Marine-Derived Penicillium Species as Producers of Cytotoxic Metabolites

Since the discovery of penicillin, Penicillium has become one of the most attractive fungal genera for the production of bioactive molecules. Marine-derived Penicillium has provided numerous excellent pharmaceutical leads over the past decades. In this review, we focused on the cytotoxic metabolites * (* Cytotoxic potency was referred to five different levels in this review, extraordinary (IC50/LD50: <1 μM or 0.5 μg/mL); significant (IC50/LD50: 1~10 μM or 0.5~5 μg/mL); moderate (IC50/LD50: 10~30 μM or 5~15 μg/mL); mild (IC50/LD50: 30~50 μM or 15~25 μg/mL); weak (IC50/LD50: 50~100 μM or 25~50 μg/mL). The comparative potencies of positive controls were referred when they were available). produced by marine-derived Penicillium species, and on their cytotoxicity mechanisms, biosyntheses, and chemical syntheses.


Introduction
The oceans, which occupy more than 70% of the earth's surface, undoubtedly support vast habitats and serve as prolific resources of various living organisms. Compared to terrestrial organisms, marine organisms often produce highly potent metabolites with unique structures to enable them to adapt to extremely challenging environments [1]. Developments and improvements made in biotechnology have led to a new era of bioprospecting for new marine products. Revolutionary target screening methods have improved the efficiency of drug discovery. In addition, leading edge genomics of biological symbiosis offer more opportunities to discover drug candidates and precursors. Marine endozoic microorganisms represent a new frontier in the discovery of pharmaceutical agents [2]. In particular, marine-derived fungi are excellent producers of biologically active secondary metabolites. Since the isolation of the broad-spectrum antibiotic, cephalosporin C from the marine-derived fungus Acremonium chrysogenum, thousands of bioactive metabolites have been discovered and evaluated [3].
Cancer is the second leading cause of death. Lung, prostate, colorectal, and digestive tract cancer are commonly encountered in males, whereas breast, lung, and cervical cancer are the major causes of female death. Marine microorganisms produce limited amounts of highly efficient toxic substances to protect their hosts from enemies, and these substances have been investigated as potential anticancer drug precursors. In particular, marine-derived Penicillium species represent a major source of cytotoxic metabolites. In this review, we list all cytotoxic or antitumor secondary metabolites isolated from marine-derived Penicillium species and classify them into distinct chemical groups. In addition, we summarize the cytotoxicity mechanisms and proposed biosyntheses of these metabolites. Overall, more than 200 natural products and their synthetic analogues are included in this review.

Polyketides
Chromone derivatives are abundantly present in nature and are considered potential immunomodulatory, anticancer, and anti-inflammatory agents. Chromone scaffolds were reported to possess outstanding pharmacological properties [86].

Conclusions
The rapid development of marine biotechnology and ever increasing needs of industrial applications resulted in the emergence of marine natural products as alternative drug sources in the early 1990s [146]. Marine-associated microorganisms are sensitive to culture conditions; therefore, strains living in extremely competitive environments tend to provide high potency leads (compound 154 in this review inhibited OVCAR-3 cell line at nanomolar concentrations). Furthermore, the activation of silent gene clusters may activate new biosynthetic pathways that produce compounds with novel structure, which provide equally valid leads (compounds 44 and 45, which have unique skeletons, had cytotoxic effects in the five cancer cell lines with IC 50 values of~10 µM). Interestingly, the halogenation of compound 31, which was completely inactive, produces compound 32, which exhibited a much greater potency (compound 32 had significant cytotoxicity in 22Rv1 cells at nanomolar levels) [147].
The genus Penicillium has been explored for antitumor leads in recent years [148]. However, the marine ecological diversity of this genus offers more opportunities for drug discovery. This review includes more than 200 cytotoxic or antitumor compounds isolated from marine Penicillium fungus and chemically synthesized analogues. Of these, the major metabolites are alkaloids, particularly diketopiperazine alkaloids and indole alkaloids (Appendix A, Table A1). Cytochalasan alkaloids, which are indole alkaloids, constitute a large class of mycotoxins that exhibit significant cytotoxicity against P388 cells (IC 50 < 1 µg/mL). Furthermore, a series of diketopiperazine alkaloids, gliotoxin analogues, and roquefortine analogues with remarkable cytotoxicity at nanomolar levels are potential anticancer leads. Terpenoid metabolites appear to be more effective against cancer cell lines than steroids; in particular, compounds 99, 104, and 132 were effective at nanomolar levels. Furthermore, citrinins (chromone analogues) and their derivatives, which are polyketide mycotoxins, possess excellent cytotoxic activities. Penostatins (cytotoxic polyketides) are cytotoxic to P388 cells with IC 50 values of~1 µg/mL. With the exception of 210 and 211, lipopeptides exhibited moderate cytotoxicity. In addition, the Penicillium genus can produce polyphenolic compounds (terrestrols) with pronounced cytotoxicity.
Although our review includes most of the cytotoxic metabolites described in the literature, more compounds are yet to be identified in marine Penicillium sp. Different marine hosts and environments can also affect the biosynthesis of metabolites by endozoic fungi. Notably, over 99% of the symbiotic microorganisms cannot be cultured. Further investigations may utilize metagenome libraries of the host organisms to identify more metabolites produced by symbiotic microorganisms [149]. Additionally, further studies are needed to explore the functional mechanisms of the bioactive compounds and to optimize their production.