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Poly(ADP-ribose) polymerase (PARP) inhibitors have demonstrated promising clinical activity in multiple cancers. However, resistance to PARP inhibitors remains a substantial clinical challenge. In the present study, we report that anaplastic lymphoma kinase (ALK) directly phosphorylates CDK9 at tyrosine-19 to promote homologous recombination (HR) repair and PARP inhibitor resistance. Phospho-CDK9-Tyr19 increases its kinase activity and nuclear localization to stabilize positive transcriptional elongation factor b and activate polymerase II-dependent transcription of HR-repair genes. Conversely, ALK inhibition increases ubiquitination and degradation of CDK9 by Skp2, an E3 ligase. Notably, combination of US Food and Drug Administration-approved ALK and PARP inhibitors markedly reduce tumor growth and improve survival of mice in PARP inhibitor-/platinum-resistant tumor xenograft models. Using human tumor biospecimens, we further demonstrate that phosphorylated ALK (p-ALK) expression is associated with resistance to PARP inhibitors and positively correlated with p-Tyr19-CDK9 expression. Together, our findings support a biomarker-driven, combinatorial treatment strategy involving ALK and PARP inhibitors to induce synthetic lethality in PARP inhibitor-/platinum-resistant tumors with high p-ALK–p-Tyr19-CDK9 expression.

Correlations between CEBPD expression and other important clinicopathological parameters in 79 muscle invasive UCUB receiving post-operative chemotherapy

Purpose: Suppressor T cells are one of the determinants of colorectal cancer (CRC) clinical outcome. LAP+CD4+ T cell is a recently identified subset of suppressor T cells. This study was designed to investigate their clinical relevance in patients with CRC.Experimental Design: Sixty patients with CRC and 24 healthy donors (HD) were enrolled in this study. The percentages of LAP+CD4+ T cells in peripheral blood and tumor tissue were measured. The phenotype and functional relevance of LAP+CD4+ T cells were analyzed subsequently.Results: The percentages of LAP+CD4+ T cells in peripheral blood of patients with CRC were significantly higher than HD (HD vs. CRC: 3.1% ± 0.78% vs. 8.8% ± 5.8%, P < 0.0001) and in tumor tissue when compared with nontumor tissue (nontumor vs. tumor: 3.2% ± 1.1% vs. 9.5% ± 5.5%, P = 0.0002). In addition, LAP+CD4+ T cells with effector memory (EM) phenotype were more likely to accumulate in the tumor sites than in peripheral blood. These LAP+CD4+ T cells produced significantly higher levels of IFN-γ, IL-17 and comparatively lower IL-2 and very few IL-10. LAP+CD4+ T cells could suppress the proliferation of LAP−CD4+ T cells that were partially mediated by TGF-β. Furthermore, these LAP+CD4+ T cells accumulated in tumor site and increased further in the peripheral blood in patients with metastasis.Conclusions: LAP+CD4+ T cells as a suppressor subset could accumulate in the tumor microenvironment and circulated more in the peripheral blood with tumor progression in patients with CRC. Clin Cancer Res; 18(19); 5224–33. ©2012 AACR.

Targeting immune checkpoints such as programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) has transformed cancer treatment, with durable clinical responses across a wide range of tumor types. However, a high percentage of patients fail to respond to anti–PD-1/PD-L1 treatment. A greater understanding of PD-L1 regulation is critical to improving the clinical response rate of PD-1/PD-L1 blockade. Here, we demonstrate that PD-L1 is phosphorylated and stabilized by casein kinase 2 (CK2) in cancer and dendritic cells (DC). Phosphorylation of PD-L1 at Thr285 and Thr290 by CK2 disrupted PD-L1 binding with speckle-type POZ protein, an adaptor protein of the cullin 3 (CUL3) ubiquitin E3 ligase complex, protecting PD-L1 from CUL3-mediated proteasomal degradation. Inhibition of CK2 decreased PD-L1 protein levels by promoting its degradation and resulted in the release of CD80 from DC to reactivate T-cell function. In a syngeneic mouse model, combined treatment with a CK2 inhibitor and an antibody against T-cell immunoglobulin mucin-3 (Tim-3) suppressed tumor growth and prolonged survival. These findings uncover a mechanism by which PD-L1 is regulated and suggest a potential antitumor treatment option to activate DC function by blocking the CK2–PD-L1 pathway and inhibiting Tim-3.Significance:This work identifies a role for CK2 in immunosuppression by phosphorylation and stabilization of PD-L1, identifying CK2 inhibition as an immunotherapeutic approach for treating cancer.

Correlations between expression status of pEGFR, pSTAT3, CEBPD, and ABCB1 in muscle invasive UCUB in our independent cohort containing 60 patients receiving post-operative chemotherapy

Attenuated pSTAT3, CEBPD, ABCB1 and ABCC2 mRNA levels are observed in CDDP-resistant NTUB1/P xenografted NOD/SCID mice. A and B, Following 3 weeks of drug treatment, all animals were sacrificed, and their tumors were collected for protein and mRNA analysis. Protein samples were employed for Western blot analysis by using specific antibodies. The following data present changes in pSTAT3, CEBPD, and ABCB1 levels in NTUB1/P-xenografted NOD/SCID mice treated with CDDP and gefitinib or with CDDP and S3I-201. qPCR assays were performed using the total RNA harvested from tumor samples. In this figure, *** denotes a significant difference (P < 0.001). B, evaluation of body weight of experimental mice used in Fig. 5. CTL: control.

CEBPD expression had significant worse MeFS in UCUB patients. A and B, Kaplan-Meier plots discloses the predictive significances of CEBPD immunoexpression for metastasis-free survival in UCUB.

Levels of CEBPD associates with STAT3 activity in gefitinib-treated CDDP-resistant NTUB1/P (NTP) cells. A, Western blot analysis was conducted with lysates from NTP cells treated with gefitinib following various time courses and blotted with specific antibodies. B, gefitinib and S3I-201 attenuate expression of phosphorylation of STAT3 (pY705) and CEBPD in J82 cells. Cells were treated with CDDP alone or in combination with gefitinib or S3I-201 for 24 h. Western blot was conducted with indicated antibodies, respectively.

Expression and activities of EGFR and STAT3 levels in UCUB cells. Western blot analysis was conducted with lysates from UCUB cells. TSGH8301 (TSG), TCCSUP (TCC), NTUB1 (NTU) and CDDP resistance NTUB1/P (NTP).

Overexpression of CEBPD upregulates ABCB1 and ABCC2 transcription in CDDP-resistant NTUB1/P (NTP) cells. A, RT-PCR assay was conducted with total RNA harvested from NTP cells transfected with pCDNA3/HA/CEBPD expression vectors. B, overexpression of CEBPD upregulates ABCB1 transcription containing known exon 1 (NM_000927.4) in CDDP-resistant NTP cells. RT-PCR assay was conducted with total RNA harvested from NTP cells transfected with pCDNA3/HA/CEBPD expression vectors. Upper panel shows location of forward primer (+159) in exon 1 and reverse primer (+348) in exon 2 of ABCB1 gene locus.

Gefitinib and S3I-201 attenuate ABCB1 transcripts of and enhance CDDP sensitivity of J82 cells. A, cells were treated with CDDP alone or in combination with gefitinib or S3I-201 for 24 h. A qPCR assay was conducted with specific primers for ABCB1 gene. ***, significant difference (P < 0.001). B, J82 cells were then transfected with ABCB1 reporter (-21~+917) (AB-I, as shown in Fig. 3D) and treated with CDDP alone or in combination with gefitinib or S3I-201. After 24 h. A Luciferase assay was conducted using lysates of transfectants. ***, significant difference (P < 0.001). C, gefitinib and S3I-201 attenuate ABCB1 pump activity and enhance CDDP sensitivity of J82 cells. J82 cells were treated with CDDP alone or CDDP combining gefitinib or S3I-201 for 24 h, MDR dye-loading solution was added to each well for 4 h, and fluorescence intensities were detected by ELISA reader. ***, significant difference (P < 0.001). D, J82 cells were treated with CDDP alone or CDDP combining gefitinib or S3I-201 for 24 h. Death of experimental cells was examined by PI staining. ***, significant difference (P < 0.001). E, CEBPD contributes to drug resistance of cisplatin (CDDP) in nasopharyngeal carcinoma HONE1/R (CDDP-resistant HONE) cells. HONE1/R cells were resistant to CDDP treatment. HONE1 and HONE1/R cells were treated with CDDP in dose-dependent manner. Cell viability was examined by CCK8 assay. *, significant difference (P < 0.05). **, significant difference (P < 0.01). ***, significant difference (P < 0.001). F, CDDP induces expression of CEBPD and ABCB1. HONE1/R cells were treated with CDDP for 24 h and then lysates of experimental cells were analyzed by Western blot. G, HONE1/R cells were treated with gefitinib, S3I-201 or CDDP or in combination as indicated for 24 h, MDR dye-loading solution was added to each well for 4 h and fluorescence intensities were detected by ELISA reader. ***, significant difference (P < 0.001). H, S3I-201 enhances sensitivity of CDDP in HONE1/R cells. HONE1/R cells were treated with CDDP (20 μM) alone or in combination with S3I-201 (100 μM) for 24 h and death of experimental cells was examined by PI staining. ***, significant difference (P < 0.001).

PARP1 inhibitors (PARPi) are currently used in the clinic for the treatment of ovarian and breast cancers, yet their therapeutic efficacy against hepatocellular carcinoma (HCC) has been disappointing. To ensure therapeutic efficacy of PARPi against HCC, a disease often diagnosed at intermediate to advanced stages with no effective treatment options, it is critical to identify not only biomarkers to predict PARPi resistance but also rational treatments to overcome this. Here, we report that a heterodimer of EGFR and MET interacts with and phosphorylates Y907 of PARP1 in the nucleus, which contributes to PARPi resistance. Inhibition of both EGFR and MET sensitized HCC cells to PARPi, and both EGFR and MET are known to be overexpressed in HCC. This report provides an explanation for the poor efficacy of PARPi against HCC and suggests combinatorial treatment consisting of EGFR, MET, and PARP inhibitors may be an effective therapeutic strategy in HCC.Significance:Regulation of PARP by the c-MET and EGFR heterodimer suggests a potentially effective combination therapy to sensitize HCC to PARPi.

CEBPD expression is correlated with pEGFR, pSTAT3 and ABCB1 expression in UCUB patients. Immunostaining on representative case developed metastatic disease (Case-45, Left panel) exhibits high pEGFR, pSTAT3, CEBPD, and ABCB1 expression; while the case with superior outcome (Case-32, Right panel) shows low expression of these proteins.

PDF file, 146K, Figure S1: Expressions of Cytokines in CD4+LAP+, CD4+Foxp3- and CD8+ T cells. The dot blot showed the expression of IFN-γ, IL-17, IL-2 and IL-10 on LAP+CD4+ T cells, Foxp3-CD4+ T cells and total CD8+ T cells were analyzed by FACS Calibur.

Purpose: Cisplatin (CDDP) is frequently used in combination chemotherapy with paclitaxel for treating urothelial carcinoma of the urinary bladder (UCUB). CDDP cross-resistance has been suggested to develop with paclitaxel, thus hindering successful UCUB treatment. Therefore, elucidating the mechanisms underlying CDDP-induced anticancer drug resistance is imperative and may provide an insight in developing novel therapeutic strategy.Experimental Design: Loss-of-function assays were performed to elucidate the role of the EGFR and STAT3 in CDDP-induced CCAAT/enhancer-binding protein delta (CEBPD) expression in UCUB cells. Reporter and in vivo DNA-binding assays were employed to determine whether CEBPD directly regulates ATP binding cassette subfamily B member 1 (ABCB1) and ATP binding cassette subfamily C member 2 (ABCC2) activation. Finally, a xenograft animal assay was used to examine the abilities of gefitinib and S3I-201 (a STAT3 inhibitor) to reverse CDDP and paclitaxel sensitivity.Results: CEBPD expression was maintained in postoperative chemotherapy patients, and this expression was induced by CDDP even in CDDP-resistant UCUB cells. Upon CDDP treatment, CEBPD activated ABCB1 and ABCC2. Furthermore, the EGFR/STAT3 pathway contributed to CDDP-induced CEBPD expression in UCUB cells. Gefitinib and S3I-201 treatment significantly reduced the expression of CEBPD and enhanced the sensitivity of CDDP-resistant UCUB cells to CDDP and paclitaxel.Conclusions: Our results revealed the risk of CEBPD activation in CDDP-resistant UCUB cells and suggested a therapeutic strategy for patients with UCUB or UCUB resisted to CDDP and paclitaxel by combination with either gefitinib or S3I-201. Clin Cancer Res; 23(2); 503–13. ©2016 AACR.

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Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with features of early metastasis and extensive desmoplasia which is the growth of fibrous or connective tissue. The PDAC patient outcomes have not improved in decades, albeit a plenty of therapies that target PDAC have been tested but failed. It could be due to most research focused on tumor cells, whereas the effect of the stroma and immune cells during tumor progression has been largely ignored. Bradykinin receptor B1 (BDKRB1), one of the cell-surface G protein-coupled receptors plays a crucial role in the physiological process associated with coagulation, vasodilation and promotes fibrotic diseases and inflammation in pathological conditions. Based on the analyzed results of Cancer Genome Atlas (TCGA) database, BDKRB1 mRNA is upregulated in PDAC patients. Of note, BDKRB1 mRNA level positively associates with several activated human pancreatic stellate cells (HPSCs; the main fibroblastic cells of the pancreas)-related genes such as CTGF (connective tissue growth factor), IL6 (interleukin 6), and PDGFB (platelet derived growth factor subunit B) in PDAC patients. Interestingly, the conditioned medium from des-Arg9-bradykinin treated-HPSCs can induce M2-like macrophages polarization which dictate clinically relevant immunosuppression in metastatic tumors and express immunosuppressive cytokines such as IL-10, CCL17 and CCL18, implying that activation of BDKRB1 not only stimulates PDAC growth, but also causes HPSCs activation, desmoplasia and immune suppression, followed by exacerbating tumor progression. Moreover, a chemotherapy gemcitabine remains a cornerstone of PDAC treatment in all stages of PDAC despite of the low therapeutic efficacy, we demonstrated that treatment of gemcitabine increases the expression of BDKRB1 in HPSCs and PDAC, indicating that activation of BDKRB1 could be one of the mechanisms involved in gemcitabine resistance. Here, we hypothesize BDKRB1 activation may orchestrate the tumor microenvironment which promotes PDAC progression by inducing tumor cells survival, HPSCs activation, immune suppression, and may contribute to chemotherapy resistance. The outcomes of this proposal will provide not only a novel cellular and molecular mechanism involved in progression of PDAC, but also highlight a potential therapeutic approach to improve PDAC treatment. Citation Format: Ching-Fei Li, Jie Fu, Heng-Huan Lee, Ying-Nai Wang, Yu-Yi Chu, Paul J. Chiao, Mien-Chie Hung. Investigation of the roles of BDKRB1 in pancreatic cancer progression and tumor microenvironment modulation [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr C070.

Expression of the receptor tyrosine kinase ephrin receptor A10 (EphA10), which is undetectable in most normal tissues except for the male testis, has been shown to correlate with tumor progression and poor prognosis in several malignancies, including triple-negative breast cancer (TNBC). Therefore, EphA10 could be a potential therapeutic target, likely with minimal adverse effects. However, no effective clinical drugs against EphA10 are currently available. Here, we report high expression levels of EphA10 in tumor regions of breast, lung, and ovarian cancers as well as in immunosuppressive myeloid cells in the tumor microenvironment. Furthermore, we developed anti-EphA10 monoclonal antibodies (mAbs) that specifically recognize cell surface EphA10, but not other EphA family isoforms, and target tumor regions precisely in vivo with no apparent accumulation in other organs. In syngeneic TNBC mouse models, we found that anti-EphA10 mAb clone #4 enhanced tumor regression, therapeutic response rate, and T cell-mediated antitumor immunity. Notably, the chimeric antigen receptor T cells derived from clone #4 significantly inhibited TNBC cell viability in vitro and tumor growth in vivo. Together, our findings suggest that targeting EphA10 via EphA10 mAbs and EphA10-specific chimeric antigen receptor-T cell therapy may represent a promising strategy for patients with EphA10-positive tumors.

Targeting immune checkpoints such as programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) has transformed cancer treatment, with durable clinical responses across a wide range of tumor types. However, a high percentage of patients fail to respond to anti–PD-1/PD-L1 treatment. A greater understanding of PD-L1 regulation is critical to improving the clinical response rate of PD-1/PD-L1 blockade. Here, we demonstrate that PD-L1 is phosphorylated and stabilized by casein kinase 2 (CK2) in cancer and dendritic cells (DC). Phosphorylation of PD-L1 at Thr285 and Thr290 by CK2 disrupted PD-L1 binding with speckle-type POZ protein, an adaptor protein of the cullin 3 (CUL3) ubiquitin E3 ligase complex, protecting PD-L1 from CUL3-mediated proteasomal degradation. Inhibition of CK2 decreased PD-L1 protein levels by promoting its degradation and resulted in the release of CD80 from DC to reactivate T-cell function. In a syngeneic mouse model, combined treatment with a CK2 inhibitor and an antibody against T-cell immunoglobulin mucin-3 (Tim-3) suppressed tumor growth and prolonged survival. These findings uncover a mechanism by which PD-L1 is regulated and suggest a potential antitumor treatment option to activate DC function by blocking the CK2–PD-L1 pathway and inhibiting Tim-3. Significance: This work identifies a role for CK2 in immunosuppression by phosphorylation and stabilization of PD-L1, identifying CK2 inhibition as an immunotherapeutic approach for treating cancer.

Poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi), which induce DNA damage by inhibiting PARP1 enzymatic activity and trapping PARP on the damaged DNA, are used to eliminate BRCA1/2-mutated (BRCAm) cancer. However, clinical observations suggest that BRCAm tumors develop PARPi resistance. Current strategies to overcome PARPi resistance include impeding multiple DNA repair pathways to induce excessive DNA damage. Here, we propose a novel strategy targeting oncogenic receptor tyrosine kinases to enhance PARP trapping. By developing triple-negative breast cancer (TNBC) cells with acquired talazoparib resistance, we observed a high prevalence of activated fibroblast growth factor receptor 3 (FGFR3) kinase in these cells through kinase antibody array analysis. Mass spectrometry analysis and in vitro kinase assay suggested that FGFR3 phosphorylated PARP1 at tyrosine residues 158 and 176. Biochemistry studies suggested that only PARP1 tyrosine 158 phosphorylation contributes to PARPi resistance in the cells we developed. We then developed a monoclonal antibody against tyrosine 158 phosphorylated PARP1, and found that high-level PARP1 tyrosine 158 phosphorylation positively correlated with PARPi resistance in breast cancer patient-derived xenograft models. We further demonstrated that the combination of FGFR inhibitor and PARPi delayed DNA repair with prolonged PARP trapping. Moreover, synergy between PARPi and FGFR inhibition was observed in multiple TNBC cell lines with PARPi resistance in vitro. The combination of PARPi and FGFR inhibitor also showed synergism in vivo, and treatment with the combination of PARPi and FGFR inhibitor was tolerated in mouse models. These findings reveal that PARP1 tyrosine 158 phosphorylation facilitates resolving of the PARPi-induced PARP-trapping, and that the tyrosine 158 phosphorylated PARP1 may be an effective biomarker to indicate FGFR3 mediated PARPi resistance. Citation Format: MeiKuang Chen, Yuan Gao, Weiya Xia, Yu-Han Wang, Jennifer K. Litton, Yu-Yi Chu, Funda Meric-Bernstam, Helen Piwnica-Worms, Banu Arun, Jordi Rodon Ahnert, Yongkun Wei, Wei-Chao Chang, Hung-Ling Wang, Coya Tapia, Constance T. Albarracin, Shao-Chun Wang, Ying-Nai Wang, Gabriel N. Hortobagyi, Chunru Lin, Liuqing Yang, Dihua Yu, Mien-Chie Hung. FGFR3 mediated PARP1 tyrosine 158 phosphorylation promotes PARP inhibitor resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1792.

Antibodies that target immune checkpoint proteins such as programmed cell death protein 1, programmed death ligand 1, and cytotoxic T-lymphocyte-associated antigen 4 in human cancers have achieved impressive clinical success; however, a significant proportion of patients fail to respond to these treatments. Galectin-9 (Gal-9), a β-galactoside-binding protein, has been shown to induce T-cell death and facilitate immunosuppression in the tumor microenvironment by binding to immunomodulatory receptors such as T-cell immunoglobulin and mucin domain-containing molecule 3 and the innate immune receptor dectin-1, suggesting that it may have potential as a target for cancer immunotherapy. Here, we report the development of two novel Gal-9-neutralizing antibodies that specifically react with the N-carbohydrate-recognition domain of human Gal-9 with high affinity. We also show using cell-based functional assays that these antibodies efficiently protected human T cells from Gal-9-induced cell death. Notably, in a T-cell/tumor cell coculture assay of cytotoxicity, these antibodies significantly promoted T cell-mediated killing of tumor cells. Taken together, our findings demonstrate potent inhibition of human Gal-9 by neutralizing antibodies, which may open new avenues for cancer immunotherapy.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer that lack effective therapeutic strategies. The response rate of PDAC for treatment with gemcitabine, a current first-line chemotherapeutic for this tumor, is lower than 20%. Identifying key targetable molecules that mediate gemcitabine resistance and developing novel strategies for precision PDAC medicine are urgently needed. Most PDACs have either intratumoral hypoxia or high reactive oxygen species (ROS) production; cytotoxic chemotherapy can elevate ROS production in PDACs. Although excessive ROS production leads to oxidative damage of macromolecules such as DNA, pancreatic cancer cells can survive high DNA damage stress levels. Therefore, identifying molecular mechanisms of overcoming ROS-induced stress in pancreatic cancer cells is important for developing novel therapeutic strategies. ROS-induced DNA damage is predominantly repaired via poly (ADP-ribose) polymerase 1 (PARP1)-mediated DNA repair mechanisms. A recent clinical trial reported that PARP inhibitors are effective in treating pancreatic patients carrying BRCA mutations. However, only less than 10% of pancreatic cancer patients bearing BRCA mutated tumors. Activation of the receptor tyrosine kinase c-MET positively correlates with poor prognosis for PDAC, and our previous study showed that nuclear c-MET can phosphorylate PARP1 at tyrosine 907 under ROS stimulation to promote DNA repair. As described herein, we proposed to expand PARP inhibitor-targeted therapy to more pancreatic cancer patients regardless of BRCA mutation status by combining olaparib, a PARP inhibitor, with c-MET inhibitors as we demonstrated in our previous studies in breast cancer. In this prospective study, we found that ROS-inducing chemotherapeutic drugs such as gemcitabine and doxorubicin stimulated nuclear accumulation of c-MET in BxPC-3 and L3.6pl pancreatic cancer cells. We further showed that combining a c-MET inhibitor with gemcitabine or a PARP inhibitor induced more DNA damage than monotherapy did. Moreover, we demonstrated the synergistic antitumor effects of c-MET inhibitors combined with a PARP inhibitor or gemcitabine in eliminating pancreatic cancer cells. These data suggested that accumulation of ROS in pancreatic cancer cells promotes nuclear localization of c-MET, resulting in resistance to both chemotherapy and PARP inhibitors. Our findings suggest that combining c-MET inhibitors with PARP inhibitors or gemcitabine is a novel, rational therapeutic strategy for advanced pancreatic cancer.

BackgroundThe rapid spread of the Coronavirus 2019 disease (COVID-19) had drastically impacted life all over the world. While some economies are actively recovering from this pestilence, others are experiencing fast and consistent disease spread, compelling governments to impose social distancing measures that have put a halt on routines, especially in densely populated areas.ObjectiveAiming at bringing more light on key economic and population health factors affecting the disease spread, this initial study utilizes a quantitative statistical analysis based on the most recent publicly available COVID-19 datasets.MethodsWe have applied Pearson Correlation Analysis and Clustering Analysis (X-Means Clustering) techniques on the data obtained by combining multiple datasets related to country economics, medical system & health, and COVID-19 - related statistics. The resulting dataset consisted of COVID-19 Case and Mortality Rates, Economic Statistics, and Population Public Health Statistics for 165 countries reported between 22 January 2020 and 28 March 2020. The correlation analysis was conducted with the significance level α of 0.05. The clustering analysis was guided by the value of Bayesian Information Criterion (BIC) with the bin value b = 1.0 and the cutoff factor c = 0.5, and have provided a stable split into four country-level clusters.ResultsThe study showed and explained multiple significant relationships between the COVID-19 data and other country-level statistics. We also identified and statistically profiled four major country-level clusters with relation to different aspects of COVID-19 development and country-level economic and health indicators. Specifically, this study identified potential COVID-19 under-reporting traits, as well as various economic factors that impact COVID-19 Diagnosis, Reporting, and Treatment. Based on the country clusters, we also described the four disease development scenarios, which are tightly knit to country-level economic and population health factors. Finally, we highlighted the potential limitation of reporting and measuring COVID-19 and provided recommendations on further in-depth quantitative research.ConclusionsIn this study, we first identified possible COVID-19 reporting issues and biases across different countries and regions. Second, we identified crucial factors affecting the speed of COVID-19 disease spread and provided recommendations on choosing and operating economic and health system factors when analyzing COVID-19 progression. Particularly, we discovered that the political system and compliance with international disease control norms are crucial for effective COVID-19 pandemic cessation. However, the role of some widely-adopted measures, such as GHS Health Index, might have been overestimated in lieu of multiple biases and underreporting challenges. Third, we benchmarked our findings against the widely-adopted Global Health Security (GHS) model and found that the latter might be redundant when measuring and forecasting COVID-19 spread, while its individual components could potentially serve as stronger COVID-19 indicators. Fourth, we discovered four clusters of countries characterized by different COVID-19 development scenarios, highlighting the differences of the disease reporting and progression in different economic and health system settings. Finally, we provided recommendations on sophisticated measures and research approaches to be implemented for effective outbreak measurements, evaluation and forecasting. We have supported the latter recommendations by a preliminary regression analysis based on the our-collected dataset. We believe that our work would encourage further in-depth quantitative research along the direction as well as would be of support to public policy development when addressing the COVID-19 crisis worldwide.

The limited treatment options and therapeutic failure due to acquired resistance for patients with triple-negative breast cancer (TNBC) represent a significant challenge. Inhibitors against poly (ADP-ribose) polymerase (PARP), olaparib and talazoparib, were recently approved for the treatment of metastatic breast cancer (including TNBC) in patients with germline BRCA1/2 mutations. Despite impressive response rates of ~60%, the prolongation in median progression-free survival with a PARPi is modest, suggesting the emergence of resistance. Several studies have reported that receptor tyrosine kinases (RTKs), such as c-MET (also known as hepatocyte growth factor receptor), are involved in resistance to various anti-neoplastic agents, including PARPi. However, the mechanism by which c-MET contributes to acquired resistance to PARPi in TNBC is not fully understood. In this study, we show that hyperactivated c-Met is detected in TNBC cells with acquired resistance to PARPi, and the combination of talazoparib and crizotinib (a multi-kinase inhibitor that inhibits c-MET) synergistically inhibits proliferation in these cells. Unexpectedly, depleting c-MET had limited effect on talazoparib sensitivity in PARPi-resistant cells. Interestingly, we found evidence of epidermal growth factor receptor (EGFR) hyperactivation and interaction of EGFR/c-Met in these cells. Notably, combining EGFR and PARP inhibitors resulted in greater inhibition of proliferation in c-MET-depleted TNBC cells, and combined c-MET and EGFR inhibition increased sensitivity to talazoparib in TNBC cells with acquired resistance to PARPi. Our findings suggest that combined inhibition of c-MET and EGFR could potentially re-sensitize TNBC to the cytotoxic effects of PARPi.

Specifically blocking more than one oncogenic pathway simultaneously in a cancer cell with a combination of different drugs is the mainstay of the majority of cancer treatments. Being able to do this via two targeted pathways without inducing side effects through a general mechanism, such as chemotherapy, could bring benefit to patients. In this work we describe a new dual inhibitor of the JAK-STAT and HDAC pathways through designing and developing two types of molecule based on the JAK2 selective inhibitor XL019 and the pan-HDAC inhibitor, vorinostat. Both series of compounds had examples with low nanomolar JAK2 and HDAC1/6 inhibition. In some cases good HDAC1 selectivity was achieved while retaining HDAC6 activity. The observed potency is explained through molecular docking studies of all three enzymes. One example, 69c had 16–25 fold selectivity against the three other JAK-family proteins JAK1, JAK3 and TYK2. A number of compounds had sub-micromolar potencies against a panel of 4 solid tumor cell lines and 4 hematological cell lines with the most potent compound, 45h, having a cellular IC50 of 70 nM against the multiple myeloma cell line KMS-12-BM. Evidence of both JAK and HDAC pathway inhibition is presented in Hela cells showing that both pathways are modulated. Evidence of apoptosis with two compounds in 4 sold tumor cell lines is also presented.

Previous studies demonstrated that nuclear EGFR is associated with malignancy and resistance to tyrosine kinase inhibitor (TKI). To decipher the cellular compartmentalized functions and underlying mechanism of EGFR, we generated EGFR mutants at nuclear translocation signal (NLS) and nuclear export signal (NES). The results showed that EGFR NLS mutant with decreased nuclear translocation sensitized the cancer cells to TKI and impaired invasiveness and stemness of the cancer cells. In contrast, EGFR NES mutant with increased nuclear accumulation promoted aggressiveness of the cancer cells including TKI resistance. Next, we generated genetically engineered mouse model (GEMM) to validate the resulting phenotypes of the EGFR mutants. To this end, CRISPR-Cas9-mediated genome editing approach was used for generating EGFR mutant knock-in mouse, which harbors an internalization-deficient mutation (EGFRLL/AA), pro-nuclear (NES) mutation (EGFRL749P(S)) and the dual mutations. Upon EGF stimulation nuclear EGFR (nEGFR) accumulation in the hepatocytes from EGFRL749P heterozygous mice was increased. Conversely, a significant reduction of nEGFR was observed in the hepatocyte from EGFTRLL/AA homozygous mice. We found that germline expression of EGFRL749P(S) gave rise to deficiency in breeding. However, EGFRLL/AA mutant mice displayed no gross abnormal even in homozygous offspring, suggesting that depletion of nEGFR has no significant phenotype. We failed to generate homozygotes of EGFR NES mutant mice due to embryonic lethality. Of note, the female and male founders and F1 EGFRL798P(S) mice developed tumors with huge spleen. The IHC staining results indicated that the infiltrating lymphocytes in liver, stomach and kidney were B cells. The latency of the B cell lymphomagenesis was from 17 to 23 months, and tumor incidence of F1 offspring of EGFRL749P(S) knock-in mice was 10 of 15 (66.7%). These findings indicated that single allele mutation in EGFR NES domain is capable of driving lymphomagenesis, suggesting that the EGFR NES mutant possesses a strong oncogenic activity. Surprisingly, by searching database of EGFR mutation in human tumors (Cosmic database https://cancer.sanger.ac.uk/cosmic ), we noticed that among the somatic mutations in EGFR gene the gene amplification represents only small part of the gene alternations in human cancers, and EGFR L747P and L747S mutation occur in the lung cancer patients with intrinsic TKI resistance. In addition, the majority of human B lymphocyte originated tumors harboring EGFR NES mutation are also heterozygotes. Cosmic database shows that somatic mutations at the EGFR NES region occur in the patients with plasma cell lymphoma. Taken together, our findings suggest that nEGFR contributes to malignant potential of the tumors and EGFR with NES mutation is a tumor driver. Citation Format: Lei Nie, Junwei Hou, Yu-Yi Chu, Ying-Nai Wnag, Li-Chuan Chan, Longfei Huo, Paul Chiao, Shao-Chun Wang, Mien-Chie Hung, Michael Wang. Compartmentalized functions of epidermal growth factor receptor (EGFR) in tumorigenesis and malignant phenotypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 120.

Triple-negative breast cancer (TNBC), which lacks estrogen receptor α (ERα), progesterone receptor, and human epidermal growth factor receptor 2 (HER2) expression, is closely related to basal-like breast cancer. Previously, we and others report that cyclin E/cyclin-dependent kinase 2 (CDK2) phosphorylates enhancer of zeste homolog 2 (EZH2) at T416 (pT416-EZH2). Here, we show that transgenic expression of phospho-mimicking EZH2 mutant EZH2T416D in mammary glands leads to tumors with TNBC phenotype. Coexpression of EZH2T416D in mammary epithelia of HER2/Neu transgenic mice reprograms HER2-driven luminal tumors into basal-like tumors. Pharmacological inhibition of CDK2 or EZH2 allows re-expression of ERα and converts TNBC to luminal ERα-positive, rendering TNBC cells targetable by tamoxifen. Furthermore, the combination of either CDK2 or EZH2 inhibitor with tamoxifen effectively suppresses tumor growth and markedly improves the survival of the mice bearing TNBC tumors, suggesting that the mechanism-based combination therapy may be an alternative approach to treat TNBC., EZH2 phosphorylation by CDK2 promotes progression of triple-negative breast cancer (TNBC). Here, the authors show that this signaling axis downregulates ERα, and thus combinatorial blockade of CDK2 and EZH2 sensitizes TNBC cells to tamoxifen.

Random access channel (RACH) is the first step that needs to be conducted by any devices which require data transmission in modern cellular networks. Since LTE and beyond also often used to serve IoT application due to its wide coverage and availability, conserving energy is important. This paper presents a flexible preamble allocation scheme which is applied in RACH procedure and evaluates its energy consumption. The flexible preamble allocation enables base station to fine-tune its preamble allocation, which mainly used to prioritize H2H services over IoT ones. Such access prioritization translates in more efficient signaling transmissions during RACH procedure by reducing collision. The result demonstrates that the scheme can effectively reduce energy usage compared with the standard RACH procedure with shared preamble pool.

Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) are promising targeted therapeutics for breast and ovarian cancers bearing a germline BRCA1/2 mutation (BRCA (m)), and several have already received regulatory approval in the United States. In patients with a BRCA (m) cancer, PARPi can increase the burden of unrepaired DNA double-strand breaks by blocking PARP activity and trapping PARP1 onto damaged DNA. Resistance to PARP inhibitors can block the formation of DNA double-strand breaks through BRCA-related DNA repair pathway. MET is a hyper-activated receptor tyrosine kinase expressed in multiple cancer types and the activation contributes to resistance to DNA damage-inducing therapeutic drugs. Our previous study showed that MET inhibition by pan-kinase inhibitors has synergism with PARPi in suppressing growth of breast cancer in vitro and in xenograft tumor models. In this study, we validated the inhibitory effect of novel inhibitors, HS10241 (selective MET inhibitor) and HS10160 (PARPi), to their target respectively in triple-negative breast cancer (TNBC) and high-grade serous ovarian cancer (HGSOC) cells. We further demonstrated that these two inhibitors function synergistically in eliminating TNBC and HGSOC cells; combining with HS10241 increased DNA double-strand breaks induced by HS10160 in cancer cells; and PARP1 tyrosine (Y)-907 phosphorylation (PARP1 p-Y907) can be an effective biomarker as an indicator of MET-mediated PARPi in HGSOC. Our results suggest that the combination of HS10241 and HS10160 may benefit patients bearing tumors overexpressing MET as well as those resistant to single-agent PARPi treatment.