Biology subjects

Medina, P.

Publications and source records attributed to Medina, P..

13 recordsLinked to original sources

bioRxiv2026

When lipids embrace RNA: pH-driven dynamics and mechanisms of LNP-mediated siRNA delivery

We present a comprehensive analysis of an efficient lipid nanoparticle (LNP) formulation that exhibits strong nucleic-acid delivery and potent inhibition of targeted RNAs. Using a combination of coarse-grained and atomistic molecular dynamics simulations, we characterized the pH-dependent structure of both unloaded and RNA-loaded LNPs, elucidated the mechanism of RNA encapsulation, and used these models to propose a plausible mechanism of endosomal escape. Consistent with prior experimental and computational studies, our simulations reproduce an inverted-hexagonal-type morphology in RNA-loaded LNPs, in which a hydrated core provides a polar environment suitable for accommodating RNA, whose charge is neutralized mainly by ionizable lipids that remain protonated near the RNA even at high pH, thereby bridging the RNA with the surrounding lipid environment. This structural picture is consistently observed across our multiscale simulations, with smaller self-assembled LNP mimetics reproducing the same local organization at both coarse-grained and atomistic resolution. A potential mechanism of endosomal escape emerges spontaneously from the simulations, involving stalk formation between the LNP and the endosomal membrane, followed by the opening of a water-filled pore that permits slow RNA diffusion, in line with the low efficiency and slow kinetics reported for endosomal escape. The rate-limiting step of endosomal escape arises from persistent electrostatic coupling between RNA and protonated ionizable lipids maintained by the immediate RNA-lipid environment, hindering cargo disengagement even after pore formation. This delayed release is consistent with the experimentally observed time-dependent inhibitory activity of the loaded LNP. Together, these results highlight the importance of local pK and protonation effects near the RNA, which are not captured by global apparent pK measurements.

biophysics↗

bioRxiv2025

Dually-decorated palmitate-containing lipid nanoparticles for the targeted delivery of siRNAs against HER2 and Hsp27 in HER2+ breast cancer

Ionizable lipid nanoparticles (LNPs) have enabled significant advances in oligonucleotide therapeutics, culminating in the clinical approval of an siRNA-based formulation. Unfortunately, practical use of siRNA containing LNPs is still hampered by difficulties, one of them directing LNPs toward specific cells. To improve cell-specific delivery, we have developed a new generation of palmitate-containing ionizable LNPs functionalized with two distinct ligands: i) a targeting peptide (octreotide, Oct), for selective recognition of somatostatin receptor 2 (SSTR2)-expressing tumor cells, and ii) a cell penetrating peptide (penetratin, RK16) to enhance intracellular delivery. This dually-decorated LNP was used to co-deliver two siRNAs targeting key mediators of drug resistance in HER2+ breast cancer -Hsp27 and HER2- demonstrating strong potential for combinatorial RNAi-based therapies. The Oct moiety conferred high selectivity toward malignant SSTR2-overexpresing (SSTR2OE) HER2+ BC cells, even in heterogeneous environments containing non-tumor epithelial cells. Simultaneously, the RK16 peptide facilitated enhanced intracellular delivery, resulting in improved antiproliferative activity and a notable reduction in the SSTR2-overexpressing cell population compared to non-functionalized LNPs. Taken together, the structural design and biological performance of this novel class of LNP formulation establish it as a versatile and innovative delivery platform for ON-based targeted therapies. By simply altering the targeting peptide moiety and siRNA combination, this dually-functionalized LNP system could be readily adapted for a broad range of tumors and combination treatment strategies. Finally, the inclusion of palmitic acid in the LNP composition selectively contributes to cytotoxicity in palmitate sensitive cell lines, such as SK-BR-3, offering an additional therapeutic advantage when targeting these tumor subtypes.

biochemistry↗

bioRxiv2024

A rare HSC-derived megakaryocyte progenitor accumulates via enhanced survival and contributes to exacerbated thrombopoiesis upon aging

Distinct routes of cellular production from hematopoietic stem cells (HSCs) have defined our current view of hematopoiesis. Recently, we challenged classical views of platelet generation, demonstrating that megakaryocyte progenitors (MkPs), and ultimately platelets, can be specified via an alternate and additive route of HSC-direct specification specifically during aging. This "shortcut" pathway generates hyperactive platelets likely to contribute to age-related platelet-mediated morbidities. Here, we used single-cell RNA/CITEseq to demonstrate that these age-unique, non-canonical (nc)MkPs can be prospectively defined and experimentally isolated from wild type mice. Surprisingly, this revealed that a rare population of ncMkPs also exist in young mice. Young and aged ncMkPs are functionally distinct from their canonical (c)MkP counterparts, with aged ncMkPs paradoxically and uniquely exhibiting enhanced survival and platelet generation capacity. We further demonstrate that aged HSCs generate significantly more ncMkPs than their younger counterparts, yet this is accomplished without strict clonal restriction. Together, these findings reveal significant phenotypic, functional, and aging-dependent heterogeneity among the MkP pool and uncover unique features of megakaryopoiesis throughout life, potentially offering cellular and molecular targets for mitigation of age-related adverse thrombotic events.

cell biology↗

bioRxiv2024

Pregnancy Reduces Il33+ Hybrid Progenitor Accumulation in the Aged Mammary Gland

Aging increases breast cancer risk while an early first pregnancy reduces a womans life-long risk. Several studies have explored the effect of either aging or pregnancy on mammary epithelial cells (MECs), but the combined effect of both remains unclear. Here, we interrogate the functional and transcriptomic changes at single cell resolution in the mammary gland of aged nulliparous and parous mice to discover that pregnancy normalizes age-related imbalances in lineage composition, while also inducing a differentiated cell state. Importantly, we uncover a minority population of Il33-expressing hybrid MECs with high cellular potency that accumulate in aged nulliparous mice but is significantly reduced in aged parous mice. Functionally, IL33 treatment of basal, but not luminal, epithelial cells from young mice phenocopies aged nulliparous MECs and promotes formation of organoids with Trp53 knockdown. Collectively, our study demonstrates that pregnancy blocks the age-associated loss of lineage integrity in the basal layer through a decrease in Il33+ hybrid MECs, potentially contributing to pregnancy-induced breast cancer protection.

developmental biology↗

bioRxiv2022

EZH2 endorses cell plasticity to carcinoma cells facilitating mesenchymal to epithelial transition and tumour colonization

Reversible transition between the epithelial and mesenchymal states are key aspects of carcinoma cell dissemination and the metastatic disease, and thus, characterizing the molecular basis of the epithelial to mesenchymal transition (EMT) is crucial to find druggable targets and more effective therapeutic approaches in cancer. Emerging studies suggest that epigenetic regulators might endorse cancer cells with the cell plasticity required to conduct dynamic changes in cell state during EMT. However, epigenetic mechanisms involved remain mostly unknown. Polycomb Repressive Complexes (PRCs) proteins are well-established epigenetic regulators of development and stem cell differentiation, but their role in different cancer systems is inconsistent and sometimes paradoxical. In this study, we have analysed the role of the PRC2 protein EZH2 in lung carcinoma cells. We found that besides its described role in CDKN2A-dependent cell proliferation, EZH2 upholds the epithelial state of cancer cells by repressing the transcription of hundreds of mesenchymal genes. Chemical inhibition or genetic removal of EZH2 promotes the residence of cancer cells in the mesenchymal state during reversible epithelial-mesenchymal transition. In fitting, analysis of human patient samples and tumour xenograft models indicate that EZH2 is required to efficiently repress mesenchymal genes and facilitate tumour colonization in vivo. Overall, this study discloses a novel role of PRC2 as a master regulator of EMT in carcinoma cells. This finding has important implications for the design of therapies based on EZH2 inhibitors in human cancer patients.

cancer biology↗

bioRxiv2021

Multi-hallmark long noncoding RNA maps reveal non-small cell lung cancer vulnerabilities

Long noncoding RNAs (lncRNAs) are widely dysregulated in cancer, yet their functional roles in cellular disease hallmarks remain unclear. Here we employ pooled CRISPR deletion to perturb all 831 lncRNAs in KRAS-mutant non-small cell lung cancer (NSCLC), and measure their contribution to proliferation, chemoresistance and migration across two cell backgrounds. Integrative analysis of this data outperforms conventional "dropout" screens in identifying cancer genes, while prioritising disease-relevant lncRNAs with pleiotropic and background-independent roles. Altogether 60 high-confidence oncogenic lncRNAs are active in NSCLC, the majority identified here for the first time, and which tend to be amplified and overexpressed in tumours. A follow-up antisense oligonucleotide (ASO) screen shortlisted two candidates, Cancer Hallmarks in Lung LncRNA (CHiLL 1&2), whose knockdown consistently suppressed cancer hallmarks in a variety of 2D and 3D tumour models. Molecular phenotyping reveals that CHiLL 1&2 control cellular-level phenotypes via distinct transcriptional networks converging on common oncogenic pathways. In summary, this work reveals a multi-dimensional functional lncRNA landscape underlying NSCLC that contains potential therapeutic vulnerabilities.

genomics↗

bioRxiv2020

1--4-phenylpiperazine Treatment After Brain Irradiation Preserves Cognitive Function in Mice

BackgroundNormal tissue toxicity to the CNS is an inevitable consequence of a successful radiotherapy of brain tumors or cancer metastases to the CNS. Cranial irradiation commonly leads to neurocognitive deficits that manifest months or years after treatment. Mechanistically, radiation-induced loss of neural stem/progenitor cells, neuro-inflammation and de-myelinization are contributing factors that lead to progressive cognitive decline. MethodsThe effects of Compound #5 on irradiated murine neurospheres, microglia cells and patients-derived gliomaspheres were assessed in sphere-formation assays, flow cytometry and IL-6 ELISAs, Activation of the Hedgehog pathway was studied by qRT-PCR. The in vivo effects of Compound #5 were analyzed using flow cytometry, sphere-formation assays, immune-histochemistry, behavioral testing and an intracranial mouse model of glioblastoma. ResultsWe report that 1-[(4-Nitrophenyl)sulfonyl]-4-phenylpiperazine (Compound #5) mitigates radiation-induced normal tissue toxicity in the brains of mice. Compound #5 treatment significantly increased the number of neural stem/progenitor cells after brain irradiation in female animals, inhibited radiation-induced microglia activation and expression of the pro-inflammatory cytokine interleukin-6. Behavioral testing revealed that treatment with Compound #5 after radiotherapy successfully mitigates radiation-induced decline in motor, sensory and memory function of the brain. In mouse models of glioblastoma, Compound #5 showed no toxicity and did not interfere with the growth-delaying effects of radiation. ConclusionsWe conclude that Compound #5 has the potential to mitigate cognitive decline in patients undergoing partial or whole brain irradiation without promoting tumor growth and that the use of this compound as a radiation mitigator of radiation late effects on the CNS warrants further investigation. Importance of the StudySuccessful radiotherapy of CNS malignancies inevitably lead to cognitive decline in cancer survivors and treatment options to mitigate this side effect are limited. We present evidence that a piperazine compound can prevent cognitive decline in mice after total brain irradiation without compromising the antitumor effect of radiation, suggesting that this compound could be used to mitigate radiation side effects in brain tumor patients undergoing radiotherapy.

neuroscience↗

bioRxiv2019

Genomic adaptations to aquatic and aerial life in mayflies and the origin of wings in insects

The first winged insects underwent profound morphological and functional transformations leading to the most successful animal radiations in the history of earth. Despite this, we still have a very incomplete picture of the changes in their genomes that underlay this radiation. Mayflies (Ephemeroptera) are one of the extant sister groups of all other winged insects and therefore are at a key phylogenetic position to understand this radiation. Here, we describe the genome of the cosmopolitan mayfly Cloeon dipterum and study its expression along development and in specific organs. We discover an expansion of odorant-binding proteins, some expressed specifically in the breathing gills of aquatic nymphs, suggesting a novel sensory role for gills. In contrast, as flying adults, mayflies make use of an enlarged set of opsins and utilise these visual genes in a sexually dimorphic manner, with some opsins expressed only in males. Finally, to illuminate the origin of wings, we identify a core set of deeply conserved wing-specific genes at the root of the pterygote insects. Globally, this is the first comprehensive study of the structure and expression of the genome of a paleopteran insect and shows how its genome has kept a record of its functional adaptations.

evolutionary biology↗

bioRxiv2019

The Dopamine Receptor Antagonist TFP Prevents Phenotype Conversion and Improves Survival in Mouse Models of Glioblastoma

Glioblastoma (GBM) is the deadliest adult brain cancer and all patients ultimately succumb to the disease. Radiation therapy (RT) provides survival benefit of 6 months over surgery alone but these results have not improved in decades. We report that radiation induces a glioma-initiating cell phenotype and we have identified trifluoperazine (TFP) as a compound that interferes with this phenotype conversion. TFP caused loss of radiation-induced Nanog mRNA expression, activation of GSK3 with consecutive post-translational reduction in p-Akt, Sox2 and {beta}-catenin protein levels. TFP did not alter the intrinsic radiation sensitivity of glioma-initiating cells (GICs). Continuous treatment with TFP and a single dose of radiation reduced the number of GICs in vivo and prolonged survival in syngeneic and patient-derived orthotopic xenograft (PDOX) mouse models of GBM. Our findings suggest that combination of a dopamine receptor antagonist with radiation enhances the efficacy of RT in GBM by preventing radiation-induced phenotype conversion of radiosensitive non-GICs into treatment resistant, induced GICs. SignificanceGBM is the most common and most deadly adult brain cancer. The current standard-of-care is surgery followed by RT and temozolomide, which results in a median survival time of only 15 months. The efficacy of chemotherapies and targeted therapies in GBM is very limited because most of these drugs do not pass the blood-brain-barrier. Ultimately, all patients succumb to the disease. Our study describes radiation-induced cellular plasticity as a novel resistance mechanism in GBM. We identified a dopamine receptor antagonist as a readily available, FDA-approved drug known to penetrate the blood-brain-barrier which prevents phenotype conversion of glioma cells into glioma-initiating cells and prologs survival in mouse models of GBM, thus suggesting that it will improve the efficacy of RT without increasing toxicity.

neuroscience↗

bioRxiv2019

On the distribution of tract lengths during adaptive introgression

Admixture is increasingly being recognized as an important factor in evolutionary genetics. The distribution of genomic admixture tracts, and the resulting effects on admixture linkage disequilibrium, can be used to date the timing of admixture between species or populations. However, the theory used for such prediction assumes selective neutrality despite the fact that many famous examples of admixture involve natural selection acting for or against admixture. In this paper, we investigate the effects of positive selection on the distribution of tract lengths. We develop a theoretical framework that relies on approximating the trajectory of the selected allele using a logistic function. By numerically calculating the expected allele trajectory, we also show that the approach can be extended to cases where the logistic approximation is poor due to the effects of genetic drift. Using simulations, we show that the model is highly accurate under most scenarios. We use the model to show that positive selection on average will tend to increase the admixture tract length. However, perhaps counter-intuitively, conditional on the allele frequency at the time of sampling, positive selection will actually produce shorter expected tract lengths. We discuss the consequences of our results in interpreting the timing of the introgression of EPAS1 from Denisovans into the ancestors of Tibetans.

genomics↗

bioRxiv2019

Deep data mining reveals variable abundance and distribution of microbial reproductive manipulators within and among diverse host species

Bacterial symbionts that manipulate the reproduction of their hosts are important factors in invertebrate ecology and evolution. Studying the genomic and phenotypic diversity of reproductive manipulators can improve efforts to control infectious diseases and contribute to our understanding of host-symbiont evolution. Despite the vast genomic and phenotypic diversity of reproductive manipulators, only a handful of strains are used as biological control agents because little is known about the broad scale infection frequencies and densities of these bacteria in nature. Here we develop a data mining approach to quantify the number of arthropod and nematode host species infected with Wolbachia and other reproductive manipulators such as Rickettsia and Spiroplasma. Across the entire Sequence Read Archive (SRA) database, we found reproductive manipulators infected 2,083 arthropod and 119 nematode samples, representing 240 and 8 species, respectively. After accounting for sampling and infection frequency differences among species, we estimated that Wolbachia infects approximately 44% of all arthropod species and 34% of all nematode species. In contrast, we estimated other reproductive manipulators infect 1-8% of arthropod and nematode species. Next, we explored another important biological parameter: the relative bacterial density, or titer, within hosts. We found variation in titer within and between arthropod species to be large, and that host species explains approximately 36% of variation in titer across our dataset. This suggests bacterial strain and/or host species plays a role in shaping bacterial densities within and between host species. By leveraging the model system Drosophila melanogaster, we also found a number of host SNPs associated with titer in genes potentially relevant to host interactions with Wolbachia, suggesting bacterial induced host genome evolution. Our study demonstrates that data mining is a powerful tool to understand host-symbiont co-evolution and opens an array of previously inaccessible questions for further analysis.

evolutionary biology↗

bioRxiv2018

Estimating the timing of multiple admixture pulses during local ancestry inference.

Admixture, the mixing of genetically distinct populations, is increasingly recognized as a fundamental biological process. One major goal of admixture analyses is to estimate the timing of admixture events. Whereas most methods today can only detect the most recent admixture event, here we present coalescent theory and associated software that can be used to estimate the timing of multiple admixture events in an admixed population. We extensively validate this approach and evaluate the conditions under which it can succesfully distinguish one from two-pulse admixture models. We apply our approach to real and simulated data of Drosophila melanogaster. We find evidence of a single very recent pulse of cosmopolitan ancestry contributing to African populations as well as evidence for more ancient admixture among genetically differentiated populations in sub-Saharan Africa. These results suggest our method can quantify complex admixture histories involving genetic material introduced by multiple discrete admixture pulses. The new method facilitates the exploration of admixture and its contribution to adaptation, ecological divergence, and speciation.

evolutionary biology↗

bioRxiv2017

Genetic mapping of species differences via "in vitro crosses" in mouse embryonic stem cells

Discovering the genetic changes underlying species differences is a central goal in evolutionary genetics. However, hybrid crosses between species in mammals often suffer from hybrid sterility, greatly complicating genetic dissection of trait variation. Here we describe a simple, robust and transgene-free technique to make \"in vitro crosses\" in hybrid mouse embryonic stem cells by inducing random mitotic crossovers with the drug ML216, which inhibits Bloom syndrome (BLM). Starting with an interspecific hybrid (between Mus musculus and Mus spretus) embryonic stem cell line spanning 1.5 million years of divergence, we demonstrate the feasibility of mapping enzymatic differences across species within weeks and the possibility of re-deriving whole mice. Our work shows how in vitro crosses can overcome major bottlenecks like hybrid sterility in traditional mouse breeding to address fundamental questions in evolutionary biology.\n\nImpact StatementBy mixing hybrid mouse genomes in stem cells via mitotic recombination, genetic mapping and hybrid mosaic mice can be achieved in weeks, even across species barriers.

genetics↗