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Gene Exp Pvrl2 Mm00436144 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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cynomolgus pvrl2  (Sino Biological)
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Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human <t>PVRIG/PVRL2</t> blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody
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pvrl2  (Proteintech)
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Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human <t>PVRIG/PVRL2</t> blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody
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primary antibodies against pvrl2  (ZenBio)
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Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human <t>PVRIG/PVRL2</t> blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody
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nectin family ligands, pvrl2 and pvr  (Johns Hopkins HealthCare)
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Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human <t>PVRIG/PVRL2</t> blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody
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anti-mouse pvrl2 brilliant violet 421  (Becton Dickinson)
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Proteomic analysis identifies <t>PVRL2</t> on tumor-derived exosomes. A, Schematic of exosome collection: exosomes were collected from the indicated tumor cell lines and primary tumor slices via differential centrifugations and purified by sucrose gradient. Exosomes and cells were then lysed, and their respective proteomes were analyzed by mass spectrometry. B, The numbers of shared immunoregulatory molecules identified from mass spectrometry results of the exosomes from the indicated tumor cell lines and primary tumors. Volcano plots present protein abundance differences in exosomes over in cells in PC3 ( C ) and SK-MEL-28 ( D ) cell lines as determined by label free quantitation. Proteins on the right of the volcano plot represent proteins enriched in exosomes versus cells. Proteins with a log 2 (fold change) value > 1 are over 2-fold enriched. E, Western blot analysis for PVRL2 in the cells and exosomes (exo) from the indicated tumor cell lines. A total of 30 μg of total protein was loaded for each sample. α-Tubulin was used as the loading control for cells, and Hrs as the loading control for exosomes.
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biotinylated human pvrl2 protein  (ACROBiosystems)
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Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human <t>PVRIG/PVRL2</t> blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody
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Image Search Results


Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human PVRIG/PVRL2 blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Characterization of a novel anti-PVRIG antibody with Fc-competent function that exerts strong antitumor effects via NK activation in preclinical models

doi: 10.1007/s00262-024-03671-z

Figure Lengend Snippet: Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human PVRIG/PVRL2 blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody

Article Snippet: After overnight incubation at 4 °C, the plates were blocked with PBS containing 1% BSA and 0.05% Tween-20 for 1 h. Serially diluted antibody and 2 μg/mL cynomolgus PVRL2 His protein (#90,206-C08H SinoBiological) were added to the plates and incubated for 3 h at room temperature.

Techniques: Binding Assay, Blocking Assay, Activity Assay, Stable Transfection, Enzyme-linked Immunosorbent Assay, Incubation, Expressing

Proteomic analysis identifies PVRL2 on tumor-derived exosomes. A, Schematic of exosome collection: exosomes were collected from the indicated tumor cell lines and primary tumor slices via differential centrifugations and purified by sucrose gradient. Exosomes and cells were then lysed, and their respective proteomes were analyzed by mass spectrometry. B, The numbers of shared immunoregulatory molecules identified from mass spectrometry results of the exosomes from the indicated tumor cell lines and primary tumors. Volcano plots present protein abundance differences in exosomes over in cells in PC3 ( C ) and SK-MEL-28 ( D ) cell lines as determined by label free quantitation. Proteins on the right of the volcano plot represent proteins enriched in exosomes versus cells. Proteins with a log 2 (fold change) value > 1 are over 2-fold enriched. E, Western blot analysis for PVRL2 in the cells and exosomes (exo) from the indicated tumor cell lines. A total of 30 μg of total protein was loaded for each sample. α-Tubulin was used as the loading control for cells, and Hrs as the loading control for exosomes.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: Proteomic analysis identifies PVRL2 on tumor-derived exosomes. A, Schematic of exosome collection: exosomes were collected from the indicated tumor cell lines and primary tumor slices via differential centrifugations and purified by sucrose gradient. Exosomes and cells were then lysed, and their respective proteomes were analyzed by mass spectrometry. B, The numbers of shared immunoregulatory molecules identified from mass spectrometry results of the exosomes from the indicated tumor cell lines and primary tumors. Volcano plots present protein abundance differences in exosomes over in cells in PC3 ( C ) and SK-MEL-28 ( D ) cell lines as determined by label free quantitation. Proteins on the right of the volcano plot represent proteins enriched in exosomes versus cells. Proteins with a log 2 (fold change) value > 1 are over 2-fold enriched. E, Western blot analysis for PVRL2 in the cells and exosomes (exo) from the indicated tumor cell lines. A total of 30 μg of total protein was loaded for each sample. α-Tubulin was used as the loading control for cells, and Hrs as the loading control for exosomes.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Derivative Assay, Purification, Mass Spectrometry, Quantitation Assay, Western Blot

PVRL2 promotes tumor growth through an immune-dependent mechanism. Average tumor volume over time following subcutaneous injection of 1 × 10 6 WT or Pvrl2 KO MC38 ( A ), TRAMP-C2 ( C ), and B16F10 ( D ) cells in C57BL/6 mice, and CT26 ( B ) in BALB/cJ mice. Error bars represent SEM. AUCs of the MC38 ( E ), CT26 ( F ), TRAMP-C2 ( G ), and B16F10 ( H ) tumors from A – D calculated at day when the first mouse reached endpoint: day 23, 18, 60, and 15, respectively. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 ( I ), CT26 ( J ), TRAMP-C2 ( K ), and B16F10 ( L ) WT and Pvrl2 KO cells in NCG mice. Error bars represent SEM. AUCs of the MC38 ( M ), CT26 ( N ), TRAMP-C2 ( O ), and B16F10 ( P ) tumors from I – L calculated at day when the first mouse reached endpoint: day 20, 15, 47, and 14, respectively. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: PVRL2 promotes tumor growth through an immune-dependent mechanism. Average tumor volume over time following subcutaneous injection of 1 × 10 6 WT or Pvrl2 KO MC38 ( A ), TRAMP-C2 ( C ), and B16F10 ( D ) cells in C57BL/6 mice, and CT26 ( B ) in BALB/cJ mice. Error bars represent SEM. AUCs of the MC38 ( E ), CT26 ( F ), TRAMP-C2 ( G ), and B16F10 ( H ) tumors from A – D calculated at day when the first mouse reached endpoint: day 23, 18, 60, and 15, respectively. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 ( I ), CT26 ( J ), TRAMP-C2 ( K ), and B16F10 ( L ) WT and Pvrl2 KO cells in NCG mice. Error bars represent SEM. AUCs of the MC38 ( M ), CT26 ( N ), TRAMP-C2 ( O ), and B16F10 ( P ) tumors from I – L calculated at day when the first mouse reached endpoint: day 20, 15, 47, and 14, respectively. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Injection

Exosomal PVRL2 partially rescues the phenotype of Pvrl2 KO tumors. A, Schematic of the experiment: 1 × 10 6 MC38 Pvrl2 KO cells were injected into C57BL/6 mice, and starting from the same day, exosomes collected from MC38 WT and Pvrl2 KO cells were injected into the mice through tail vein according to the indicated timeline. B, Average tumor volume over time following the injection of MC38 Pvrl2 KO tumors along with no exosome injection, MC38 WT exosomes, and MC38 Pvrl2 KO exosomes as indicated in A . Error bars represent SEM. C, AUCs of the tumor growth in B on day 41. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. D, Mouse survival curves following injections as described in B . P values are calculated by log-rank test. E, AUCs of the growth of MC38 Pvrl2 KO tumors with no exosome injection or WT exosome injection from B , in comparison with the MC38 WT tumors from A on day 23. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: Exosomal PVRL2 partially rescues the phenotype of Pvrl2 KO tumors. A, Schematic of the experiment: 1 × 10 6 MC38 Pvrl2 KO cells were injected into C57BL/6 mice, and starting from the same day, exosomes collected from MC38 WT and Pvrl2 KO cells were injected into the mice through tail vein according to the indicated timeline. B, Average tumor volume over time following the injection of MC38 Pvrl2 KO tumors along with no exosome injection, MC38 WT exosomes, and MC38 Pvrl2 KO exosomes as indicated in A . Error bars represent SEM. C, AUCs of the tumor growth in B on day 41. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. D, Mouse survival curves following injections as described in B . P values are calculated by log-rank test. E, AUCs of the growth of MC38 Pvrl2 KO tumors with no exosome injection or WT exosome injection from B , in comparison with the MC38 WT tumors from A on day 23. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Injection, Comparison

PVRL2 regulates CD8 T-cell and NK-cell activation. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 WT and Pvrl2 KO MC38 cells in Rag1 KO mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A and A on day 19. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Schematic of immunophenotyping experiment: 25 days after subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in C57BL/6 mice, the tumors were collected. CD45 + cell populations were isolated from the tumor dissociates using CD45 positive magnetic selection kit. Then the isolated cells were subjected to viability dye, CD8, CD4, NK1.1, CD107a, and PD-1 staining followed by flow cytometry analysis. Flow cytometric quantification of the percentage of CD8 + ( D ), CD4 + ( E ), and NK1.1 + CD8 − (NK cells; F ) populations, respectively, in the CD45 + cells isolated from the MC38 WT ( n = 3) and Pvrl2 KO ( n = 4) tumors as indicated in C . P values are calculated by unpaired t test. Line represents mean. Quantification of the percentage of CD107a + ( G ) and PD-1 + ( H ) cells among the CD8 + T cell ( D ) population. P values are calculated by unpaired t test. Line represents mean. I, Quantification of the percentage of CD107a + cells among the NK cell ( F ) population. P value is calculated by unpaired t test. Line represents mean. J, Schematic of experiment design in K – O : After subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in C57BL/6 WT or Rag1 KO mice, the mice were treated with anti-CD8, CD4, or NK1.1 depleting antibodies or isotype control at the indicated serial doses and schedule. Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 mice with anti-CD8 ( K ) and anti-CD4 ( L ) depleting antibodies or isotype control as indicated in J . Error bars represent SEM. M, AUCs of the MC38 tumors from ( K ) and ( L ) on day 17. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. N, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in Rag1 KO mice with anti-NK1.1 depleting antibody or isotype control as indicated in J . Error bars represent SEM. O, AUCs of the MC38 tumors from N on day 16. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: PVRL2 regulates CD8 T-cell and NK-cell activation. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 WT and Pvrl2 KO MC38 cells in Rag1 KO mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A and A on day 19. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Schematic of immunophenotyping experiment: 25 days after subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in C57BL/6 mice, the tumors were collected. CD45 + cell populations were isolated from the tumor dissociates using CD45 positive magnetic selection kit. Then the isolated cells were subjected to viability dye, CD8, CD4, NK1.1, CD107a, and PD-1 staining followed by flow cytometry analysis. Flow cytometric quantification of the percentage of CD8 + ( D ), CD4 + ( E ), and NK1.1 + CD8 − (NK cells; F ) populations, respectively, in the CD45 + cells isolated from the MC38 WT ( n = 3) and Pvrl2 KO ( n = 4) tumors as indicated in C . P values are calculated by unpaired t test. Line represents mean. Quantification of the percentage of CD107a + ( G ) and PD-1 + ( H ) cells among the CD8 + T cell ( D ) population. P values are calculated by unpaired t test. Line represents mean. I, Quantification of the percentage of CD107a + cells among the NK cell ( F ) population. P value is calculated by unpaired t test. Line represents mean. J, Schematic of experiment design in K – O : After subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in C57BL/6 WT or Rag1 KO mice, the mice were treated with anti-CD8, CD4, or NK1.1 depleting antibodies or isotype control at the indicated serial doses and schedule. Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 mice with anti-CD8 ( K ) and anti-CD4 ( L ) depleting antibodies or isotype control as indicated in J . Error bars represent SEM. M, AUCs of the MC38 tumors from ( K ) and ( L ) on day 17. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. N, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in Rag1 KO mice with anti-NK1.1 depleting antibody or isotype control as indicated in J . Error bars represent SEM. O, AUCs of the MC38 tumors from N on day 16. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Activation Assay, Injection, Isolation, Selection, Staining, Flow Cytometry

PVRL2 functions through a PVRIG independent mechanism. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 mice and Pvrig KO mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A on day 24. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Mouse survival curves following subcutaneous injections as described in A . P values are calculated by log rank test. D, Average tumor volume over time following subcutaneous injection of 1 × 10 6 TRAMP-C2 WT and Pvrl2 KO cells in WT C57BL/6 and Pvrig KO mice. Error bars represent SEM. E, AUCs of the TRAMP-C2 tumors from D on day 58. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. F, Mouse survival curves following injections as described in D . P values are calculated by log-rank test. G, Average tumor volume over time following subcutaneous injection of 1 × 10 6 B16F10 WT and Pvrl2 KO cells in WT C57BL/6 mice and Pvrig KO mice. Error bars represent SEM. H, AUCs of the B16F10 tumors from G on day 15. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. I, Mouse survival curves following subcutaneous injections as described in G . P values are calculated by log-rank test. J, Schematic of experiment design in K : NK cells were isolated from WT C57BL/6 or Pvrig KO mice. After 7–9 days in vitro stimulation with IL2, NK cells were cocultured at 1:1 TRAMP WT or Pvrl2 KO tumor cells for 4 hours. Then NK cell lysis was evaluated by live/dead cell dye followed by flow cytometry. K, Percentage lysis of TRAMP-C2 WT and Pvrl2 KO cells after coculturing with WT or Pvrig KO NK cells at 1:1 ratio. Dots represent individual replicates. 4 replicates per condition in each experiment ( n = 2 and n = 3 independent experiments for WT NK cells and Pvrig KO NK cells, respectively). P values are calculated by unpaired t test. Line represents mean.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: PVRL2 functions through a PVRIG independent mechanism. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 mice and Pvrig KO mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A on day 24. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Mouse survival curves following subcutaneous injections as described in A . P values are calculated by log rank test. D, Average tumor volume over time following subcutaneous injection of 1 × 10 6 TRAMP-C2 WT and Pvrl2 KO cells in WT C57BL/6 and Pvrig KO mice. Error bars represent SEM. E, AUCs of the TRAMP-C2 tumors from D on day 58. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. F, Mouse survival curves following injections as described in D . P values are calculated by log-rank test. G, Average tumor volume over time following subcutaneous injection of 1 × 10 6 B16F10 WT and Pvrl2 KO cells in WT C57BL/6 mice and Pvrig KO mice. Error bars represent SEM. H, AUCs of the B16F10 tumors from G on day 15. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. I, Mouse survival curves following subcutaneous injections as described in G . P values are calculated by log-rank test. J, Schematic of experiment design in K : NK cells were isolated from WT C57BL/6 or Pvrig KO mice. After 7–9 days in vitro stimulation with IL2, NK cells were cocultured at 1:1 TRAMP WT or Pvrl2 KO tumor cells for 4 hours. Then NK cell lysis was evaluated by live/dead cell dye followed by flow cytometry. K, Percentage lysis of TRAMP-C2 WT and Pvrl2 KO cells after coculturing with WT or Pvrig KO NK cells at 1:1 ratio. Dots represent individual replicates. 4 replicates per condition in each experiment ( n = 2 and n = 3 independent experiments for WT NK cells and Pvrig KO NK cells, respectively). P values are calculated by unpaired t test. Line represents mean.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Injection, Isolation, In Vitro, Lysis, Flow Cytometry

PVRL2 loss and TIGIT blockade function cooperatively to inhibit tumor growth. A, Schematic of experiment design in B – G : After subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 or Pvrig KO mice, starting on day 4, mice were treated with serial doses of anti-TIGIT blocking antibody or isotype control at the indicated doses and schedule. B, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT cells with and without anti-TIGIT treatment in WT C57BL/6 mice. Error bars represent SEM. C, AUCs of the MC38 tumors from B on day 26. Dots represent individual mice. P value is calculated by unpaired t test. Error bars represent SD. D, Mouse survival curves following injections as described in B . P value is calculated by log-rank test. E, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells with and without anti-TIGIT treatment in Pvrig KO mice. Error bars represent SEM. F, AUCs of the MC38 tumors from E on day 26. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. G, Mouse survival curves following injections as described in E . P values are calculated by log-rank test.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: PVRL2 loss and TIGIT blockade function cooperatively to inhibit tumor growth. A, Schematic of experiment design in B – G : After subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells in WT C57BL/6 or Pvrig KO mice, starting on day 4, mice were treated with serial doses of anti-TIGIT blocking antibody or isotype control at the indicated doses and schedule. B, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT cells with and without anti-TIGIT treatment in WT C57BL/6 mice. Error bars represent SEM. C, AUCs of the MC38 tumors from B on day 26. Dots represent individual mice. P value is calculated by unpaired t test. Error bars represent SD. D, Mouse survival curves following injections as described in B . P value is calculated by log-rank test. E, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT and Pvrl2 KO cells with and without anti-TIGIT treatment in Pvrig KO mice. Error bars represent SEM. F, AUCs of the MC38 tumors from E on day 26. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. G, Mouse survival curves following injections as described in E . P values are calculated by log-rank test.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Injection, Blocking Assay

Combined loss of PVRL2 and PVR loss does not further inhibit tumor growth. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT, Pvrl2 KO, Pvr KO and Pvrl2 ; Pvr KO cells in WT C57BL/6 mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A on day 23. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Mouse survival curves following injections as described in A . P values are calculated by log-rank test. D, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT, Pvrl2 KO, Pvr KO and Pvrl2 ; Pvr KO cells in Rag1 KO mice. Error bars represent SEM. E, AUCs of the MC38 tumors from D on day 19. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. F, Mouse survival curves following injections as described in D . P values are calculated by log-rank test. G, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 Pvr KO cells with and without anti-TIGIT treatment in WT C57BL/6 mice. Error bars represent SEM. H, AUCs of the tumors from G on day 26. Dots represent individual mice. P value is calculated by unpaired t test. Error bars represent SD. I, Mouse survival curves following injections as described in G . P value is calculated by log-rank test.

Journal: Cancer Immunology Research

Article Title: PVRL2 Suppresses Antitumor Immunity through PVRIG- and TIGIT-independent Pathways

doi: 10.1158/2326-6066.CIR-23-0722

Figure Lengend Snippet: Combined loss of PVRL2 and PVR loss does not further inhibit tumor growth. A, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT, Pvrl2 KO, Pvr KO and Pvrl2 ; Pvr KO cells in WT C57BL/6 mice. Error bars represent SEM. B, AUCs of the MC38 tumors from A on day 23. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. C, Mouse survival curves following injections as described in A . P values are calculated by log-rank test. D, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 WT, Pvrl2 KO, Pvr KO and Pvrl2 ; Pvr KO cells in Rag1 KO mice. Error bars represent SEM. E, AUCs of the MC38 tumors from D on day 19. Dots represent individual mice. P values are calculated by unpaired t test. Error bars represent SD. F, Mouse survival curves following injections as described in D . P values are calculated by log-rank test. G, Average tumor volume over time following subcutaneous injection of 1 × 10 6 MC38 Pvr KO cells with and without anti-TIGIT treatment in WT C57BL/6 mice. Error bars represent SEM. H, AUCs of the tumors from G on day 26. Dots represent individual mice. P value is calculated by unpaired t test. Error bars represent SD. I, Mouse survival curves following injections as described in G . P value is calculated by log-rank test.

Article Snippet: Flow antibodies: Anti-mouse CD8 Brilliant Violet 605 (53-6.7; BioLegend, catalog no. 100744), anti-mouse CD4 Brilliant Violet 421 (GK1.5; BioLegend, catalog no. 100437), anti-mouse NK1.1 PE (PK136; BioLegend, catalog no. 108707), anti-mouse NK1.1 Brilliant Violet 711 (PK136; BioLegend, catalog no. 108745), anti-mouse CD107a (LAMP-1) Brilliant Violet 711 (1D4B; BioLegend, catalog no. 121631), anti-mouse PD-1 (CD279) APC (J43; BD Biosciences, catalog no. 562671), anti-mouse PVRL2 Brilliant Violet 421 (BD Biosciences, catalog no. 748046), anti-mouse PVR PE (TX56; BioLegend, catalog no. 131507), anti-mouse PVR APC (TX56; BioLegend, catalog no. 131509), anti-mouse TIGIT PE (1G9; BioLegend, catalog no. 142104), anti-mouse CD96 APC (3.3; BioLegend, catalog no. 131711), anti-mouse DNAM-1 Brilliant Violet 785 (TX42.1; BioLegend, catalog no. 133611), anti-mouse CD98 PE (4F2; BioLegend, catalog no. 128207), anti-mouse PD-L1 Super Bright 780 (MIH5; Invitrogen, catalog no. 78-5982-82), Brilliant Violet 605 Rat IgG2a, k Isotype control (RTK2758; BioLegend, catalog no. 400539), Brilliant Violet 421 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400639), PE Mouse IgG2a, κ Isotype control (MOPC-173; BioLegend, catalog no. 400211), Brilliant Violet 711 Rat IgG2b, κ Isotype control (RTK4530; BioLegend, catalog no. 400653), APC Hamster IgG2, κ Isotype control (B81-3; BD Biosciences, catalog no. 562169).

Techniques: Injection

Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human PVRIG/PVRL2 blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Characterization of a novel anti-PVRIG antibody with Fc-competent function that exerts strong antitumor effects via NK activation in preclinical models

doi: 10.1007/s00262-024-03671-z

Figure Lengend Snippet: Binding affinity and ligand blocking activity of IBI352g4a. a Affinity of BMK and IBI352g4a for human PVRIG and cynoPVRIG assayed using FortéBio. b Human PVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/hPVRIG stable cell line. c CynoPVRIG cell-based binding was conducted for BMK and IBI352g4a using the GS-CHO/cynoPVRIG stable cell line. d ELISA-based human PVRIG/PVRL2 blocking assay. Biotinylated human PVRL2 cells were incubated with increasing amounts of IBI352g4a or BMK in precoated human PVRIG plates. After incubation and washing, biotinylated human PVRL2 was detected via streptavidin (HRP). e ELISA-based cynoPVRIG/PVRL2 blocking assay. CynoPVRL2 protein (his tag) was incubated with increasing amounts of IBI352g4a or BMK in precoated cynomolgus PVRIG plates. After incubation and washing, the expression of cynomolgus monkey PVRL2 was detected with an HRP-conjugated antibody

Article Snippet: After overnight incubation at 4 °C, the plates were blocked with PBS containing 1% BSA and 0.05% Tween-20 for 1 h. Serially diluted antibody and 2 μg/mL biotinylated human PVRL2 protein (Acro Biosystems, Inc., Newark, Delaware, USA) were added to the plates, which were subsequently incubated for 3 h at room temperature.

Techniques: Binding Assay, Blocking Assay, Activity Assay, Stable Transfection, Enzyme-linked Immunosorbent Assay, Incubation, Expressing