIVE: We conducted a combinational quantitative small-molecule high-throughput screening (qHTS) to identify potential candidates to enhance the treatment effectiveness of sorafenib. METHODS: First, using a Hep3B human HCC cell line, 7051 approved drugs and bioactive compounds were screened, then the primary hits were tested with/without 0.5 µM sorafenib respectively, the compound has the half maximal Inhibitory Concentration (IC50) shift value greater than 1.5 was thought to have the synergistic effect with sorafenib. Furthermore, the MEK inhibitor PD198306 was selected for the further mechanistic study. RESULTS: 12 effective compounds were identified, including kinase inhibitors targeting MEK, AURKB, CAMK, ROCK2, BRAF, PI3K, AKT and EGFR, and a µ-opioid receptor agonist and a Ltype calcium channel blocker. The mechanistic research of the combination of sorafenib plus PD198306 showed that the two compounds synergistically inhibited MEK-ERK and mTORC1- 4EBP1 and induced apoptosis in HCC cells, which can be attributed to the transcriptional and posttranslational regulation of MCL-1 and BIM. CONCLUSION: Small-molecule qHTS identifies MEK inhibitor PD1938306 as a potent sorafenib enhancer, together with several novel combination strategies that are valuable for further studies.

Jinhua Qinggan granules attenuates acute lung injury by promotion of neutrophil apoptosis and inhibition of TLR4/MyD88/NF-κB pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Acute lung injury (ALI) is one of the fatal complications of respiratory virus infections such as influenza virus and coronavirus, which has high clinical morbidity and mortality. Jinhua Qinggan granules (JHQG) has been approved by China Food and Drug Administration in the treatment of H1N1 influenza and mild or moderate novel coronavirus disease 2019 (COVID-19), which is an herbal formula developed based on Maxingshigan decoction and Yinqiao powder that have been used to respiratory diseases in China for thousands of years. However, the underlying mechanism of JHQG in treating infectious diseases remains unclear. AIM OF THE STUDY: This study investigated the effects of JHQG on neutrophil apoptosis and key signaling pathways in lipopolysaccharide (LPS) -induced ALI mice in order to explore its mechanism of anti-inflammation. MATERIALS AND METHODS: The effect of JHQG on survival rate was observed in septic mouse model by intraperitoneal injection of LPS (20 mg/kg). To better pharmacological evaluation, the mice received an intratracheal injection of 5 mg/kg LPS. Lung histopathological changes, wet-to-dry ratio of the lungs, and MPO activity in the lungs and total protein concentration, total cells number, TNF-α, IL-1ß, IL-6, and MIP-2 levels in BALF were assessed. Neutrophil apoptosis rate was detected by Ly6G-APC/Annexin V-FITC staining. Key proteins associated with apoptosis including caspase 3/7 activity, Bcl-xL and Mcl-1 were measured by flow cytometry and confocal microscope, respectively. TLR4 receptor and its downstream signaling were analyzed by Western blot assay and immunofluorescence, respectively. RESULTS: JHQG treatment at either 6 or 12 g/kg/day resulted in 20% increase of survival in 20 mg/kg LPS-induced mice. In the model of 5 mg/kg LPS-induced mice, JHQG obviously decreased the total protein concentration in BALF, wet-to-dry ratio of the lungs, and lung histological damage. It also attenuated the MPO activity and the proportion of Ly6G staining positive neutrophils in the lungs, as well as the MIP-2 levels in BALF were reduced. JHQG inhibited the expression of Mcl-1 and Bcl-xL and enhanced caspase-3/7 activity, indicating that JHQG partially acted in promoting neutrophil apoptosis via intrinsic mitochondrial apoptotic pathway. The levels of TNF-α, IL-1ß, and IL-6 were significantly declined in LPS-induced mice treated with JHQG. Furthermore, JHQG reduced the protein expression of TLR4, MyD88, p-p65 and the proportion of nuclei p65, suggesting that JHQG treatment inhibited TLR4/MyD88/NF-κB pathway. CONCLUSION: JHQG reduced pulmonary inflammation and protected mice from LPS-induced ALI by promoting neutrophil apoptosis and inhibition of TLR4/MyD88/NF-κB pathway, suggesting that JHQG may be a promising drug for treatment of ALI.

Acute toxicity and regenerative dose finding of an extract of Miconia ferruginata DC. in a mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a severe disease with no cure caused by a genetic abnormality, promoting progressive muscle degeneration. Corticosteroids are used drugs in treatment associated with adverse effects. The extract of Miconia ferruginata (Melastomataceae) (MF) has demonstrated potent antioxidant and anti-inflammatory potential in vitro. This study used a DMD model (mdx) to determine the toxic dose of this plant and found a possible non-toxic dose with therapeutic effects. The mdx groups received an intraperitoneal injection of 0 (control group), 50, 100, 200, 300, and 2000 mg kg-1 of the aqueous leaf extract following a single-dose acute toxicity protocol and were observed for 14 days. The range of toxicity of the extract and LD50 were determined. Histopathological analysis, the quantification of fibrosis, and immunohistochemical analysis of the tissues were performed. The results demonstrated that 2000 mg kg-1 was highly toxic, inducing histopathological changes in the tissues evaluated, with 100% mortality in 48 hours. The other doses caused no behavioral changes or signs of toxicity. The MF extract led reduction in histopathological changes, fibrosis, and inflammation, a reduction in HSP70 and an increase in MCL-1 proteins. Doses of 50-200 mg kg-1 demonstrated regenerative tissue and anti-inflammatory potential.

Druggable host gene dependencies in primary effusion lymphoma.

Kaposi's sarcoma-associated herpesvirus (KSHV) causes primary effusion lymphoma (PEL). Here, we review what is known about human gene essentiality in PEL-derived cell lines. We provide an updated list of PEL-specific human gene dependencies, based on the improved definition of core essential genes across human cancer types. The requirements of PEL cell lines for interferon regulatory factor 4 (IRF4), basic leukine zipper ATF-like transcription factor (BATF), G1/S cyclin D2 (CCND2), CASP8 and FADD like apoptosis regulator (CFLAR), MCL1 apoptosis regulator (MCL1), and murine double minute 2 (MDM2) have been confirmed experimentally. KSHV co-opts IRF4 and BATF to drive superenhancer (SE)-mediated expression of IRF4 itself, MYC, and CCND2. IRF4 dependency of SE-mediated gene expression is shared with Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) and human T-cell leukemia virus type 1-transformed adult T-cell leukemia/lymphoma (ATLL) cell lines, as well as several B-cell lymphomas of nonviral etiology. LCLs and ATLL cell lines similarly share dependencies on CCND2 and CFLAR with PEL, but also have distinct gene dependencies. Genetic dependencies could be exploited for therapeutic intervention in PEL and other cancers.

A MCL-1-targeted photosensitizer to combat triple-negative breast cancer with enhanced photodynamic efficacy, sensitization to ROS-induced damage, and immune response.

As growth factor receptor-2 (HER-2), progesterone receptor (PR) and estrogen receptor (ER) are scarce in triple-negative breast cancer (TNBC), it is a great challenge to combat TNBC with high tumor specificity and therapeutic efficacy. Most traditional treatments including surgical resection, chemotherapy, and radiotherapy would more or less cause serious side effects and drug resistance. Photodynamic therapy (PDT) has huge potential in the treatment of TNBC for minimal invasiveness, low toxicity, less drug resistance and high spatiotemporal selectivity. Inspired by the advantages of small-molecule-targeted PDT and the sensitization effect of myeloid cell leukemia-1 (MCL-1) inhibitor, a novel photosensitizer BC-Pc was designed by conjugating MCL-1 inhibitor with zinc phthalocyanines. Owning to 3-chloro-6-methyl-1-benzothiophene-2-carboxylic acid (BC) moiety, BC-Pc exhibits the high affinity towards MCL-1 and reduce its self-aggregation in TNBC cells. Therefore, MCL-1 targeted BC-Pc showed remarkable intracellular fluorescence and ROS generation in TNBC cells. Additionally, BC-Pc can selectively sensitize TNBC cells to ROS-induced damage, resulting in improved therapeutic effect to TNBC cells and negligible toxicity to normal cells. More importantly, BC-Pc can effectively inhibit the migration and invasion of TNBC cells, and enhance immune response, all of which will be beneficial to eradicate TNBC. To the best of our knowledge, BC-Pc is the novel MCL-targeted photosensitizer, which owns the amplified ROS-induced lethality and anticancer immune response for TNBC. Overall, our study provides a promising strategy to achieve targeting and highly efficient therapy of TNBC.

Transcriptional signatures of the BCL2 family for individualized acute myeloid leukaemia treatment.

BACKGROUND: Although anti-apoptotic proteins of the B-cell lymphoma-2 (BCL2) family have been utilized as therapeutic targets in acute myeloid leukaemia (AML), their complicated regulatory networks make individualized therapy difficult. This study aimed to discover the transcriptional signatures of BCL2 family genes that reflect regulatory dynamics, which can guide individualized therapeutic strategies. METHODS: From three AML RNA-seq cohorts (BeatAML, LeuceGene, and TCGA; n = 451, 437, and 179, respectively), we constructed the BCL2 family signatures (BFSigs) by applying an innovative gene-set selection method reflecting biological knowledge followed by non-negative matrix factorization (NMF). To demonstrate the significance of the BFSigs, we conducted modelling to predict response to BCL2 family inhibitors, clustering, and functional enrichment analysis. Cross-platform validity of BFSigs was also confirmed using NanoString technology in a separate cohort of 47 patients. RESULTS: We established BFSigs labeled as the BCL2, MCL1/BCL2, and BFL1/MCL1 signatures that identify key anti-apoptotic proteins. Unsupervised clustering based on BFSig information consistently classified AML patients into three robust subtypes across different AML cohorts, implying the existence of biological entities revealed by the BFSig approach. Interestingly, each subtype has distinct enrichment patterns of major cancer pathways, including MAPK and mTORC1, which propose subtype-specific combination treatment with apoptosis modulating drugs. The BFSig-based classifier also predicted response to venetoclax with remarkable performance (area under the ROC curve, AUROC = 0.874), which was well-validated in an independent cohort (AUROC = 0.950). Lastly, we successfully confirmed the validity of BFSigs using NanoString technology. CONCLUSIONS: This study proposes BFSigs as a biomarker for the effective selection of apoptosis targeting treatments and cancer pathways to co-target in AML.

Enzalutamide Overcomes Dihydrotestosterone-Induced Chemoresistance in Triple- Negative Breast Cancer Cells via Apoptosis.

BACKGROUND: Triple-negative breast cancer is challenging to treat due to its heterogeneity and lack of therapeutic targets. Hence, systemic chemotherapy is still the mainstay in TNBC treatment. Unfortunately, patients commonly develop chemoresistance. Androgen signalling through its receptor is an essential player in breast cancer, where it has been shown to confer chemoresistance to TNBC cells. OBJECTIVE: The objective of the study was to elucidate the mechanistic effects of enzalutamide in the chemoresponse of TNBC cells to doxorubicin through the apoptosis pathway. METHODS: MDA-MB-231 and MDA-MB-453 cells were used as model systems of TNBC. Cell viability and apoptosis were investigated upon treatment of cells with doxorubicin in the presence of dihydrotestosterone (DHT) and/or enzalutamide. Caspase 3/7 activity and TUNEL assays were performed to assess the induction of apoptosis. The expression of apoptosis-regulatory genes was assayed by qPCR for the detection of expression changes. RESULTS: Enzalutamide decreased the viability of MDA-MB-231 and MDA-MB- 453 cells and reduced DHT-induced chemoresistance of both cell lines. It also increased the chemosensitivity towards doxorubicin in MDA-MB-231 cells. Increasing DNA degradation and caspase 3/7 activity were concomitant with these outcomes. Moreover, enzalutamide downregulated the expression of the anti-apoptosis genes, mcl1 and bcl2, in MDA-MB-231 cells, while increasing the expression of the pro-apoptotic gene bid. On the other hand, DHT upregulated the expression of the anti-apoptosis genes, mcl1 and bcl2, in both cell lines. CONCLUSION: DHT increased the expression of the anti-apoptosis genes mcl1 and bcl2 in the TNBC cells, presumably leading to cell survival via the prevention of doxorubicin-induced apoptosis. On the other hand, enzalutamide may sensitize the cells to doxorubicin through downregulation of the bid/bcl2/mcl1 axis that normally activates the executive caspases, caspase 3/7. The activities of the latter enzymes were apparent in DNA degradation at the late stages of apoptosis.

Beyond BCL-2 Inhibition in Acute Myloid Leukemia: Other Approaches to Leverage the Apoptotic Pathway.

BCL-2 inhibition has transformed the therapeutic landscape of acute myeloid leukemia (AML) but is not curative for the majority of patients. Consequently, there has been growing interest in targeting other facets of the apoptotic machinery to improve outcomes. These approaches include targeting the intrinsic and extrinsic apoptotic pathway, inducing apoptosis via p53 activation, and others. Targeting the intrinsic apoptotic pathway includes MCL-1 antagonism and BCL-xL inhibition. MCL-1 can be targeted via direct inhibitors as well as via indirect mechanisms to downregulate MCL-1 including inhibition of cyclin dependent kinases and Nedd8 activating enzyme. The extrinsic apoptotic pathway could be harnessed via inhibition of inhibitor of apoptosis proteins, agonism of the TNF-related apoptosis-inducing ligand receptors and inhibiting FLICE-like inhibitor protein. Approaches inducing p53-mediated apoptosis are being evaluated using inhibitors of MDM2, dual inhibitor of MDM2/X in TP53 wild-type AML and p53 reactivators in TP53-mutant myeloid disorders. Several such agents are in early clinical development and rationale combinations of these agents may help improving outcomes for patients with AML.