Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Jul 31;13(8):4733-53.
doi: 10.3390/md13084733.

Antitumor Effects and Related Mechanisms of Penicitrinine A, a Novel Alkaloid with a Unique Spiro Skeleton from the Marine Fungus Penicillium citrinum

Affiliations

Antitumor Effects and Related Mechanisms of Penicitrinine A, a Novel Alkaloid with a Unique Spiro Skeleton from the Marine Fungus Penicillium citrinum

Qin-Ying Liu et al. Mar Drugs. .

Abstract

Penicitrinine A, a novel alkaloid with a unique spiro skeleton, was isolated from a marine-derived fungus Penicillium citrinum. In this study, the isolation, structure and biosynthetic pathway elucidation of the new compound were described. This new compound showed anti-proliferative activity on multiple tumor types. Among them, the human malignant melanoma cell A-375 was confirmed to be the most sensitive. Morphologic evaluation, apoptosis rate analysis, Western blot and real-time quantitative PCR (RT-qPCR) results showed penicitrinine A could significantly induce A-375 cell apoptosis by decreasing the expression of Bcl-2 and increasing the expression of Bax. Moreover, we investigated the anti-metastatic effects of penicitrinine A in A-375 cells by wound healing assay, trans-well assay, Western blot and RT-qPCR. The results showed penicitrinine A significantly suppressed metastatic activity of A-375 cells by regulating the expression of MMP-9 and its specific inhibitor TIMP-1. These findings suggested that penicitrinine A might serve as a potential antitumor agent, which could inhibit the proliferation and metastasis of tumor cells.

Keywords: anti-metastatic; anticancer activity; apoptosis; human malignant melanoma cell A-375; marine-derived fungus; penicitrinine A.

PubMed Disclaimer

Figures

Figure 1
Figure 1
Key COSY, HMBC and NOESY correlations of penicitrinine A.
Figure 2
Figure 2
Chemical structure of penicitrinine A.
Figure 3
Figure 3
Plausible biosynthetic pathway of penicitrinine A.
Figure 4
Figure 4
Tumor cell-growth inhibitory activity of penicitrinine A. Twenty-three tumor cell lines were treated with penicitrinine A for 48 h at concentrations from 12.5 to 100 μM. The data are presented as means ± SD from five independent experiments.
Figure 5
Figure 5
The viability of A-375 cells treated with penicitrinine A (A) or 5-Fu (B) was determined by RTCA assay.
Figure 6
Figure 6
Penicitrinine A induced significant apoptotic morphological changes. (A) After exposed to 12.5, 25 and 50 μM penicitrinine A for 24 h, A-375 cells were stained by Hoeschest 33258 or AO/EB. Photos were taken under an inverted fluorescence microscope; (B) Quantification of the proportion of apoptotic cells detected by Hoechst 33258 staining and AO/EB staining. The values (means ± SD, n = 3) differed significantly (* p < 0.05; ** p < 0.01). The third column in Figure A was amplified from the second column for ten fold.
Figure 7
Figure 7
Penicitrinine A induced A-375 cells apoptosis. (A) A-375 cells were treated with penicitrinine A (12.5 μM, 25 μM and 50 μM) for 24 h, stained by annexin-V/PI and analyzed by flow cytometry, 5-Fu (50 μM, 100 μM and 200 μM) as a positive control; (B) Densitometry of cell counts. The values (means ± SD, n = 3) differed significantly (* p < 0.05; ** p < 0.01).
Figure 8
Figure 8
Western blot and RT-qPCR analysis of the apoptosis-associated molecules in A-375 cells. (A) After cells were treated with different concentrations of penicitrinine A for 24 h, western blot analysis was performed using antibodies against Bcl-2, Bax and β-actin; (B) Densitometric analysis of Bcl-2 and Bax at protein levels; (C) A-375 cells were treated with 0, 12.5, 25 and 50 µM penicitrinine A for 24 h. The mRNA levels from whole cell lysates were analyzed by RT-qPCR, GAPDH was used as a loading control; (D) Summary of the ratio of Bcl-2 to Bax as demonstrated by histograms. Data are expressed as mean ± SEM (n = 3). * p < 0.05, ** p < 0.01 vs. the control group.
Figure 9
Figure 9
Effects of penicitrinine A on the wound healing migration of A-375 cells. A wound was introduced by scraping confluent cell layers with a pipet tip. A-375 cells were incubated with penicitrinine A (5, 10 or 20 μM) for 24 h. Photos of the wounds were taken at 0 and 24 h under an inverted microscope.
Figure 10
Figure 10
Effect of penicitrinine A on the transwell invasion assay of A-375 cells. (A) A-375 cells incubated with penicitrinine A (5, 10, or 20 μM) were plated in the upper chambers of Matrigel-coated transwell insert for 24 h. The lower chamber contained 20% FBS as a chemoattractant in the medium. After incubated for 24 h, cells were fixed by methanol, stained with crystal violet, and observed under microscope; (B) The invaded A-375 cells were counted in five random fields in each treatment, and data were calculated from three independent experiments. Data are presented as mean ± SD of three independent experiments. * p < 0.05, ** p < 0.01 compared with the untreated control.
Figure 11
Figure 11
RT-qPCR and western blot analysis of the metastatic-associated molecules in A-375 cells. (A) After cells were treated with different concentration of penicitrinine A for 24 h, western blot analysis was performed using antibodies against TIMP-1, MMP-9 and β-actin; (B) Densitometric analysis of MMP-9 and TIMP-1 at protein levels; (C) A-375 cells were treated with 0, 5, 10 and 20 µM penicitrinine A for 24 h. The mRNA level from whole cell lysates was analyzed by RT-qPCR, GAPDH was used as a loading control. Data are expressed as mean ± SEM (n = 3). * p < 0.05, ** p < 0.01 vs. the control group.

References

    1. Landis S.H., Murray T., Bolden S., Wingo P.A. Cancer statistics, 1999. CA-Cancer J. Clin. 1999;49:8–31. doi: 10.3322/canjclin.49.1.8. - DOI - PubMed
    1. Rigel D.S. Epidemiology of melanoma. Semin. Cutan. Med. Surg. 2010;29:204–209. doi: 10.1016/j.sder.2010.10.005. - DOI - PubMed
    1. Ott P.A., Hodi F.S., Robert C. CTLA-4 and PD-1/PD-L1 blockade: New immunotherapeutic modalities with durable clinical benefit in melanoma patients. Clin. Cancer Res. 2013;19:5300–5309. doi: 10.1158/1078-0432.CCR-13-0143. - DOI - PubMed
    1. Luke J.J., Schwartz G.K. Chemotherapy in the management of advanced cutaneous malignant melanoma. Clin. Dermatol. 2013;31:290–297. doi: 10.1016/j.clindermatol.2012.08.016. - DOI - PMC - PubMed
    1. Tao L.-Y., Zhang J.-Y., Liang Y.-J., Chen L.-M., Zheng L.-S., Wang F., Mi Y.-J., She Z.-G., To K.K.W., Lin Y.-C. Anticancer effect and structure-activity analysis of marine products isolated from metabolites of mangrove fungi in the South China Sea. Mar. Drugs. 2010;8:1094–1105. doi: 10.3390/md8041094. - DOI - PMC - PubMed

Publication types

MeSH terms

LinkOut - more resources