Legal aspects of patenting time.

Times of embryonic development are naturally occurring phenomena and, therefore, under the caveat that all things in nature without human intervention are not patentable, should not be patentable.

The US patent system is broken.

In discussions in this issue of RBM Online relating to a patent granted to Auxogyn, Professor Renee Reijo Pera claims this is not about a "broken patent system". I shall demonstrate how dysfunctional the patent system has become and how much in denial 'inventor' Renee Reijo Pera is about her invention differing from a naturally occurring phenomenon.

Inhibition of human spermatozoa-zona pellucida binding by a combinatorially derived peptide from a synthetic target.

Intact zona-free human oocytes were screened using a combinatorial peptide library selection protocol. Pieczenik Peptide Sequence 1 (PPS1) HEHRKRG binds human spermatozoa. A complementary and unique binding sequence HNSSLSPLATPA (PPS2) was developed from the first PPS1 ligand that binds to the human zona pellucida or oolemma. Cytoplasm-free zonae from unfertilized eggs were obtained and used as an assay system to test the effects of exposure to these two ligands. Spermatozoa were inserted into evacuated zonae and their behaviour and binding activity were assessed at regular intervals. The behaviour of spermatozoa exposed to PPS1 and unlabelled spermatozoa injected into unexposed zonae was similar as far as binding was concerned (50 and 54% binding), but PPS1 exposed spermatozoa had higher motility and displacement, marked by their escape from the zona pellucida. Zonae exposed to PPS2 inhibited the interaction between injected spermatozoa and the inside of the zona when compared with controls (8.3 and 53.8% attached respectively, P < 0.001). The sperm-zona pellucida interaction described in this paper is applied as a functional assay for molecular interactions of sperm binding and can be used to assess function for potential surface markers on gametes. It is shown here that a unique binding ligand (PPS2) can be synthesized from another complimentary ligand (PPS1) without the need for a known intermediate substrate. PPS1 and PPS2 may have properties that can be used to target processes involved in conception and assisted reproduction. A movie sequence taken approximately 30 min after injection of spermatozoa into empty human zonae pellucidae shows behaviour of non-manipulated spermatozoa into zonae not exposed or exposed to ligand. This may be purchased for viewing on the Internet at www.rbmonline.com/Article/2159 (free to web subscribers).

A positive-selection function for microRNA: an adaptor hypothesis revisited.

A hypothesis for coding different combinations of mRNA segments by a novel postulated function of microRNAs (miRNA) is presented. In this hypothesis, miRNA act as both coding adaptors and ligating enzymes. This hypothesis postulates an enzyme-substrate intermediate comprising a RNA triplex structure consisting of a miRNA adaptor and two mRNA configured as two turns of an A-form RNA helix. Postulating the existence of differentially expressed sets of adaptors (codes) may explain biological functions such as imprinting, differentiation, immunological escape and immunological tolerance, inter alia.

Combinatorial peptide library binding of mammalian spermatozoa identifies a ligand (HIPRT) in the axin protein: putative identification of a sperm surface axin binding protein and intriguing developmental implications.

The identification of components in cell-cell interactions is an important research goal in reproductive and developmental biology. Such interactions are critical to gamete development, fertilization, implantation and basic development. Several proteins involved with sperm-oocyte interaction and other developmentally important phenomena have been identified. However, these are obviously only a subset of the molecular components involved in such complex cell-cell interactions. One method that has been used to identify binding partners for particular molecular targets is the use of combinatorial libraries accessible on phage surfaces. For the most part, this technique has mainly been applied to screen specific target moieties. However, in some cases whole-cell screening has been attempted. This study describes the first report of screening intact, living mammalian gametes using a proprietary whole-cell combinatorial library binding and analysis protocol. Results from the first screening protocol of mouse spermatozoa strongly identified a putative sperm-binding ligand using proprietary bioinformatic analysis. This amino acid sequence (HIPRT) precisely corresponds with a previously characterized highly conserved protein-protein interaction site in the axin protein. This sequence is found within the binding site for a known sperm surface protein, glycogen synthase kinase-3. This result not only provides proof of the utility of this technique to identify cell surface ligands in mammalian gametes, but it also suggests a potential role for spermatozoa in facilitating developmental axis formation in mammalian embryos.

Are the universes of antibodies and antigens symmetrical?

This paper discusses the assumed infinite numbers of antigens and antibodies, and the nature of their interactions, in relation to reproductive immunology. Classical models offered by Pauling, Burnet and Crick are discussed. The author's own model is described, based on selectionist principles, proposing that the numbers are defined, discrete and not infinite. The binding specificity of monoclonal antibodies of four to six amino acids limits the number on the total number of antibody-binding peptides. This limitation on size enables the total number of these peptides to define 'one-hit' or 'two-hit' protein sequences in databases of known proteins. Estimate of the combinations of variable heavy and light chains of different numerical magnitude is 2.5 x 10(6). The author proposes the equivalence principle, where equivalent numbers of antigens and antibodies exist and theorizing a limited number of antigen-antibody pairs in the magnitude of 3.2 x 10(6) or slightly less. The nature of various protein-ligand interactions, indicated by their limited numbers of interactions, clarifies why a particular amino acid is encoded by a particular codon and the nature of encoding by tRNA. In relation to reproductive phenomena, specific proteins for contact with sperm ligands were designed using these parameters.