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Advances in genome and tissue engineering have spurred significant progress and opportunity for innovation in cancer modeling. Human induced pluripotent stem cells (iPSCs) are an established and powerful tool to study cellular processes in the context of disease-specific genetic backgrounds; however, their application to cancer has been limited by the resistance of many transformed cells to undergo successful reprogramming. Here, we review the status of human iPSC modeling of solid tumors in the context of genetic engineering, including how base and prime editing can be incorporated into "bottom-up" cancer modeling, a term we coined for iPSC-based cancer models using genetic engineering to induce transformation. This approach circumvents the need to reprogram cancer cells while allowing for dissection of the genetic mechanisms underlying transformation, progression, and metastasis with a high degree of precision and control. We also discuss the strengths and limitations of respective engineering approaches and outline experimental considerations for establishing future models.

Significant challenges have gone into the design of smart hydrogels, with numerous potential applications in the industrial, cosmetic, and biomedical fields. Herein, we report the synthesis of novel 4-arm self-assembling peptide-polyethylene glycol (PEG) hybrid star-shaped polymers and their comprehensive hydrogel properties. β-sheet-forming oligopeptides with alternating hydrophobic Leu/ionizable Glu repeats and Cys residues were successfully conjugated to 4-arm PEG

In 2021, the World Health Organization reclassified glioblastoma, the most common form of adult brain cancer, into isocitrate dehydrogenase (IDH)-wild-type glioblastomas and grade IV IDH mutant (G4 IDHm) astrocytomas. For both tumor types, intratumoral heterogeneity is a key contributor to therapeutic failure. To better define this heterogeneity, genome-wide chromatin accessibility and transcription profiles of clinical samples of glioblastomas and G4 IDHm astrocytomas were analyzed at single-cell resolution. These profiles afforded resolution of intratumoral genetic heterogeneity, including delineation of cell-to-cell variations in distinct cell states, focal gene amplifications, as well as extrachromosomal circular DNAs. Despite differences in IDH mutation status and significant intratumoral heterogeneity, the profiled tumor cells shared a common chromatin structure defined by open regions enriched for nuclear factor 1 transcription factors (NFIA and NFIB). Silencing of NFIA or NFIB suppressed in vitro and in vivo growths of patient-derived glioblastomas and G4 IDHm astrocytoma models. These findings suggest that despite distinct genotypes and cell states, glioblastoma/G4 astrocytoma cells share dependency on core transcriptional programs, yielding an attractive platform for addressing therapeutic challenges associated with intratumoral heterogeneity.

Diffuse midline glioma (DMG) is a leading cause of brain tumor death in children. In addition to hallmark H3.3K27M mutations, significant subsets also harbor alterations of other genes, such asTP53andPDGFRA. Despite the prevalence of H3.3K27M, the results of clinical trials in DMG have been mixed, possibly due to the lack of models recapitulating its genetic heterogeneity. To address this gap, we developed human iPSC-derived tumor models harboring TP53R248Qwith or without heterozygous H3.3K27M and/or PDGFRAD842Voverexpression. The combination of H3.3K27M and PDGFRAD842Vresulted in more proliferative tumors when gene-edited neural progenitor (NP) cells were implanted into mouse brains compared to NP with either mutation alone. Transcriptomic comparison of tumors and their NP cells of origin identified conserved JAK/STAT pathway activation across genotypes as characteristic of malignant transformation. Conversely, integrated genome-wide epigenomic and transcriptomic analyses, as well as rational pharmacologic inhibition, revealed targetable vulnerabilities unique to the TP53R248Q; H3.3K27M; PDGFRAD842Vtumors and related to their aggressive growth phenotype. These includeAREG-mediated cell cycle control, altered metabolism, and vulnerability to combination ONC201/trametinib treatment. Taken together, these data suggest that cooperation between H3.3K27M and PDGFRA influences tumor biology, underscoring the need for better molecular stratification in DMG clinical trials.

Synthesis of amino acid-derived vinyl polymers with precisely controlled hydropathy by ultra-rapid reversible addition fragmentation chain transfer polymerization and their thermoresponsive behavior in water.

Background Heterozygous TERT (telomerase reverse transcriptase) promoter mutations (TPMs) facilitate TERT expression and are the most frequent mutation in glioblastoma (GBM). A recent analysis revealed this mutation is one of the earliest events in gliomagenesis. However, no appropriate human models have been engineered to study the role of this mutation in the initiation of these tumors. Method We established GBM models by introducing the heterozygous TPM in human induced pluripotent stem cells (hiPSCs) using a two-step targeting approach in the context of GBM genetic alterations, CDKN2A/B and PTEN deletion, and EGFRvIII overexpression. The impact of the mutation was evaluated through the in vivo passage and in vitro experiment and analysis. Results Orthotopic injection of neuronal precursor cells (NPCs) derived from hiPSCs with the TPM into immunodeficient mice did not enhance tumorigenesis compared to TERT promoter wild type NPCs at initial in vivo passage presumably due to relatively long telomeres. However, the mutation recruited GA-Binding Protein and engendered low-level TERT expression resulting in enhanced tumorigenesis and maintenance of short telomeres upon secondary passage as observed in human GBM. These results provide the first insights regarding increased tumorigenesis upon introducing a TPM compared to isogenic controls without TPMs. Conclusion Our novel GBM models presented the growth advantage of heterozygous TPMs for the first time in the context of GBM driver mutations relative to isogenic controls, thereby allowing for the identification and validation of TERT promoter-specific vulnerabilities in a genetically accurate background.

スチレン(St)とアクリル酸アラニンメチルエステル(NAAMe)のATRP重合により、２種類のブロック鎖長をもつ目的のジブロックポリマー(PSt-b-PNAAMe(m, n))を合成した。SORP法により微粒子化に成功し、(m, n)=(400, 25)は(220, 55)に比べて分布の狭いナノ粒子を与えた。このナノ粒子は、温度に応答して約60nm程度の可逆的な粒径変化を示した。, By polymerization of styrene and N-acryloyl alanine methyl ester (NAAMe) through atom transfer radical polymerization (ATRP), the desired block polymers, polystyrene (PSt)-b-PNAAMe (m, n), were successfully obtained. Block lengths of PSt-b-PNAAMe (m, n) were evaluated to be m=400, n=25 and m=220, n=55, and polydispersity indices were controlled within 1.2. Polymer nanoparticles were prepared through SORP method. PSt-b-PNAAMe (m, n=400, 25) spontaneously formed nanoparticles with relatively narrow size distribution but PSt-b-PNAAMe (m, n =220, 55) did not probably due to the difference in their amphiphilicity. The particle size of PST-b-PNAAMe was varied reversibly with the surrounding temperature below and above the LCST of the PNAAMe block segment.

Heterozygous TERT (Telomerase reverse transcriptase) promoter mutations (TPMs) facilitate TERT expression and are the most frequent mutation in glioblastoma (GBM). A recent analysis revealed this mutation is one of the earliest events in gliomagenesis, however no appropriate human models have been engineered to study the role of this mutation in the initiation of these tumors. To address this, we established GBM models by introducing the heterozygous TPM in human induced pluripotent stem cells (hiPSCs) using a two-step targeting approach in the context of GBM genetic alterations, CDKN2A/B and PTEN deletion, and EGFRvIII overexpression. Orthotopic injection of neuronal precursor cells (NPCs) derived from hiPSCs with TPM into immunodeficient mice did not enhance tumorigenesis compared to TERT promoter wild type (TPW) NPCs at initial in vivo passage which we attribute to relatively long telomeres. We further show that the TPM mutation recruited GA-Binding Protein (GABPA) and engendered low-level TERT expression, resulting in enhanced tumorigenesis upon secondary passage and maintenance of short telomere length as has been reported in human GBM. RNA sequencing of harvested tumors grown as secondary spheres demonstrated upregulated proliferation and mitosis pathway signatures in TPM cells, consistent with their increased in vitro proliferation relative to TPW counterparts. Finally, when secondary TPM and TPW sphere cultures were reinjected into mice, only the TPM led to tumor formation. In summary, our novel GBM model illustrates a growth advantage imparted by heterozygous TPMs in the context of GBM driver mutations relative to isogenic controls, and thereby allows for the identification and validation of TERT promoter-specific vulnerabilities in a genetically accurate background.

Background The goal was to determine whether the addition of temozolomide (TMZ) to the standard treatment of high-dose methotrexate (HD-MTX) and whole-brain radiotherapy (WBRT) for primary central nervous system lymphoma (PCNSL) improves survival. Methods An open-label, randomized, phase III trial was conducted in Japan, enrolling immunocompetent patients aged 20–70 years with histologically confirmed, newly diagnosed PCNSL. After administration of HD-MTX, patients were randomly assigned to receive WBRT (30 Gy) ± 10 Gy boost (arm A) or WBRT ± boost with concomitant and maintenance TMZ for 2 years (arm B). The primary endpoint was overall survival (OS). Results Between September 29, 2014 and October 15, 2018, 134 patients were enrolled, of whom 122 were randomly assigned and analyzed. At the planned interim analysis, 2-year OS was 86.8% (95% confidence interval [CI]: 72.5–94.0%) in arm A and 71.4% (56.0–82.2%) in arm B. The hazard ratio was 2.18 (95% CI: 0.95–4.98), with the predicted probability of showing the superiority of arm B at the final analysis estimated to be 1.3%. The study was terminated early due to futility. O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation status was measured in 115 tumors, and it was neither prognostic nor predictive of TMZ response. Conclusions This study failed to demonstrate the benefit of concomitant and maintenance TMZ in newly diagnosed PCNSL.

Among children and adolescents in the United States (0 to 19 years old), brain and other central nervous system tumors are the second most common types of cancers, surpassed in incidence only by leukemias. Despite significant progress in the diagnosis and treatment modalities, brain cancer remains the leading cause of death in the pediatric population. There is an obvious unfulfilled need to streamline the therapeutic strategies and improve survival for these patients. For that purpose, preclinical models play a pivotal role. Numerous models are currently used in pediatric brain tumor research, including genetically engineered mouse models, patient-derived xenografts and cell lines, and newer models that utilize novel technologies such as genome engineering and organoids. Furthermore, extensive studies by the Children's Brain Tumor Network (CBTN) researchers and others have revealed multiomic landscapes of variable pediatric brain tumors. Combined with such integrative data, these novel technologies have enabled numerous applicable models. Genome engineering, including CRISPR/Cas9, expanded the flexibility of modeling. Models generated through genome engineering enabled studying particular genetic alterations in clean isogenic backgrounds, facilitating the dissection of functional mechanisms of those mutations in tumor biology. Organoids have been applied to study tumor-to-tumor-microenvironment interactions and to address developmental aspects of tumorigenesis, which is essential in some pediatric brain tumors. Other modalities, such as humanized mouse models, could potentially be applied to pediatric brain tumors. In addition to current valuable models, such novel models are anticipated to expedite functional tumor biology study and establish effective therapeutics for pediatric brain tumors.

The Spalt-like 4 transcription factor (SALL4) plays an essential role in controlling the pluripotent property of embryonic stem cells via binding to AT-rich regions of genomic DNA, but structural details on this binding interaction have not been fully characterized. Here, we present crystal structures of the zinc finger cluster 4 (ZFC4) domain of SALL4 (SALL4

Spalt-like 4 (SALL4) plays an essential role in controlling the pluripotent property of embryonic stem cells (ESCs) via binding to AT-rich regions of genomic DNA. Here we present crystal structures of the the zinc finger cluster 4 (ZFC4) domain of SALL4 (SALL4ZFC4) bound with different double stranded DNAs containing a conserved AT-rich motif. In the structures, two zinc fingers of SALL4ZFC4 coordinatively recognize an AATA tetranucleotide. We also solved the DNA-bound structures of SALL3ZFC4 and SALL4ZFC1. These structures illuminate a common recognition mode for AT-rich DNA by the SALL family proteins. The DNA binding activity is essential for SALL4 function as DNA-binding defective mutants of mouse Sall4 failed to repress aberrant gene expression in Sall4-/- mESCs. Thus, these analyses provide new insights into the mechanisms of action underlying SALL family in controlling cell fate via preferentially targeted to AT-rich sites within genomic DNAs during cell differentiation.

Natural biopolymers exhibit smart functions and extraordinary physical properties because of their optimized and rational molecular structures. Understanding and mimicking such architectural principles on a nanoscale offer promising opportunities for designing novel functional polymer materials. Herein, we report unique nano/microfilm materials fabricated using spider silk-inspired peptide multiblock hybrid copolymers. The multiblock copolymers are composed of self-assembling oligopeptides (Ala, Gly, Val, and Leu-based peptides) and non-crystalline flexible polypropylene glycol. Self-supporting microfilms are easily obtained by casting from polymer solutions via β-sheet network formation, and their structural and mechanical properties are characterized comprehensively. The resulting spider silk-like specific nanostructure comprises balanced crystalline and non-crystalline regions; additionally, it exhibits reversible interactions among peptide blocks based on multiple hydrogen bonds. This enables mechanical toughness and self-healing characteristics in the thin films. Moreover, such film characteristics, including healing efficiency, can be modulated by manipulating the sequence and length of the peptide blocks. Additionally, these hybrid multiblock copolymers are used to prepare free-standing and ultraflexible nanofilms, which demonstrate excellent followability to nanoscale roughness based on film thickness and serve as a nano-coating with ion barrier function. The biomimetic approach presented herein is a facile and effective method for the development of high-performance polymer films and is promising in various fields, such as industrial, nanotechnological, and biomedical material fields.

In this paper, we report the synthesis and characterization of novel peptide based hybrid graft copolymers that can form injectable hydrogels. These hybrid graft copolymers are composed of a biocompatible poly(2-methacryloyloxyethyl phosphorylcholine) main chain and self-assembling oligoalanine-block-poly(ethylene glycol) grafts (grafting ratio (m) = 6, 10, 20, 28, 32, and 40%), which are readily synthesized using a macromonomer by combining the solid phase peptide synthesis and conventional radical polymerization methods. Conformational analyses under a wide range of dilute aqueous solutions (pH 3–10 and 4–70 °C) revealed stable β-sheet formation. With an increase in solution concentration, the graft copolymers, except those with a low grafting ratio of m = 6, formed hydrogels displaying shear-thinning and self-healing behaviors because of the reversible self-assembly property of peptide grafts. Interestingly, the mechanical properties of hybrid graft copolymers were strongly dependent on the grafting density of the hybrid copolymers. Thus, it was observed that an increase in grafting ratio strengthened the inter-polymer network. These features enabled the graft-type hybrids to be used as injectable hydrogels for 3D cell scaffolds and controlled-release applications.

Background There are ongoing clinical trials exploring the efficacy of dopamine receptor D2 (DRD2) inhibition against glioblastomas, the most common primary brain tumor. Here we examine potential molecular determinants of this efficacy. Methods The Cancer Genome Atlas glioblastoma database and other published mRNA profiles were used to analyze the DRD2 and epidermal growth factor receptor (EGFR) expression pattern. In vitro and in vivo responses to DRD2 inhibitors were determined using patient-derived xenograft (PDX) glioblastoma models. Immunohistochemical studies were performed on clinically annotated glioblastoma samples derived from patients treated with ONC201. Results Analysis of clinical glioblastoma specimens derived from independent patient cohorts revealed an inverse correlation between EGFR and DRD2 mRNA expression, with implication that signaling mediated by these proteins shares overlapping functions. In independent panels of PDX glioblastoma lines, high EGFR expression was associated with poor in vitro and in vivo response to DRD2 inhibitors, including haloperidol and ONC201. Moreover, ectopic expression of a constitutively active EGFR, variant (v)III, suppressed glioblastoma sensitivity to ONC201. DRD2 expression positively correlated with expression of rate-limiting enzymes for dopamine synthesis as well as dopamine secretion, suggesting contribution of autocrine DRD2 signaling. Analysis of specimens from patients treated with ONC201 (n = 15) showed an inverse correlation between the intensity of EGFR staining and clinical response. The median overall survival for patients with high and low EGFR staining was 162 and 373 days, respectively (0.037). Conclusions High EGFR expression is a determinant of poor glioblastoma response to DRD2. This finding should inform future clinical trial designs.

A wind-and-react MgB 2 solenoid magnet for klystrons has been developed. While the current normal-conducting (Cu) magnet consumes 20 kW per magnet, this MgB$_2$ magnet consumes less than 3 kW in refrigerator power. The conduction-cooled half coil of the magnet is 337 mm in inner diameter; the winding pack, 19.4 mm wide × 136.6 mm high, uses 2.7 km of 10 filament circular conductor, which is insulated with glass 0.83 mm in diameter, and is reacted after being wound onto a stainless steel bobbin. The coil has Cu plates of 0.2 mm in thickness between each coil layer and on the inner and outer sides. The magnet has two coils and produces 0.8 T in the center and its stored energy is 11.8 kJ. Together with the above-mentioned coil structure, these coils can consume stored energy in itself at quench without a special quench protection system. A performance test of the magnet was successful.

A novel water-soluble amino acid derived vinyl polymer whose block sequence was designed to achieve a gradient thermoresponsiveness along a chain was accurately prepared through an ultrarapid reversible addition-fragmentation chain-transfer polymerization. The polymer exhibited unique temperature-regulated self-assembly in water, leading to multiple nanostructural transformations including disassembly-to-ordered and ordered-to-ordered transitions. The morphologies were drastically changed by heating the solution from 4 °C (soluble form) to 20 °C (spherical micelle) to 70 °C (vesicle). Moreover, such transitions exhibited hysteresis upon cooling, namely, from 70 °C (vesicle) to 20 °C (wormlike micelle) to 4 °C (soluble form). In this polymer system, the specific monomer sequence contributed to the self-assembly behavior. These findings provide significant insight into the design of new thermoresponsive nanomaterials with potential applications in biomedical chemistry.

Collagen is an essential structural protein in animal tissues and plays key roles in cellular modulation. We investigated methods to discover collagen model peptides (CMPs) that would self-assemble into triple helices and then grow into supramolecular organizations with diverse morphological features, which would be valuable as biomaterials. This challenging undertaking was achieved by placing azobenzene groups on the ends of the CMPs, (GPO)n (n = 3–10), Azo-(GPO)n. In a dilute aqueous solution (80 μM), CD spectra indicated that the Azo-(GPO)n (n > 4) formed triple helices due to strong hydrophobic azobenzene interactions, and that helix stability was increased with the peptide segment length. The resulting triple helices induced a specific azobenzene orientation through turned and twisted configurations as shown by CD spectra. TEM observations for the same solutions disclosed the morphologies for the Azo-CMPs. Azo-(GPO)3, having the shortest peptide segment, showed no nanostructure, both Azo-(GPO)4 and Azo-(GPO)5 provided consistent well-developed nanofiber structures resembling the natural collagen fibers, and Azo-(GPO)ns (n = 6–10) grew into flexible rod-like micelle fibers. In addition, alkyl chain-attached CmAzo-(GPO)5 displayed a toroidal morphology, and Azp-deg-(GPO)5 having a hydrophilic spacer assembled into a bilayer vesicle structure. These diverse morphological features are considered to be due to the characteristics of the pre-organized triple helix units. Photo-isomerization of the azobenzene moiety brought about the disappearance of such characteristic nano-architectures. When the solution concentration was increased up to 1 wt%, only Azo-(GPO)4 and Azo-(GPO)5 spontaneously formed hydrogels exhibiting a satisfactory gel-to-sol transition upon UV irradiation.

Herein, we describe the hierarchical self-assembly accompanying self-sorting of collagen-inspired peptides (CPs). The two amphiphilic CPs used in this study contained an azobenzene (Az) moiety at the

Histone H3 K27M mutation is a hallmark mutation for H3 K27M-mutant diffuse midline glioma (DMG), but targeting this mutation has yet to achieve a significant survival benefit in clinical trials. Recent analyses revealed alterations in several genes, such as NF1 and PDGFRA, are observed in substantial subpopulations of H3 K27M-mutant DMG patients in addition to H3 mutation and recurrent TP53 mutations, indicating patient-to-patient tumor heterogeneity and the potential necessity of tailored target therapy for the treatment of this disease. Here, using our human induced pluripotent stem cells (iPSC)-derived glioma avatar platform, we designed DMG models by introducing TP53R248Q with or without heterozygous H3 K27M mutation in combination with further genetic modifications of either NF1 knockout or PDGFRAD842V overexpression to recapitulate DMG subpopulations. Mice with TP53R248Q; H3F3AK27M (QM) tumors survived significantly longer than those harboring QM;NF1-/- (QMN) tumors and QM; PDGFRAD842Voe (QMP) tumors. RNA-sequencing of those induced DMG (iDMG) neurospheres revealed altered patterns of upregulation of MAPK pathway genes both in QMN and QMP-iDMG neurospheres compared to their H3 wildtype counterparts with the same combinations of genetic alterations, suggesting that those additional mutations modifies the oncogenic signaling associated with H3 K27M mutation. Further, differential expression analysis comparing QMN and QMP-iDMG neurospheres revealed 405 differentially expressed genes. Gene set enrichment analysis showed upregulation of transcriptional programs related to mesenchymal signature in QMN-iDMG neurospheres and proneural signature in QMP-iDMG neurosphere as expected. These data show that NF1 deletion and PDGFRAD842V overexpression significantly alter gene expression in H3 K27M-mutant iDMG tumors, potentially opening up a new therapeutic avenue in these devastating tumors with patient-to-patient heterogeneity. Further work using these models will shed light on the development of tailored therapy based on detailed genetic information on each patient sample, such as combining targeted kinase inhibition with HDAC inhibitors that have shown promise in the clinic.

INTRODUCTION Lactate dehydrogenase A (LDHA) encodes an enzyme that catalyzes the inter-conversion between pyruvate and lactate in glycolysis. Here, we demonstrate that LDHA mediates a novel role in DNA repair independent of this metabolic function. METHODS siRNA screen, The Cancer Genome Atlas (TCGA) survival analysis, ionizing radiation (IR), g-H2AX, and chromatin assays, site-directed mutagenesis. RESULTS In an orthogonal siRNA-informatic screen to identify genes 1) when silenced caused IR sensitivity in patient-derived glioblastoma lines and 2) lowered expression is associated with improved survival in TCGA, LDHA surfaced as the top candidate. The survival association was validated by LDHA immunohistochemical staining in an independent collection of glioblastoma samples. In vitro and in vivo, silencing of LDHA sensitized glioblastoma lines to IR and enhanced radiation-induced g-H2AX accumulation. Such sensitization was not observed after treatment with an LDHA inhibitor, suggesting the metabolic function of LDHA is distinct from its role in DNA repair. Supporting this hypothesis, truncation mutations that suppressed the LDHA glycolysis function minimally affected its role in DNA repair. Mechanistically, cytoplasmic LDHA translocates into the nucleus in response to IR. This translocation was associated with subsequent chromatin transition into an open conformation and enhanced homologous recombination. CONCLUSION The novel LDHA function in DNA repair suggests intricate crosstalks between glycolytic metabolism and DNA repair, offering a new platform for glioblastoma therapeutic development.

INTRODUCTION In 2021, the World Health Organization (WHO) reclassified glioblastoma, the most common form of adult brain cancer, into isocitrate dehydrogenase (IDH) wild-type glioblastomas and grade IV IDH mutant (G4 IDHm) astrocytomas. For both tumor types, intra-tumoral heterogeneity is a key contributor to therapeutic failure. METHODS we applied integrated genome-wide chromatin accessibility (snATACseq) and transcription (snRNAseq) profiles to clinical specimens derived IDHwt glioblastomas and G4 IDHm) astrocytomas, with goal of therapeutic target discovery. RESULTS The integrated analysis achieved resolution of intra-tumoral heterogeneity not previously possible, providing a molecular landscape of extensive regional and cellular variability. snATACseq delineated focal amplification down to an ~40 KB resolution. The snRNA analysis elucidated distinct cell types and cell states (neural progenitor/oligodendrocyte cell-like or astrocyte/mesenchymal cell-like) that were superimposable onto the snATACseq landscape. Paired-seq (parallel snATACseq and snRNAseq using the same clinical sample) provided high resolution delineation of extrachromosomal circular DNA (ecDNA), harboring oncogenes including CCND1 and EGFR. Importantly, the copy number of ecDNA genes correlated closely with the level of RNA expression. Integrated analysis across all specimens profiled suggests that IDHm grade 4 astrocytoma and IDHwt glioblastoma cells shared a common chromatin structure defined by open regions enriched for Nuclear Factor 1 transcription factors (NFIA and NFIB). Silencing of NF1A or NF1B suppressed in vitro and in vivo growth of patient-derived IDHwt glioblastomas and G4 IDHm astrocytoma models that mimic distinct glioblastoma cell states. CONCLUSION Our findings suggest despite distinct genotypes and cell states, glablastoma/G4 astrocytoma cells share dependency on core transcriptional programs, yielding an attractive platform for addressing therapeutic challenges associated with intra-tumoral heterogeneity.

There has been a resurgence of interest in brachytherapy as a treatment for glioblastoma, with several currently ongoing clinical trials. To provide a foundation for the analysis of these trials, we analyze the Surveillance, Epidemiology, and End Results (SEER) database to determine whether receipt of brachytherapy conveys a survival benefit independent of traditional prognostic factors. We identified 60,456 glioblastoma patients, of whom 362 underwent brachytherapy. We grouped patients based on receipt of brachytherapy and compared clinical and demographic variables between groups using Student’s t-test and Pearson’s chi-squared test. We assessed survival using Kaplan–Meier curves and Cox proportional hazards models. Median overall survival was 16 months in patients who received brachytherapy compared to 9 months in those who did not (log-rank p

Controlling the surface properties of engineered materials to enhance or reduce their cellular affinities remains a significant challenge in the field of biomaterials. We describe a universal technique for modulating the cytocompatibilities of two-dimensional (2D) and three-dimensional (3D) materials using a novel photocleavable peptide-grafted poly(2-hydroxyethyl methacrylate) (PHEMA) hybrid. The reversible addition-fragmentation chain transfer copolymerization of HEMA and propargyl acrylate was successfully controlled. The resultant alkyne-containing PHEMA was then used to modify the azide-terminated oligopeptides [Arg-Gly-Asp-Ser (RGDS)] with a photolabile 3-amino-3-(2-nitrophenyl)propanoic acid moiety via the copper-catalyzed alkyne-azide click chemistry. This strategy was readily used to decorate the surfaces of both hydrophilic and hydrophobic materials with RGDS peptides due to the high film-forming abilities of the PHEMA unit. The resultant thin film acted as an effective scaffold for improving cell adhesion and growth of NIH/3T3 fibroblasts and MC3T3-E1 osteoblast-like cells in vitro. In addition, UV irradiation of the surface led to the detachment of cells from the material surface accompanied by the photocleavage of RGDS grafts and enabled the 2D-patterning of cells and cell sheet engineering. The applicability of this system to 3D materials was investigated, and the cell adhesion was remarkably enhanced on a 3D-printed poly(lactic acid) object. This facile, biocompatible, and photoprocessable peptide-vinyl polymer hybrid system is valuable for its ability to advance the fields of tissue engineering, cell chips, and regenerative medicine.

Precision oncology hinges on linking tumour genotype with molecularly targeted drugs1; however, targeting the frequently dysregulated metabolic landscape of cancer has proven to be a major challenge2. Here we show that tissue context is the major determinant of dependence on the nicotinamide adenine dinucleotide (NAD) metabolic pathway in cancer. By analysing more than 7,000 tumours and 2,600 matched normal samples of 19 tissue types, coupled with mathematical modelling and extensive in vitro and in vivo analyses, we identify a simple and actionable set of 'rules'. If the rate-limiting enzyme of de novo NAD synthesis, NAPRT, is highly expressed in a normal tissue type, cancers that arise from that tissue will have a high frequency of NAPRT amplification and be completely and irreversibly dependent on NAPRT for survival. By contrast, tumours that arise from normal tissues that do not express NAPRT highly are entirely dependent on the NAD salvage pathway for survival. We identify the previously unknown enhancer that underlies this dependence. Amplification of NAPRT is shown to generate a pharmacologically actionable tumour cell dependence for survival. Dependence on another rate-limiting enzyme of the NAD synthesis pathway, NAMPT, as a result of enhancer remodelling is subject to resistance by NMRK1-dependent synthesis of NAD. These results identify a central role for tissue context in determining the choice of NAD biosynthetic pathway, explain the failure of NAMPT inhibitors, and pave the way for more effective treatments.

Significant efforts have been dedicated toward designing conjugated sequence-controlled peptides and synthetic vinyl polymers as a new class of polymeric materials with ordered structures and specific biofunctions. The creation of novel synthetic strategies that enables the precise incorporation of multiple tailored peptides along a vinyl polymer backbone remains highly challenging. Herein, we report a useful method for preparing multiblock architectures composed of alternately aligned sequential peptides and various vinyl polymers through nitroxide-mediated polymerization (NMP). A new cyclic oligopeptide was developed containing a 2,2,5-trimethyl-4-phenyl-3-azahexane-3-nitroxide (TIPNO)-derived alkoxyamine bond in the framework. Controlled radical polymerization in a homogeneous liquid phase using this peptide initiator successfully provided a well-defined multiblock hybrid polymer in one step, in which chain extension and multimerization proceeded simultaneously. This method was applied to the polymerizations of a wide range of vinyl monomers, including styrene, acrylamides, acrylates and acrylonitrile. Expanding the compatibility of monomers with this approach is valuable for constructing complex multiblock architectures with structural and functional diversity.

A novel multiblock architecture composed of an alternating β-sheet forming oligopeptide and a thermo-responsive glycine-derived vinyl polymer was synthesized. The polymer exhibited lower critical solution temperature (LCST) behavior in water, unlike the behavior of a glycine-derived homopolymer, and formed nanoparticles through protein-mimicking folding driven by thermal cycle-induced β-sheet formation.

Diffuse intrinsic pontine glioma (DIPG) is a subset of high-grade glioma that occurs predominantly in children and has no cure. Up to 80% of DIPG harbor a heterozygous point mutation that results in a lysine 27 to methionine substitution in histone variant H3.3 (H3.3K27M). However, H3.3K27M alone is insufficient for tumorigenesis in existing DIPG models, suggesting that it interacts with co-occurring oncogenic mutations. NF1 deletion co-occurs with TP53 mutation and H3.3K27M in DIPG, but the impact of these mutations individually and together are understudied. To address this gap, we designed a model of DIPG based on human induced pluripotent stem cells (iPSC) edited via CRISPR to express either wild-type or deleted NF1 in conjunction with heterozygous H3.3K27M and a loss of function mutation in TP53. Edited iPSC were chemically differentiated into neural progenitor cells (iNPC), which upon implantation into the brainstems of immunodeficient mice formed diffusely invasive tumors that were histologically consistent with high-grade glioma. Mice with TP53 mut;H3.3K27M tumors survived significantly (p Citation Format: Kasey Skinner, Tomoyuki Koga, Shunichiro Miki, Robert F. Gruener, R. Stephanie Huang, Frank Furnari, Ryan Miller. NF1 deletion potentiates tumorigenesis and activates expression of cancer-related kinases in an iPSC-based model of H3.3K27M diffuse intrinsic pontine glioma [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 LB067.

Precision medicine in oncology leverages clinical observations of exceptional response. Toward an understanding of the molecular features that define this response, we applied an integrated, multiplatform analysis of RNA profiles derived from clinically annotated glioblastoma samples. This analysis suggested that specimens from exceptional responders are characterized by decreased accumulation of microglia/macrophages in the glioblastoma microenvironment. Glioblastoma-associated microglia/macrophages secreted interleukin 11 (IL11) to activate STAT3-MYC signaling in glioblastoma cells. This signaling induced stem cell states that confer enhanced tumorigenicity and resistance to the standard-of-care chemotherapy, temozolomide (TMZ). Targeting a myeloid cell restricted an isoform of phosphoinositide-3-kinase, phosphoinositide-3-kinase gamma isoform (PI3Kγ), by pharmacologic inhibition or genetic inactivation disrupted this signaling axis by reducing microglia/macrophage-associated IL11 secretion in the tumor microenvironment. Mirroring the clinical outcomes of exceptional responders, PI3Kγ inhibition synergistically enhanced the anti-neoplastic effects of TMZ in orthotopic murine glioblastoma models. Moreover, inhibition or genetic inactivation of PI3Kγ in murine glioblastoma models recapitulated expression profiles observed in clinical specimens isolated from exceptional responders. Our results suggest key contributions from tumor-associated microglia/macrophages in exceptional responses and highlight the translational potential for PI3Kγ inhibition as a glioblastoma therapy.

The highly lethal brain cancer glioblastoma (GBM) poses a daunting challenge because the blood-brain barrier renders potentially druggable amplified or mutated oncoproteins relatively inaccessible. Here, we identify SMPD1, an enzyme that regulates the conversion of sphingomyelin to ceramide and a critical regulator of plasma membrane structure and organization, as an actionable drug target in glioblastoma. We show that the safe and highly brain-penetrant antidepressant fluoxetine, potently inhibits SMPD1 activity, killing GBMs, in vitro and in patient-derived xenografts, through inhibition of EGFR signaling and via activation of lysosomal stress. Combining fluoxetine with the chemotherapeutic agent temozolomide, a standard of care for GBM patients, causes massive increases in GBM cell death, and complete and long-lived tumor regression in mice. Incorporation of real-world evidence from electronic medical records from insurance databases, reveals significantly increased survival in glioblastoma patients treated with fluoxetine, which was not seen in patients treated with other SSRI anti-depressants. These results nominate the repurposing of fluoxetine as a potentially safe and promising therapy for GBM patients and suggest prospective randomized clinical trials.

Summary The highly lethal brain cancer glioblastoma (GBM) poses a daunting challenge because the blood-brain barrier renders potentially druggable amplified or mutated oncoproteins relatively inaccessible. Here, we identify sphingomyelin phosphodiesterase 1 (SMPD1), an enzyme that regulates the conversion of sphingomyelin to ceramide, as an actionable drug target in GBM. We show that the highly brain-penetrant antidepressant fluoxetine potently inhibits SMPD1 activity, killing GBMs, through inhibition of epidermal growth factor receptor (EGFR) signaling and via activation of lysosomal stress. Combining fluoxetine with temozolomide, a standard of care for GBM, causes massive increases in GBM cell death and complete tumor regression in mice. Incorporation of real-world evidence from electronic medical records from insurance databases reveals significantly increased survival in GBM patients treated with fluoxetine, which was not seen in patients treated with other selective serotonin reuptake inhibitor (SSRI) antidepressants. These results nominate the repurposing of fluoxetine as a potentially safe and promising therapy for patients with GBM and suggest prospective randomized clinical trials.