CD38-Directed Vincristine Nanotherapy for Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) is the most common malignancy in children. Although intensive chemotherapy greatly improved the survival rate, it is often accompanied by severe and lifelong side effects as a result of weak ALL selectivity. The intensive and poorly selective chemotherapy is also detrimental to patients' immune system. There is an urgent need to develop more selective and less toxic chemotherapy for ALL. Here, we report daratumumab-polymersome-vincristine (DP-VCR) as a CD38-directed nanotherapy for ALL. DP-VCR showed selective uptake in CD38-positive 697 and Nalm-6-Luc ALL cells and potent anti-ALL activity with an IC50 as low as 0.06 nM VCR, which was 13.7-fold more potent than free VCR. In contrast, no toxicity to human peripheral blood mononuclear cells was detected for DP-VCR even at 108.3 nM VCR. The apoptotic assays confirmed a high selectivity of DP-VCR to CD38-positive ALL cells. DP-VCR exhibited superior treatment of both 697 and Nalm-6-Luc orthotopic ALL models to all controls, as revealed by significant survival benefit and marked reduction of leukemia burden in bone marrow, blood, spleen, and liver. Importantly, DP-VCR induced few side effects. DP-VCR emerges as a safe and potent nanotherapy for CD38-positive ALL.

Small, Smart, and LDLR-Specific Micelles Augment Sorafenib Therapy of Glioblastoma.

Targeted molecular therapy, for example, with sorafenib (SF) is considered as a new and potent strategy for glioblastoma (GBM) that remains hard to treat today. Several clinical trials with SF, as monotherapy or combination therapy with current treatments, have not met the clinical endpoints, likely as a result of the blood-brain barrier (BBB) and inferior GBM delivery. Here, we designed and explored small, smart, and LDLR-specific micelles to load SF (LDLR-mSF) and to improve SF therapy of GBM by enhancing BBB penetration, GBM accumulation, and cell uptake. LDLR-mSF with 2.5% ApoE peptide functionality based on poly(ethylene glycol)-poly(ε-caprolactone-co-dithiolane trimethylene carbonate)-mefenamate exhibited nearly quantitative SF loading, small size (24 nm), high colloidal stability, and glutathione-activated SF release. The in vitro and in vivo studies certified that LDLR-mSF greatly enhanced BBB permeability and U-87 MG cell uptake and caused 10.6- and 12.9-fold stronger anti-GBM activity and 6.0- and 2.5-fold higher GBM accumulation compared with free SF and non-LDLR mSF controls, respectively. The treatment of an orthotopic human GBM tumor model revealed that LDLR-mSF at a safe dosage of 15 mg of SF/kg significantly retarded tumor progression and improved the survival rate by inducing tumor cell apoptosis and inhibiting tumor angiogenesis. These small, smart, and LDLR-specific micelles provide a potential solution to enhance targeted molecular therapy of GBM.

Biodegradable Polymersomes with Structure Inherent Fluorescence and Targeting Capacity for Enhanced Photo-Dynamic Therapy.

Biodegradable nanostructures displaying aggregation-induced emission (AIE) are desirable from a biomedical point of view, due to the advantageous features of loading capacity, emission brightness, and fluorescence stability. Herein, biodegradable polymers comprising poly (ethylene glycol)-block-poly(caprolactone-gradient-trimethylene carbonate) (PEG-P(CLgTMC)), with tetraphenylethylene pyridinium-TMC (PAIE) side chains have been developed, which self-assembled into well-defined polymersomes. The resultant AIEgenic polymersomes are intrinsically fluorescent delivery vehicles. The presence of the pyridinium moiety endows the polymersomes with mitochondrial targeting ability, which improves the efficiency of co-encapsulated photosensitizers and improves therapeutic index against cancer cells both in vitro and in vivo. This contribution showcases the ability to engineer AIEgenic polymersomes with structure inherent fluorescence and targeting capacity for enhanced photodynamic therapy.

Molecular Programming of Biodegradable Nanoworms via Ionically Induced Morphology Switch toward Asymmetric Therapeutic Carriers.

Engineering biodegradable nanostructures with precise morphological characteristics is a key objective in nanomedicine. In particular, asymmetric (i.e., nonspherical) nanoparticles are desirable due to the advantageous effects of shape in a biomedical context. Using molecular engineering, it is possible to program unique morphological features into the self-assembly of block copolymers (BCPs). However, the criteria of biocompatibility and scalability limit progress due to the prevalence of nondegradable components and the use of toxic solvents during fabrication. To address this shortfall, a robust strategy for the fabrication of morphologically asymmetric nanoworms, comprising biodegradable BCPs, has been developed. Modular BCPs comprising poly (ethylene glycol)-block-poly(caprolactone-gradient-trimethylene carbonate) (PEG-PCLgTMC), with a terminal chain of quaternary ammonium-TMC (PTMC-Q), undergo self-assembly via direct hydration into well-defined nanostructures. By controlling the solution ionic strength during hydration, particle morphology switches from spherical micelles to nanoworms (of varying aspect ratio). This ionically-induced switch is driven by modulation of chain packing with salts screening interchain repulsions, leading to micelle elongation. Nanoworms can be loaded with cytotoxic cargo (e.g., doxorubicin) at high efficiency, preferentially interact with cancer cells, and increase tumor penetration. This work showcases the ability to program assembly of BCPs and the potential of asymmetric nanosystems in anticancer drug delivery.

GE11-Directed Functional Polymersomal Doxorubicin as an Advanced Alternative to Clinical Liposomal Formulation for Ovarian Cancer Treatment.

Ovarian cancer as a recurrent disease is often refractory to treatment including pegylated liposomal doxorubicin hydrochloride (Lipo-Dox). Here, GE11 peptide-modified reversibly cross-linked polymersomal doxorubicin (GE11-PS-Dox) was investigated as an advanced treatment for SKOV3 human ovarian tumors, which overexpress epidermal growth factor receptor (EGFR). The in vitro experiments using SKOV3 cancer cells demonstrated that GE11-PS-Dox induced obviously higher cellular uptake, Dox delivery to the nuclei, and antitumor activity than the nontargeted PS-Dox and Lipo-Dox controls. In vivo biodistribution experiments displayed 2.5-fold higher tumor accumulation for GE11-PS-Dox as compared to Lipo-Dox. Notably, GE11-PS-Dox could effectively suppress the progression of SKOV3 tumors and cause little adverse effects at 12 mg of Dox equiv/kg, leading to a remarkably increased survival rate of 100% over 78 days. In contrast, continued tumor growth and body weight loss were discerned for Lipo-Dox treated mice at 6 mg of Dox equiv/kg. Moreover, a single dose of GE11-PS-Dox at 60 mg of Dox equiv/kg showed also effective treatment and low toxicity toward SKOV3-tumor bearing mice. GE11-directed reversibly cross-linked polymersomal doxorubicin has emerged as an advanced alternative to Lipo-Dox for treatment of EGFR-overexpressing ovarian cancers.

Screening for tumor suppressors: Loss of ephrin receptor A2 cooperates with oncogenic KRas in promoting lung adenocarcinoma.

Lung adenocarcinoma, a major form of non-small cell lung cancer, is the leading cause of cancer deaths. The Cancer Genome Atlas analysis of lung adenocarcinoma has identified a large number of previously unknown copy number alterations and mutations, requiring experimental validation before use in therapeutics. Here, we describe an shRNA-mediated high-throughput approach to test a set of genes for their ability to function as tumor suppressors in the background of mutant KRas and WT Tp53. We identified several candidate genes from tumors originated from lentiviral delivery of shRNAs along with Cre recombinase into lungs of Loxp-stop-Loxp-KRas mice. Ephrin receptorA2 (EphA2) is among the top candidate genes and was reconfirmed by two distinct shRNAs. By generating knockdown, inducible knockdown and knockout cell lines for loss of EphA2, we showed that negating its expression activates a transcriptional program for cell proliferation. Loss of EPHA2 releases feedback inhibition of KRAS, resulting in activation of ERK1/2 MAP kinase signaling, leading to enhanced cell proliferation. Intriguingly, loss of EPHA2 induces activation of GLI1 transcription factor and hedgehog signaling that further contributes to cell proliferation. Small molecules targeting MEK1/2 and Smoothened hamper proliferation in EphA2-deficient cells. Additionally, in EphA2 WT cells, activation of EPHA2 by its ligand, EFNA1, affects KRAS-RAF interaction, leading to inhibition of the RAS-RAF-MEK-ERK pathway and cell proliferation. Together, our studies have identified that (i) EphA2 acts as a KRas cooperative tumor suppressor by in vivo screen and (ii) reactivation of the EphA2 signal may serve as a potential therapeutic for KRas-induced human lung cancers.

ATN-161 Peptide Functionalized Reversibly Cross-Linked Polymersomes Mediate Targeted Doxorubicin Delivery into Melanoma-Bearing C57BL/6 Mice.

PHSCN peptide (licensed as ATN-161) is an effective α5ß1 integrin inhibitor that has advanced to phase II clinical trials to treat solid tumors. Here we developed ATN-161 functionalized self-cross-linkable and intracellularly de-cross-linkable polymersomes (ATN/SCID-Ps) for highly efficient and targeted delivery of doxorubicin hydrochloride (DOX·HCl) into B16F10 melanoma-bearing C57BL/6 mice. ATN/SCID-Ps exhibited a high loading capacity of DOX·HCl. The size of DOX-loaded ATN/SCID-Ps (DOX-ATN/SCID-Ps) decreased from 150 to 88 nm with increasing ATN surface densities from 0 to 100% (mol/mol). DOX-ATN/SCID-Ps were robust with low drug leakage under physiological conditions while quickly releasing DOX with the addition of 10 mM glutathione. MTT assay results displayed that DOX-ATN/SCID-Ps induced ATN density-dependent antitumor activity to α5ß1 integrin overexpressing B16F10 melanoma cells, in which 56% ATN-161 was optimal. Flow cytometry and CLSM studies revealed significantly more efficient internalization and cytoplasmic DOX release in B16F10 cells for DOX-ATN/SCID-Ps than for DOX-SCID-Ps (nontargeting control) as well as clinically used pegylated liposomal doxorubicin (DOX-LPs). DOX-ATN/SCID-Ps displayed a long blood circulation time (elimination half-life = 4.13 h) and 4 times higher DOX accumulation in B16F10 bearing C57BL/6 mice than DOX-LPs. Interestingly, DOX-ATN/SCID-Ps exhibited a superior maximum-tolerated dose of over 100 mg DOX·HCl/kg, 10 times higher than DOX-LPs. Remarkably, DOX-ATN/SCID-Ps could significantly inhibit the growth of aggressive B16F10 melanoma with little adverse effects via either multiple or single injection of total dosage of 100 mg DOX·HCl/kg, resulting in greatly improved survival rates as compared to DOX-LPs. ATN/SCID-Ps are appealing nanovehicles for targeted chemotherapy of α5ß1 integrin positive solid tumors.

IKK biology.

The inhibitor of nuclear factor-κB (IκB) kinase (IKK) complex is the master regulator of the NF-κB signaling pathway. The activation of the IKK complex is a tightly regulated, highly stimulus-specific, and target-specific event that is essential for the plethora of functions attributed to NF-κB. More recently, NF-κB-independent roles of IKK members have brought increased complexity to its biological function. This review highlights some of the major advances in the studies of the process of IKK activation and the biological roles of IKK family members, with a focus on NF-κB-independent functions. Understanding these complex processes is essential for targeting IKK for therapeutics.

Inflammatory tales of liver cancer.

Cell culture studies have established NF-kappaB's critical role in cancer cell survival and proliferation and led to the clinical use of NF-kappaB inhibitors. However, a paper in this issue of Cancer Cell reveals an anticancer function for NF-kappaB in a mouse model where NF-kappaB activity is lost specifically in hepatocytes. These studies suggest careful examination of NF-kappaB inhibitors as a therapeutic modality for cancer.

A Lung Cancer Mouse Model Database.

Lung cancer, the leading cause of cancer mortality, exhibits diverse histological subtypes and genetic complexities. Numerous preclinical mouse models have been developed to study lung cancer, but data from these models are disparate, siloed, and difficult to compare in a centralized fashion. Here we established the Lung Cancer Mouse Model Database (LCMMDB), an extensive repository of 1,354 samples from 77 transcriptomic datasets covering 974 samples from genetically engineered mouse models (GEMMs), 368 samples from carcinogen-induced models, and 12 samples from a spontaneous model. Meticulous curation and collaboration with data depositors have produced a robust and comprehensive database, enhancing the fidelity of the genetic landscape it depicts. The LCMMDB aligns 859 tumors from GEMMs with human lung cancer mutations, enabling comparative analysis and revealing a pressing need to broaden the diversity of genetic aberrations modeled in GEMMs. Accompanying this resource, we developed a web application that offers researchers intuitive tools for in-depth gene expression analysis. With standardized reprocessing of gene expression data, the LCMMDB serves as a powerful platform for cross-study comparison and lays the groundwork for future research, aiming to bridge the gap between mouse models and human lung cancer for improved translational relevance.

Transferrin-guided intelligent nanovesicles augment the targetability and potency of clinical PLK1 inhibitor to acute myeloid leukemia.

Acute myeloid leukemia (AML) remains a most lethal hematological malignancy, partly because of its slow development of targeted therapies compared with other cancers. PLK1 inhibitor, volasertib (Vol), is among the few molecular targeted drugs granted breakthrough therapy status for AML; however, its fast clearance and dose-limiting toxicity greatly restrain its clinical benefits. Here, we report that transferrin-guided polymersomes (TPs) markedly augment the targetability, potency and safety of Vol to AML. Vol-loaded TPs (TPVol) with 4% transferrin exhibited best cellular uptake, effective down-regulation of p-PLK1, p-PTEN and p-AKT and superior apoptotic activity to free Vol in MV-4-11 leukemic cells. Intravenous injection of TPVol gave 6-fold higher AUC than free Vol and notable accumulation in AML-residing bone marrow. The efficacy studies in orthotopic MV-4-11 leukemic model demonstrated that TPVol significantly reduced leukemic cell proportions in periphery blood, bone marrow, liver and spleen, effectively enhanced mouse survival rate, and impeded bone loss. This transferrin-guided nano-delivery of molecular targeted drugs appears to be an interesting strategy towards the development of novel treatments for AML.

Phosphorylation of p53 by IkappaB kinase 2 promotes its degradation by beta-TrCP.

Functional inactivation of p53 and constitutive activation of the NF-kappaB pathway has been associated with several human cancers. In this study, we show that IkappaB kinase 2 (IKK2/IKKbeta), which is critical for NF-kappaB activation, also phosphorylates p53. Phosphorylation of p53 at serines 362 and 366 by IKK2 leads to its recruitment to and ubiquitination by beta-TrCP1. Degradation of ubiquitinated p53 is independent of Mdm2, because it occurs in both wild-type and Mdm2(-/-) cells. SiRNA-mediated reduction in the levels of beta-TrCP1 and other members of the SCF(beta-TrCP1)E3 ubiquitin ligase complex or overexpression of a dominant negative form of beta-TrCP1 enhances p53 stability. Substitutions at Ser-362 and 366 of p53 by alanines (p53 AA) result in reduced phosphorylation of p53 by IKK2, decreased association with beta-TrCP1, and thus increased stability of p53 and expression of p53 target genes such as p21, altering the G1 phase of the cell cycle. Our results identify IKK2 and beta-TrCP1 as novel regulators of the p53 pathway and suggest that blocking of IKK2 and beta-TrCP1 could be a means of regulating p53 stability and thereby modulating its biological activity.

Polymersome-mediated cytosolic delivery of cyclic dinucleotide STING agonist enhances tumor immunotherapy.

Cyclic dinucleotides (CDNs) as stimulator of interferon genes (STING) agonists capable of inducing strong antitumor innate immune response are highly promising for tumor immunotherapy. The efficacy of these CDNs is, however, reduced greatly by their fast clearance, poor cell uptake and inefficient cytosolic transportation. Here, we report that reduction-responsive biodegradable chimaeric polymersomes (CPs) markedly enhance tumor retention and cytosolic delivery of a synthetic CDN, ADU-S100, and bolster STING pathway activation in the tumor microenvironment and tumor draining lymph nodes, giving significantly better tumor repression and survival of B16F10 melanoma-bearing mice compared with free CDN control. The superiority of CPs-mediated CDN delivery is further verified in combination therapy with low-dose fractionated radiation, which brings about clearly stronger and longer-term immunotherapeutic effects and protection against tumor re-challenge. The development of nano-STING agonists that are able to overcome the delivery barriers of CDNs represents an effective strategy to potentiate cancer immunotherapy.

Immunotherapy of Malignant Glioma by Noninvasive Administration of TLR9 Agonist CpG Nano-Immunoadjuvant.

Immunotherapy with toll like receptor 9 (TLR9) agonist CpG ODN offers an emergent strategy to treat life-threatening malignant glioma. CpG is typically applied invasively by intracranial and intrathecal administration which induces not only poor compliance and lessened potency but also possibly strong adverse effects and immunotoxicity. Here, it is reported that immunotherapy of murine LCPN glioma is greatly boosted by polymersome-steered intravenous and intranasal brain delivery of CpG. CpG is efficiently loaded in apolipoprotein E peptide-directed polymersomes to give blood-brain barrier permeable and glioma and cervical lymph node-homing CpG nano-immunoadjuvant (t-NanoCpG) which strongly stimulates the maturation of dendritic cells, antigen cross-presentation, and production of proinflammatory cytokines in vivo. Intriguingly, both intravenous and intranasal administration of t-NanoCpG brings about significant survival benefits in murine LCPN glioma-bearing mice while free CpG and nontargeted CpG nano-immunoadjuvant (NanoCpG) afford modest therapeutic effects. Moreover, combination of t-NanoCpG with radiotherapy further boosts the immunotherapeutic effects leading to more improved survival rate of mice. This intelligent brain-permeable nano-immunoadjuvant provides a new, minimally invasive and highly potent strategy for immunotherapy of glioma.

Systemic administration of polymersomal oncolytic peptide LTX-315 combining with CpG adjuvant and anti-PD-1 antibody boosts immunotherapy of melanoma.

Oncolytic peptide LTX-315 while showing clinical promise in treating solid tumors is limited to intratumoral administration, which is not applicable for inaccessible or metastatic tumors. The cationic and amphipathic nature of oncolytic peptides engenders formidable challenges to developing systems for their systemic delivery. Here, we describe cRGD-functionalized chimaeric polymersomes (cRGD-CPs) as a robust systemic delivery vehicle for LTX-315, which in combination with CpG adjuvant and anti-PD-1 boost immunotherapy of malignant B16F10 melanoma in mice. cRGD-CPs containing 14.9 wt% LTX-315 (cRGD-CPs-L) exhibited a size of 53 nm, excellent serum stability, and strong and selective killing of B16F10 cells (versus L929 fibroblasts) in vitro, which provoked similar immunogenic effects to free LTX-315 as revealed by release of danger-associated molecular pattern molecules. The systemic administration of cRGD-CPs-L gave a notable tumor accumulation of 4.8% ID/g and significant retardation of tumor growth. More interestingly, the treatment of B16F10 tumor-bearing mice was further boosted by co-administration of polymersomal CpG and anti-PD-1 antibody, in which two out of seven mice were cured as a result of strong immune response and long-term immune memory protection. The immunotherapeutic effect was evidenced by secretion of IL-6, IFN-γ and TNF-α, tumor infiltration of CD8+ CTLs and Th, and induction of TEM and TCM in spleen. This study opens a new avenue to oncolytic peptides, which enables durable immunotherapy of tumors via systemic administration.

Genetic and pharmacological interrogation of cancer vulnerability using a multiplexed cell line screening platform.

The multiplexed cancer cell line screening platform PRISM demonstrated its utility in testing hundreds of cell lines in a single run, possessing the potential to speed up anti-cancer drug discovery, validation and optimization. Here we described the development and implementation of a next-generation PRISM platform combining Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9-mediated gene editing, cell line DNA barcoding and next-generation sequencing to enable genetic and/or pharmacological assessment of target addiction in hundreds of cell lines simultaneously. Both compound and CRISPR-knockout PRISM screens well recapitulated the results from individual assays and showed high consistency with a public database.

A6 peptide-tagged, ultra-small and reduction-sensitive polymersomal vincristine sulfate as a smart and specific treatment for CD44+ acute myeloid leukemia.

Acute myeloid leukemia (AML) is a severe blood malignancy associated with a high relapse rate. The current clinical chemotherapy is typically perplexed with serious side effects. Here, A6 peptide-tagged, small and reduction-sensitive polymersomal vincristine sulfate (A6-cPS-VCR) is reported as a novel, smart and specific treatment for CD44 positive AML. A6-cPS-VCR stably loaded with 3.3 wt% VCR displays a size of ≈ 31 nm and pronounced selectivity toward CD44-overexpressed MV4-11 leukemia cells. Intriguingly, A6-cPS-VCR effectively represses the outgrowth of orthotopic MV4-11 AML in vivo, as revealed by significant reduction of leukemia burdens in the circulation, bone marrow, liver and spleen, and significantly extends the median survival time of MV4-11 AML-bearing mice. In addition to active targetability and therapeutic benefits, A6-cPS-VCR has the advantage of easy fabrication, rendering it potentially interesting for clinical translation.

α3ß1 Integrin-Targeting Polymersomal Docetaxel as an Advanced Nanotherapeutic for Nonsmall Cell Lung Cancer Treatment.

Docetaxel (DTX) widely used for treating nonsmall cell lung cancer (NSCLC) patients is associated with dose-limiting side effects, especially neurotoxicity and myelosuppression. Here, we have developed cyclic cNGQGEQc peptide-directed polymersomal docetaxel (cNGQ-PS-DTX) as a targeted and multifunctional formulation for NSCLC. cNGQ-PS-DTX carrying 8.1 wt % DTX had a size of 93 nm, neutral surface charge, high stability, and glutathione-triggered DTX release behavior. Cytotoxicity studies demonstrated a clearly better antitumor activity of cNGQ-PS-DTX in α3ß1 integrin overexpressing A549 human lung cancer cells than free DTX and nontargeted PS-DTX. cNGQ-PS-DTX showed a remarkably high tolerability (over 8 times better than free DTX) and slow elimination in mice. Importantly, cNGQ-PS-DTX exhibited greatly improved tumor accumulation and higher suppression of subcutaneous and orthotopic A549 xenografts as compared to PS-DTX and free DTX controls. α3ß1 integrin-targeting polymersomal docetaxel emerges as an advanced nanotherapeutic for NSCLC treatment.

GE11 peptide-installed chimaeric polymersomes tailor-made for high-efficiency EGFR-targeted protein therapy of orthotopic hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) remains a leading malignancy with a high mortality and little improvement in treatments. Protein drugs though known for their extraordinary potency and specificity have rarely been investigated for HCC therapy owing to lack of appropriate delivery systems. Here, we designed GE11 peptide-installed chimaeric polymersomes (GE11-CPs) for high-efficiency EGFR-targeted protein therapy of orthotopic SMMC-7721 HCC-bearing nude mice. GE11-CPs were assembled from poly(ethylene glycol)-b-poly(trimethylene carbonate-co-dithiolane trimethylene carbonate)-b-poly(aspartic acid) (PEG-P(TMC-DTC)-PAsp) and GE11-functionalized PEG-P(TMC-DTC), which allowed efficient loading and protection of proteins in the watery interior and fine-tuning of GE11 densities at the surface. CPs with short PAsp segments (degree of polymerization (DP) = 5, 10 and 15) exhibited a protein loading efficiency of 60%-72% and glutathione-responsive protein release. Saporin-loaded GE11-CPs had a size of 36 - 62 nm depending on GE11 densities and DP of PAsp. Notably, GE11-CPs with 10% GE11 revealed greatly enhanced uptake in SMMC-7721 cells, boosting the anticancer potency of saporin for over 3-folds compared with non-targeted control (half-maximal inhibitory concentration (IC50) = 11.0 versus 36.3 nM). The biodistribution studies using Cy5-labeled cytochrome C as a model protein demonstrated about 3-fold higher accumulation of GE11-CPs formulation than CPs counterpart in both subcutaneous and orthotopic SMMC-7721 tumor models. Notably, saporin-loaded GE11-CPs revealed low toxicity, effective tumor inhibition and significant improvement of survival rate compared with PBS and non-targeted groups (median survival time: 99 versus 37 and 42 days). EGFR-targeted chimaeric polymersomes carrying proteins appear an interesting HCC treatment modality.