Journal: Redox Biology

Article Title: Combination of YAP inhibition and photodynamic therapy induces dual DNA damage and activates STING pathway to enhance immunotherapy in uveal melanoma

doi: 10.1016/j.redox.2025.103965

Figure Lengend Snippet: Activation of the cGAS-STING pathway by HANP/VP via YAP-inhibition and PDT in vitro. (A) The phosphorylation levels of representative proteins of the cGAS-STING pathway were analyzed using Western blot ( left ) and quantified ( right ). After HANP/VP treatment without laser irradiation, the pSTING/STING, pTBK1/TBK1, and pIRF3/IRF3 ratio were significantly increased compared to the untreated cells; the phosphorylation levels were further increased after laser irradiation, n = 3/group. (B) qPCR was used to detect IFN-β1 mRNA levels. Compared with the control group, HANP/VP without Laser treatment significantly increased IFN-β1 expression in B16F10 cells, while HANP/VP combined with laser irradiation further elevated IFN-β1 mRNA levels. When cells were pretreated with ethidium bromide (EB, 120 ng/mL), IFN-β1 mRNA levels decreased in the HANP/VP with Laser irradiation group. Following H151 treatment, IFN-β1 mRNA levels decreased to control group levels regardless of laser irradiation status. n = 3/group. Statistical significance was determined by one-way ANOVA (∗∗ p < 0.01, ∗∗∗ p < 0.001).

Article Snippet: The mouse melanoma cell line B16F10 was purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Activation Assay, Inhibition, In Vitro, Phospho-proteomics, Western Blot, Irradiation, Control, Expressing

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: Thirty-two 10.5 week-old male C57BL/6J mice were inoculated with 250,000 B16F10 melanoma cells by tail vein injection. Two mice died of uncertain causes, and the remaining mice were euthanized on day 14 following tumor inoculation. Groups were: A. untreated; B. rhARSB (0.2 ng/ml IV on days 2, 7, and 12); C. Pembrolizumab (5 mg/ml IP on days 2,7, and 12); and D. combined rhARSB and Pembrolizumab treatments. E . Compared to saline-treated control, the rhARSB, Pembrolizumab, and the combined rhARSB and Pembrolizumab groups had significantly fewer tumors (unpaired t-test, two-tailed, unequal standard variation, p<0.05). Bar = 0.5 cm. (ARSB=arylsulfatase B=N-acetylgalactosamine-4-sulfatase; rh=recombinant human).

Article Snippet: B16 F10 cells were cultured in DMEM (ATCC 30-2002) with 10% FBS under the recommended conditions.

Techniques: Injection, Saline, Control, Two Tailed Test, Recombinant

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: A. Cleaved caspase-3 increased following rhARSB (p=2.5×10 -7 ) or Pembrolizumab (p=1.2×10 -5 ) and was further Increased by their combination (p=1.2×10 -7 ) in treated B16F10 pulmonary melanomas. All p-values are calculated by unpaired t-test, two-sided with unequal standard deviation. B. In treated B16F10 melanomas, COP1 mRNA expression increased following rhARSB, but not following Pembrolizumab treatment. C. BCL2 mRNA expression declined following either rhARSB (p-1.7×10 or Pembrolizumab and declined further by their combination in the B16F10 pulmonary melanomas. D. Cleaved caspase-3 was inversely correlated with Bcl2 (r = -0.97; Spearman’s correlation coefficient). E. Cleaved capase-3 was directly correlated with COP1 in the pulmonary melanomas.(r=0.85, Spearman’s correlation coefficient). F. In cultured human A375 melanoma cells, cleaved caspase-3 increased following rhARSB and was further increased by the combination of rhARSB with activated PBMC [PBMC(Ac)] (p=0.007). Pembrolizumab alone or Pembrolizumab with unactivated PBMC did not increase cleaved caspase-3. However, cleaved caspase 3 was increased by the combination of Pembrolizumab with PBMC(Ac) (p=0.002) and further increased by rhARSB, Pembrolizumab, and activated PBMC (p=0.009). G. In the A375 cells, COP1 increased following rhARSB, but was unaffected by Pembrolizumab or by PBMC, either activated or not activated. H. Inverse to the effect on COP1, rhARSB reduced the expression of BCL2. Neither Pembrolizumab alone nor PBMC alone affected BCL2 expression, but Pembro with activated PBMC reduced BCL2 (p=4.1×10 -5 ). BCL2 was further reduced by the combination of rhARSB + Pembrolizumab + PBMC(Ac) in the A375 cells (p=4.8×10 -9 ). I. Granzyme B increased following the combinations of rhARSB and PBMC(Ac) (p=5.3×10 -10 ) and Pembrolizumab and PBMC(Ac) (p=1.3×10 -7 ). Synergistic effect was evident following treatment by rhARSB + Pembrolizumab + and PBMC(Ac) in the A375 cells. [PBMC(Ac)=activated peripheral blood mononuclear cells; COP1=constitutive photomorphogenic protein 1; Pembro=Pembrolizumab; PBMC=peripheral blood mononuclear cells; rhARSB=recombinant human arylsulfatase B]

Article Snippet: B16 F10 cells were cultured in DMEM (ATCC 30-2002) with 10% FBS under the recommended conditions.

Techniques: Standard Deviation, Expressing, Cell Culture, Recombinant

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: A. In B16F10 pulmonary melanoma tissue, mRNA expression of MCP1 and IL-10 increased following treatment by Pembrolizumab or rhARSB, and the increase was enhanced by their combination. In contrast, IL-17α expression was increased by Pembrolizumab, but not by rhARSB. B. KC, the mouse analog of IL-8, and IL-6 were reduced by both rhARSB and Pembrolizumab and further reduced by their combination. In contrast, TNFα was increased by Pembrolizumab and reduced by rhARSB, with overall reduction by their combination. C. FGF2, VEGF, and EGF were reduced by ARSB, but not by Pembrolizumab, and the combination of ARSB and Pembrolizumab did not lead to additional decline. [ARSB=arylsulfatase B=N-acetylgalactosamine-4-sulfatase; EGF=epidermal growth factor; bFGF=basic fibroblast growth factor; IL=interleukin; KC=CXCL1=keratinocyte-derived cytokine; rh=recombinant human; MCP1=CCL2= monocyte chemoattractant protein-1; VEGF=vascular endothelial growth factor]

Article Snippet: B16 F10 cells were cultured in DMEM (ATCC 30-2002) with 10% FBS under the recommended conditions.

Techniques: Expressing, Derivative Assay, Recombinant

Journal: bioRxiv

Article Title: CAR-T-drug conjugate against solid tumor

doi: 10.64898/2026.01.02.696502

Figure Lengend Snippet: (a) Schematic illustration of the mouse anti-CA9 CAR-T generation and treatment strategy in the syngeneic Colon26-CA9 colon cancer model in BALB/c mice. (b) Flow cytometry showing the surface CA9 expression level of Colon26 cell line with or without human CA9 overexpression. (c) Upper panel: Schematic representation of the CAR-T treatment strategy in the syngeneic Colon26 colon cancer model in BALB/c mice. Lower panel: Growth curves showing tumor volume measured over time under different treatments: conventional CA9 CAR-T (n = 7), MMAE CA9 CAR-T-D-C (n = 8), or PBS vehicle control (n = 5). Data are presented as mean ± SEM. Statistical comparisons were performed using two-way ANOVA for each group against PBS vehicle control group. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. (d) Overall survival analysis of Colon26-CA9 tumors in BALB/c mice following treatments with PBS, CAR-T and CAR-T-D-C. Statistical comparisons were made using the Log-rank (Mantel-Cox) test for each treatment group against the vehicle control group. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. (e) Tumor-infiltrating immune cell composition in Colon26-CA9 tumors following treatments with PBS vehicle control, CAR-T and CAR-T-D-C. Statistical comparisons were made with one-way ANOVA. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. (f) Upper panel: Schematic representation of the OT-I T cell treatment strategy in the syngeneic mouse B16F10-OVA melanoma model. Lower panel: Growth curves of B16F10-OVA tumors in C57BL/6 mice following different treatments: OT-I T cells (n = 4), OT-I T-D-C (n = 4), or PBS vehicle control (n = 5). Data are presented as mean ± SEM. Statistical comparisons were performed using two-way ANOVA for each group against PBS. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. (g) Overall survival analysis of the B16F10-OVA tumor model treated with OT-I T cells, OT-1-D-Cs, or PBS vehicle control. Statistical comparisons were made using the Log-rank (Mantel-Cox) test for each treatment group against the vehicle control group. Statistical significance is denoted as follows: ∗∗p < 0.01; ∗∗∗p < 0.001. (h) Tumor-infiltrating immune cell composition in B16F10 tumors following treatments with PBS vehicle control, CAR-T and CAR-T-D-C. Statistical comparisons were made with one-way ANOVA. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001.

Article Snippet: Expi293F, HEK293FT, Lenti-X 293T, A498, ACHN, HT29, Colon26, and B16F10 cell lines were obtained from commercial sources (ThermoFisher, American Type Culture Collection (ATCC), Takara) and were periodically tested negative for mycoplasma contamination.

Techniques: Flow Cytometry, Expressing, Over Expression, Control

Journal: bioRxiv

Article Title: CAR-T-drug conjugate against solid tumor

doi: 10.64898/2026.01.02.696502

Figure Lengend Snippet: (a) Schematics representation the CAR-T-D-C treatment strategy in a dual tumor model of mouse Colon26 colon cancer to study the systemic anti-tumor response. (b-d) Growth curve showing Colon26 tumor volume measured overtime following treatments with PBS vehicle control, CAR-T and CAR-T-D-C. ( b ) Primary tumor Colon26-CA9 with human CA9 overexpression; ( c ) Secondary tumor Colon26 without human CA9 antigen. (d) Combined analysis of primary and secondary tumors. Statistical comparisons were made using two-way ANOVA. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗∗p < 0.0001. (e) Survival of mice with bilateral Colon26-CA9 (primary) and Colon26 (secondary) tumors following CAR-T, CAR-T-D-C, or PBS vehicle control treatment. (f) IFN-γ secretion ability of the splenocytes isolated from tumor bearing-mice treated with CAR-T, CAR-T-D-C or PBS vehicle, measured by ELISPOT assay. Statistical comparisons were made with one-way ANOVA. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. (g) Schematics representation of the OT-I T cell treatment strategy in a dual tumor model with B16F10-OVA (primary) and B16F10 (secondary) in C57BL/6 mice. (h-j) Growth curves of the tumor volume measured overtime with indicated treatments. ( f ) Primary tumor with the OVA antigen; ( g ) Secondary tumor without OVA antigen. Statistical comparisons were made using two-way ANOVA. ( j ) Combined analysis of primary and secondary tumors. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗∗p < 0.0001. ( k ) Survival of mice with bilateral B16F10- OVA (primary) and B16F10 (secondary) tumors following OT-I, OT-I-D-C, or PBS vehicle treatment. (l) IFN-γ secretion ability of the splenocytes isolated from B16F10 tumor bearing-mice treated with OT1-T, OT-I-D-C or PBS vehicle, measured by IFN-γ ELISPOT assay. Statistical comparisons were made using one-way ANOVA. Statistical significance is denoted as follows: ∗p < 0.05; ∗∗∗∗p < 0.0001. ( m ) Schematic summarizing the mechanism of the dual-function CAR-T-D-C approach.

Article Snippet: Expi293F, HEK293FT, Lenti-X 293T, A498, ACHN, HT29, Colon26, and B16F10 cell lines were obtained from commercial sources (ThermoFisher, American Type Culture Collection (ATCC), Takara) and were periodically tested negative for mycoplasma contamination.

Techniques: Control, Over Expression, Isolation, Enzyme-linked Immunospot

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: Thirty-two 10.5 week-old male C57BL/6J mice were inoculated with 250,000 B16F10 melanoma cells by tail vein injection. Two mice died of uncertain causes, and the remaining mice were euthanized on day 14 following tumor inoculation. Groups were: A. untreated; B. rhARSB (0.2 ng/ml IV on days 2, 7, and 12); C. Pembrolizumab (5 mg/ml IP on days 2,7, and 12); and D. combined rhARSB and Pembrolizumab treatments. E . Compared to saline-treated control, the rhARSB, Pembrolizumab, and the combined rhARSB and Pembrolizumab groups had significantly fewer tumors (unpaired t-test, two-tailed, unequal standard variation, p<0.05). Bar = 0.5 cm. (ARSB=arylsulfatase B=N-acetylgalactosamine-4-sulfatase; rh=recombinant human).

Article Snippet: B16F10 mouse melanoma cells were purchased (ATCC CRL-6475) and cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin.

Techniques: Injection, Saline, Control, Two Tailed Test, Recombinant

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: A. Cleaved caspase-3 increased following rhARSB (p=2.5×10 -7 ) or Pembrolizumab (p=1.2×10 -5 ) and was further Increased by their combination (p=1.2×10 -7 ) in treated B16F10 pulmonary melanomas. All p-values are calculated by unpaired t-test, two-sided with unequal standard deviation. B. In treated B16F10 melanomas, COP1 mRNA expression increased following rhARSB, but not following Pembrolizumab treatment. C. BCL2 mRNA expression declined following either rhARSB (p-1.7×10 or Pembrolizumab and declined further by their combination in the B16F10 pulmonary melanomas. D. Cleaved caspase-3 was inversely correlated with Bcl2 (r = -0.97; Spearman’s correlation coefficient). E. Cleaved capase-3 was directly correlated with COP1 in the pulmonary melanomas.(r=0.85, Spearman’s correlation coefficient). F. In cultured human A375 melanoma cells, cleaved caspase-3 increased following rhARSB and was further increased by the combination of rhARSB with activated PBMC [PBMC(Ac)] (p=0.007). Pembrolizumab alone or Pembrolizumab with unactivated PBMC did not increase cleaved caspase-3. However, cleaved caspase 3 was increased by the combination of Pembrolizumab with PBMC(Ac) (p=0.002) and further increased by rhARSB, Pembrolizumab, and activated PBMC (p=0.009). G. In the A375 cells, COP1 increased following rhARSB, but was unaffected by Pembrolizumab or by PBMC, either activated or not activated. H. Inverse to the effect on COP1, rhARSB reduced the expression of BCL2. Neither Pembrolizumab alone nor PBMC alone affected BCL2 expression, but Pembro with activated PBMC reduced BCL2 (p=4.1×10 -5 ). BCL2 was further reduced by the combination of rhARSB + Pembrolizumab + PBMC(Ac) in the A375 cells (p=4.8×10 -9 ). I. Granzyme B increased following the combinations of rhARSB and PBMC(Ac) (p=5.3×10 -10 ) and Pembrolizumab and PBMC(Ac) (p=1.3×10 -7 ). Synergistic effect was evident following treatment by rhARSB + Pembrolizumab + and PBMC(Ac) in the A375 cells. [PBMC(Ac)=activated peripheral blood mononuclear cells; COP1=constitutive photomorphogenic protein 1; Pembro=Pembrolizumab; PBMC=peripheral blood mononuclear cells; rhARSB=recombinant human arylsulfatase B]

Article Snippet: B16F10 mouse melanoma cells were purchased (ATCC CRL-6475) and cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin.

Techniques: Standard Deviation, Expressing, Cell Culture, Recombinant

Journal: medRxiv

Article Title: Effects of Arylsulfatase B and Pembrolizumab in Combination on Progression of Metastatic Melanoma in the B16F10 Syngeneic Mouse Model

doi: 10.64898/2026.01.02.25343295

Figure Lengend Snippet: A. In B16F10 pulmonary melanoma tissue, mRNA expression of MCP1 and IL-10 increased following treatment by Pembrolizumab or rhARSB, and the increase was enhanced by their combination. In contrast, IL-17α expression was increased by Pembrolizumab, but not by rhARSB. B. KC, the mouse analog of IL-8, and IL-6 were reduced by both rhARSB and Pembrolizumab and further reduced by their combination. In contrast, TNFα was increased by Pembrolizumab and reduced by rhARSB, with overall reduction by their combination. C. FGF2, VEGF, and EGF were reduced by ARSB, but not by Pembrolizumab, and the combination of ARSB and Pembrolizumab did not lead to additional decline. [ARSB=arylsulfatase B=N-acetylgalactosamine-4-sulfatase; EGF=epidermal growth factor; bFGF=basic fibroblast growth factor; IL=interleukin; KC=CXCL1=keratinocyte-derived cytokine; rh=recombinant human; MCP1=CCL2= monocyte chemoattractant protein-1; VEGF=vascular endothelial growth factor]

Article Snippet: B16F10 mouse melanoma cells were purchased (ATCC CRL-6475) and cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin.

Techniques: Expressing, Derivative Assay, Recombinant