APLF facilitates interstrand DNA crosslink repair and replication fork protection to confer cisplatin resistance.

Replication stress converts the stalled forks into reversed forks, which is an important protection mechanism to prevent fork degradation and collapse into poisonous DNA double-strand breaks (DSBs). Paradoxically, the mechanism also acts in cancer cells to contribute to chemoresistance against various DNA-damaging agents. PARP1 binds to and is activated by stalled forks to facilitate fork reversal. Aprataxin and polynucleotide kinase/phosphatase-like factor (APLF) binds to PARP1 through the poly(ADP-ribose) zinc finger (PBZ) domain and is known to be involved in non-homologous end joining (NHEJ). Here, we identify a novel function of APLF involved in interstrand DNA crosslink (ICL) repair and fork protection. We demonstrate that PARP1 activity facilitates the APLF recruitment to stalled forks, enabling the FANCD2 recruitment to stalled forks. The depletion of APLF sensitizes cells to cisplatin, impairs ICL repair, reduces the FANCD2 recruitment to stalled forks, and results in nascent DNA degradation by MRE11 nucleases. Additionally, cisplatin-resistant cancer cells show high levels of APLF and homologous recombination-related gene expression. The depletion of APLF sensitizes cells to cisplatin and results in fork instability. Our results reveal the novel function of APLF to facilitate ICL repair and fork protection, thereby contributing to cisplatin-resistant phenotypes of cancer cells.

Prussian blue analog with separated active sites to catalyze water driven enhanced catalytic treatments.

Chemodynamic therapy (CDT) uses the Fenton or Fenton-like reaction to yield toxic ‧OH following H2O2 → ‧OH for tumoral therapy. Unfortunately, H2O2 is often taken from the limited endogenous supply of H2O2 in cancer cells. A water oxidation CoFe Prussian blue (CFPB) nanoframes is presented to provide sustained, external energy-free self-supply of ‧OH from H2O to process CDT and/or photothermal therapy (PTT). Unexpectedly, the as-prepared CFPB nanocubes with no near-infrared (NIR) absorption is transformed into CFPB nanoframes with NIR absorption due to the increased Fe3+-N ≡ C-Fe2+ composition through the proposed proton-induced metal replacement reactions. Surprisingly, both the CFPB nanocubes and nanoframes provide for the self-supply of O2, H2O2, and ‧OH from H2O, with the nanoframe outperforming in the production of ‧OH. Simulation analysis indicates separated active sites in catalyzation of water oxidation, oxygen reduction, and Fenton-like reactions from CFPB. The liposome-covered CFPB nanoframes prepared for controllable water-driven CDT for male tumoral mice treatments.

Atomically dispersed golds on degradable zero-valent copper nanocubes augment oxygen driven Fenton-like reaction for effective orthotopic tumor therapy.

Herein, we employ a galvanic replacement approach to create atomically dispersed Au on degradable zero-valent Cu nanocubes for tumor treatments on female mice. Controlling the addition of precursor HAuCl4 allows for the fabrication of different atomic ratios of AuxCuy. X-ray absorption near edge spectra indicates that Au and Cu are the predominant oxidation states of zero valence. This suggests that the charges of Au and Cu remain unchanged after galvanic replacement. Specifically, Au0.02Cu0.98 composition reveals the enhanced •OH generation following O2 → H2O2 → •OH. The degradable Au0.02Cu0.98 released Cu+ and Cu2+ resulting in oxygen reduction and Fenton-like reactions. Simulation studies indicate that Au single atoms boot zero-valent copper to reveal the catalytic capability of Au0.02Cu0.98 for O2 → H2O2 → •OH as well. Instead of using endogenous H2O2, H2O2 can be sourced from the O2 in the air through the use of nanocubes. Notably, the Au0.02Cu0.98 structure is degradable and renal-clearable.

BMP2 promotes lung adenocarcinoma metastasis through BMP receptor 2-mediated SMAD1/5 activation.

Bone morphogenetic protein 2 (BMP2) is highly overexpressed in human non-small cell lung cancer (NSCLC) and correlates with tumor stage and metastatic burden. Although several lines of evidence suggest that BMP2 promotes cell migration and invasiveness in vitro, the in vivo role of BMP2 in the metastasis of lung adenocarcinoma cells remains less well understood. Here, we revealed that BMP2 is highly overexpressed in lung adenocarcinoma patients with lymph node metastasis compared with patients without lymph node metastasis. Using an in vivo orthotopic mouse model, we clearly demonstrated that BMP2 promotes lung adenocarcinoma metastasis. The depletion of BMP2 or its receptor BMPR2 significantly reduced cell migration and invasiveness. We further identified that BMP2/BMPR2-mediated cell migration involves the activation of the SMAD1/5/8 signaling pathway, independent of the KRAS signaling pathway. Significantly, the depletion of SMAD1/5/8 or the inhibition of SMAD1/5/8 by LDN193189 inhibitor significantly reduced cell migration. These findings show that BMP2 promotes NSCLC metastasis, indicating that targeting the BMP2 signaling pathway may represent a potential therapeutic strategy for treating patients with metastatic NSCLC.

Polyaniline-Based Glyco-Condensation on Au Nanoparticles Enhances Immunotherapy in Lung Cancer.

Lung cancer is considered among the deadliest cancers with a poor prognosis. Au@PG nanoparticles (NPs) are gold (Au)-based NPs featuring a polyaniline-based glyco structure (PG) generated from the polymerization of ortho-nitrophenyl-ß-d-galactopyranoside (ONPG) with promising M1 macrophage polarization activity, resulting in tumor remodeling and from a cold to a hot microenvironment, which promotes the cytotoxic T cell response and tumor inhibition. The combination of Au@PG NPs and anti-programmed cell death protein 1 (PD-1) therapy improved tumor inhibition and immunosuppression, accompanied by the secretion of immunogenic cytokines. A one-pot synthetic method was developed to achieve glyco-condensation during the formation of Au@PG NPs, which induced macrophage polarization more efficiently than Au@glucose, Au@mannose, and Au@galactose NPs. The switch from M2 to M1 macrophages was dependent on NP size, with smaller Au@PG NPs performing better than larger ones, with effectiveness ranked as follows: 32.2 nm ≈ 29.8 nm < 26.4 nm < 18.3 nm. Cellular uptake by endocytosis induced size-dependent endoplasmic reticulum (ER) stress, which resulted in the activation of spleen tyrosine kinase (SYK), leading to immune modulations and macrophage polarization. Our results suggested the promising potential of Au@PG NPs in lung cancer immunotherapy.

Chemical Structure and Shape Enhance MR Imaging-Guided X-ray Therapy Following Marginative Delivery.

Ineffective site-specific delivery has seriously impeded the efficacy of nanoparticle-based drugs to a disease site. Here, we report the preparation of three different shapes (sphere, scroll, and oblate) to systematically evaluate the impact of the marginative delivery on the efficacy of magnetic resonance (MR) imaging-guided X-ray irradiation at a low dose of 1 Gy. In addition to the shape effect, the therapeutic efficacy is investigated for the first time to be strongly related to the structure effect that is associated with the chemical activity. The enhanced particle-vessel wall interaction of both the flat scroll and oblate following margination dynamics leads to greater accumulation in the lungs, resulting in superior performance over the sphere against lung tumor growth and suppression of lung metastasis. Furthermore, the impact of the structural discrepancy in nanoparticles on therapeutic efficacy is considered. The tetragonal oblate reveals that the feasibility of the charge-transfer process outperforms the orthorhombic scroll and cubic sphere to suppress tumors. Finally, surface area is also a crucial factor affecting the efficacy of X-ray treatments from the as-prepared particles.

Synergistic Action of Immunotherapy and Nanotherapy against Cancer Patients Infected with SARS-CoV-2 and the Use of Artificial Intelligence.

Since 2019, the SARS-CoV-2 pandemic has caused a huge chaos throughout the world and the major threat has been possessed by the immune-compromised individuals involving the cancer patients; their weakened immune response makes them vulnerable and susceptible to the virus. The oncologists as well as their patients are facing many problems for their treatment sessions as they need to postpone their surgery, chemotherapy, or radiotherapy. The approach that could be adopted especially for the cancer patients is the amalgamation of immunotherapy and nanotherapy which can reduce the burden on the healthcare at this peak time of the infection. There is also a need to predict or analyze the data of cancer patients who are at a severe risk of being exposed to an infection in order to reduce the mortality rate. The use of artificial intelligence (AI) could be incorporated where the real time data will be available to the physicians according to the different patient's clinical characteristics and their past treatments. With this data, it will become easier for them to modify or replace the treatment to increase the efficacy against the infection. The combination of an immunotherapy and nanotherapy will be targeted to treat the cancer patients diagnosed with SARS-CoV-2 and the AI will act as icing on the cake to monitor, predict and analyze the data of the patients to improve the treatment regime for the most vulnerable patients.

A Narrative Review of Implementing Precision Oncology in Metastatic Castration-Resistant Prostate Cancer in Emerging Countries.

The therapeutic landscape of metastatic castration-resistant prostate cancer (mCRPC) has evolved considerably with the introduction of newer agents, such as poly-ADP ribose polymerase (PARP) inhibitors targeting DNA damage repair mutations. Combining and sequencing novel and existing therapies appropriately is necessary for optimizing the management of mCRPC and ensuring better treatment outcomes. The purpose of this review is to provide evidence-based answers to key clinical questions on treatment selection, treatment sequencing patterns, and factors influencing treatment decisions in the management of mCRPC in the era of PARP inhibitors. This article can also serve as a comprehensive guide to clinicians for optimizing genetic testing and counseling and management of patients with mCRPC. Although the PROfound study has validated the concept of PARP sensitivity across multiple genes associated with homologous recombination repair (HRR) in mCRPC and highlighted the importance of genomic testing in this at-risk patient population, it still remains unclear how patients with rarer HRR mutations will respond to PARP inhibitors. Therefore, real-world data obtained through registry-based randomized controlled trials in the future may help produce robust scientific evidence for supporting optimal clinician decision-making in the management of mCRPC.

Transferrin Modified GSH Sensitive Hyaluronic Acid Derivative Micelle to Deliver HSP90 Inhibitors to Enhance the Therapeutic Efficacy of Brain Cancers.

Herein, GSH-sensitive hyaluronic acid-poly(lactic-co-glycolic acid) (HA-SS-PLGA) was synthesized. Surface modification of PLGA with hyaluronic acid produced a highly stable micelle at physiological pH while a micelle was destabilized at a higher GSH level. Fluorescence microscopy results showed that rhodamine-encapsulated micelle was taken up by brain cancer cells, while competitive inhibition was observed in the presence of free HA and free transferrin. In vitro cytotoxicity results revealed that transferrin-targeted nanoformulated AUY922 (TF-NP-AUY922) shows higher cytotoxicity than either free AUY922 or non-targeted AUY922-loaded micelles (NP-AUY922). In comparison to the control groups, free AUY922, TF-NP-AUY922 or NP-AUY922 treatment revealed the upregulation of HSP70, while the expression of HSP90 client proteins was simultaneously depleted. In addition, the treatment group induced caspase-dependent PARP cleavage and the upregulation of p53 expression, which plays a key role in apoptosis of brain cancer cells. In vivo and ex vivo biodistribution studies showed that cypate-loaded micelle was taken up and accumulated in the tumor regions. Furthermore, in vivo therapeutic efficacy studies revealed that the AUY922-loaded micelle significantly suppressed tumor growth in comparison to the free AUY922, or control groups using tumor-bearing NOD-SCID mice. Moreover, biochemical index and histological analysis revealed synthesized micelle does not show any significant cytotoxicity to the selected major organs. Overall, a synthesized micelle is the best carrier for AUY922 to enhance the therapeutic efficiency of brain cancer.

Synchronization of Nanoparticle Sensitization and Radiosensitizing Chemotherapy through Cell Cycle Arrest Achieving Ultralow X-ray Dose Delivery to Pancreatic Tumors.

Pancreatic cancer is among the leading causes of cancer-related death and remains a formidable therapeutic challenge. To date, surgical resection and chemotherapy have been the standards of care. Methotrexate (MTX), which is recognized as a refractory drug for pancreatic cells, was conjugated to the surface of LiYF4:Ce3+ nanoparticles (NP-MTX) through a photocleavable linker molecule. When LiYF4:Ce3+ NPs are stimulated by X-rays, they emit light, which induces the photocleavage of the photolabile linker molecule to release MTX. MTX can target pancreatic tumors, which overexpress folic acid (FA) receptors and are internalized into the cell through receptor-mediated endocytosis. The synergistic effect of the NP-MTX treatment initiated by X-ray irradiation occurs due to the combination of nanoparticle sensitization and the radiosensitizing chemotherapy of the photocleaved MTX molecule. This dual sensitization effect mediated by NP-MTX enabled 40% dose enhancement, which corresponded with an increase in the generation of cytotoxic cellular reactive oxygen species (ROS) and enhanced S phase arrest within the cell cycle. The delivery of an ultralow radiation dose of 0.1 Gy resulted in the photocleavage of MTX from NP-MTX, and this strategy demonstrated in vivo efficacy against AsPC-1 and PANC-1 xenografted pancreatic tumors.

Study on the pathophysiological mechanism responsible for lower urinary tract symptoms associated with prostate cancer using an animal model.

OBJECTIVES: To investigate the pathophysiological mechanism leading to lower urinary tract symptoms in prostate cancer (PCa) by using an animal model. METHODS: An orthotopic PCa model in mice was established by injection of human DU145 cells into the prostate gland lateral lobe of NOD.CB17-Prkdcscid /NcrCrlBltw (NOD-SCID) mice. Cancer growth was quantified by a luciferase-based in vivo imaging system (IVIS) serially every 7 days. Comparisons were made for urodynamic parameters, bladder histology, and biological markers until the sixth week. Bladder wall structural changes were assessed by the bladder wall thickness and degree of fibrosis. Biomarker expressions in bladder tissue including muscarinic acetylcholine receptor 2 (M2 ), transient receptor potential cation channel subfamily V member 4 (TRPV4), BCL2-associated X protein (Bax), and caspase3 were evaluated by immunohistochemical staining and immunofluorescence confocal laser scanning microscopy. RESULTS: DU145 cell growth in the prostate was successfully monitored by a luciferase-based IVIS. after orthotopic injection. Using our injection technique, no anatomical obstruction of the bladder outlet and urethra was noted up to 6 weeks after injection. The presence of PCa induced changes in urinary bladder histology, biomarkers, and urodynamic parameters. Cystometry showed features of detrusor overactivity with increased voiding frequency and high-amplitude voiding contractions from the fourth week onward. Histological analyses 4 weeks after DU145 injection demonstrated detrusor thickening and bladder wall fibrosis. Immunohistochemistry showed increased expressions of bladder M2 , TRPV4, Bax, and caspase3 in the PCa mice as early as in the first or second week. CONCLUSIONS: PCa can induce bladder microenvironment changes involving neural receptors and biological mediators leading to histological and functional alterations even in the absence of overt anatomical obstruction.

1550 nm excitation-responsive upconversion nanoparticles to establish dual-photodynamic therapy against pancreatic tumors.

The second near-infrared biological window b (NIR-IIb, 1500-1700 nm) is recently considered as the promising region for deeper tissue penetration. Herein, a nanocarrier for 1550 nm light-responsive dual-photodynamic therapy (PDT) is developed to efficiently boost singlet oxygen (1O2) generation. The dual-photosensitizers (PSs), rose bengal (RB) and chlorin e6 (Ce6), are carried by the silica-coated core-shell LiYbF4:Er@LiGdF4 upconversion nanoparticles (UCNPs), forming UCNP/RB,Ce6. Following 1550 nm laser irradiation, the upconversion emission of UCNP/RB,Ce6 in both green (∼550 nm) and red (∼670 nm) colors is fully utilized to activate RB and Ce6, respectively. The simultaneous triggering of dual-PS generates an abundant amount of 1O2 resulting in boosted PDT efficacy. This dual-PDT nanocarrier presents an enhanced anticancer effect under single dose treatment in comparison with the single-PS ones from in vitro and in vivo treatments. The marriage between the boosted dual-PDT and 1550 nm light excitation is anticipated to provide a new avenue for non-invasive therapy.

Cancer Immunotherapy and Application of Nanoparticles in Cancers Immunotherapy as the Delivery of Immunotherapeutic Agents and as the Immunomodulators.

In the last few decades, cancer immunotherapy becomes an important tactic for cancer treatment. However, some immunotherapy shows certain limitations including poor therapeutic targeting and unwanted side effects that hinder its use in clinics. Recently, several researchers are exploring an alternative methodology to overcome the above limitations. One of the emerging tracks in this field area is nano-immunotherapy which has gone through rapid progress and revealed considerable potentials to solve limitations related to immunotherapy. Targeted and stimuli-sensitive biocompatible nanoparticles (NPs) can be synthesized to deliver immunotherapeutic agents in their native conformations to the site of interest to enhance their antitumor activity and to enhance the survival rate of cancer patients. In this review, we have discussed cancer immunotherapy and the application of NPs in cancer immunotherapy, as a carrier of immunotherapeutic agents and as a direct immunomodulator.

The HLTF-PARP1 interaction in the progression and stability of damaged replication forks caused by methyl methanesulfonate.

Human HLTF participates in the lesion-bypass mechanism through the fork reversal structure, known as template switching of post-replication repair. However, the mechanism by which HLTF promotes the replication progression and fork stability of damaged forks remains unclear. Here, we identify a novel protein-protein interaction between HLTF and PARP1. The depletion of HLTF and PARP1 increases chromosome breaks, further reduces the length of replication tracks, and concomitantly increases the number of stalled forks after methyl methanesulfonate treatment according to a DNA fiber analysis. The progression of replication also depends on BARD1 in the presence of MMS treatment. By combining 5-ethynyl-2'-deoxyuridine with a proximity ligation assay, we revealed that the HLTF, PARP1, and BRCA1/BARD1/RAD51 proteins were initially recruited to damaged forks. However, prolonged stalling of damaged forks results in fork collapse. HLTF and PCNA dissociate from the collapsed forks, with increased accumulation of PARP1 and BRCA1/BARD1/RAD51 at the collapsed forks. Our results reveal that HLTF together with PARP1 and BARD1 participates in the stabilization of damaged forks, and the PARP1-BARD1 interaction is further involved in the repair of collapse forks.

Combination Delivery of Alpha-Tocopheryl Succinate and Curcumin Using a GSH-Sensitive Micelle (PAH-SS-PLGA) to Treat Pancreatic Cancer.

Pancreatic cancer is one of the highest causes of mortality throughout the world; thus, it requires an effective treatment strategy. Some chemotherapeutic agents used in the clinics or under clinical trials are hydrophobic and have poor aqueous solubility; consequently, they also have minimal systemic bioavailability. Nanoparticle-based drug delivery tactics have the potential for overcoming these limitations and enhancing their therapeutic efficacy. Herein, a glutathione (GSH)-sensitive micelle (PAH-SS-PLGA) was synthesized for the combined delivery of alpha-tocopheryl succinate (TOS) and curcumin to improve its therapeutic efficacy. The chemical structures of PAH-SS-PLGA were analyzed using Proton Nuclear Magnetic Resonance (1H-NMR) and Fourier Transform Infrared (FTIR) spectroscopy, whereas the particle size, zeta potential, and surface morphology were observed using dynamic light scattering (DLS) and transmission electron microscopy (TEM). In vitro drug release results revealed that more TOS and curcumin were released in the presence of GSH (5 mM) than the physiological pH value. Fluorescence microscopy images revealed that nanoformulated curcumin/rhodamine was uptaken by PAN02 pancreatic cancer cells. In vitro cytotoxicity assays showed higher cytotoxicity for nanoformulated TOS and/or curcumin than free TOS and/or curcumin. In addition, higher cytotoxicity was observed for combination drugs than free drugs alone. Most interestingly, at all tested concentrations of nanoformulated drugs (PAH-SS-PLGA, TOS, and curcumin), the calculated combination index (CI) value was less than one, which shows that TOS and curcumin have a synergistic effect on cellular proliferation inhibition. Overall, synthesized co-polymers are the best carriers for combination drugs, TOS, and curcumin, because they enhance the therapeutic efficacy and improve pancreatic cancer treatments.

Ramucirumab plus docetaxel versus placebo plus docetaxel in patients with locally advanced or metastatic urothelial carcinoma after platinum-based therapy (RANGE): overall survival and updated results of a randomised, double-blind, phase 3 trial.

BACKGROUND: Ramucirumab-an IgG1 vascular endothelial growth factor receptor 2 antagonist-plus docetaxel was previously reported to improve progression-free survival in platinum-refractory, advanced urothelial carcinoma. Here, we report the secondary endpoint of overall survival results for the RANGE trial. METHODS: We did a randomised, double-blind, phase 3 trial in patients with advanced or metastatic urothelial carcinoma who progressed during or after platinum-based chemotherapy. Patients were enrolled from 124 investigative sites (hospitals, clinics, and academic centres) in 23 countries. Previous treatment with one immune checkpoint inhibitor was permitted. Patients were randomly assigned (1:1) using an interactive web response system to receive intravenous ramucirumab 10 mg/kg or placebo 10 mg/kg volume equivalent followed by intravenous docetaxel 75 mg/m2 (60 mg/m2 in Korea, Taiwan, and Japan) on day 1 of a 21-day cycle. Treatment continued until disease progression, unacceptable toxicity, or other discontinuation criteria were met. Randomisation was stratified by geographical region, Eastern Cooperative Oncology Group performance status at baseline, and visceral metastasis. Progression-free survival (the primary endpoint) and overall survival (a key secondary endpoint) were assessed in the intention-to-treat population. The study is registered with ClinicalTrials.gov, NCT02426125; patient enrolment is complete and the last patient on treatment is being followed up for safety issues. FINDINGS: Between July 20, 2015, and April 4, 2017, 530 patients were randomly allocated to ramucirumab plus docetaxel (n=263) or placebo plus docetaxel (n=267) and comprised the intention-to-treat population. At database lock (March 21, 2018) for the final overall survival analysis, median follow-up was 7·4 months (IQR 3·5-13·9). In our sensitivity analysis of investigator-assessed progression-free survival at the overall survival database lock, median progression-free survival remained significantly improved with ramucirumab compared with placebo (4·1 months [95% CI 3·3-4·8] vs 2·8 months [2·6-2·9]; HR 0·696 [95% CI 0·573-0·845]; p=0·0002). Median overall survival was 9·4 months (95% CI 7·9-11·4) in the ramucirumab group versus 7·9 months (7·0-9·3) in the placebo group (stratified HR 0·887 [95% CI 0·724-1·086]; p=0·25). Grade 3 or worse treatment-related treatment-emergent adverse events in 5% or more of patients and with an incidence more than 2% higher with ramucirumab than with placebo were febrile neutropenia (24 [9%] of 258 patients in the ramucirumab group vs 16 [6%] of 265 patients in the placebo group) and neutropenia (17 [7%] of 258 vs six [2%] of 265). Serious adverse events were similar between groups (112 [43%] of 258 patients in the ramucirumab group vs 107 [40%] of 265 patients in the placebo group). Adverse events related to study treatment and leading to death occurred in eight (3%) patients in the ramucirumab group versus five (2%) patients in the placebo group. INTERPRETATION: Additional follow-up supports that ramucirumab plus docetaxel significantly improves progression-free survival, without a significant improvement in overall survival, for patients with platinum-refractory advanced urothelial carcinoma. Clinically meaningful benefit might be restricted in an unselected population. FUNDING: Eli Lilly and Company.

Ramucirumab plus docetaxel versus placebo plus docetaxel in patients with locally advanced or metastatic urothelial carcinoma after platinum-based therapy (RANGE): a randomised, double-blind, phase 3 trial.

BACKGROUND: Few treatments with a distinct mechanism of action are available for patients with platinum-refractory advanced or metastatic urothelial carcinoma. We assessed the efficacy and safety of treatment with docetaxel plus either ramucirumab-a human IgG1 VEGFR-2 antagonist-or placebo in this patient population. METHODS: We did a randomised, double-blind, phase 3 trial in patients with advanced or metastatic urothelial carcinoma who progressed during or after platinum-based chemotherapy. Patients were enrolled from 124 sites in 23 countries. Previous treatment with one immune-checkpoint inhibitor was permitted. Patients were randomised (1:1) using an interactive web response system to receive intravenous docetaxel 75 mg/m2 plus either intravenous ramucirumab 10 mg/kg or matching placebo on day 1 of repeating 21-day cycles, until disease progression or other discontinuation criteria were met. The primary endpoint was investigator-assessed progression-free survival, analysed by intention-to-treat in the first 437 randomised patients. This study is registered with ClinicalTrials.gov, number NCT02426125. FINDINGS: Between July, 2015, and April, 2017, 530 patients were randomly allocated either ramucirumab plus docetaxel (n=263) or placebo plus docetaxel (n=267). Progression-free survival was prolonged significantly in patients allocated ramucirumab plus docetaxel versus placebo plus docetaxel (median 4·07 months [95% CI 2·96-4·47] vs 2·76 months [2·60-2·96]; hazard ratio [HR] 0·757, 95% CI 0·607-0·943; p=0·0118). A blinded independent central analysis was consistent with these results. An objective response was achieved by 53 (24·5%, 95% CI 18·8-30·3) of 216 patients allocated ramucirumab and 31 (14·0%, 9·4-18·6) of 221 assigned placebo. The most frequently reported treatment-emergent adverse events, regardless of causality, in either treatment group (any grade) were fatigue, alopecia, diarrhoea, decreased appetite, and nausea. These events occurred predominantly at grade 1-2 severity. The frequency of grade 3 or worse adverse events was similar for patients allocated ramucirumab and placebo (156 [60%] of 258 vs 163 [62%] of 265 had an adverse event), with no unexpected toxic effects. 63 (24%) of 258 patients allocated ramucirumab and 54 (20%) of 265 assigned placebo had a serious adverse event that was judged by the investigator to be related to treatment. 38 (15%) of 258 patients allocated ramucirumab and 43 (16%) of 265 assigned placebo died on treatment or within 30 days of discontinuation, of which eight (3%) and five (2%) deaths were deemed related to treatment by the investigator. Sepsis was the most common adverse event leading to death on treatment (four [2%] vs none [0%]). One fatal event of neutropenic sepsis was reported in a patient allocated ramucirumab. INTERPRETATION: To the best of our knowledge, ramucirumab plus docetaxel is the first regimen in a phase 3 study to show superior progression-free survival over chemotherapy in patients with platinum-refractory advanced urothelial carcinoma. These data validate inhibition of VEGFR-2 signalling as a potential new therapeutic treatment option for patients with urothelial carcinoma. FUNDING: Eli Lilly and Company.

Chronic treatment with cisplatin induces chemoresistance through the TIP60-mediated Fanconi anemia and homologous recombination repair pathways.

The Fanconi anemia pathway in coordination with homologous recombination is essential to repair interstrand crosslinks (ICLs) caused by cisplatin. TIP60 belongs to the MYST family of acetyltransferases and is involved in DNA repair and regulation of gene transcription. Although the physical interaction between the TIP60 and FANCD2 proteins has been identified that is critical for ICL repair, it is still elusive whether TIP60 regulates the expression of FA and HR genes. In this study, we found that the chemoresistant nasopharyngeal carcinoma cells, derived from chronic treatment of cisplatin, show elevated expression of TIP60. Furthermore, TIP60 binds to the promoters of FANCD2 and BRCA1 by using the chromatin immunoprecipitation experiments and promote the expression of FANCD2 and BRCA1. Importantly, the depletion of TIP60 significantly reduces sister chromatid exchange, a measurement of HR efficiency. The similar results were also shown in the FNACD2-, and BRCA1-deficient cells. Additionally, these TIP60-deficient cells encounter more frequent stalled forks, as well as more DNA double-strand breaks resulting from the collapse of stalled forks. Taken together, our results suggest that TIP60 promotes the expression of FA and HR genes that are important for ICL repair and the chemoresistant phenotype under chronic treatment with cisplatin.

Combined Interactions of Plant Homeodomain and Chromodomain Regulate NuA4 Activity at DNA Double-Strand Breaks.

DNA double-strand breaks (DSBs) represent one of the most threatening lesions to the integrity of genomes. In yeast Saccharomyces cerevisiae, NuA4, a histone acetylation complex, is recruited to DSBs, wherein it acetylates histones H2A and H4, presumably relaxing the chromatin and allowing access to repair proteins. Two subunits of NuA4, Yng2 and Eaf3, can interact in vitro with methylated H3K4 and H3K36 via their plant homeodomain (PHD) and chromodomain. However, the roles of the two domains and how they interact in a combinatorial fashion are still poorly characterized. In this study, we generated mutations in the PHD and chromodomain that disrupt their interaction with methylated H3K4 and H3K36. We demonstrate that the combined mutations in both the PHD and chromodomain impair the NuA4 recruitment, reduce H4K12 acetylation at the DSB site, and confer sensitivity to bleomycin that induces DSBs. In addition, the double mutant cells are defective in DSB repair as judged by Southern blot and exhibit prolonged activation of phospho-S129 of H2A. Cells harboring the H3K4R, H3K4R, K36R, or set1Δ set2Δ mutant that disrupts H3K4 and H3K36 methylation also show very similar phenotypes to the PHD and chromodomain double mutant. Our results suggest that multivalent interactions between the PHD, chromodomain, and methylated H3K4 and H3K36 act in a combinatorial manner to recruit NuA4 and regulate the NuA4 activity at the DSB site.