The CENP-A nucleosome: where and when it happens during the inner kinetochore's assembly.

CENP-A is an essential histone variant that replaces the canonical H3 at the centromeres and marks these regions epigenetically. The CENP-A nucleosome is the specific building block of centromeric chromatin, and it is recognized by CENP-C and CENP-N, two components of the constitutive centromere-associated network (CCAN), the first protein layer of the kinetochore. Recent proposals of the yeast and human (h)CCAN structures position the assembly on exposed DNA, suggesting an elusive spatiotemporal recognition. We summarize the data on the structural organization of the CENP-A nucleosome and the binding of CENP-C and CENP-N. The latter posits an apparent contradiction in engaging the CENP-A nucleosome versus the CCAN. We propose a reconciliatory model for the assembly of CCAN on centromeric chromatin.

Novel Insights into the Biological Activity of Croton lechleri Twigs Extracts and Advancements in Their Sustainable Recovery.

Sangre de drago, the sap of Croton lechleri Müll. Arg. tree, has been used for centuries in traditional medicine owing to its diverse biological activities. Extracts derived from different parts of the species contain a multitude of phytochemicals with varied applications. Twigs, however, are among the least studied parts of the plant. This study unveils new biological activities of Croton lechleri twig extracts recovered by applying Soxhlet and advanced green techniques. For all extracts, total phenolic content and antioxidant activity were determined. Subsequently, four were selected, and their cytotoxic effects were assessed on both normal (HaCat) and malignant melanoma (A375) skin cell lines using the MTT assay and trypan blue exclusion assay. All showed dose-dependent cytotoxicity, with the Soxhlet ethanol extract demonstrating the highest selectivity towards A375 cells over HaCat cells. The extracts induced apoptosis and necrosis, as confirmed by Annexin V/PI dual-labeling and flow cytometry, highlighting their ability to trigger programmed cell death in cancer cells. The selective inhibition of cell cycle progression in A375 compared to HaCat observed both for Soxhlet ethanol and pressurized ethanol extracts induces cell cycle arrest at multiple points, primarily in the G1 and G2/M phases, and significantly reduces DNA synthesis as evidenced by the decrease in the S-phase population, confirmed by the EdU assay. Consequently, the Soxhlet extract composition was analyzed using LC-MS, which revealed their richness in polyphenolic compounds, particularly flavonoids from the flavonol subclass.

Analysis of the Ibotenic Acid, Muscimol, and Ergosterol Content of an Amanita Muscaria Hydroalcoholic Extract with an Evaluation of Its Cytotoxic Effect against a Panel of Lung Cell Lines In Vitro.

The fungus Amanita muscaria is universally recognizable for its iconic appearance; it is also widely regarded as poisonous, inedible, and even deadly. In spite of that, there have been documented cases of use of A. muscaria-containing preparations against various diseases, including cancer, to no apparent ill effect. The search for compounds that can be used to treat cancer among various plants and fungi has been intensifying in recent years. In light of this, we describe an HPLC HILIC analytical method for the evaluation of the content of the anticancer compound ergosterol (ERG) and the neuroactive alkaloids ibotenic acid (IBO) and muscimol (MUS) that contribute significantly to the unpleasant physiological syndrome associated with A. muscaria consumption. A 'homemade' A. muscaria tincture made using 80-proof rye vodka as the solvent, an A. muscaria extract made with a standardized water-ethanol solution as the solvent, and fractions obtained from the second extract via liquid-liquid extraction with nonpolar solvents were analyzed. The study also presents the results of capillary zone electrophoresis with contactless conductivity detection and UHPLC-MS/MS analyses of the IBO and MUS content of the two native A. muscaria extracts and an evaluation of the standardized extract's cytotoxic effect against a small panel of lung cell cultures in vitro. Our results show that the standardized extract has a significant cytotoxic effect and does not contain the compounds of interest in any significant quantity.

Hollow spherical nucleic acid structures based on polymer-coated phospholipid vesicles.

A feasible one pot synthesis of hollow spherical nucleic acids (SNAs) using phospholipid liposomes is reported. These constructs are synthesized in a chemically straightforward process involving formation of unilamellar liposomes, coating the liposomes with a thin cross-linked polymeric layer, and grafting the latter with short (about 20 bases) DNA oligonucleotide strands. They consist of vesicular cores, composed of readily available phospholipid (1,2-dipalmitoyl-sn-glycero-phosphocholine), whereas the strands are deliberately arranged on the surface of the vesicular entities. The initial vesicular structure and morphology are preserved during the coating and grafting reactions. The novel hollow/vesicular SNAs are characterized with a hydrodynamic radius and radius of gyration of 78.3 and 88.5 nm, respectively, and moderately negative (-14.2 mV) ζ potential. They carry thousands (5868) of oligonucleotide strands per vesicle, which are not strongly radially oriented and adopt an unextended conformation as anticipated from the smaller value of the grafting density compared to the critical grafting density at the transition to brush conformation. The constructs are practically devoid of toxicity and exhibit high binding affinity to complementary nucleic acids. Unlike any other nucleic acid structural motif, they cross the cell membrane and enter cells without the need of transfection agents.

Cationic (Co)polymers Based on N-Substituted Polyacrylamides as Carriers of Bio-macromolecules: Polyplexes, Micelleplexes, and Spherical Nucleic Acidlike Structures.

Novel N-substituted polyacrylamides bearing a cycle with two tertiary amines, poly(4-methyl-piperazin-1-yl)-propenone (PMPP) and its block copolymers with polylactide (PMPP-b-PLA), are synthesized and characterized. The homopolymers are water-soluble, whereas the block copolymers self-assemble in aqueous solution into a small size (Rh around 30 nm), are narrowly distributed, and exhibit core-shell micelles with good colloidal stability. Both the homopolymers and copolymer micelles are positively charged (ζ-potentials in the 13.8-17.6 mV range), which are employed for formation of electrostatic complexes with oppositely charged DNA. Complexes (polyplexes, micelleplexes, and spherical nucleic acidlike structures) in a wide range of N/P (amino to phosphate groups) ratios are prepared with short (115 bp) and long (2000 bp) DNA. The behavior and physicochemical properties of the resulting nanocarriers of DNA are strongly dependent on the polymer/polymer micelles' characteristics and the DNA chain length. All systems exhibit low cytotoxicity and good cellular uptake ability and show promise for gene delivery and regulation.

Functional Polyglycidol-Based Block Copolymers for DNA Complexation.

Gene therapy is an attractive therapeutic method for the treatment of genetic disorders for which the efficient delivery of nucleic acids into a target cell is critical. The present study is aimed at evaluating the potential of copolymers based on linear polyglycidol to act as carriers of nucleic acids. Functional copolymers with linear polyglycidol as a non-ionic hydrophilic block and a second block bearing amine hydrochloride pendant groups were prepared using previously synthesized poly(allyl glycidyl ether)-b-polyglycidol block copolymers as precursors. The amine functionalities were introduced via highly efficient radical addition of 2-aminoethanethiol hydrochloride to the alkene side groups. The modified copolymers formed loose aggregates with strongly positive surface charge in aqueous media, stabilized by the presence of dodecyl residues at the end of the copolymer structures and the hydrogen-bonding interactions in polyglycidol segments. The copolymer aggregates were able to condense DNA into stable and compact nanosized polyplex particles through electrostatic interactions. The copolymers and the corresponding polyplexes showed low to moderate cytotoxicity on a panel of human cancer cell lines. The cell internalization evaluation demonstrated the capability of the polyplexes to successfully deliver DNA into the cancer cells.

One-pot synthesis of oligonucleotide-grafted polymeric nanoparticles.

A feasible one-pot approach for constructing oligonucleotide-grafted polymeric nanoparticles is reported. The approach involves formation of mesoglobules from a thermoresponsive polymer, coating of the mesoglobules with a cross-linked polymeric shell, and grafting the latter with oligonucleotide strands. Dynamic and static light scattering are used to parameterize the novel constructs. They are relatively large structures with hydrodynamic radii and molar masses reaching 200 nm and 150.0 × 106 g mol-1, respectively. The oligonucleotide-grafted polymeric nanoparticles are of spherical morphology and moderately negative (-12.4 to -19.1 mV) ζ potential as revealed by AFM, TEM, and electrophoretic light scattering. In accordance with their large size, they are found to carry thousands of oligonucleotide strands per particle. The novel constructs are thermoresponsive. They undergo reversible collapse upon heating and swelling upon cooling, which is associated with changes in the grafting density and, hence, the conformation of the oligonucleotide strands from unextended at room temperature to a more extended one at elevated temperatures. The versatility of the approach is demonstrated by varying the type of the cross-linked shell and content of the oligonucleotide strands and, hence, the grafting density. Appropriate diversification and modifications are suggested as well.

Cellular Pharmacology of Palladinum(III) Hematoporphyrin IX Complexes: Solution Stability, Antineoplastic and Apoptogenic Activity, DNA Binding, and Processing of DNA-Adducts.

Two paramagnetic PdIII complexes of hematoporphyrin IX ((7,12-bis(1-hydroxyethyl)-3,8,13,17-tetramethyl-21H-23H-porphyn-2,18-dipropionic acid), Hp), namely a dinuclear one [PdIII2(Hp-3H)Cl3(H2O)5]·2PdCl2, Pd1 and a mononuclear metalloporphyrin type [PdIII(Hp-2H)Cl(H2O)]·H2O, Pd2 have been synthesized reproducibly and isolated as neutral compounds at different reaction conditions. Their structure and solution stability have been assayed by UV/Vis and EPR spectroscopy. The compounds researched have shown in vitro cell growth inhibitory effects at micromolar concentration against a panel of human tumor cell lines. A DNA fragmentation test in the HL-60 cell line has indicated that Pd1 causes comparable proapoptotic effects with regard to cisplatin but at substantially higher concentrations. Pd1 and cisplatin form intra-strand guanine bis-adducts as the palladium complex is less capable of forming DNA adducts. This demonstrates its cisplatin-dissimilar pharmacological profile. The test for efficient removal of DNA-adducts by the NER synthesis after modification of pBS plasmids with either cisplatin or Pd1 has manifested that the lesions induced by cisplatin are far better recognized and repaired compared those of Pd1. The study on the recognition and binding of the HMGB-1 protein to cisplatin or Pd1 modified DNA probes have shown that HMG proteins are less involved in the palladium agent cytotoxicity.

Antimycobacterial activity generated by the amide coupling of (-)-fenchone derived aminoalcohol with cinnamic acids and analogues.

Aminoethyl substituted 2-endo-fenchol prepared from (-)-fenchone was used as scaffold for the synthesis of series of 31 amide structures by N-acylation applying cinnamic acids and analogues. The evaluation of their in vitro activity against Mycobacterium tuberculosis H37Rv showed for some of them promising activity-up to 0.2 µg/ml, combined with relatively low cytotoxicity of the selected active compounds.

Interaction of nucleolin with ribosomal RNA genes and its role in RNA polymerase I transcription.

Nucleolin is a multi-functional nucleolar protein that is required for ribosomal RNA gene (rRNA) transcription in vivo, but the mechanism by which nucleolin modulates RNA polymerase I (RNAPI) transcription is not well understood. Nucleolin depletion results in an increase in the heterochromatin mark H3K9me2 and a decrease in H4K12Ac and H3K4me3 euchromatin histone marks in rRNA genes. ChIP-seq experiments identified an enrichment of nucleolin in the ribosomal DNA (rDNA) coding and promoter region. Nucleolin is preferentially associated with unmethylated rRNA genes and its depletion leads to the accumulation of RNAPI at the beginning of the transcription unit and a decrease in UBF along the coding and promoter regions. Nucleolin is able to affect the binding of transcription termination factor-1 on the promoter-proximal terminator T0, thus inhibiting the recruitment of TIP5 and HDAC1 and the establishment of a repressive heterochromatin state. These results reveal the importance of nucleolin for the maintenance of the euchromatin state and transcription elongation of rDNA.

pH-responsive niosome-based nanocarriers of antineoplastic agents.

Differences in pH between the tumour interstitium and healthy tissues can be used to induce conformational changes in the nanocarrier structure, thereby triggering drug release at the desired site. In the present study, novel pH-responsive nanocarriers were developed by modifying conventional niosomes with hexadecyl-poly(acrylic acid)n copolymers (HD-PAAn). Niosomal vesicles were prepared by the thin film hydration method using Span 60, Span 60/Tween 60 and cholesterol as main constituents, and HD-PAA modifiers of different concentrations (0.5, 1, 2.5, 5 mol%). Next, two model substances, a water-soluble fluorescent dye (calcein) and a hydrophobic agent with pronounced antineoplastic activity (curcumin), were loaded in the aqueous core and hydrophobic membrane of the elaborated niosomes, respectively. Physicochemical properties of blank and loaded nanocarriers such as hydrodynamic diameter (Dh), size distribution, zeta potential, morphology and pH-responsiveness were investigated in detail. The cytotoxicity of niosomal curcumin was evaluated against human malignant cell lines of different origins (MJ, T-24, HUT-78), and the mechanistic aspects of proapoptotic effects were elucidated. The formulation composed of Span 60/Tween 60/cholesterol/2.5% HD-PAA17 exhibited optimal physicochemical characteristics (Dh 302 nm; ζ potential -22.1 mV; high curcumin entrapment 83%), pH-dependent drug release and improved cytotoxic and apoptogenic activity compared to free curcumin.

The chaperone like function of the nonhistone protein HMGB1.

Almost all essential nuclear processes as replication, repair, transcription and recombination require the chromatin template to be correctly unwound and than repackaged. The major strategy that the cell uses to overcome the nucleosome barrier is the proper removal of the histone octamer and subsequent deposition onto DNA. Important factors in this multi step phenomenon are the histone chaperones that can assemble nucleosome arrays in vitro in the absence of ATP. The nonhistone protein HMGB1 is a good candidate for a chaperone as its molecule consists of two DNA binding motives, Box's A and B, and a long nonstructured C tail highly negatively charged. HMGB1 protein is known as a nuclear "architectural" factor for its property to bind preferentially to distorted DNA structures and was reported to kink the double helix. Our experiments show that in the classical stepwise dialysis method for nucleosome assembly the addition of HMGB1 protein stimulates more than two times the formation of middle-positioned nucleosomes. The stimulation effect persists in dialysis free experiment when the reconstitution is possible only in the presence of a chaperone. The addition of HMGB1 protein strongly enhanced the formation of a nucleosome in a dose dependant manner. Our results show that the target of HMGB1 action as a chaperone is the DNA fragment not the histone octamer. One possible explanation for the stimulating effect of HMGB1 is the "architectural" property of the protein to associate with the middle of the DNA fragment and to kink it. The acquired V shaped DNA structure is probably conformationals more favorable to wrap around the prefolded histone octamer. We tested also the role of the post-synthetic acetylation for the chaperone function of HMGB1 protein. The presence of an acetyl groups at Lys 2 decreases strongly the stimulating effect of the protein in the stepwise salt dialysis experiment and the same tendency persisted in the dialysis free experiment.

A novel pH sensitive water soluble fluorescent nanomicellar sensor for potential biomedical applications.

Herein we report on the synthesis and sensor activity of a novel pH sensitive probe designed as highly water-soluble fluorescent micelles by grafting of 1,8-naphthalimide-rhodamine bichromophoric FRET system (RNI) to the PMMA block of a well-defined amphiphilic diblock copolymer-poly(methyl methacrylate)-b-poly(methacrylic acid) (PMMA48-b-PMAA27). The RNI-PMMA48-b-PMAA27 adduct is capable of self-assembling into micelles with a hydrophobic PMMA core, containing the anchored fluorescent probe, and a hydrophilic shell composed of PMAA block. Novel fluorescent micelles are able to serve as a highly sensitive pH probe in water and to internalize successfully HeLa and HEK cells. Furthermore, they showed cell specificity and significantly higher photostability than that of a pure organic dye label such as BODIPY. The valuable properties of the newly prepared fluorescent micelles indicate the high potential of the probe for future biological and biomedical applications.

Chromatin regulators in DNA replication and genome stability maintenance during S-phase.

The duplication of genetic information is central to life. The replication of genetic information is strictly controlled to ensure that each piece of genomic DNA is copied only once during a cell cycle. Factors that slow or stop replication forks cause replication stress. Replication stress is a major source of genome instability in cancer cells. Multiple control mechanisms facilitate the unimpeded fork progression, prevent fork collapse and coordinate fork repair. Chromatin alterations, caused by histone post-translational modifications and chromatin remodeling, have critical roles in normal replication and in avoiding replication stress and its consequences. This text reviews the chromatin regulators that ensure DNA replication and the proper response to replication stress. We also briefly touch on exploiting replication stress in therapeutic strategies. As chromatin regulators are frequently mutated in cancer, manipulating their activity could provide many possibilities for personalized treatment.

Micellar Form of a Ferrocene-Containing Camphor Sulfonamide with Improved Aqueous Solubility and Tumor Curing Potential.

The discovery of new anticancer drugs with а higher, more specific activity and diminished side effects than the conventional chemotherapeutic agents is a tremendous challenge to contemporary medical research and development. To achieve a pronounced efficacy, the design of antitumor agents can combine various biologically active subunits in one molecule, which can affect different regulatory pathways in cancer cells. We recently demonstrated that a newly synthesized organometallic compound, a ferrocene-containing camphor sulfonamide (DK164), possesses promising antiproliferative activity against breast and lung cancer cells. However, it still encounters the problem of solubility in biological fluids. In this work, we describe a novel micellar form of DK164 with significantly improved solubility in aqueous medium. DK164 was embedded in biodegradable micelles based on a poly(ethylene oxide)-b-poly(α-cinnamyl-ε-caprolactone-co-ε-caprolactone)-b-poly(ethylene oxide) triblock copolymer (PEO113-b-P(CyCL3-co-CL46)-b-PEO113), and the physicochemical parameters (size, size distribution, zeta potential, encapsulation efficiency) and biological activity of the obtained system were studied. We used cytotoxicity assays and flow cytometry to determine the type of cell death, as well as immunocytochemistry to assess the influence of the encapsulated drug on the dynamics of cellular key proteins (p53 and NFkB) and the process of autophagy. According to our results, the micellar form of the organometallic ferrocene derivate (DK164-NP) exhibited several advantages compared to the free substance, such as higher metabolic stability, better cellular uptake, improved bioavailability, and long-term activity, maintaining nearly the same biological activity and anticancer properties of the drug.

Antitumor Activity of Bioactive Compounds from Rapana venosa against Human Breast Cell Lines.

This study is the first report describing the promising antitumor activity of biologically active compounds isolated from the hemolymph of marine snail Rapana venosa-a fraction with Mw between 50 and 100 kDa and two structural subunits (RvH1 and RvH2), tested on a panel of human breast cell lines-six lines of different molecular subtypes of breast cancer MDA-MB-231, MDA-MB-468, BT-474, BT-549, SK-BR-3, and MCF-7 and the non-cancerous MCF-10A. The fraction with Mw 50-100 kDa (HRv 50-100) showed good antitumor activity manifested by a significant decrease in cell viability, altered morphology, autophagy, and p53 activation in treated cancer cells. An apparent synergistic effect was observed for the combination of HRv 50-100 with cis-platin for all tested cell lines. The combination of HRv 50-100 with cisplatin and/or tamoxifen is three times more effective compared to treatment with classical chemotherapeutics alone. The main proteins in the active fraction, with Mw at ~50 kDa, ~65 kDa, ~100 kDa, were identified by MALDI-MS, MS/MS analyses, and bioinformatics. Homology was established with known proteins with antitumor potential detected in different mollusc species: peroxidase-like protein, glycoproteins Aplysianin A, L-amino acid oxidase (LAAO), and the functional unit with Mw 50 kDa of RvH. Our study reveals new perspectives for application of HRv 50-100 as an antitumor agent used alone or as a booster in combination with different chemotherapies.

The effect of PKC phosphorylation on the "architectural" properties of HMGB1 protein.

High mobility group box (HMGB)1 protein acts as an architectural element, promoting the assembly of active nucleoprotein complexes due to its ability to bend DNA and to bind preferentially to distorted DNA structures. The behavior of HMGB1 as an "architect" of chromatin defines it as an important factor in many cellular processes such as repair, replication and remodeling. It was shown that the post-synthetic acetylation of HMGB1 at Lys2 modulated its essential properties as a structure-specific nuclear protein. We studied the role of PKC phosphorylation on the "architectural" properties of HMGB1, (i) the effect for the formation of a stable complex with DNA damaged by the anti-tumour drug cis-platinum and (ii) the influence on the ability of HMGB1 protein to bend short DNA fragments. PKC-phosphorylated recombinant HMGB1 increased about an order of magnitude its affinity to cis-platinated DNA, a finding that has already been reported for in vivo acetylated protein. Regarding the effect on the protein's DNA bending ability, it was enhanced upon phosphorylation as demonstrated by the stimulation of DNA circularization. We showed also that PKC phosphorylated the recombinant protein in vitro simultaneously at two target sites. Our results demonstrate that the PKC phosphorylation of HMGB1 has a considerable effect on the fundamental properties of the protein; therefore this post-synthetic modification may serve as a modulator of the HMGB1 participation in different nuclear processes.

In Vitro Anticancer Activity of Two Ferrocene-Containing Camphor Sulfonamides as Promising Agents against Lung Cancer Cells.

The successful design of antitumour drugs often combines in one molecule different biologically active subunits that can affect various regulatory pathways in the cell and thus achieve higher efficacy. Two ferrocene derivatives, DK-164 and CC-78, with different residues were tested for cytotoxic potential on non-small lung cancer cell lines, A549 and H1299, and non-cancerous MRC5. DK-164 demonstrated remarkable selectivity toward cancer cells and more pronounced cytotoxicity against A549. The cytotoxicity of CC-78 toward H1299 was even higher than that of the well-established anticancer drugs cisplatin and tamoxifen, but it did not reveal any noticeable selective effect. DK-164 showed predominantly pro-apoptotic activity in non-small cell lung carcinoma (NSCLC) cells, while CC-78 caused accidental cell death with features characteristic of necrosis. The level of induced autophagy was similar for both substances in cancer cells. DK-164 treatment of A549, H1299, and MRC5 cells for 48 h significantly increased the fluorescence signal of the NFkB (nuclear factor 'kappa-light-chain-enhancer' of activated B-cells) protein in the nucleus in all three cell lines, while CC-78 did not provoke NFkB translocation in any of the tested cell lines. Both compounds caused a significant transfer of the p53 protein in the nucleus of A549 cells but not in non-cancerous MRC5 cells. In A549, DK-164 generated oxidative stress close to the positive control after 48 h, while CC-78 had a moderate effect on the cellular redox status. In the non-cancerous cells, MRC5, both compounds produced ROS similar to the positive control for the same incubation period. The different results related to the cytotoxic potential of DK-164 and CC-78 associated with the examined cellular mechanisms induced in lung cancer cells might be used to conclude the specific functions of the various functional groups in the ferrocene compounds, which can offer new perspectives for the design of antitumour drugs.

In vitro pharmacological study of monomeric platinum(III) hematoporphyrin IX complexes.

Three stable mononuclear hematoporphyrin IX ((7,12-bis(1-hydroxyethyl)-3,8,13,17-tetramethyl-21H-23H-porphyn-2,18-dipropionic acid), Hp) complexes of Pt(III), namely cis-[ Pt(III)(NH(3))(2)(Hp(-3H))(H(2)O)(2)].H(2)O 1, [Pt(III)(Hp(-3H))(H(2)O)(2)].H(2)O 2 and [Pt(III)((O,O)Hp(-2H))Cl(H(2)O)(3)] 3 with distorted octahedral structure and (d(z)2)(1) ground state have been tested in vitro for antineoplastic activity in a panel of tumor cell lines. The novel platinum(III) complexes showed cytotoxic activity in a concentration-dependent manner with IC(50) values comparable to those of referent cytotoxic agent cisplatin together with lower cytotoxicity against renal cells. Further detailed evaluation of the active analogue 2 and the less active complex 3 showed that their potency greatly correlates with the ability to induce apoptosis and to bind DNA. Despite the structural dissimilarities between complex 2 and cisplatin, their DNA-adducts were equally effectively recognized and repaired by the nucleotide excision repair system. Complex 2 showed quite superior ability to accumulate in K-562 cells relative to cisplatin.

Physicochemical Properties and Biological Performance of Polymethacrylate Based Gene Delivery Vector Systems: Influence of Amino Functionalities.

Physicochemical characteristics and biological performance of polyplexes based on two identical copolymers bearing tertiary amino or quaternary ammonium groups are evaluated and compared. Poly(2-(dimethylamino)ethyl methacrylate)-b-poly(oligo(ethylene glycol) methyl ether methacrylate) block copolymer (PDMAEMA-b-POEGMA) is synthesized by reversible addition fragmentation chain transfer polymerization. The tertiary amines of PDMAEMA are converted to quaternary ammonium groups by quaternization with methyl iodide. The two copolymers spontaneously formed well-defined polyplexes with DNA. The size, zeta potential, molar mass, aggregation number, and morphology of the polyplex particles are determined. The parent PDMAEMA-b-POEGMA exhibits larger buffering capacity, whereas the corresponding quaternized copolymer (QPDMAEMA-b-POEGMA) displays stronger binding affinity to DNA, yielding invariably larger in size and molar mass particles bearing greater number of DNA molecules per particle. Experiments revealed that QPDMAEMA-b-POEGMA is more effective in transfecting pEGFP-N1 than the parent copolymer, attributed to the larger size, molar mass, and DNA cargo, as well as to the effective cellular traffic, which dominated over the enhanced ability for endo-lysosomal escape of PDMAEMA-b-POEGMA.