Therapeutic targeting Tudor domains in leukemia via CRISPR-Scan Assisted Drug Discovery.

Epigenetic dysregulation has been reported in multiple cancers including leukemias. Nonetheless, the roles of the epigenetic reader Tudor domains in leukemia progression and therapy remain unexplored. Here, we conducted a Tudor domain-focused CRISPR screen and identified SGF29, a component of SAGA/ATAC acetyltransferase complexes, as a crucial factor for H3K9 acetylation, ribosomal gene expression, and leukemogenesis. To facilitate drug development, we integrated the CRISPR tiling scan with compound docking and molecular dynamics simulation, presenting a generally applicable strategy called CRISPR-Scan Assisted Drug Discovery (CRISPR-SADD). Using this approach, we identified a lead inhibitor that selectively targets SGF29's Tudor domain and demonstrates efficacy against leukemia. Furthermore, we propose that the structural genetics approach used in our study can be widely applied to diverse fields for de novo drug discovery.

METTL16 promotes liver cancer stem cell self-renewal via controlling ribosome biogenesis and mRNA translation.

BACKGROUND: While liver cancer stem cells (CSCs) play a crucial role in hepatocellular carcinoma (HCC) initiation, progression, recurrence, and treatment resistance, the mechanism underlying liver CSC self-renewal remains elusive. We aim to characterize the role of Methyltransferase 16 (METTL16), a recently identified RNA N6-methyladenosine (m6A) methyltransferase, in HCC development/maintenance, CSC stemness, as well as normal hepatogenesis. METHODS: Liver-specific Mettl16 conditional KO (cKO) mice were generated to assess its role in HCC pathogenesis and normal hepatogenesis. Hydrodynamic tail-vein injection (HDTVi)-induced de novo hepatocarcinogenesis and xenograft models were utilized to determine the role of METTL16 in HCC initiation and progression. A limiting dilution assay was utilized to evaluate CSC frequency. Functionally essential targets were revealed via integrative analysis of multi-omics data, including RNA-seq, RNA immunoprecipitation (RIP)-seq, and ribosome profiling. RESULTS: METTL16 is highly expressed in liver CSCs and its depletion dramatically decreased CSC frequency in vitro and in vivo. Mettl16 KO significantly attenuated HCC initiation and progression, yet only slightly influenced normal hepatogenesis. Mechanistic studies, including high-throughput sequencing, unveiled METTL16 as a key regulator of ribosomal RNA (rRNA) maturation and mRNA translation and identified eukaryotic translation initiation factor 3 subunit a (eIF3a) transcript as a bona-fide target of METTL16 in HCC. In addition, the functionally essential regions of METTL16 were revealed by CRISPR gene tiling scan, which will pave the way for the development of potential inhibitor(s). CONCLUSIONS: Our findings highlight the crucial oncogenic role of METTL16 in promoting HCC pathogenesis and enhancing liver CSC self-renewal through augmenting mRNA translation efficiency.

A novel class of inhibitors that disrupts the stability of integrin heterodimers identified by CRISPR-tiling-instructed genetic screens.

The plasma membrane is enriched for receptors and signaling proteins that are accessible from the extracellular space for pharmacological intervention. Here we conducted a series of CRISPR screens using human cell surface proteome and integrin family libraries in multiple cancer models. Our results identified ITGAV (integrin αV) and its heterodimer partner ITGB5 (integrin ß5) as the essential integrin α/ß pair for cancer cell expansion. High-density CRISPR gene tiling further pinpointed the integral pocket within the ß-propeller domain of ITGAV for integrin αVß5 dimerization. Combined with in silico compound docking, we developed a CRISPR-Tiling-Instructed Computer-Aided (CRISPR-TICA) pipeline for drug discovery and identified Cpd_AV2 as a lead inhibitor targeting the ß-propeller central pocket of ITGAV. Cpd_AV2 treatment led to rapid uncoupling of integrin αVß5 and cellular apoptosis, providing a unique class of therapeutic action that eliminates the integrin signaling via heterodimer dissociation. We also foresee the CRISPR-TICA approach to be an accessible method for future drug discovery studies.

Hematologic DNMT3A reduction and high-fat diet synergize to promote weight gain and tissue inflammation.

During aging, blood cell production becomes dominated by a limited number of variant hematopoietic stem cell (HSC) clones. Differentiated progeny of variant HSCs are thought to mediate the detrimental effects of such clonal hematopoiesis on organismal health, but the mechanisms are poorly understood. While somatic mutations in DNA methyltransferase 3A (DNMT3A) frequently drive clonal dominance, the aging milieu also likely contributes. Here, we examined in mice the interaction between high-fat diet (HFD) and reduced DNMT3A in hematopoietic cells; strikingly, this combination led to weight gain. HFD amplified pro-inflammatory pathways and upregulated inflammation-associated genes in mutant cells along a pro-myeloid trajectory. Aberrant DNA methylation during myeloid differentiation and in response to HFD led to pro-inflammatory activation and maintenance of stemness genes. These findings suggest that reduced DNMT3A in hematopoietic cells contributes to weight gain, inflammation, and metabolic dysfunction, highlighting a role for DNMT3A loss in the development of metabolic disorders.

SOD1 is a synthetic lethal target in PPM1D-mutant leukemia cells.

The DNA damage response is critical for maintaining genome integrity and is commonly disrupted in the development of cancer. PPM1D (protein phosphatase, Mg2+/Mn2+ dependent 1D) is a master negative regulator of the response; gain-of-function mutations and amplifications of PPM1D are found across several human cancers making it a relevant pharmacologic target. Here, we used CRISPR/Cas9 screening to identify synthetic-lethal dependencies of PPM1D, uncovering superoxide dismutase-1 (SOD1) as a potential target for PPM1D-mutant cells. We revealed a dysregulated redox landscape characterized by elevated levels of reactive oxygen species and a compromised response to oxidative stress in PPM1D-mutant cells. Altogether, our results demonstrate the protective role of SOD1 against oxidative stress in PPM1D-mutant leukemia cells and highlight a new potential therapeutic strategy against PPM1D-mutant cancers.

RNA m6A reader YTHDF2 facilitates precursor miR-126 maturation to promote acute myeloid leukemia progression.

As the most common internal modification of mRNA, N6-methyladenosine (m6A) and its regulators modulate gene expression and play critical roles in various biological and pathological processes including tumorigenesis. It was reported previously that m6A methyltransferase (writer), methyltransferase-like 3 (METTL3) adds m6A in primary microRNAs (pri-miRNAs) and facilitates its processing into precursor miRNAs (pre-miRNAs). However, it is unknown whether m6A modification also plays a role in the maturation process of pre-miRNAs and (if so) whether such a function contributes to tumorigenesis. Here, we found that YTHDF2 is aberrantly overexpressed in acute myeloid leukemia (AML) patients, especially in relapsed patients, and plays an oncogenic role in AML. Moreover, YTHDF2 promotes expression of miR-126-3p (also known as miR-126, as it is the main product of precursor miR-126 (pre-miR-126)), a miRNA that was reported as an oncomiRNA in AML, through facilitating the processing of pre-miR-126 into mature miR-126. Mechanistically, YTHDF2 recognizes m6A modification in pre-miR-126 and recruits AGO2, a regulator of pre-miRNA processing, to promote the maturation of pre-miR-126. YTHDF2 positively and negatively correlates with miR-126 and miR-126's downstream target genes, respectively, in AML patients, and forced expression of miR-126 could largely rescue YTHDF2/Ythdf2 depletion-mediated suppression on AML cell growth/proliferation and leukemogenesis, indicating that miR-126 is a functionally important target of YTHDF2 in AML. Overall, our studies not only reveal a previously unappreciated YTHDF2/miR-126 axis in AML and highlight the therapeutic potential of targeting this axis for AML treatment, but also suggest that m6A plays a role in pre-miRNA processing that contributes to tumorigenesis.

Association between estimated pulse wave velocity and all-cause death in patients with heart failure.

BACKGROUND: Arteriosclerosis has been proven to be a risk factor for the development of heart failure and readmission. ePWV is a novel non-invasive and simple indicator of arterial stiffness, and this study aims to investigate its relationship with all-cause mortality rate in patients with heart failure. METHODS: This study is a cohort study that included 1272 patients with heart failure from NHANES data from 1999 to 2018. The ePWV was divided into three groups, and the cumulative mortality rate of heart failure patients was calculated using KM survival curves. The relationship between ePWV and all-cause mortality rate in heart failure patients was represented by a smoothed curve fitting. COX regression analysis was used to assess the association between ePWV and all-cause mortality rate in heart failure patients. RESULTS: The average age of the study population was 67.8 ± 12.6 years, with 862 males and 650 females. During the 12-month follow-up period, there were 790 cases of all-cause mortality. Cox regression analysis was used to validate the relationship between ePWV and all-cause mortality rate in patients with heart failure. Patients with higher levels of ePWV tended to have a higher all-cause mortality rate. After adjustment for multiple factors, an increase in ePWV was positively associated with all-cause mortality rate (HR = 1.17, 95% confidence interval (CI): (1.12, 1.22)). Compared to the lowest tertile, the multivariable-adjusted HR and 95% CI for the highest tertile of ePWV were 1.81 (95% CI: (1.45, 2.27)). Additionally, a smoothed curve fitting was used to observe the relationship between ePWV and mortality rate, where the curve demonstrated a positive correlation between ePWV and all-cause mortality rate. Furthermore, KM survival curves indicated that all-cause mortality rate increased with the increase in ePWV. Subgroup analysis suggested a correlation between ePWV and mortality rate. CONCLUSION: Our study shows that ePWV is positively associated with all-cause mortality in patients with heart failure.

Single-Layer Hexagonal Boron Nitride Nanopores as High-Performance Ionic Gradient Power Generators.

Atomically thin two-dimensional (2D) materials have emerged as promising candidates for efficient energy harvesting from ionic gradients. However, the exploration of robust 2D atomically thin nanopore membranes, which hold sufficient ionic selectivity and high ion permeability, remains challenging. Here, the single-layer hexagonal boron nitride (hBN) nanopores are demonstrated as various high-performance ion-gradient nanopower harvesters. Benefiting from the ultrathin atomic thickness and large surface charge (also a large Dukhin number), the hBN nanopore can realize fast proton transport while maintaining excellent cation selectivity even in highly acidic environments. Therefore, a single hBN nanopore achieves the pure osmosis-driven proton-gradient power up to ≈3 nW under 1000-fold ionic gradient. In addition, the robustness of hBN membranes in extreme pH conditions allows the ionic gradient power generation from acid-base neutralization. Utilizing 1 m HCl/KOH, the generated power can be promoted to an extraordinarily high level of ≈4.5 nW, over one magnitude higher than all existing ionic gradient power generators. The synergistic effects of ultrathin thickness, large surface charge, and excellent chemical inertness of 2D single-layer hBN render it a promising membrane candidate for harvesting ionic gradient powers, even under extreme pH conditions.

Mitigating stimulated Brillouin scattering in multimode fibers with focused output via wavefront shaping.

The key challenge for high-power delivery through optical fibers is overcoming nonlinear optical effects. To keep a smooth output beam, most techniques for mitigating optical nonlinearities are restricted to single-mode fibers. Moving out of the single-mode paradigm, we show experimentally that wavefront-shaping of coherent input light to a highly multimode fiber can increase the power threshold for stimulated Brillouin scattering (SBS) by an order of magnitude, whilst simultaneously controlling the output beam profile. The SBS suppression results from an effective broadening of the Brillouin spectrum under multimode excitation, without broadening of transmitted light. Strongest suppression is achieved with selective mode excitation that gives the broadest Brillouin spectrum. Our method is efficient, robust, and applicable to continuous waves and pulses. This work points toward a promising route for mitigating detrimental nonlinear effects in optical fibers, enabling further power scaling of high-power fiber systems for applications to directed energy, remote sensing, and gravitational-wave detection.

SRCAP mutations drive clonal hematopoiesis through epigenetic and DNA repair dysregulation.

Somatic mutations accumulate in all cells with age and can confer a selective advantage, leading to clonal expansion over time. In hematopoietic cells, mutations in a subset of genes regulating DNA repair or epigenetics frequently lead to clonal hematopoiesis (CH). Here, we describe the context and mechanisms that lead to enrichment of hematopoietic stem cells (HSCs) with mutations in SRCAP, which encodes a chromatin remodeler that also influences DNA repair. We show that SRCAP mutations confer a selective advantage in human cells and in mice upon treatment with the anthracycline-class chemotherapeutic doxorubicin and bone marrow transplantation. Furthermore, Srcap mutations lead to a lymphoid-biased expansion, driven by loss of SRCAP-regulated H2A.Z deposition and increased DNA repair. Altogether, we demonstrate that SRCAP operates at the intersection of multiple pathways in stem and progenitor cells, offering a new perspective on the functional impact of genetic variants that promote stem cell competition in the hematopoietic system.

Near-Infrared Imaging of Colonic Adenomas In Vivo Using Orthotopic Human Organoids for Early Cancer Detection.

Colorectal cancer is a leading cause of cancer-related morbidity and mortality worldwide. Premalignant lesions that are flat and subtle in morphology are often missed in conventional colonoscopies. Patient-derived adenoma colonoids with high and low cMet expression and normal colonoids were implanted orthotopically in the colon of immunocompromised mice to serve as a preclinical model system. A peptide specific for cMet was labeled with IRDye800, a near-infrared (NIR) fluorophore. This peptide was administered intravenously, and in vivo imaging was performed using a small animal fluorescence endoscope. Quantified intensities showed a peak target-to-background ratio at ~1 h after intravenous peptide injection, and the signal cleared by ~24 h. The peptide was stable in serum with a half-life of 3.6 h. Co-staining of adenoma and normal colonoids showed a high correlation between peptide and anti-cMet antibody. A human-specific cytokeratin stain verified the presence of human tissues implanted among surrounding normal mouse colonic mucosa. Peptide biodistribution was consistent with rapid renal clearance. No signs of acute toxicity were found on either animal necropsy or serum hematology and chemistries. Human colonoids provide a clinically relevant preclinical model to evaluate the specific uptake of a NIR peptide to detect premalignant colonic lesions in vivo.

Diverse Roles of Protein Palmitoylation in Cancer Progression, Immunity, Stemness, and Beyond.

Protein S-palmitoylation, a type of post-translational modification, refers to the reversible process of attachment of a fatty acyl chain-a 16-carbon palmitate acid-to the specific cysteine residues on target proteins. By adding the lipid chain to proteins, it increases the hydrophobicity of proteins and modulates protein stability, interaction with effector proteins, subcellular localization, and membrane trafficking. Palmitoylation is catalyzed by a group of zinc finger DHHC-containing proteins (ZDHHCs), whereas depalmitoylation is catalyzed by a family of acyl-protein thioesterases. Increasing numbers of oncoproteins and tumor suppressors have been identified to be palmitoylated, and palmitoylation is essential for their functions. Understanding how palmitoylation influences the function of individual proteins, the physiological roles of palmitoylation, and how dysregulated palmitoylation leads to pathological consequences are important drivers of current research in this research field. Further, due to the critical roles in modifying functions of oncoproteins and tumor suppressors, targeting palmitoylation has been used as a candidate therapeutic strategy for cancer treatment. Here, based on recent literatures, we discuss the progress of investigating roles of palmitoylation in regulating cancer progression, immune responses against cancer, and cancer stem cell properties.

The NCOR-HDAC3 co-repressive complex modulates the leukemogenic potential of the transcription factor ERG.

The ERG (ETS-related gene) transcription factor is linked to various types of cancer, including leukemia. However, the specific ERG domains and co-factors contributing to leukemogenesis are poorly understood. Drug targeting a transcription factor such as ERG is challenging. Our study reveals the critical role of a conserved amino acid, proline, at position 199, located at the 3' end of the PNT (pointed) domain, in ERG's ability to induce leukemia. P199 is necessary for ERG to promote self-renewal, prevent myeloid differentiation in hematopoietic progenitor cells, and initiate leukemia in mouse models. Here we show that P199 facilitates ERG's interaction with the NCoR-HDAC3 co-repressor complex. Inhibiting HDAC3 reduces the growth of ERG-dependent leukemic and prostate cancer cells, indicating that the interaction between ERG and the NCoR-HDAC3 co-repressor complex is crucial for its oncogenic activity. Thus, targeting this interaction may offer a potential therapeutic intervention.

TP63 fusions drive multicomplex enhancer rewiring, lymphomagenesis, and EZH2 dependence.

Gene fusions involving tumor protein p63 gene (TP63) occur in multiple T and B cell lymphomas and portend a dismal prognosis for patients. The function and mechanisms of TP63 fusions remain unclear, and there is no target therapy for patients with lymphoma harboring TP63 fusions. Here, we show that TP63 fusions act as bona fide oncogenes and are essential for fusion-positive lymphomas. Transgenic mice expressing TBL1XR1::TP63, the most common TP63 fusion, develop diverse lymphomas that recapitulate multiple human T and B cell lymphomas. Here, we identify that TP63 fusions coordinate the recruitment of two epigenetic modifying complexes, the nuclear receptor corepressor (NCoR)-histone deacetylase 3 (HDAC3) by the N-terminal TP63 fusion partner and the lysine methyltransferase 2D (KMT2D) by the C-terminal TP63 component, which are both required for fusion-dependent survival. TBL1XR1::TP63 localization at enhancers drives a unique cell state that involves up-regulation of MYC and the polycomb repressor complex 2 (PRC2) components EED and EZH2. Inhibiting EZH2 with the therapeutic agent valemetostat is highly effective at treating transgenic lymphoma murine models, xenografts, and patient-derived xenografts harboring TP63 fusions. One patient with TP63-rearranged lymphoma showed a rapid response to valemetostat treatment. In summary, TP63 fusions link partner components that, together, coordinate multiple epigenetic complexes, resulting in therapeutic vulnerability to EZH2 inhibition.

Bioenergetic alteration in gastrointestinal cancers: The good, the bad and the ugly.

Cancer cells exhibit metabolic reprogramming and bioenergetic alteration, utilizing glucose fermentation for energy production, known as the Warburg effect. However, there are a lack of comprehensive reviews summarizing the metabolic reprogramming, bioenergetic alteration, and their oncogenetic links in gastrointestinal (GI) cancers. Furthermore, the efficacy and treatment potential of emerging anticancer drugs targeting these alterations in GI cancers require further evaluation. This review highlights the interplay between aerobic glycolysis, the tricarboxylic acid (TCA) cycle, and oxidative phosphorylation (OXPHOS) in cancer cells, as well as hypotheses on the molecular mechanisms that trigger this alteration. The role of hypoxia-inducible transcription factors, tumor suppressors, and the oncogenetic link between hypoxia-related enzymes, bioenergetic changes, and GI cancer are also discussed. This review emphasizes the potential of targeting bioenergetic regulators for anti-cancer therapy, particularly for GI cancers. Emphasizing the potential of targeting bioenergetic regulators for GI cancer therapy, the review categorizes these regulators into aerobic glycolysis/ lactate biosynthesis/transportation and TCA cycle/coupled OXPHOS. We also detail various anti-cancer drugs and strategies that have produced pre-clinical and/or clinical evidence in treating GI cancers, as well as the challenges posed by these drugs. Here we highlight that understanding dysregulated cancer cell bioenergetics is critical for effective treatments, although the diverse metabolic patterns present challenges for targeted therapies. Further research is needed to comprehend the specific mechanisms of inhibiting bioenergetic enzymes, address side effects, and leverage high-throughput multi-omics and spatial omics to gain insights into cancer cell heterogeneity for targeted bioenergetic therapies.

Chiroptical Response Inversion and Enhancement of Room-Temperature Exciton-Polaritons Using 2D Chirality in Perovskites.

Although chiral semiconductors have shown promising progress in direct circularly polarized light (CPL) detection and emission, they still face potential challenges. A chirality-switching mechanism or approach integrating two enantiomers is needed to discriminate the handedness of a given CPL; additionally, a large material volume is required for sufficient chiroptical interaction. These two requirements pose significant obstacles to the simplification and miniaturization of the devices. Here, room-temperature chiral polaritons fulfilling dual-handedness functions and exhibiting a more-than-two-order enhancement of the chiroptical signal are demonstrated, by embedding a 40 nm-thick perovskite film with a 2D chiroptical effect into a Fabry-Pérot cavity. By mixing chiral perovskites with different crystal structures, a pronounced 2D chiroptical effect is accomplished in the perovskite film, featured by an inverted chiroptical response for counter-propagating CPL. This inversion behavior matches the photonic handedness switch during CPL circulation in the Fabry-Pérot cavity, thus harvesting giant enhancement of the chiroptical response. Furthermore, affected by the unique quarter-wave-plate effects, the polariton emission achieves a chiral dissymmetry of ±4% (for the emission from the front and the back sides). The room-temperature polaritons with the strong dissymmetric chiroptical interaction shall have implications on a fundamental level and future on-chip applications for biomolecule analysis and quantum computing.

MLL3 regulates the CDKN2A tumor suppressor locus in liver cancer.

Mutations in genes encoding components of chromatin modifying and remodeling complexes are among the most frequently observed somatic events in human cancers. For example, missense and nonsense mutations targeting the mixed lineage leukemia family member 3 (MLL3, encoded by KMT2C) histone methyltransferase occur in a range of solid tumors, and heterozygous deletions encompassing KMT2C occur in a subset of aggressive leukemias. Although MLL3 loss can promote tumorigenesis in mice, the molecular targets and biological processes by which MLL3 suppresses tumorigenesis remain poorly characterized. Here, we combined genetic, epigenomic, and animal modeling approaches to demonstrate that one of the mechanisms by which MLL3 links chromatin remodeling to tumor suppression is by co-activating the Cdkn2a tumor suppressor locus. Disruption of Kmt2c cooperates with Myc overexpression in the development of murine hepatocellular carcinoma (HCC), in which MLL3 binding to the Cdkn2a locus is blunted, resulting in reduced H3K4 methylation and low expression levels of the locus-encoded tumor suppressors p16/Ink4a and p19/Arf. Conversely, elevated KMT2C expression increases its binding to the CDKN2A locus and co-activates gene transcription. Endogenous Kmt2c restoration reverses these chromatin and transcriptional effects and triggers Ink4a/Arf-dependent apoptosis. Underscoring the human relevance of this epistasis, we found that genomic alterations in KMT2C and CDKN2A were associated with similar transcriptional profiles in human HCC samples. These results collectively point to a new mechanism for disrupting CDKN2A activity during cancer development and, in doing so, link MLL3 to an established tumor suppressor network.

Suppressing transverse mode instability through multimode excitation in a fiber amplifier.

High-power fiber laser amplifiers have enabled an increasing range of applications in industry, science, and defense. The power scaling for fiber amplifiers is currently limited by transverse mode instability. Most techniques for suppressing the instability are based on single- or few-mode fibers in order to output a clean collimated beam. Here, we study theoretically using a highly multimode fiber amplifier with many-mode excitation for efficient suppression of thermo-optical nonlinearity and instability. We find that the mismatch of characteristic length scales between temperature and optical intensity variations across the fiber generically leads to weaker thermo-optical coupling between fiber modes. Consequently, the transverse mode instability (TMI) threshold power increases linearly with the number of equally excited modes. When the frequency bandwidth of a coherent seed laser is narrower than the spectral correlation width of the multimode fiber, the amplified light maintains high spatial coherence and can be transformed to any target pattern or focused to a diffraction-limited spot by a spatial mask at either the input or output end of the amplifier. Our method simultaneously achieves high average power, narrow spectral width, and good beam quality, which are required for fiber amplifiers in various applications.

Chimeric Human Papillomavirus-16 Virus-like Particles Presenting HIV-1 P18I10 Peptide: Expression, Purification, Bio-Physical Properties and Immunogenicity in BALB/c Mice.

Human papillomavirus (HPV) vaccines based on HPV L1 virus-like particles (VLPs) are already licensed but not accessible worldwide. About 38.0 million people were living with HIV in 2020 and there is no HIV vaccine yet. Therefore, safe, effective, and affordable vaccines against both viruses are an urgent need. In this study, the HIV-1 P18I10 CTL peptide from the V3 loop of HIV-1 gp120 glycoprotein was inserted into the HPV16 L1 protein to construct chimeric HPV:HIV (L1:P18I10) VLPs. Instead of the traditional baculovirus expression vector/insect cell (BEVS/IC) system, we established an alternative mammalian 293F cell-based expression system using cost-effective polyethylenimine-mediated transfection for L1:P18I10 protein production. Compared with conventional ultracentrifugation, we optimized a novel chromatographic purification method which could significantly increase L1:P18I10 VLP recovery (~56%). Chimeric L1:P18I10 VLPs purified from both methods were capable of self-assembling to integral particles and shared similar biophysical and morphological properties. After BALB/c mice immunization with 293F cell-derived and chromatography-purified L1:P18I10 VLPs, almost the same titer of anti-L1 IgG (p = 0.6409) was observed as Gardasil anti-HPV vaccine-immunized mice. Significant titers of anti-P18I10 binding antibodies (p < 0.01%) and P18I10-specific IFN-γ secreting splenocytes (p = 0.0002) were detected in L1:P18I10 VLP-immunized mice in comparison with licensed Gardasil-9 HPV vaccine. Furthermore, we demonstrated that insertion of HIV-1 P18I10 peptide into HPV16 L1 capsid protein did not affect the induction in anti-L1 antibodies. All in all, we expected that the mammalian cell expression system and chromatographic purification methods could be time-saving, cost-effective, scalable platforms to engineer bivalent VLP-based vaccines against HPV and HIV-1.