An update on the potential for male contraception : emerging options

The human population continues to grow and is estimated to rise to 10.1 billion by the end of the century. Therefore, there is still an unmet need for safe and highly effective contraceptive options for both men and women. Current options available to men include withdrawal, condoms, and vasectomy. Methods in development fall into two categories: hormonal and nonhormonal. This review will provide an overview of the testosterone combinations and immunocontraception of hormonal targets. Nonhormonal immunocontraception of sperm proteins will also be examined, together with the use of agents to disrupt other sperm-associated targets and pathways. The categories focused on include epididymal proteins, testicular kinases, epigenetic reader proteins, opioids, lonidamine derivatives, retinoic acid, microRNAs associated with spermatogenesis, and plant extracts. Considering these developments, the number of options available to men is likely to increase in the near future

Immunization of Female Mice with a Plasmid DNA Vaccine Coding EightRepeats of Gonadotrophin Releasing Hormone (Gnrh-I) and Eight T-HelperEpitopes Suppress Fertility In Vivo

Induction of an appropriate immune response against gonadotrophin releasing hormone (GnRH-I) disrupt fertility,reduce fecundity and regress tumours of reproductive system.To disrupt fertility a plasmid DNA vaccine wasengineered coding eight repeats of GnRH-I and eight T-helper epitopes. Translation efficiency of the vaccine wasevaluated in undifferentiated COS1 cells and found to release GnRH-I fusion protein in culture supernatant. Swissalbino female mice (N=24) were immunized with 50μg plasmid DNA construct in study weeks 0, 3, 6, 9 and 12.Group 2 mice were primed with the plasmid DNA in hemagglutinating virus of japanese envelope (HVJE) vector andsubsequent boosts were carried out in phosphate buffer saline. Group 3 mice were immunized with the plasmid DNAin non-ionic surfactant vesicles (NISV) and Group 1 was served as untreated control. The effect of immunization wasstudied in terms of anti-GnRH-I antibody response (OD value at A540 ± SD), suppression of ovarian folliculogenesis,altered uterine histoarchitecture and impaired fertility in vivo in mating trials. In study week 24 OD values of anti-GnRH-I antibody response were 0.982 ± 0.231 in Group 3 mice, followed by 0.783 ± 0.191 in Group 2 in comparisonwith no response in Group 1 controls (0.237 ± 0.147). Results of mating trials showed conception failure invaccinated mice; 51, 18 and 05 pups were seen in the uteri of Groups 1, 2 and 3 mice respectively. There wassignificant (p>0.001) reduction in the weight of ovaries in Group 2 (8.50 ± 2.38 mg) and Group 3 (7.25 ± 0.95 mg)mice compared to Group 1 control (15.00 ± 1.41 mg). Significant reduction of ovarian folliculogenesis was seen inGroup 2 (p>0.001) and Group 3 mice (p>0.01). In conclusion, the plasmid DNA vaccine delivered in female micewith HVJE and NISV induced significantly (p>0.001) higher levels of anti-GnRH-I antibody response, suppressedovarian and uterine function and impaired fertility in vivo

Comparison of the physical characteristics of monodisperse non-ionic surfactant vesicles (NISV) prepared using different manufacturing methods

Non-ionic surfactant vesicles (NISV) are synthetic membrane vesicles formed by self-assembly of a non-ionic surfactant, often in a mixture with cholesterol and a charged chemical species. Different methods can be used to manufacture NISV, with the majority of these requiring bulk mixing of two phases. This mixing process is time-consuming and leads to the preparation of large and highly dispersed vesicles, which affects the consistency of the final product and could hinder subsequent regulatory approval. In this study, we have compared the physical characteristics of NISV prepared using two conventional methods (thin-film hydration method and heating method) with a recently introduced microfluidic method. The resulting particles from these methods were assessed for their physical characteristics and in vitro cytotoxicity. Through microfluidics, nano-sized NISV were prepared in seconds, through rapid and controlled mixing of two miscible phases (lipids dissolved in alcohol and an aqueous medium) in a microchannel, without the need of a size reduction step, as required for the conventional methods. Stability studies over two months showed the particles were stable regardless of the method of preparation and there were no differences in terms of EC50 on A375 and A2780 cell lines. However, this work demonstrates the flexibility and ease of applying lab-on-chip microfluidics for the preparation of NISV that could be used to significantly improve formulation research and development, by enabling the rapid manufacture of a consistent end-product, under controlled conditions

Development of a mucosal vaccine delivery system

This thesis was previously held under moratorium from 15th August 2014 until 15th August 2019.Vaccine delivery systems that target the mucosal immune system carry important advantages in terms of accessibility and acceptability. Lipid based nanoparticles with and without the incorporation of bile salts were formulated by novel preparation methods in order to provide suitable immunogen carriers for delivery to mucosal surfaces. The nanoparticles were physically characterised by determination of size, zeta potential, and morphological appearance using a variety of analytical tools. A new method for the analysis of the chemical lipid components by HPLC separation was implemented, which can be adopted for quality control in future. The adjuvant and delivery properties of the nanoparticles were evaluated with different immunogens incuding non-pathogenic peptides and immunogenic proteins originating from pathogenic organisms. In particular, the reproductive self-hormone gonadotrophin releasing hormone (GnRH) conjugated with different carrier proteins was used as an immunogen in in vivo studies. Various mucosal administration routes were compared against parenteral vaccination, including oral, nasal and vaginal. Systemic and local specific antibodies (IgG, IgG subclasses and IgA) were evaluated in body fluids obtained from immunised mice. The effect of immunisation was evaluated by assessment of reproductive hormone levels after immunisation. In addition, pathogen-derived immunogens were used to evaluate whether the lipid nanoparticles were suitable for vaccination via mucosal surfaces. The results showed that lipid-based nanoparticles enhanced antigen delivery via subcutaneous and nasal routes with nasal delivery proving to be successful in terms of producing systemic and mucosal antibodies. Whereas, vaginal and oral routes showed a very low immune responses when using GnRH conjugates. However, oral administration of pathogen-derived antigens showed promising results. The possible reasons for the limited capability of orally delivered nanoparticles were investigated using LC/MS analysis and fluorescent multiphoton microscopy. The study suggests that lipid based nanoparticles possess adjuvant properties and can be potential candidate for enabling vaccine delivery via the nasal route.Vaccine delivery systems that target the mucosal immune system carry important advantages in terms of accessibility and acceptability. Lipid based nanoparticles with and without the incorporation of bile salts were formulated by novel preparation methods in order to provide suitable immunogen carriers for delivery to mucosal surfaces. The nanoparticles were physically characterised by determination of size, zeta potential, and morphological appearance using a variety of analytical tools. A new method for the analysis of the chemical lipid components by HPLC separation was implemented, which can be adopted for quality control in future. The adjuvant and delivery properties of the nanoparticles were evaluated with different immunogens incuding non-pathogenic peptides and immunogenic proteins originating from pathogenic organisms. In particular, the reproductive self-hormone gonadotrophin releasing hormone (GnRH) conjugated with different carrier proteins was used as an immunogen in in vivo studies. Various mucosal administration routes were compared against parenteral vaccination, including oral, nasal and vaginal. Systemic and local specific antibodies (IgG, IgG subclasses and IgA) were evaluated in body fluids obtained from immunised mice. The effect of immunisation was evaluated by assessment of reproductive hormone levels after immunisation. In addition, pathogen-derived immunogens were used to evaluate whether the lipid nanoparticles were suitable for vaccination via mucosal surfaces. The results showed that lipid-based nanoparticles enhanced antigen delivery via subcutaneous and nasal routes with nasal delivery proving to be successful in terms of producing systemic and mucosal antibodies. Whereas, vaginal and oral routes showed a very low immune responses when using GnRH conjugates. However, oral administration of pathogen-derived antigens showed promising results. The possible reasons for the limited capability of orally delivered nanoparticles were investigated using LC/MS analysis and fluorescent multiphoton microscopy. The study suggests that lipid based nanoparticles possess adjuvant properties and can be potential candidate for enabling vaccine delivery via the nasal route

Optimizing efficacy of mucosal vaccines

In general, there are only a few vaccines administered via mucosal routes, as the mucosal immune system presents numerous hurdles, including diversity in mucosal surface structure, complexity in immune cell interaction and limitations in experimental methodology. This therefore necessitates a range of strategies to be used for each target area. With reference to the three main routes of delivery and associated mucosal surfaces (oral/intestinal, nasal/respiratory and female genital tract), this review examines how coadministration of immune-stimulatory molecules, adjuvants, delivery systems and mucoadhesives are used to improve mucosal vaccine efficacy. Key considerations to the development of next-generation mucosal vaccines include improved efficacy and safety, technological advancements in medical devices to enable convenience and better administration, as well as reduced manufacturing costs

Assessment of the antigen-specific antibody response induced by mucosal administration of a GnRH conjugate entrapped in lipid nanoparticles

Vaccines administered parenterally have been developed against gonadotrophin releasing hormone (GnRH) for anti-fertility and anti-cancer purposes. The aim of this study was to demonstrate whether mucosal delivery using GnRH immunogens entrapped in lipid nanoparticles (LNP) could induce anti-GnRH antibody titres. Immunogens consisting of KLH (keyhole limpet haemocyanin) conjugated to either GnRH-I or GnRH-III analogues were entrapped in LNP. Loaded non-ionic surfactant vesicles (NISV) were administered subcutaneously, while nasal delivery was achieved using NISV in xanthan gum and oral delivery using NISV containing deoxycholate (bilosomes). NISV and bilosomes had similar properties: they were spherical, in the nanometre size range, with a slightly negative zeta potential and surface properties that changed with protein loading and inclusion of xanthan gum. Following immunisation in female BALB/c mice, systemic antibody responses were similar for both GnRH-I and GnRH-III immunisation. Only nasal delivery proved to be successful in terms of producing systemic and mucosal antibodies

An update on developments in female hormonal contraception

The human population continues to grow in some parts of the world, which has severe impact on resources, health and the environment. Individually, contraception enables women to choose their optimal family size and birth spacing, while in resource-poor countries it can help lift families out of poverty. While the oral contraceptive pill revolutionised female contraceptive options, there was a price to pay in terms of increased health risks. Today, improved formulations have been developed, together with non-oral hormonal technologies. This review will examine the history of female contraceptive research and provide an update on the status and future direction of new products

Mucosal and systemic immune responses following mucosal immunisation of tetanus toxoid entrapped in lipid nanoparticles prepared by microwave reactor

In this study, the use of a microwave reactor, which allowed high input of energy into a pressurised system in a short period of time, was investigated for preparation of lipid nanoparticles (LNPs). The aim was to optimise the formulation process by reducing manufacturing time. Two types of LNPs were prepared; non-ionic surfactant vesicles (NISV) and bilosomes (modified NISV incorporating bile salts), with a model antigen (tetanus toxoid, TT) and the immune response induced after mucosal (nasal and oral, respectively) administration was assessed. The TT loaded LNPs were characterised in terms of particle size, size distribution, morphology, and entrapment efficiency. Immunisation was evaluated by lethal challenge with tetanus toxin in an animal model. The efficiency of vaccination was evaluated by measuring the anti-TT IgG antibody levels in the vaccinated animals. Bilosomes formed by this method showed an immunogen entrapment efficiency of ∼ 30% which was significantly (

Investigating the influence of ubiquinone blood level on the abilities of children with specific learning disorder

Abstract Background Ubiquinone has antioxidant properties and has been linked to cognitive performance in some neuropsychiatric disorders. Its role in specific learning disorder manifestations has not been previously investigated. Therefore, the aim of this study was to measure the blood levels of ubiquinone in a group of children with specific learning disorder in comparison to typically developing children and to investigate the correlation between ubiquinone levels in children with specific learning disorder and some of their intellectual capabilities, reading, spelling and writing performance.  Methods The study included 71 native Arabic speaking children: 31 in the specific learning disorder group and 40 in the typically developing (TD) group. The abilities of the children with specific learning disorder were evaluated by the Stanford-Binet Intelligence Scale-4th edition, the Dyslexia Assessment Test, and the Illinois Test of Psycholinguistic Abilities. The level of ubiquinone was measured in both groups by ELISA. Correlation between some aptitudes of children with specific learning disorder and the ubiquinone level was performed. Results The blood levels of ubiquinone in the children with specific learning disorder group were less than those in the TD group. Correlation analysis revealed a significant positive correlation between ubiquinone and the scores of backward digit span abilities. Conclusions Ubiquinone has a role in the auditory working memory performance of children with specific learning disorder (with impairment in reading). The decreased levels of ubiquinone in this sample of children with specific learning disorder could have participated in the pathogenesis of this disorder

Evaluation of immunocastration conjugates based on GnRH linked to carrier molecules in a male rodent model

Gonadotropin-releasing hormone (GnRH) stimulates the pituitary gland to secrete sex hormones. In the present study, we evaluated different conjugates of GnRH to abrogate sex hormone secretion in a male rat model. Firstly, GnRH-I was conjugated to keyhole limpet haemocyanin (KLH) using either whole-sequence GnRH-I or an analogue of GnRH-I (CHWSYGLRPG-NH2) using glutaraldehyde cross linkage. Six-week-old Sprague-Dawley male rats (n=6) were immunized intramuscularly with the conjugates adsorbed onto alum, equivalent to 50 μg of GnRH peptide and administered in weeks 1, 3, 5 and 7. The study was concluded in week 15. Comparison was made with untreated controls and previously established anti-fertility conjugates: CHWSYGLRPG-NH2 (GnRH-I) or CHWSHDWKPG-NH2 (analogue of lamprey GnRH-III, lGnRH-III) linked to tetanus toxoid (using a heterobifunctional reagent to achieve cross linkage). Antibody production, hormone levels and testicular diameter changes were assessed, together with, sperm movement and effects on organ weights. Similar high levels of antibody secretion were observed in all the immunized groups, although whole GnRHI- KLH produced a sustained level of production for an additional week. Similarly, testosterone levels were significantly (p<0.05) reduced in all immunized groups. There were no significant changes observed in body weight and testicular diameter of immunized animals compared with the untreated controls. However, in terms of sperm motility and sperm number, the best anti-fertility effects were observed with lGnRH-III-TT and GnRH-I-KLH and to a lesser extent whole GnRH-I-KLH. These groups also showed significant increase in kidney weight. Finally, considering all the above-mentioned subjects in addition to availability and easier and cheaper way of preparation,we came to this conclusion that whole GnRH-IKLH satisfactorily met most of our favourite criteria and could be used in immunocastration vaccine production purposes successfully