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Science (New York, N.Y.) Nov 2023The global replacement of histones with protamines in sperm chromatin is widespread in animals, including insects, but its actual function remains enigmatic. We show...
The global replacement of histones with protamines in sperm chromatin is widespread in animals, including insects, but its actual function remains enigmatic. We show that in the paternal effect mutant (), sperm chromatin retains germline histones H3 and H4 genome wide without impairing sperm viability. However, after fertilization, sperm chromosomes are targeted by the egg chromosomal passenger complex and engage into a catastrophic premature division in synchrony with female meiosis II. We show that encodes a rapidly evolving transition protein specifically required for the eviction of (H3-H4) tetramers from spermatid DNA after the removal of H2A-H2B dimers. Our study thus reveals an unsuspected role of histone eviction from insect sperm chromatin: safeguarding the integrity of the male pronucleus during female meiosis.
Topics: Animals; Female; Male; Chromatin; Drosophila Proteins; Fertilization; Histones; Spermatozoa; Drosophila melanogaster; Amidine-Lyases; DNA Packaging; Paternal Inheritance
PubMed: 37943933
DOI: 10.1126/science.adh0037 -
Nanomaterials (Basel, Switzerland) Jun 2021Macromolecular biomolecules are currently dethroning classical small molecule therapeutics because of their improved targeting and delivery properties. Protamine-a small... (Review)
Review
Macromolecular biomolecules are currently dethroning classical small molecule therapeutics because of their improved targeting and delivery properties. Protamine-a small polycationic peptide-represents a promising candidate. In nature, it binds and protects DNA against degradation during spermatogenesis due to electrostatic interactions between the negatively charged DNA-phosphate backbone and the positively charged protamine. Researchers are mimicking this technique to develop innovative nanopharmaceutical drug delivery systems, incorporating protamine as a carrier for biologically active components such as DNA or RNA. The first part of this review highlights ongoing investigations in the field of protamine-associated nanotechnology, discussing the self-assembling manufacturing process and nanoparticle engineering. Immune-modulating properties of protamine are those that lead to the second key part, which is protamine in novel vaccine technologies. Protamine-based RNA delivery systems in vaccines (some belong to the new class of mRNA-vaccines) against infectious disease and their use in cancer treatment are reviewed, and we provide an update on the current state of latest developments with protamine as pharmaceutical excipient for vaccines.
PubMed: 34200384
DOI: 10.3390/nano11061508 -
Pharmaceutics Jun 2021Protamine is a natural cationic peptide mixture mostly known as a drug for the neutralization of heparin and as a compound in formulations of slow-release insulin.... (Review)
Review
Protamine is a natural cationic peptide mixture mostly known as a drug for the neutralization of heparin and as a compound in formulations of slow-release insulin. Protamine is also used for cellular delivery of nucleic acids due to opposite charge-driven coupling. This year marks 60 years since the first use of Protamine as a transfection enhancement agent. Since then, Protamine has been broadly used as a stabilization agent for RNA delivery. It has also been involved in several compositions for RNA-based vaccinations in clinical development. Protamine stabilization of RNA shows double functionality: it not only protects RNA from degradation within biological systems, but also enhances penetration into cells. A Protamine-based RNA delivery system is a flexible and versatile platform that can be adjusted according to therapeutic goals: fused with targeting antibodies for precise delivery, digested into a cell penetrating peptide for better transfection efficiency or not-covalently mixed with functional polymers. This manuscript gives an overview of the strategies employed in protamine-based RNA delivery, including the optimization of the nucleic acid's stability and translational efficiency, as well as the regulation of its immunostimulatory properties from early studies to recent developments.
PubMed: 34198550
DOI: 10.3390/pharmaceutics13060877 -
EMBO Reports Jun 2023Spermatozoa have a unique genome organization. Their chromatin is almost completely devoid of histones and is formed instead of protamines, which confer a high level of...
Spermatozoa have a unique genome organization. Their chromatin is almost completely devoid of histones and is formed instead of protamines, which confer a high level of compaction and preserve paternal genome integrity until fertilization. Histone-to-protamine transition takes place in spermatids and is indispensable for the production of functional sperm. Here, we show that the H3K79-methyltransferase DOT1L controls spermatid chromatin remodeling and subsequent reorganization and compaction of the spermatozoon genome. Using a mouse model in which Dot1l is knocked-out (KO) in postnatal male germ cells, we found that Dot1l-KO sperm chromatin is less compact and has an abnormal content, characterized by the presence of transition proteins, immature protamine 2 forms and a higher level of histones. Proteomic and transcriptomic analyses performed on spermatids reveal that Dot1l-KO modifies the chromatin prior to histone removal and leads to the deregulation of genes involved in flagellum formation and apoptosis during spermatid differentiation. As a consequence of these chromatin and gene expression defects, Dot1l-KO spermatozoa have less compact heads and are less motile, which results in impaired fertility.
Topics: Animals; Male; Cell Differentiation; Chromatin; Chromatin Assembly and Disassembly; Gene Expression; Histones; Proteomics; Semen; Spermatogenesis; Spermatozoa; Mice
PubMed: 37099396
DOI: 10.15252/embr.202256316 -
Drugs in Context 2022Due to worldwide increases in obesity and average maternal age, the incidence of gestational diabetes mellitus (GDM) is increasing. The primary treatment of GDM is... (Review)
Review
Due to worldwide increases in obesity and average maternal age, the incidence of gestational diabetes mellitus (GDM) is increasing. The primary treatment of GDM is medical nutrition therapy but approximately 15-30% of individuals need pharmacotherapy to reach blood glucose goals to minimize the adverse consequences of hyperglycaemia. In the past, regular and neutral protamine Hagedorn insulin were the mainstays of pharmacological treatment for GDM due to their well-established safety; however, because they are administered as injections and require strict timing of doses and meals to minimize hypoglycaemia, alternatives are often sought. The research around the treatment of GDM continues to evolve as insulin analogues and oral agents are studied in clinical trials. The short-term and long-term effects of treatment choices on both mothers and progeny are being evaluated, and this narrative review summarizes the current state of information available regarding the treatment of GDM.
PubMed: 35775071
DOI: 10.7573/dic.2021-9-12 -
Expert Review of Vaccines 2023mRNA vaccines have been developed as a promising cancer management. It is noted that specification of the antigen sequence of the target antigen is necessary for the... (Review)
Review
INTRODUCTION
mRNA vaccines have been developed as a promising cancer management. It is noted that specification of the antigen sequence of the target antigen is necessary for the design and manufacture of an mRNA vaccine.
AREAS COVERED
The steps involved in preparing the mRNA-based cancer vaccines are isolation of the mRNA cancer from the target protein using the nucleic acid RNA-based vaccine, sequence construction to prepare the DNA template, transcription for protein translation from DNA into mRNA strand, 5' cap addition and poly(A) tailing to stabilize and protect the mRNA from degradation and purification process to remove contaminants produced during preparation.
EXPERT OPINION
Lipid nanoparticles, lipid/protamine/mRNA nanoparticles, and cell-penetrating peptides have been used to formulate mRNA vaccine and to ensure vaccine stability and delivery to the target site. Delivery of the vaccine to the target site will trigger adaptive and innate immune responses. Two predominant factors of the development of mRNA-based cancer vaccines are intrinsic influence and external influence. In addition, research relating to the dosage, route of administration, and cancer antigen types have been observed to positively impact the development of mRNA vaccine.
Topics: Humans; Cancer Vaccines; Vaccines; Neoplasms; Immunity, Innate; mRNA Vaccines; Nanoparticles; RNA, Messenger
PubMed: 37401128
DOI: 10.1080/14760584.2023.2232450 -
Nature Communications Dec 2023Idiopathic fertility disorders are associated with mutations in various genes. Here, we report that coiled-coil glutamate-rich protein 1 (CCER1), a germline-specific and...
Idiopathic fertility disorders are associated with mutations in various genes. Here, we report that coiled-coil glutamate-rich protein 1 (CCER1), a germline-specific and intrinsically disordered protein (IDP), mediates postmeiotic spermatid differentiation. In contrast, CCER1 deficiency results in defective sperm chromatin compaction and infertility in mice. CCER1 increases transition protein (Tnp1/2) and protamine (Prm1/2) transcription and mediates multiple histone epigenetic modifications during the histone-to-protamine (HTP) transition. Immiscible with heterochromatin in the nucleus, CCER1 self-assembles into a polymer droplet and forms a liquid-liquid phase-separated condensate in the nucleus. Notably, we identified loss-of-function (LoF) variants of human CCER1 (hCCER1) in five patients with nonobstructive azoospermia (NOA) that were absent in 2713 fertile controls. The mutants led to premature termination or frameshift in CCER1 translation, and disrupted condensates in vitro. In conclusion, we propose that nuclear CCER1 is a phase-separated condensate that links histone epigenetic modifications, HTP transitions, chromatin condensation, and male fertility.
Topics: Male; Humans; Mice; Animals; Histones; Protamines; Semen; Chromatin; Spermatozoa; Spermatogenesis; Fertility; Infertility, Male
PubMed: 38081819
DOI: 10.1038/s41467-023-43480-z -
Molecules (Basel, Switzerland) Jan 2023As antibody-drug conjugates have become a very important modality for cancer therapy, many site-specific conjugation approaches have been developed for generating... (Review)
Review
As antibody-drug conjugates have become a very important modality for cancer therapy, many site-specific conjugation approaches have been developed for generating homogenous molecules. The selective antibody coupling is achieved through antibody engineering by introducing specific amino acid or unnatural amino acid residues, peptides, and glycans. In addition to the use of synthetic cytotoxins, these novel methods have been applied for the conjugation of other payloads, including non-cytotoxic compounds, proteins/peptides, glycans, lipids, and nucleic acids. The non-cytotoxic compounds include polyethylene glycol, antibiotics, protein degraders (PROTAC and LYTAC), immunomodulating agents, enzyme inhibitors and protein ligands. Different small proteins or peptides have been selectively conjugated through unnatural amino acid using click chemistry, engineered C-terminal formylglycine for oxime or click chemistry, or specific ligation or transpeptidation with or without enzymes. Although the antibody protamine peptide fusions have been extensively used for siRNA coupling during early studies, direct conjugations through engineered cysteine or lysine residues have been demonstrated later. These site-specific antibody conjugates containing these payloads other than cytotoxic compounds can be used in proof-of-concept studies and in developing new therapeutics for unmet medical needs.
Topics: Cytotoxins; Antibodies; Immunoconjugates; Proteins; Peptides; Amino Acids; Antineoplastic Agents; Polysaccharides
PubMed: 36770585
DOI: 10.3390/molecules28030917 -
Journal of Hematology & Oncology Dec 2022Acute myeloid leukemia (AML) is a fatal clonal hematopoietic malignancy, which results from the accumulation of several genetic aberrations in myeloid progenitor cells,...
BACKGROUND
Acute myeloid leukemia (AML) is a fatal clonal hematopoietic malignancy, which results from the accumulation of several genetic aberrations in myeloid progenitor cells, with a worldwide 5-year survival prognosis of about 30%. Therefore, the development of more effective therapeutics with novel mode of action is urgently demanded. One common mutated gene in the AML is the DNA-methyltransferase DNMT3A whose function in the development and maintenance of AML is still unclear. To specifically target "undruggable" oncogenes, we initially invented an RNAi-based targeted therapy option that uses the internalization capacity of a colorectal cancer specific anti-EGFR-antibody bound to cationic protamine and the anionic siRNA. Here, we present a new experimental platform technology of molecular oncogene targeting in AML.
METHODS
Our AML-targeting system consists of an internalizing anti-CD33-antibody-protamine conjugate, which together with anionic molecules such as siRNA or ibrutinib-Cy3.5 and cationic free protamine spontaneously assembles into vesicular nanocarriers in aqueous solution. These nanocarriers were analyzed concerning their physical properties and relevant characteristics in vitro in cell lines and in vivo in xenograft tumor models and patient-derived xenograft leukemia models with the aim to prepare them for translation into clinical application.
RESULTS
The nanocarriers formed depend on a balanced electrostatic combination of the positively charged cationic protamine-conjugated anti-CD33 antibody, unbound cationic protamine and the anionic cargo. This nanocarrier transports its cargo safely into the AML target cells and has therapeutic activity against AML in vitro and in vivo. siRNAs directed specifically against two common mutated genes in the AML, the DNA-methyltransferase DNMT3A and FLT3-ITD lead to a reduction of clonal growth in vitro in AML cell lines and inhibit tumor growth in vivo in xenotransplanted cell lines. Moreover, oncogene knockdown of DNMT3A leads to increased survival of mice carrying leukemia patient-derived xenografts. Furthermore, an anionic derivative of the approved Bruton's kinase (BTK) inhibitor ibrutinib, ibrutinib-Cy3.5, is also transported by this nanocarrier into AML cells and decreases colony formation.
CONCLUSIONS
We report important results toward innovative personalized, targeted treatment options via electrostatic nanocarrier therapy in AML.
Topics: Humans; Mice; Animals; Protamines; Static Electricity; RNA, Small Interfering; Leukemia, Myeloid, Acute; Methyltransferases; DNA
PubMed: 36457063
DOI: 10.1186/s13045-022-01390-5 -
JACS Au Dec 2022Fondaparinux, a clinically approved anticoagulant pentasaccharide for the treatment of thrombotic diseases, displays better efficacy and biosafety than other...
Fondaparinux, a clinically approved anticoagulant pentasaccharide for the treatment of thrombotic diseases, displays better efficacy and biosafety than other heparin-based anticoagulant drugs. However, there is no suitable antidote available for fondaparinux to efficiently manage its potential bleeding risks, thereby precluding its widespread use. Herein, we describe a convergent and stereocontrolled approach to efficiently synthesize an aminopentyl-functionalized pentasaccharide, which is further used to prepare fondaparinux-based biotin conjugates and clusters. Biological activity evaluation demonstrates that the anticoagulant activity of the fondaparinux-based biotin conjugate and trimer is, respectively, neutralized by avidin and protamine as effective antidotes. This work suggests that our synthetic biotin conjugate and trimer have potential for the development of neutralizable and safe anticoagulant drugs.
PubMed: 36590263
DOI: 10.1021/jacsau.2c00537