Poly(2-oxazoline)-based polymeric micelle platform for drug delivery

Polymeric micelles (PMs) have been extensively utilized as drug delivery platform. Particularly, potent hydrophobic small molecules were encapsulated in the PMs to alleviate toxicity issues and improve therapeutic outcomes. We attempt to provide detailed information on PMs for hydrophobic small molecules, such as the design of block copolymers (BCP) and current clinical outcomes from PMs. In particular, we aim to describe advanced analytical approaches for elucidating molecular interactions for effective solubilization. This dissertation includes a novel computer-aided strategy for rational design of PMbased delivery systems for poorly soluble drugs. We have developed novel descriptors of drug polymer complexes that were employed to build models to predict both drug loading efficiency (LE) and loading capacity (LC). These models were used for virtual screening of drug libraries and eight drugs for the experimental validation. Three putative true positive as well as three putative negative hits were confirmed (implying 75% prediction accuracy). The success of the computational strategy suggests its broad utility for rational design of drug delivery systems. This dissertation involves the study of poly(2-oxazoline) micelles (POx) for treatment of medulloblastoma. For patients with SHH-subgroup medulloblastoma, SHH-pathway inhibition may be more effective and less toxic than current non-targeted therapy. We successfully solubilized SHH-pathway inhibitor, vismodegib, in POx micelles (POx-vismo) and showed the PM formulation improved drug efficacy, demonstrated in the treatment of medulloblastoma animal model. Mechanistic studies revealed that POx-vismo decreased vismodegib binding to serum proteins and improved brain and tumor drug penetration without penetration of the nanoparticle carrier into the CNS. This dissertation also includes the development of novel poly(2-oxazoline)-based block copolymer with the aromatic heterocyclic side chains and demonstration of its application as a drug delivery platform. The copolymer was synthesized via the condensation of N,N dimethylbiguanide with the methyl ester side chain in poly(2-methoxycarboxyethyl-2-oxazoline) block (PMestOx). Successful encapsulation into these micelles has been demonstrated for several poorly soluble drugs. The capability of this new copolymer to solubilize a uniquely diverse set of active pharmaceutical ingredients suggests potential applications in drug delivery. In summary, poly(2-oxazoline)-based PM platforms are versatile drug delivery platform and exhibit the broad potential for ideal drug delivery of therapeutic small molecules.Doctor of Philosoph

ATR maintains chromosomal integrity during postnatal cerebellar neurogenesis and is required for medulloblastoma formation

Microcephaly and medulloblastoma may both result from mutations that compromise genomic stability. We report that ATR, which is mutated in the microcephalic disorder Seckel syndrome, sustains cerebellar growth by maintaining chromosomal integrity during postnatal neurogenesis. Atr deletion in cerebellar granule neuron progenitors (CGNPs) induced proliferation-associated DNA damage, p53 activation, apoptosis and cerebellar hypoplasia in mice. Co-deletions of either p53 or Bax and Bak prevented apoptosis in Atr-deleted CGNPs, but failed to fully rescue cerebellar growth. ATR-deficient CGNPs had impaired cell cycle checkpoint function and continued to proliferate, accumulating chromosomal abnormalities. RNA-Seq demonstrated that the transcriptional response to ATR-deficient proliferation was highly p53 dependent and markedly attenuated by p53 co-deletion. Acute ATR inhibition in vivo by nanoparticle-formulated VE-822 reproduced the developmental disruptions seen with Atr deletion. Genetic deletion of Atr blocked tumorigenesis in medulloblastoma-prone SmoM2 mice. Our data show that p53-driven apoptosis and cell cycle arrest – and, in the absence of p53, non-apoptotic cell death – redundantly limit growth in ATR-deficient progenitors. These mechanisms may be exploited for treatment of CGNP-derived medulloblastoma using ATR inhibition

Nanoformulated Remdesivir with Extremely Low Content of Poly(2-oxazoline)-Based Stabilizer for Aerosol Treatment of COVID-19

The rise of the novel virus SARS-CoV2 which causes the disease known as COVID-19 has led to a global pandemic claiming millions of lives. With no clinically approved treatment for COVID-19, physicians initially struggled to treat the disease, and a need remains for improved antiviral therapies in this area. It is conceived early in the pandemic that an inhalable formulation of the drug remdesivir which directly targets the virus at the site of infection could improve therapeutic outcomes in COVID-19. A set of requirements are developed that would be conducive to rapid drug approval: 1) try to use GRAS reagents 2) minimize excipient concentration and 3) achieve a working concentration of 5 mg/mL remdesivir to obtain a deliverable dose which is 5-10% of the IV dose. In this work, it is discovered that Poly(2-oxazoline) block copolymers can stabilize drug nanocrystal suspensions and provide suitable formulation characteristics for aerosol delivery while maintaining antiviral efficacy. The authors believe POx block copolymers can be used as a semi-ubiquitous stabilizer for the rapid development of nanocrystal formulations for new and existing diseases

Inhibition of UCH-L1 Deubiquitinating Activity with Two Forms of LDN-57444 Has Anti-Invasive Effects in Metastatic Carcinoma Cells

Normally ubiquitin C-terminal hydrolase L1 (UCH-L1) is expressed in the central nervous and reproductive systems of adults, but its de novo expression has been detected in many human cancers. There is a growing body of evidence that UCH-L1 de-ubiquitinating (DUB) activity plays a major pro-metastatic role in certain carcinomas. Here we tested anti-metastatic effects of the small-molecule inhibitor of UCH-L1 DUB activity, LDN-57444, in cell lines from advanced oral squamous cell carcinoma (OSCC) as well as invasive nasopharyngeal (NP) cell lines expressing the major pro-metastatic gene product of Epstein–Barr virus (EBV) tumor virus, LMP1. To overcome the limited aqueous solubility of LDN-57444 we developed a nanoparticle formulation of LDN-57444 by incorporation of the compound in polyoxazoline micellear nanoparticles (LDN-POx). LDN-POx nanoparticles were equal in effects as the native compound in vitro. Our results demonstrate that inhibition of UCH-L1 DUB activity with LDN or LDN-POx inhibits secretion of exosomes and reduces levels of the pro-metastatic factor in exosomal fractions. Both forms of UCH-L1 DUB inhibitor suppress motility of metastatic squamous carcinoma cells as well as nasopharyngeal cells expressing EBV pro-metastatic Latent membrane protein 1 (LMP1) in physiological assays. Moreover, treatment with LDN and LDN-POx resulted in reduced levels of pro-metastatic markers, a decrease of carcinoma cell adhesion, as well as inhibition of extra-cellular vesicle (ECV)-mediated transfer of viral invasive factor LMP1. We suggest that soluble inhibitors of UCH-L1 such as LDN-POx offer potential forms of treatment for invasive carcinomas including EBV-positive malignancies