11 research outputs found
Biodegradable PEG-poly(ω-pentadecalactone- co - p -dioxanone) nanoparticles for enhanced and sustained drug delivery to treat brain tumors
Intracranial delivery of therapeutic agents is limited by penetration beyond the blood-brain barrier (BBB)
and rapid metabolism of the drugs that are delivered. Convection-enhanced delivery (CED) of drugloaded
nanoparticles (NPs) provides for local administration, control of distribution, and sustained
drug release. While some investigators have shown that repeated CED procedures are possible, longer
periods of sustained release could eliminate the need for repeated infusions, which would enhance
safety and translatability of the approach. Here, we demonstrate that nanoparticles formed from
poly(ethylene glycol)-poly(u-pentadecalactone-co-p-dioxanone) block copolymers [PEG-poly(PDL-co-
DO)] are highly efficient nanocarriers that provide long-term release: small nanoparticles (less than
100 nm in diameter) continuously released a radiosensitizer (VE822) over a period of several weeks
in vitro, provided widespread intracranial drug distribution during CED, and yielded significant drug
retention within the brain for over 1 week. One advantage of PEG-poly(PDL-co-DO) nanoparticles is that
hydrophobicity can be tuned by adjusting the ratio of hydrophobic PDL to hydrophilic DO monomers,
thus making it possible to achieve a wide range of drug release rates and drug distribution profiles. When
administered by CED to rats with intracranial RG2 tumors, and combined with a 5-day course of fractionated
radiation therapy, VE822-loaded PEG-poly(PDL-co-DO) NPs significantly prolonged survival
when compared to free VE822. Thus, PEG-poly(PDL-co-DO) NPs represent a new type of versatile
nanocarrier system with potential for sustained intracranial delivery of therapeutic agents to treat brain
tumors
SAMHD1 promotes DNA end resection to facilitate DNA repair by homologous recombination
DNA double-strand break (DSB) repair by homologous recombination (HR) is initiated by CtIP/MRN-mediated DNA end resection to maintain genome integrity. SAMHD1 is a dNTP triphosphohydrolase, which restricts HIV- 1 infection, and mutations are associated with Aicardi-Goutières syndrome and cancer. We show that SAMHD1 has a dNTPase-independent function in promoting DNA end resection to facilitate DSB repair by HR. SAMHD1 deficiency or Vpx-mediated degradation causes hypersensitivity to DSB-inducing agents, and SAMHD1 is recruited to DSBs. SAMHD1 complexes with CtIP via a conserved C-terminal domain and recruits CtIP to DSBs to facilitate end resection and HR. Significantly, a cancer-associated mutant with impaired CtIP interaction, but not dNTPase-inactive SAMHD1, fails to rescue the end resection impairment of SAMHD1 depletion. Our findings define a dNTPase-independent function for SAMHD1 in HR-mediated DSB repair by facilitating CtIP accrual to promote DNA end resection, providing insight into how SAMHD1 promotes genome integrity
Loss of ATRX confers DNA repair defects and PARP inhibitor sensitivity
Alpha Thalassemia/Mental Retardation Syndrome X-Linked (ATRX) is mutated frequently in gliomas and represents a potential target for cancer therapies. ATRX is known to function as a histone chaperone that helps incorporate histone variant, H3.3, into the genome. Studies have implicated ATRX in key DNA damage response (DDR) pathways but a distinct role in DNA repair has yet to be fully elucidated. To further investigate the function of ATRX in the DDR, we created isogenic wild-type (WT) and ATRX knockout (KO) model cell lines using CRISPR-based gene targeting. These studies revealed that loss of ATRX confers sensitivity to poly(ADP)-ribose polymerase (PARP) inhibitors, which was linked to an increase in replication stress, as detected by increased activation of the ataxia telangiectasia and Rad3-related (ATR) signaling axis. ATRX mutations frequently co-occur with mutations in isocitrate dehydrogenase-1 and -2 (IDH1/2), and the latter mutations also induce HR defects and PARP inhibitor sensitivity. We found that the magnitude of PARP inhibitor sensitivity was equal in the context of each mutation alone, although no further sensitization was observed in combination, suggesting an epistatic interaction. Finally, we observed enhanced synergistic tumor cell killing in ATRX KO cells with ATR and PARP inhibition, which is commonly seen in HR-defective cells. Taken together, these data reveal that ATRX may be used as a molecular marker for DDR defects and PARP inhibitor sensitivity, independent of IDH1/2 mutations. These data highlight the important role of common glioma-associated mutations in the regulation of DDR, and novel avenues for molecularly guided therapeutic intervention
Antioxidant Status and Lipid Peroxidation in Type II Diabetes Mellitus with and without Complications
Ubiquitin-Fused and/or Multiple Early Genes from Cottontail Rabbit Papillomavirus as DNA Vaccines
Human papillomavirus (HPV) vaccines have the potential to prevent cervical cancer by preventing HPV infection or treating premalignant disease. We previously showed that DNA vaccination with the cottontail rabbit papillomavirus (CRPV) E6 gene induced partial protection against CRPV challenge and that the vaccine's effects were greatly enhanced by priming with granulocyte-macrophage colony-stimulating factor (GM-CSF). In the present study, two additional strategies for augmenting the clinical efficacy of CRPV E6 vaccination were evaluated. The first was to fuse a ubiquitin monomer to the CRPV E6 protein to enhance antigen processing and presentation through the major histocompatibility complex class I pathway. Rabbits vaccinated with the wild-type E6 gene plus GM-CSF or with the ubiquitin-fused E6 gene formed significantly fewer papillomas than the controls. The papillomas also required a longer time to appear and grew more slowly. Finally, a significant proportion of the papillomas subsequently regressed. The ubiquitin-fused E6 vaccine was significantly more effective than the wild-type E6 vaccine plus GM-CSF priming. The second strategy was to vaccinate with multiple CRPV early genes to increase the breadth of the CRPV-specific response. DNA vaccines encoding the wild-type CRPV E1-E2, E6, or E7 protein were tested alone and in all possible combinations. All vaccines and combinations suppressed papilloma formation, slowed papilloma growth, and stimulated subsequent papilloma regression. Finally, the two strategies were merged and a combination DNA vaccine containing ubiquitin-fused versions of the CRPV E1, E2, and E7 genes was tested. This last vaccine prevented papilloma formation at all challenge sites in all rabbits, demonstrating complete protection
Characterization of Cardiac Glycoside Natural Products as Potent Inhibitors of DNA Double-Strand Break Repair by a Whole-Cell Double Immunofluorescence Assay
Small-molecule inhibitors of DNA
repair pathways are being intensively
investigated as primary and adjuvant chemotherapies. We report the
discovery that cardiac glycosides, natural products in clinical use
for the treatment of heart failure and atrial arrhythmia, are potent
inhibitors of DNA double-strand break (DSB) repair. Our data suggest
that cardiac glycosides interact with phosphorylated mediator of DNA
damage checkpoint protein 1 (phospho-MDC1) or E3 ubiquitin–protein
ligase ring finger protein 8 (RNF8), two factors involved in DSB repair,
and inhibit the retention of p53 binding protein 1 (53BP1) at the
site of DSBs. These observations provide an explanation for the anticancer
activity of this class of compounds, which has remained poorly understood
for decades, and provide guidance for their clinical applications.
This discovery was enabled by the development of the first high-throughput
unbiased cellular assay to identify new small-molecule inhibitors
of DSB repair. Our assay is based on the fully automated, time-resolved
quantification of phospho-SER139-H2AX (γH2AX) and 53BP1 foci,
two factors involved in the DNA damage response network, in cells
treated with small molecules and ionizing radiation (IR). This primary
assay is supplemented by robust secondary assays that establish lead
compound potencies and provide further insights into their mechanisms
of action. Although the cardiac glycosides were identified in an evaluation
of 2366 small molecules, the assay is envisioned to be adaptable to
larger compound libraries. The assay is shown to be compatible with
small-molecule DNA cleaving agents, such as bleomycin, neocarzinostatin
chromophore, and lomaiviticin A, in place of IR
PPM1D mutations silence NAPRT gene expression and confer NAMPT inhibitor sensitivity in glioma.
Genetic transformation of a Corynebacterial symbiont from the Chagas disease vector Triatoma infestans
Insect-borne diseases have experienced a troubling resurgence in recent years. Emergence of resistance to pesticides greatly hampers control efforts. Paratransgenesis, or the genetic transformation of bacterial symbionts of disease vectors, is an alternative to traditional approaches. Previously, we developed paratransgenic lines of Rhodniusprolixus, a vector of Chagas disease in Central America. Here, we report identification of a Corynebacterial species as a symbiont of Triatoma infestans, a leading vector of Chagas disease in South America. We have modified this bacterium to produce an immunologically active single chain antibody fragment, termed rDB3. This study establishes the basis for generating paratransgenic T infestans as a strategy for control of Chagas disease. Published by Elsevier Inc