Journal: bioRxiv

Article Title: Targeting Langerhans cells using a modular mannosylated nucleic acid-based vaccine platform

doi: 10.1101/2025.06.04.657560

Figure Lengend Snippet: Delivery of gp100 on mannosylated HJ scaffolds into moLCs for antigen presentation. A ) Reaction scheme for gp100-functionalization of Q2 and Q4 with NHS-DBCO ( 1 ), followed by SPAAC reaction to azide-modified ( pink ) Val-Cit-PAB-linked ( yellow) gp100 ( red ) ( 2 ). B ) Schematic representation of gp100-conjugated HJs employed for antigen presentation. C ) Visual representation of the antigen-presentation assay using moLCs incubated with gp100-conjugated HJs to facilitate receptor-mediated endocytosis ( I ), followed by release of gp100 in the lysosomes ( II ) and presentation of gp100 on HLA-A2 ( III ). Subsequently, moLCs were co-cultured with wildtype- or gp100 Jurkat cells to verify antigen presentation by T cell activation. D ) IL-2 concentration (pg/mL) in the co-culture media by ELISA. Results shown from four independent human donors were expressed by the mean ± SD. Statistical analysis was conducted by a one-way ANOVA followed by a Tukey’s post hoc test. E ) Antigen-presentation assay comparing HJ-3xTriMan-1xgp100 assembled with either a cleavable linker ( Val-Cit ) or a non-cleavable linker. Results shown from three independent human donors were expressed by the mean ± SD. Statistical analysis was conducted by a two-way t test. p-value * = <0.05, ** = <0.01, *** = <0.001, ****= <0.0001.

Article Snippet: Antigenic peptide gp100 154-162 (sequence: KTWGQYWQV, specificity: HLA-A*02:01, GENAXXON bioscience) was conjugated to the oligonucleotide strands in a 2-step one-pot reaction.

Techniques: Immunopeptidomics, Modification, Incubation, Cell Culture, Activation Assay, Concentration Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Synthetic Melanin Acts as Efficient Peptide Carrier in Cancer Vaccine Strategy

doi: 10.3390/ijms232314975

Figure Lengend Snippet: Conventional dendritic cells (cDCs) are required for immune response to melanin-based vaccines. ( A , B ) ELISpot analysis of splenocytes from the indicated mouse strain, 7 days after immunization with [gp100-Mel + CpG] vaccine. ( B ) B6-B6 chimera treated with DT and zDC-B6 not exposed to DT represent positive control. Data are presented as the normalized number of IFNγ spot forming cells (SFCs) per 10 5 CD8 + T cells, based on flow cytometry analysis. Each point represents an individual mouse (pooled data from three independent experiments). Bars = mean values ± SEM. ** p < 0.01.

Article Snippet: The endotoxin-free synthetic pOVA30 (SMLVLLPKKVSGLKQLESIINFEKLTKWTS) and gp100 (KVPRNQDWL) peptides, containing H2-2b epitopes (underlined) [ ], and peptides VIYRYYGL and Acetyl-R-VIYRYYGL were purchased from Genosphere Biotechnologies (Paris, France).

Techniques: Enzyme-linked Immunospot, Positive Control, Flow Cytometry

Journal: International Journal of Molecular Sciences

Article Title: Synthetic Melanin Acts as Efficient Peptide Carrier in Cancer Vaccine Strategy

doi: 10.3390/ijms232314975

Figure Lengend Snippet: L-DOPA acts as a carrier for peptides, and Langerin + cells carry melanin–peptide aggregates into lymph nodes. ( A ) ELISpot analysis of splenocytes 7 days after immunization with [gp100-Mel + CpG] vaccine in Langerin DTR mice treated with DT (diphtheria toxin) prior to immunization, 3 days after immunization, or not treated with DT (positive control, mice with Langerin + cells not depleted). Data are presented as normalized number of IFNγ spot-forming cells (SFCs) per 10 5 CD8 + T cells based on flow cytometry analysis. ( B ) ELISpot analysis of splenocytes on day 7, after immunizations on day 0 with different vaccine formulations. Each point represents an individual mouse (n = 8 mice/group with pooled data from 2 different experiments). Bars = mean values ± SEM. * p < 0.5, *** p < 0.001, ns = not significant.

Article Snippet: The endotoxin-free synthetic pOVA30 (SMLVLLPKKVSGLKQLESIINFEKLTKWTS) and gp100 (KVPRNQDWL) peptides, containing H2-2b epitopes (underlined) [ ], and peptides VIYRYYGL and Acetyl-R-VIYRYYGL were purchased from Genosphere Biotechnologies (Paris, France).

Techniques: Enzyme-linked Immunospot, Positive Control, Flow Cytometry

Journal: International Journal of Molecular Sciences

Article Title: Synthetic Melanin Acts as Efficient Peptide Carrier in Cancer Vaccine Strategy

doi: 10.3390/ijms232314975

Figure Lengend Snippet: Biodistribution of pOVA30-Mel nanoaggregates after subcutaneous injection in mice. ( A ) Transmission electronic microscopy (TEM) image of peptide–melanin nanoaggregates (gp100). ( B ) Outline of the experiment: Balb/c mice were injected subcutaneously at the right hind leg with [pOVA30-Mel + CpG] or [pOVA30 + CpG] as negative control, and [ 18 F]MEL050 CT/PET imaging was performed 1, 2, 7, 14, and 21 days, as well as 3 months later. ( C ) Melanin–peptide kinetics in the right inguinal lymph node post-injection of the vaccine versus negative control, expressed as normalized values of [ 18 F]MEL050 (% ID/g) to background radioactivity (contralateral lymph node). ( D ) Representative images of the [ 18 F]MEL050 max signal into the right inguinal lymph node (red arrow), 2 days and 3 months post-injection of [pOVA30-Mel + CpG. ( E ) ELISpot analysis of splenocytes 1, 2, and 4 months after immunization with [gp100-Mel + CpG] vaccine (injections on days 0 and 14). Results are presented as mean ± standard error of the mean (SEM) (data of four mice per group pooled from two different experiments); Mel = melanin; * = p < 0.05.

Article Snippet: The endotoxin-free synthetic pOVA30 (SMLVLLPKKVSGLKQLESIINFEKLTKWTS) and gp100 (KVPRNQDWL) peptides, containing H2-2b epitopes (underlined) [ ], and peptides VIYRYYGL and Acetyl-R-VIYRYYGL were purchased from Genosphere Biotechnologies (Paris, France).

Techniques: Injection, Transmission Assay, Microscopy, Negative Control, Imaging, Radioactivity, Enzyme-linked Immunospot

Journal: Frontiers in Immunology

Article Title: IRE1α Activation in Bone Marrow-Derived Dendritic Cells Modulates Innate Recognition of Melanoma Cells and Favors CD8 + T Cell Priming

doi: 10.3389/fimmu.2018.03050

Figure Lengend Snippet: Inhibition of IRE1α endonuclease function reduces the cross-presentation of a melanoma-endogenous antigen in vitro . (A) FL-DCs were preincubated with 50 μM 4μ8C or DMSO for 6 h and pulsed with 100 μg/ml MEL for the last 5 h of culture. Alternatively, cells were pulsed with 2.5 μM human gp100 peptide for the last 20 min of culture. Cells were counted, fixed and 5 × 10 4 FL-DCs were cocultured with 5 × 10 4 pmel-1 CD8 + T cells. Pmel-1 CD8 + T cell activation was quantified by expression of CD69 on day 1 through flow cytometry. Data in graph shows seven independent experiments. (B) FL-DCs were treated as in (A) but were not fixed and 2 × 10 4 FL-DCs were cultured with 5 × 10 4 CFSE-labeled pmel-1 CD8 + T cells. Proliferation was quantified on day 3 by flow cytometry. Data in graph shows three independent experiments. (C) GM-CSF BMDCs were treated and cocultured as in (A) . Data in graph shows six independent experiments. (D) GM-CSF BMDCs were treated and cocultured as in (B) . Data in graph shows four independent experiments. (E) FL-DCs were treated as in (B) but were cultured with 5 × 10 4 CellTrace Violet-labeled CTV = CD4 + T cells isolated from Trp1 mice. Proliferation was measured on day 5 by flow cytometry. Data in graph shows two independent experiments of (A) . Each symbol in the graphs represents data derived from one independent experiment. For all error bars represent mean ± SEM. * p < 0.05, ** p < 0.01 (paired Student's t -test).

Article Snippet: Human gp100 peptide (hgp100 25−33 , KVPRNQDWL) and Mouse TRP-1 peptide (TRP-1 106−130 , SGHNCGTCRPGWRGAACNQKILTVR) were purchased from Genetel Laboratories LLC.

Techniques: Inhibition, In Vitro, Activation Assay, Expressing, Flow Cytometry, Cell Culture, Labeling, Isolation, Derivative Assay