Beyond patents: Scientific knowledge, and access to vaccine

Knowledge is a public good. Patents provide property rights in knowledge, which gives the patentee the right to exclude others from utilising the knowledge for the life of the patent. Patents in the field of pharmaceuticals are controversial because of the importance of the knowledge which they exclude others from using. Patents have come under significant criticism for this very reason – with some going as far as to claim that patent protection on pharmaceutical products as the cause of developing states having poor or limited access to life-saving pharmaceutical products. Most of the academic literature regarding access to medicines goes this same way. This paper challenges this viewpoint, and considers the barriers to generic access to medicines beyond patents. This paper looks beyond intellectual property rights to determine what other mechanisms exist that allow innovative vaccine manufacturers to control access to knowledge regarding their products which can act as a barrier to the utilisation of knowledge in the pharmaceutical industry, in a similar manner to intellectual property rights. This paper takes a case study approach considering non-patent-related barriers to access to medicines, focusing on pandemic influenza vaccines and the role of proprietary, non-patented knowledge. This paper concludes that manufacturers have an exclusive monopoly, not because of their intellectual property rights, but because the knowledge required to make the drug is not accessible to generic manufacturers, and highlights why this is the case. This paper argues that it is not the patent protection which is the barrier to introducing generic pandemic influenza vaccines, but rather it is the inaccessibility of knowledge which is not in the public domain, or the inability of manufacturers in developing states to utilise this knowledge, which is the true barrier in this field

Investigation of Translational Reprogramming during Transient and Stable Expression of Monoclonal Antibodies in Chinese Hamster Ovary Cells

Translational reprogramming and mRNA translation e ciency greatly in uence global protein synthesis, cell proliferation and growth; important parameters in de ning recombinant protein expression yields. Polysome pro ling is a widely-used technique to analyse mRNA transla- tion and its e ciency that provides a snapshot of ribosomes loaded on mRNA transcripts at any particular time. A higher number of polysomes present on a given mRNA suggests that the mRNA is being more heavily translated than those mRNAs with few ribosomes. Fur- ther, a large pool of sub-polysomes (40S, 60S and 80S) compared to polysomes in a sample suggests low translational activity. Here, polysome pro ling has been applied to investigate translational reprogramming in multiple recombinant monoclonal antibody (mAb)-producing Chinese hamster ovary (CHO) cell lines, and to determine how reprogramming re ects the ability of such cells to proliferate and make recombinant proteins in stable and transient mAb expression systems, in batch and fed-batch culture mode. The impact of culture temperature on the polysome pro le and hence on reprogramming was also investigated in transient studies. Polysome pro ling revealed reprogramming di ered between recombinant cell lines. Those with the highest global translational e ciency generally had the fastest cell speci c growth rates, although total ribosome capacity did not directly relate to those with the fastest growth rates or mAb productivities. This suggests it is the ability to utilise available machinery that determines protein synthetic capacity. Recombinant cell lines with higher cell speci c produc- tivities generally maintained a higher polysome to monosome (P:M) ratio during stationary phase and had elevated recombinant mRNA copy numbers localised to translationally active heavy polysomes. In transient systems, the P:M ratio was maintained longer at reduced tem- perature cultivation and related to higher mAb yields being obtained. A number of endogenous transcripts were found to be more or less abundant on polysomes at di erent times of culture, indicative of changes in the cellular requirements of the encoded proteins. Such transcripts could be potential cell engineering targets to help tune the needs of the cell to the demands of a culture process or recombinant protein, or alternatively their untranslated regions harnessed to help preferentially load target mRNAs onto ribosomes. When upstream open reading frames (uORFs) or alternative translation start sites were engineered into recombinant transcripts a range of mAb expressions were observed allowing the tuning of mAb expression, including improvement over a standard untranslated region used industrially as a control. The ndings described in this thesis therefore reveal insights into the mechanisms involved in translational regulation and reprogramming in CHO cells during bioprocessing. These can be utilised for further improvement via targeted cell engineering strategies, cell line screen- ing approaches or modi cation of recombinant transcripts for enhanced industrial host and recombinant cell lines

PharmaSUG 2015 -Paper PO02

ABSTRACT Breaking the vertical and/or horizontal axis can simplify the figure, improve aesthetics, and save space. Two SAS samples 48330 and 38765 have provided examples to break the vertical axis. However, using ENTRY and DRAWLINE statements can create a much better break

Determination of Bioprocessing Variables that Influence Particle and Aggregation Formation of Therapeutic IgGs

One of the dominant classes of protein based drugs, which continues to contribute to the growth and success of biologics, are recombinant monoclonal antibody (mAb) therapies. In preparing recombinant antibodies for biotherapeutic applications, the stability and maintenance of this is of utmost importance; not just for the activity of the molecule but also in terms of the safety of the drug and this must be demonstrated to regulatory authorities before a biotherapeutic receives approval for use. One such measure of protein integrity, quality or stability is the assessment of soluble aggregates and sub-visible particles present in formulations after stressed stability studies. The presence of such particles and aggregates in recombinant biotheraputics is potentially of concern in the manufacture of such biologics due to the reported immunogenic responses observed in patients when these biologics are administered; which is believed to be associated with the presence of aggregated antibody species. In addition to this, the in vivo efficacy of the biologic is inevitably compromised upon formation of such species. As such the regulatory bodies require that aggregate and particle levels are monitored and reported in the specification of a biologic. The manufacturing of recombinant mAbs using mammalian expression systems involves three general stages; upstream, where the target molecule is cloned and expressed in the system of choice; downstream, where the recombinant protein material is recovered from the fermentation supernatant and formulation whereby the molecule is ‘formulated’ to maintain its stability and for delivery to patients. Throughout this process the mAb molecules experience a wide variety of stresses such as temperature, high concentrations, interactions with host cell impurities, varying pH and shear stresses that could all potentially influence a molecules susceptibility to aggregate or form particles. Whilst there has been much research at the amino acid level to determine how primary sequence influences propensity of molecules to aggregate, there has been little work investigating how the whole manufacturing process might influence the susceptibility of mAbs to aggregate and form particles. The focus of this work was therefore to investigate whether three variables during manufacturing of mAbs; feeding during fermentation, harvest day and the exclusion/inclusion of an additional wash step during purification, influence the levels of aggregates and particles in antibody formulations post-temperature stressing. Atomic force microscopy was also used to investigate the mechanism of mAb particle formation and aggregation using a model system under forced aggregation conditions. The results from these studies confirmed that washing of the protein A column during mAb purification can influence subsequent particle formation. The inclusion of a specific wash step during protein A purification was effective at lowering particle levels in formulations of a model mAb compared to those samples which were purified without. The wash step was shown to remove mAb fragments and molecular chaperones and other host cell proteins from the Chinese hamster ovary cell host. The re-introduction of these species back into mAb solutions resulted in a subsequent increase in particle formation suggesting that species present in the material removed by the wash fraction influenced the particle numbers observed. With regard to culture conditions and feeding of the cells, batch cultures produced a less stable molecule compared to the fed-batch cultures as determined by the propensity to form particles and this could be related to the intracellular stress being experienced by cells in during culture. Cell lysis is influenced by the intracellular stress levels of the cultures, which is in-turn influenced by the feeding regime and harvest day. The levels of mAb fragments and sub-visible particles increased in fed-batch and batch Mab-184 samples, as culture viability decreases; which was mirrored by changes in the expression of certain stress inducible genes. Purification of the samples with the wash step reduces the particle levels to similar levels across all harvest days suggesting that the wash step is not only removing misfolded protein released into the culture supernatant upon cell lysis, but also removes species that are present in the cell culture supernatant which contribute to antibody particle formation. These results highlight the need for better monitoring of intracellular stress levels during CHO culture for informing harvest decisions as the intracellular state of the cells influence the quality of the final therapeutic product. Finally, using AFM particle formation of a model mAb is characteristic of a coagulation mechanism after one hour of temperature stress. Collectively together these studies demonstrate the importance of both upstream and downstream processing decisions on the subsequent propensity of mAbs to form particles and hence the need to consider the whole bioprocess to obtain the best possible product with regard to aggregate and particle amounts