Uptake of biosimilars for TNF-α inhibitors adalimumab and etanercept following the best-value biological medicine initiative in Ireland.

Background There is over 10 years of clinical experience and evidence to show that biosimilar medicines can be used as safely and effectively in approved therapeutic indications as their originator biological medicines. In Ireland, biosimilar medicine uptake has been very slow, and savings to the health service will only be realised through fostering a competitive biological medicine market. Objective The objective of this study was to investigate the utilisation of biosimilars following a 'best-value biological' medicine initiative for adalimumab and etanercept in the Irish healthcare setting. Methods Data was extracted from the National High Tech claims database and High Tech ordering and management hub for the following drugs; adalimumab (Humira®, Amgevita®, Hulio®, Idacio®, and Imraldi®) and etanercept (Enbrel® and Benepali®). Main outcome measure: uptake of the best-value biological medicines. Results In June 2019, just over 90 patients had been initiated on, or switched to a best-value biological for adalimumab or etanercept. Over the next 12 months this increased to over 8500 patients. With the best-value biologicals accounting for approximately 50 % of market share in June 2020, the combined estimated savings and avoided costs are €22.7 million to date. The gain-share prescribing incentive has raised over €3.6 million for the specialties to invest back into patient care. Conclusion Against the background of a finite healthcare budget, this study shows that increasing use of biosimilars can create the financial savings and space to invest in new innovative therapies for the benefit of many patients.

Experimental characterization of the association of ß-cyclodextrin and eight novel cyclodextrin derivatives with two guest compounds.

We investigate the binding of native ß-cyclodextrin (ß-CD) and eight novel ß-CD derivatives with two different guest compounds, using isothermal calorimetry and 2D NOESY NMR. In all cases, the stoichiometry is 1:1 and binding is exothermic. Overall, modifications at the 3' position of ß-CD, which is at the secondary face, weaken binding by several kJ/mol relative to native ß-CD, while modifications at the 6' position (primary face) maintain or somewhat reduce the binding affinity. The variations in binding enthalpy are larger than the variations in binding free energy, so entropy-enthalpy compensation is observed. Characterization of the bound conformations with NOESY NMR shows that the polar groups of the guests may be situated at either face, depending on the host molecule, and, in some cases, both orientations are populated. The present results were used in the SAMPL7 blinded prediction challenge whose results are detailed in the same special issue of JCAMD.

Gait in ducks (Anas platyrhynchos) and chickens (Gallus gallus) - similarities in adaptation to high growth rate.

Genetic selection for increased growth rate and muscle mass in broiler chickens has been accompanied by mobility issues and poor gait. There are concerns that the Pekin duck, which is on a similar selection trajectory (for production traits) to the broiler chicken, may encounter gait problems in the future. In order to understand how gait has been altered by selection, the walking ability of divergent lines of high- and low-growth chickens and ducks was objectively measured using a pressure platform, which recorded various components of their gait. In both species, lines which had been selected for large breast muscle mass moved at a slower velocity and with a greater step width than their lighter conspecifics. These high-growth lines also spent more time supported by two feet in order to improve balance when compared with their lighter, low-growth conspecifics. We demonstrate that chicken and duck lines which have been subjected to intense selection for high growth rates and meat yields have adapted their gait in similar ways. A greater understanding of which components of gait have been altered in selected lines with impaired walking ability may lead to more effective breeding strategies to improve gait in poultry.

An evaluation of an attempt to change the snacking habits of pre-school children using social marketing.

OBJECTIVES: To demonstrate the feasibility and value of social marketing over a wide geographical footprint, and to improve the snacking habits of pre-school children. STUDY DESIGN: Two cross-sectional studies before and approximately 3 months after a social marketing intervention targeted into the least affluent areas using convenience sampling. METHODS: Based on market research and directed at the least affluent areas, a brand was created ('Snack Right') and an information leaflet was distributed, supported by a media launch and events at children's centres in targeted areas. This evaluation of some aspects of the project is based on a questionnaire delivered before and after the events. RESULTS: There were several differences in pre- and post-event responses consistent with the messages delivered, for example increased spending on fruit (but not vegetables) and more positive attitudes towards fruits and vegetables. Some ambiguities were exposed, for example towards snacking, which have implications for the nutritional knowledge of health professionals. The Snack Right brand was recalled by a very high proportion of respondents at follow-up. CONCLUSIONS: This project has shown that social marketing is a viable tool at a subregional level and has the potential to change attitudes, knowledge and behaviour.

Pilonidal sinus of the penis.

A pilonidal sinus is a subcutaneous sinus containing hair. It is most commonly found in the natal cleft of hirsute men. Here we describe the unusual finding of a pilonidal sinus arising on the male foreskin.

Downregulation of Bcl-2 sensitises interferon-resistant renal cancer cells to Fas.

Interferon alpha (IFNalpha) is used to treat patients with advanced renal cell carcinoma (RCC) despite limited clinical benefit. IFNalpha can induce Fas receptor-mediated apoptosis by direct activation of pro-caspase-8 followed by activation of caspase-3. Alternative, indirect activation of caspase-3 via mitochondrial release of cytochrome c can occur and may explain the rescue from Fas-activated cell death by the antiapoptotic members of the Bcl-2 family. In this study, we examined G3139, a novel antisense compound targeting Bcl-2, in combination with IFNalpha. Human RCC lines (SK-RC-44 and SK-RC-07) were treated with IFNalpha, G3139 or a combination of the two. Fas-mediated cytotoxicity was induced by anti-Fas mAb, CH11. An analysis of Bcl-2, Fas and the cleavage of PARP was performed. IFNalpha induced Fas and Bcl-2 in SK-RC-44 and SK-RC-07. IFNalpha sensitised SK-RC-44 to anti-Fas and induced PARP cleavage confirming that IFNalpha has a cytotoxic effect on RCC lines by induction of the Fas antigen. Cytotoxicity was not evident in SK-RC-07 cells treated with IFNalpha. G3139 induced a specific downregulation of Bcl-2 in SK-RC-07 cells, which were then sensitised to anti-Fas after treatment with IFNalpha. Taken together, these results suggest that Fas-dependent pathways as well as alternative pathways, which can be inhibited by Bcl-2, exist in renal cell carcinoma. G3139 in combination with IFNalpha is a potential therapy in patients with metastatic renal cell carcinoma.

Potential bias in NMR relaxation data introduced by peak intensity analysis and curve fitting methods.

We present an evaluation of the accuracy and precision of relaxation rates calculated using a variety of methods, applied to data sets obtained for several very different protein systems. We show that common methods of data evaluation, such as the determination of peak heights and peak volumes, may be subject to bias, giving incorrect values for quantities such as R1 and R2. For example, one common method of peak-height determination, using a search routine to obtain the peak-height maximum in successive spectra, may be a source of significant systematic error in the relaxation rate. The alternative use of peak volumes or of a fixed coordinate position for the peak height in successive spectra gives more accurate results, particularly in cases where the signal/noise is low, but these methods have inherent problems of their own. For example, volumes are difficult to quantitate for overlapped peaks. We show that with any method of sampling the peak intensity, the choice of a 2- or 3-parameter equation to fit the exponential relaxation decay curves can dramatically affect both the accuracy and precision of the calculated relaxation rates. In general, a 2-parameter fit of relaxation decay curves is preferable. However, for very low intensity peaks a 3 parameter fit may be more appropriate.

The effect of antisense Bcl-2 oligonucleotides on Bcl-2 protein expression and apoptosis in human bladder transitional cell carcinoma.

PURPOSE: Bcl-2 is an important determinant of transitional cell carcinoma of the bladder recurrence and progression as well as a factor in patient response to chemotherapy or radiotherapy. We determined Bcl-2 down-regulation after antisense oligonucleotide therapy and synergism with mitomycin C in transitional cell carcinoma of the bladder. MATERIALS AND METHODS: Bcl-2 protein was quantified using flow cytometry and immunohistochemistry in 4 bladder cancer cell lines, in bladder washings from 6 patients with carcinoma in situ and in 16 patient tumor samples. The synergistic effects of antisense oligonucleotides G3139 and 2009, and mitomycin C were investigated in 4 cell lines, while 2009 down-regulation was examined in 20 tumor explants in an ex vivo model. RESULTS: Bcl-2 protein expression was found in all 4 cell lines and in 5 of the 6 cell populations derived from patients with carcinoma in situ. Of the 16 tumors 7 were classified positive by frozen section immunohistochemistry and quantitative flow cytometry. G3139 and 2009 down-regulated Bcl-2 protein expression in all 4 cell lines and 2009 down-regulated Bcl-2 protein expression in half of the Bcl-2 positive tumor specimens. There was only evidence in 1 cell line, T24/83, that Bcl-2 protein expression down-regulation enhanced mitomycin C induced apoptotic cell death. CONCLUSIONS: Bcl-2 was expressed in a significant proportion of bladder tumors and in carcinoma in situ. Therefore, antisense oligonucleotides represent a viable strategy for Bcl-2 protein down-regulation. However, it may not always translate into an increased level of mitomycin C induced apoptosis in transitional cell carcinoma of the bladder.

SANE (Structure Assisted NOE Evaluation): an automated model-based approach for NOE assignment.

A reliable automated approach for assignment of NOESY spectra would allow more rapid determination of protein structures by NMR. In this paper we describe a semi-automated procedure for complete NOESY assignment (SANE, Structure Assisted NOE Evaluation), coupled to an iterative procedure for NMR structure determination where the user is directly involved. Our method is similar to ARIA [Nilges et al. (1997) J. Mol. Biol., 269, 408-422], but is compatible with the molecular dynamics suites AMBER and DYANA. The method is ideal for systems where an initial model or crystal structure is available, but has also been used successfully for ab initio structure determination. Use of this semi-automated iterative approach assists in the identification of errors in the NOE assignments to short-cut the path to an NMR solution structure.

Antisense Bcl-2 oligonucleotide uptake in human transitional cell carcinoma.

OBJECTIVES: Antisense oligonucleotides (AO) downregulate Bcl-2 protein expression in various tumours if good target cell uptake is achieved. In this study, uptake of FITC labelled AO (FITC-AO) directed at Bcl-2 was examined in: (1) the RT4 bladder tumour cell line; (2) normal pig urothelium, and (3) human superficial bladder tumours. METHODS: In the RT4 cell line, uptake of FITC-AO, FITC-scrambled and FITC-sense oligonucleotides were quantified by flow cytometry at 4-hour intervals over 24 h. Uptake of FITC-AO was assessed in normal pig urothelium by flow cytometry after FITC-AO was infused for 1 h. Uptake of FITC AO was assessed in samples from 14 human superficial bladder tumours which were maintained in an ex vivo model. In samples from 6 tumours, uptake at 4 h was assessed using fluorescence microscopy. In samples from 8 separate tumours uptake every 4 h within the first 24-hour incubation period was assessed by flow cytometry. RESULTS: In the RT4 cell line the FITC-AO, FITC-scrambled and FITC-sense oligonucleotide uptake was similar. Disaggregated cells from the normal urothelium of the 3 pigs exhibited 33, 46 and 51% of cells staining positively for FITC-AO as determined by flow cytometry. All 6 tumour samples had detectable intracellular FITC-AO by fluorescence microscopy at 4 h. In the 8 tumours examined over the 24-hour incubation period, there was a range of percentages of positively staining cells. However, most tumours had a monotonic increase in intracellular fluorescence intensity that plateaued 16 h post-infusion. CONCLUSION: Antisense Bcl-2 oligonucleotides were readily taken up by superficial bladder cancer cells but the heterogeneous uptake in tumour samples needs to be considered when assessing the bioavailability of these drugs.

Inherent flexibility in a potent inhibitor of blood coagulation, recombinant nematode anticoagulant protein c2.

Nematode anticoagulant proteins (NAPs) from the hematophagous nematode Ancylostoma caninum inhibit blood coagulation with picomolar inhibition constants, and have been targeted as novel pharmaceutical agents. NAP5 and NAP6 inhibit factor Xa by binding to its active site, whereas NAPc2 binds to factor Xa at a different, as yet unidentified, site and the resultant binary complex inhibits the tissue factor-factor VIIa complex. We have undertaken NMR studies of NAPc2, including the calculation of a solution structure, and found that the protein is folded, with five disulfide bonds, but is extremely flexible, especially in the acidic loop. The Halpha secondary shifts and 3JHNHalpha coupling constants indicate the presence of some beta structure and a short helix, but the intervening loops are highly conformationally heterogeneous. Heteronuclear NOE measurements showed the presence of large amplitude motions on a subnanosecond timescale at the N-terminus and C-terminus and in the substrate-binding loop, indicating that the conformational heterogeneity observed in the NMR structures is due to flexibility of the polypeptide chain in these regions. Flexibility may well be an important factor in the physiological function of NAPc2, because it must interact with other proteins in the inhibition of blood coagulation. We suggest that this inhibitor is likely to become structured on binding to factor Xa, because the inhibition of the tissue factor-factor VIIa complex requires both NAPc2 and factor Xa.

Cold-knife conization versus conization by the loop electrosurgical excision procedure: a randomized, prospective study.

OBJECTIVE: Our purpose was to compare the diagnostic ability and treatment efficacy of conization by the loop electrosurgical excision procedure with cold-knife conization. STUDY DESIGN: One hundred eighty women who required conization for diagnosis and treatment of cervical dysplasia or microinvasive cervical carcinoma were prospectively enrolled in a randomized clinical trial to receive either cold-knife conization or conization by the loop electrosurgical excision procedure. Conization complications, rate of lesion clearance, and therapeutic outcome were assessed for the 2 study groups. RESULTS: There were no statistically significant differences in the complication rate (P = 1.00), the rate of lesion clearance (P =.18), or the rate of disease recurrence (P =.13) between the 2 study groups. The mean follow-up was 11.2 months in the cold-knife conization group and 10.4 months in the loop-excision conization group. CONCLUSION: Cold-knife conization and loop-excision conization yield similar diagnostic and therapeutic results.

Genetics and biology of gynecologic cancer.

Gynecologic epithelial tumors can be grouped into two major categories depending on whether they are derived embryologically from squamous epithelium of the urogenital sinus or from müllerian ducts. Ovarian carcinomas appear morphologically similar to those arising in müllerian-derived organs, and molecular genetic defects present in tumors from these different sources appear to reflect more their histologic subtypes than their organ of origin. The possibility that ovarian epithelial tumors arise from remnants of müllerian ducts in the vicinity of the ovary therefore merits further investigation. Recent advances in our understanding of the state of clonality of various gynecologic tumors, of the influence of age and ovulatory activity on their genetic characteristics, and of their overall molecular genetic features, provide important clues about their initial underlying mechanisms. Novel strategies based on these advances are being tested for their potential utility in treating and monitoring gynecologic tumors.

Detection of ovarian cancer cells: comparison of a telomerase assay and cytologic examination.

BACKGROUND: Telomerase is an enzyme essential for the normal replication of chromosomes. Telomerase activity is absent in most somatic cells in adults, but it is usually expressed in cancer cells, including ovarian carcinoma cells. Our principal goal was to compare the sensitivity of a telomerase assay, i.e., the telomeric repeat amplification protocol (TRAP) assay, with that of cytologic examination in detecting cancer cells in the peritoneal cavity of patients with ovarian carcinoma. METHODS: TRAP assays and cytologic examinations were performed on peritoneal washings and ascitic fluids from 42 patients with active ovarian carcinoma. Control specimens included washings from 29 patients with benign ovarian diseases and ascitic fluids from 14 patients with liver failure. We also evaluated the stability of telomerase in ascitic fluids left unprocessed at room temperature as well as the ability of the TRAP assay to detect cancer cells in mixtures containing large numbers of normal cells. RESULTS: Specimens from 37 (88%) of the 42 patients with ovarian carcinoma tested positive for telomerase. Cytologic examination detected cancer cells in only 27 of the telomerase-positive specimens (i.e., in specimens from 64% of the 42 patients). This difference of 24% (95% confidence interval = 17%-30%) in sensitivity between the two tests was statistically significant (two-sided P = .002). Specimens from five of the patients with ovarian carcinoma were cytologically negative and telomerase negative. All 43 control specimens were cytologically negative, but the TRAP assay detected telomerase in two of them. Telomerase activity was detected in unprocessed samples left at room temperature for 5 days and in mixtures containing a small number of cancer cells and a 2000- to 10000-fold excess of normal cells. CONCLUSIONS: Assaying for telomerase is more sensitive than cytologic examination in detecting cancer cells in the peritoneal cavity of patients with ovarian carcinoma.