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Summer Scholar Publications

Stable nuclear expression of ATP8 and ATP6 genes rescues a mtDNA Complex V null mutant.

Nucleic Acids Res. 2016 Sep 4.

Stable nuclear expression of ATP8 and ATP6 genes rescues a mtDNA Complex V null mutant.

Boominathan A, Vanhoozer S, Basisty N, Powers K, Crampton AL, Wang X, Friedricks N, Schilling B, Brand MD, O'Connor MS.

Abstract

Abstract:

We explore the possibility of re-engineering mitochondrial genes and expressing them from the nucleus as an approach to rescue defects arising from mitochondrial DNA mutations. We have used a patient cybrid cell line with a single point mutation in the overlap region of the ATP8 and ATP6 genes of the human mitochondrial genome. These cells are null for the ATP8 protein, have significantly lowered ATP6 protein levels and no Complex V function. Nuclear expression of only the ATP8 gene with the ATP5G1 mitochondrial targeting sequence appended restored viability on Krebs cycle substrates and ATP synthesis capabilities but, failed to restore ATP hydrolysis and was insensitive to various inhibitors of oxidative phosphorylation. Co-expressing both ATP8 and ATP6 genes under similar conditions resulted in stable protein expression leading to successful integration into Complex V of the oxidative phosphorylation machinery. Tests for ATP hydrolysis / synthesis, oxygen consumption, glycolytic metabolism and viability all indicate a significant functional rescue of the mutant phenotype (including re-assembly of Complex V) following stable co-expression of ATP8 and ATP6. Thus, we report the stable allotopic expression, import and function of two mitochondria encoded genes, ATP8 and ATP6, resulting in simultaneous rescue of the loss of both mitochondrial proteins.

Automation of CAR-T Cell Adoptive Immunotherapy Bioprocessing.

BioProcess International 14(4)s.

Automation of CAR-T Cell Adoptive Immunotherapy Bioprocessing.

Bure K, Ball A, Biagioni K, Mehta S, Choudhary R, Arshad Z, Pettitt DA, Holländer G, Al-Mossawi H, Faulstich F, Reeve B, Smith JA, Brindley DA.

Abstract

Abstract:

Continued clinical efficacy demonstrations of cell-based immunotherapies (iTx) such as chimeric antigen receptor T cell (CAR-T) therapies has made the prospect increasingly likely of an immunotherapy product achieving conditional market authorization in the short term. For example, Novartis and the University of Pennsylvania’s lead candidate (CTL019) for treating a range of hematological malignancies received breakthrough status from the US Food and Drug Administration (FDA) in 2014, permitting access to an expedited drug development pathway for high unmet medical needs. Then in March 2015, the European Medicines Agency’s (EMA’s) Pediatric Committee agreed on a pediatric investigation plan for tisagenlecleucel-T...

Open Access Could Transform Drug Discovery: A Case Study of JQ1.

Expert Opin Drug Discov. 2016 Mar;11(3):321-32.

Open Access Could Transform Drug Discovery: A Case Study of JQ1.

Arshad Z, Smith J, Roberts M, Lee WH, Davies B, Bure K, Hollander GA, Dopson S, Bountra C, Brindley D.

Abstract

Abstract:

The cost to develop a new drug from target discovery to market is a staggering $1.8 billion, largely due to the very high attrition rate of drug candidates and the lengthy transition times during development. Open access is an emerging model of open innovation that places no restriction on the use of information and has the potential to accelerate the development of new drugs. Areas Covered: To date, no quantitative assessment has yet taken place to determine the effects and viability of open access on the process of drug translation. This need is addressed within this study. The literature and intellectual property landscapes of the drug candidate JQ1, which was made available on an open access basis when discovered, and conventionally developed equivalents that were not are compared using the Web of Science and Thomson Innovation software, respectively. Expert opinion: Results demonstrate that openly sharing the JQ1 molecule led to a greater uptake by a wider and more multi-disciplinary research community. A comparative analysis of the patent landscapes for each candidate also found that the broader scientific diaspora of the publically released JQ1 data enhanced innovation, evidenced by a greater number of downstream patents filed in relation to JQ1. The authors' findings counter the notion that open access drug discovery would leak commercial intellectual property. On the contrary, JQ1 serves as a test case to evidence that open access drug discovery can be an economic model that potentially improves efficiency and cost of drug discovery and its subsequent commercialization.

Mitochondrial Dysfunction Induces Senescence with a Distinct Secretory Phenotype.

Cell Metab. 23(2):303-14.

Mitochondrial Dysfunction Induces Senescence with a Distinct Secretory Phenotype.

Wiley CD, Velarde MC, Lecot P, Liu S, Sarnoski EA, Freund A, Shirakawa K, Lim HW, Davis SS, Ramanathan A, Gerencser AA, Verdin E, Campisi J.

Abstract

Abstract:

Cellular senescence permanently arrests cell proliferation, often accompanied by a multi-faceted senescence-associated secretory phenotype (SASP). Loss of mitochondrial function can drive age-related declines in the function of many post-mitotic tissues, but little is known about how mitochondrial dysfunction affects mitotic tissues. We show here that several manipulations that compromise mitochondrial function in proliferating human cells induce a senescence growth arrest with a modified SASP that lacks the IL-1-dependent inflammatory arm. Cells that underwent mitochondrial dysfunction-associated senescence (MiDAS) had lower NAD+/NADH ratios, which caused both the growth arrest and prevented the IL-1-associated SASP through AMPK-mediated p53 activation. Progeroid mice that rapidly accrue mtDNA mutations accumulated senescent cells with a MiDAS SASP in vivo, which suppressed adipogenesis and stimulated keratinocyte differentiation in cell culture. Our data identify a distinct senescence response and provide a mechanism by which mitochondrial dysfunction can drive aging phenotypes.

Decision-Support Tools for Monoclonal Antibody and Cell Therapy Bioprocessing: Current Landscape and Development Opportunities.

BioProcess Executive 13(11).

Decision-Support Tools for Monoclonal Antibody and Cell Therapy Bioprocessing: Current Landscape and Development Opportunities.

Rekhi R, Smith JA, Arshad Z, Roberts M, Bountra C, Bingham I, Bure K, Brindley DA.

Abstract

Abstract:

Industrial-scale manufacturers in a number of fields — from automobiles to biotherapeutics — have long relied on powerful computational and mathematical tools to aid in the scale-up, optimization, quality control, and monitoring of product development. Typical process pathways are highly multifactorial, with numerous branch points, feedback steps, instrumental attributes, and target parameters. Moreover, margins for error are minimal for most industrial processes, requiring high standards of precision from industrial and operational pathways. For those reasons, the complexity of process engineering and process pathway design necessitates that modeling and decision-support tools (DSTs) be used to ensure high-quality and economically viable end products.

A Quantitative Assessment of Factors Affecting the Technological Development and Adoption of Companion Diagnostics

Front. Genet. 6:357.

A Quantitative Assessment of Factors Affecting the Technological Development and Adoption of Companion Diagnostics

Luo D, Smith JA, Meadows NA, Manescu K, Bure K, Davies B, Horne R, DiGiusto DL, Brindley DA.

Abstract

Abstract:

Rapid innovation in (epi)genetics and biomarker sciences is driving a new drug development and product development pathway, with the personalized medicine era dominated by biologic therapeutics and companion diagnostics. Companion diagnostics (CDx) are tests and assays that detect biomarkers and specific mutations to elucidate disease pathways, stratify patient populations, and target drug therapies. CDx can substantially influence the development and regulatory approval for certain high-risk biologics. However, despite the increasingly important role of companion diagnostics in the realization of personalized medicine, in the United States, there are only twenty-three Food and Drug Administration (FDA) approved companion diagnostics on the market for eleven unique indications. Personalized medicines have great potential, yet their use is currently constrained. A major factor for this may lie in the increased complexity of the companion diagnostic and corresponding therapeutic development and adoption pathways. Understanding the market dynamics of companion diagnostic/therapeutic (CDx/Rx) pairs is important to further development and adoption of personalized medicine. Therefore, data collected on a variety of factors may highlight incentives or disincentives driving the development of companion diagnostics. Statistical analysis for thirty-six hypotheses resulted in two significant relationships and thirty-four non-significant relationships. The sensitivity of the companion diagnostic was the only factor that significantly correlated with the price of the companion diagnostic. This result indicates that while there is regulatory pressure for the diagnostic and pharmaceutical industry to collaborate and co-develop companion diagnostics for the approval of personalized therapeutics, there seems to be a lack of parallel economic collaboration to incentivize development of companion diagnostics.

Extracellular Vesicles: Commercial Potential As Byproducts of Cell Manufacturing for Research and Therapeutic Use.

BioProcess International. 13(4).

Extracellular Vesicles: Commercial Potential As Byproducts of Cell Manufacturing for Research and Therapeutic Use.

Smith JA, Ng KS, Mead BE, Dopson S, Reeve B, Edwards J, Wood MJA, Carr AJ, Bure K, Karp JM, Brindley DA.

Abstract

Abstract:

Extracellular vesicles (EVs) are emerging as a potential alternative to some stem-cell–derived therapeutics. Sometimes called exosomes, they are small, secreted vesicles that can possess similar therapeutic mechanisms to whole cells, possibly representing the active pharmaceutical ingredient. In the past 15 years, academic and industry interest in EVs has exponentially increased as mounting evidence demonstrates their role in physiology and pathology as well as their therapeutic potential.

In light of growing efforts in using EVs for research and therapy, optimizing EV manufacturing is important. However, many challenges come with their characterization, scalable manufacture, and regulatory status. Here, we briefly review the biology and therapeutic application of EVs, discuss associated challenges, and suggest how the biotechnology industry could play an important role in overcoming those challenges. Many cell manufacturing companies currently produce EVs but discard them as waste, thereby losing a potentially valuable resource with multiple purposes in a market that’s otherwise rich with an exorbitant cost of goods.

Generating iPSCs: Translating Cell Reprogramming Science into Scalable and Robust Biomanufacturing Strategies.

Cell Stem Cell. 2015 Jan 8;16(1):13-7.

Generating iPSCs: Translating Cell Reprogramming Science into Scalable and Robust Biomanufacturing Strategies.

Silva M, Daheron L, Hurley H, Bure K, Barker R, Carr AJ, Williams D, Kim HW, French A, Coffey PJ, Cooper-White JJ, Reeve B, Rao M, Snyder EY, Ng KS, Mead BE, Smith JA, Karp JM, Brindley DA, Wall I.

Abstract

Abstract:

Induced pluripotent stem cells (iPSCs) have the potential to transform drug discovery and healthcare in the 21(st) century. However, successful commercialization will require standardized manufacturing platforms. Here we highlight the need to define standardized practices for iPSC generation and processing and discuss current challenges to the robust manufacture of iPSC products.

p53-dependent release of Alarmin HMGB1 is a central mediator of senescent phenotypes.

J Cell Biol 2013;201(4):613-29.

p53-dependent release of Alarmin HMGB1 is a central mediator of senescent phenotypes.

Davalos AR, Kawahara M, Malhotra GK, Schaum N, Huang J, Ved U, Beausejour CM, Coppe JP, Rodier F, Campisi J.

Abstract

Abstract:

Cellular senescence irreversibly arrests proliferation in response to potentially oncogenic stress. Senescent cells also secrete inflammatory cytokines such as IL-6, which promote age-associated inflammation and pathology. HMGB1 (high mobility group box 1) modulates gene expression in the nucleus, but certain immune cells secrete HMGB1 as an extracellular Alarmin to signal tissue damage. We show that nuclear HMGB1 relocalized to the extracellular milieu in senescent human and mouse cells in culture and in vivo. In contrast to cytokine secretion, HMGB1 redistribution required the p53 tumor suppressor, but not its activator ATM. Moreover, altered HMGB1 expression induced a p53-dependent senescent growth arrest. Senescent fibroblasts secreted oxidized HMGB1, which stimulated cytokine secretion through TLR-4 signaling. HMGB1 depletion, HMGB1 blocking antibody, or TLR-4 inhibition attenuated senescence-associated IL-6 secretion, and exogenous HMGB1 stimulated NF-κB activity and restored IL-6 secretion to HMGB1-depleted cells. Our findings identify senescence as a novel biological setting in which HMGB1 functions and link HMGB1 redistribution to p53 activity and senescence-associated inflammation.