Marine Streptomyces-Derived Novel Alkaloids Discovered in the Past Decade

Natural alkaloids originating from actinomycetes and synthetic derivatives have always been among the important suppliers of small-molecule drugs. Among their biological sources, Streptomyces is the highest and most extensively researched genus. Marine-derived Streptomyces strains harbor unconventional metabolic pathways and have been demonstrated to be efficient producers of biologically active alkaloids; more than 60% of these compounds exhibit valuable activity such as antibacterial, antitumor, anti-inflammatory activities. This review comprehensively summarizes novel alkaloids produced by marine Streptomyces discovered in the past decade, focusing on their structural features, biological activity, and pharmacological mechanisms. Future perspectives on the discovery and development of novel alkaloids from marine Streptomyces are also provided.


Introduction
Extensive marine habitats differ greatly from the land in terms of temperature, pressure and inorganic salt content, thereby providing a wealth of ecological and biogenetic diversity [1].Benefiting from technological advances in deep-sea resource extraction and microbial culture methods, the number of new microbial species with unique metabolisms has constantly expanded in recent years [2].Natural products of marine microbial origin are more likely to have novel skeletons and significant pharmacological activity [3].It is estimated that at least 30,000 compounds with therapeutic potential have been isolated from marine microorganisms; some of these substances have been used as lead compounds or biomaterials in new therapies such as drug-resistant cancer treatment [4,5].Actinomycetes are one of the largest phyla of bacterial groups and are ubiquitous in both terrestrial and marine ecosystems [6].Their biosynthetic gene clusters (BGCs) have well-known abilities in the metabolization of complex natural products [7].Streptomyces, as the largest and most advanced genus of actinomycetes, is the source of 60% of natural antibiotics, and classic examples widely are used in clinical practice, including erythromycin, streptomycin and rifamycin [8,9].In the past thirty years, the number of novel metabolites produced with marine Streptomyces as a percentage of the total source has increased from 23.0% to 40.1% per decade (Figure 1), suggesting that these Streptomyces strains play an increasingly important role in the production of new natural products.
Alkaloids are the main chemical constituents in the secondary metabolites (SMs) of actinomycetes, and one of the compounds known to have the highest degree of druggability [10].Most of these nitrogen-containing molecules have complex ring structures with promising pharmacological activity [11].It has been demonstrated that marine alkaloids have widely clinical application value in the treatment of cancer, microbial infection, cardiovascular disease, inflammation, etc [12].With the development of microbial genomics and metabonomics, the biosynthetic potential of marine Streptomyces has been deeply explored in the past decade.This review first provides a comprehensive overview of all new alkaloids produced by marine Streptomyces strains reported between January 2013 and June 2023.Three core databases (Web of Science, SciFinder and Dictionary of Natural Products) were used to search the targeted literature pertaining to specific topics related to marine and Streptomyces as well as novel or new alkaloids.According to their chemical structures, these metabolites (1-261) are grouped into nine types including indole, pyrrole, oxazole and thiazole, pyridine, pyrazine and piperazine, phenazine and phenoxazine, indolizidine and pyrrolizidine, amide and miscellaneous alkaloids.It is notable that indole, pyrrole, pyridine and amide are the major types, accounting for 72.9% of marine Streptomyces-derived novel alkaloids (Figure 2a).Detailed information for these substances is summarized in Table S1.In addition, Streptomyces strains isolated from marine sediments have a numerical advantage (Figure 2b); the number of new alkaloids produced by marine Streptomyces had a short-lived upward trend, but has fallen in the last two years (Figure 2c).Alkaloids are the main chemical constituents in the secondary metabolites (SMs) of actinomycetes, and one of the compounds known to have the highest degree of druggability [10].Most of these nitrogen-containing molecules have complex ring structures with promising pharmacological activity [11].It has been demonstrated that marine alkaloids have widely clinical application value in the treatment of cancer, microbial infection, cardiovascular disease, inflammation, etc [12].With the development of microbial genomics and metabonomics, the biosynthetic potential of marine Streptomyces has been deeply explored in the past decade.This review first provides a comprehensive overview of all new alkaloids produced by marine Streptomyces strains reported between January 2013 and June 2023.Three core databases (Web of Science, SciFinder and Dictionary of Natural Products) were used to search the targeted literature pertaining to specific topics related to marine and Streptomyces as well as novel or new alkaloids.According to their chemical structures, these metabolites (1-261) are grouped into nine types including indole, pyrrole, oxazole and thiazole, pyridine, pyrazine and piperazine, phenazine and phenoxazine, indolizidine and pyrrolizidine, amide and miscellaneous alkaloids.It is notable that indole, pyrrole, pyridine and amide are the major types, accounting for 72.9% of marine Streptomyces-derived novel alkaloids (Figure 2a).Detailed information for these substances is summarized in Table S1.In addition, Streptomyces strains isolated from marine sediments have a numerical advantage (Figure 2b); the number of new alkaloids produced by marine Streptomyces had a short-lived upward trend, but has fallen in the last two years (Figure 2c).Alkaloids are the main chemical constituents in the secondary metabolites (SMs) of actinomycetes, and one of the compounds known to have the highest degree of druggability [10].Most of these nitrogen-containing molecules have complex ring structures with promising pharmacological activity [11].It has been demonstrated that marine alkaloids have widely clinical application value in the treatment of cancer, microbial infection, cardiovascular disease, inflammation, etc [12].With the development of microbial genomics and metabonomics, the biosynthetic potential of marine Streptomyces has been deeply explored in the past decade.This review first provides a comprehensive overview of all new alkaloids produced by marine Streptomyces strains reported between January 2013 and June 2023.Three core databases (Web of Science, SciFinder and Dictionary of Natural Products) were used to search the targeted literature pertaining to specific topics related to marine and Streptomyces as well as novel or new alkaloids.According to their chemical structures, these metabolites (1-261) are grouped into nine types including indole, pyrrole, oxazole and thiazole, pyridine, pyrazine and piperazine, phenazine and phenoxazine, indolizidine and pyrrolizidine, amide and miscellaneous alkaloids.It is notable that indole, pyrrole, pyridine and amide are the major types, accounting for 72.9% of marine Streptomyces-derived novel alkaloids (Figure 2a).Detailed information for these substances is summarized in Table S1.In addition, Streptomyces strains isolated from marine sediments have a numerical advantage (Figure 2b); the number of new alkaloids produced by marine Streptomyces had a short-lived upward trend, but has fallen in the last two years (Figure 2c).

Indoles
Indoles are bicyclic alkaloids that usually use tryptophan or tryptamine as biosynthetic precursors [13].They are common and grow rapidly in the SMs of marine-derived microorganisms with a wide range of biological activities [14].Compounds 1-55 have been described as indole alkaloid derivatives of marine Streptomyces origin, including bisindole, indole sesquiterpenoid, and other miscellaneous indoles.

Bisindoles
Bisindole alkaloids represent a family formed by the oxidation and polymerization of two L-tryptophan molecules, which have more pronounced biological activities compared to the indole monomer structure [15].These alkaloids have been reported to have cytotoxic, antibacterial, and antiviral activities; bisindole derivatives containing triazine groups have been used as targeted pyruvate dehydrogenase kinase (PDK) inhibitors in the treatment of pancreatic ductal adenocarcinoma [16,17].This chapter highlights marine Streptomycesderived staurosporine analogues and chlorinated bisindoles.

Halogenated Bisindoles
To the best of our knowledge, all bisindoles from marine Streptomyces are chlorinated (25-39, Figure 4).Indimicins A−E (25-29) and lynamicins F-G (30 and 31) were obtained from a deep-sea-derived Streptomyces sp.SCSIO 03032 by solid phase extraction with XAD-16 resin [28].These compounds had unusual 1 ′ ,3 ′ -dimethyl-2 ′ -hydroindole structures.Only dimethyl-substituted indimicin B ( 26) was seen to have antitumor activity against the MCF-7 cell line with an IC 50 value of 10.0 µM.In order to characterize the function of gene spmH, which was predicted to be an L-Trp 5-halogenase, the authors inactivated this gene in strain SCSIO 03032 and obtained four bisindoles without a halogen substituent named spiroindimicins G-H (32)(33) and indimicins F-G (34-35) [29].It was confirmed that spmH functioned as halogenase and acted in early biosynthesis using L-Trp as a substrate.Compounds 32 and 33 showed various degrees of cytotoxicity against four cancer cell lines (SF-268, MCF-7, HepG2 and A549) and the presence or absence of chlorine atoms had no significant effect on the cytotoxic activity.Two non-typical bisindole spiroindimicins E and F (36 and 37) were purified from the metabolites of strain MP131-18 with cluster 36 being predicted to be the gene cluster responsible for bisindole biosynthesis [30].Only compound 36 showed weak cytotoxic activity against T24 bladder cancer cells, while 36-37 had no antimicrobial activity due to this type of effect was proportional to the amount of halogenation.In addition, Streptomyces sp.SCSIO 11791 produced two chlorinated bisindoles (38 and 39), which displayed moderate cytotoxicity against four tumor cells with IC50 values ranging from 2.9 µM to 19.4 µM [31].Compound 38 additionally exhibited cytotoxic activity against MDA-MB- Two non-typical bisindole spiroindimicins E and F (36 and 37) were purified from the metabolites of strain MP131-18 with cluster 36 being predicted to be the gene cluster responsible for bisindole biosynthesis [30].Only compound 36 showed weak cytotoxic Mar.Drugs 2024, 22, 51 5 of 26 activity against T24 bladder cancer cells, while 36-37 had no antimicrobial activity due to this type of effect was proportional to the amount of halogenation.In addition, Streptomyces sp.SCSIO 11791 produced two chlorinated bisindoles (38 and 39), which displayed moderate cytotoxicity against four tumor cells with IC 50 values ranging from 2.9 µM to 19.4 µM [31].Compound 38 additionally exhibited cytotoxic activity against MDA-MB-231 and NCI-H460 cell lines as well as inhibition of Gram-positive bacteria.The structureactivity relationship suggested that the substitution of the C-6 ′′ position by the chlorine atom was more beneficial to the biological activity.

Indole Sesquiterpenoids
The first bacterial-derived indole sesquiterpenoid alkaloid, named xiamycin, was isolated from an endophytic Streptomyces sp. from B. gymnorrhiza in 2010 [32].In recent years, compounds of this family have been found in marine Streptomyces, and have attracted attention because of their antibacterial, anti-human-immunodeficiency-virus (HIV) and anti-tumor activities [33].Xiamycins C-E (40-42, Figure 5) were isolated from Streptomyces sp.HK-18 [34].Compounds 41 and 42 exhibited strong activity against the replication of porcine epidemic diarrhea virus (PEDV) in a dose-dependent manner, with EC 50 values of 0.93 µM and 2.89 µM, respectively.The mechanism of action inhibited the synthesis of key structural proteins for PEDV to prevent virus replication.The methyl ester group was an important functional group to maintain activity.Overexpression of the orf2011 gene encoding the LuxR family regulator in the Streptomyces sp.HK-18 resulted in the production of two members of the xiamycins family containing an N-O bone linked aliphatic chain structure named lipoxiamycins A and B (43 and 44, Figure 5) as well as increased production of xiamycins dimers dixiamycins A and C (45 and 46, Figure 5) [35].An anti-inflammatory assay showed that compounds 43 and 45 could significantly inhibit the production of lipopolysaccharide-induced NO with IC 50 values of 9.89 ± 0.92 and 4.12 ± 0.22 µM, respectively.

Pyrroles
Pyrrole alkaloids have more potential to be designed as lead compounds due to their property of easily forming hydrogen bonds [40].Marine pyrrole derivatives usually have more unique structures and significant pharmacological activities such as antimicrobial, antiproliferative, anti-inflammatory and antiviral activities [41].

Pyrrolones and Pyrrolidones
This part describes cases in which pyrrole or pyrrolidine pentacyclic rings are replaced by carbonyl groups.The formed pyrrolone and pyrrolidone structures are crucial heterocyclic pharmacophores in medicinal chemistry, with significant biological activities [42].The tirandamycins are a class of bacterial RNA polymerase (RNAP) inhibitors containing dienoyl tetramic acid and 2,4-pyrrolidinedione structures [43].Two tirandamycin analogues tirandamycin K and isotirandamycin B (56 and 57, Figure 7), together with two known derivatives (58 and 59, Figure 7), were produced from marine Streptomyces strains [44,45].Compound 56 was the first linear tirandamycin derivative that avoided bicyclic ketal ring formation due to the inability of the C-9S hydroxyl group to be nucleophilically attacked by C-13.In a pathogenic bacterial inhibitory activity assay, compounds 57-59 showed obvious antibacterial activity against S. agalactiae with MIC values of 2.5-5.0 µg/mL.However, compound 56 was inactive, therefore the authors deduced that the bicyclic ketal ring moiety was a necessary RNA polymerase target.

Pyrroles
Pyrrole alkaloids have more potential to be designed as lead compounds due to their property of easily forming hydrogen bonds [40].Marine pyrrole derivatives usually have more unique structures and significant pharmacological activities such as antimicrobial, antiproliferative, anti-inflammatory and antiviral activities [41].

Pyrrolones and Pyrrolidones
This part describes cases in which pyrrole or pyrrolidine pentacyclic rings are replaced by carbonyl groups.The formed pyrrolone and pyrrolidone structures are crucial heterocyclic pharmacophores in medicinal chemistry, with significant biological activities [42].The tirandamycins are a class of bacterial RNA polymerase (RNAP) inhibitors containing dienoyl tetramic acid and 2,4-pyrrolidinedione structures [43].Two tirandamycin analogues tirandamycin K and isotirandamycin B (56 and 57, Figure 7), together with two known derivatives (58 and 59, Figure 7), were produced from marine Streptomyces strains [44,45].Compound 56 was the first linear tirandamycin derivative that avoided bicyclic ketal ring formation due to the inability of the C-9S hydroxyl group to be nucleophilically attacked by C-13.In a pathogenic bacterial inhibitory activity assay, compounds 57-59 showed obvious antibacterial activity against S. agalactiae with MIC values of 2.5-5.0 µg/mL.However, compound 56 was inactive, therefore the authors deduced that the bicyclic ketal ring moiety was a necessary RNA polymerase target.
rivatives, ligiamycins A and B (68 and 69, Figure 7), were obtained by co-culture of two marine-derived strains Streptomyces sp.GET02.ST and Achromobacter sp.GET02.AC [50].A biological evaluation showed that compound 68 exhibited moderate effects against S. aureus and S. enterica (both MIC values of 16 µg/mL), while compound 69 was cytotoxic against HCT116 cancer cells (IC50 = 20.1 µM).The presence of the hydroxyl group in compound 69 had an opposite effect on antitumor activity and antibacterial activity.

Ansamycins
Ansamycins are a class of important macrocyclic lactam alkaloids obtained mainly from actinomycetes, of which the more representative include the anti-tuberculosis drug laofomycin, Hsp90 inhibitor geldamycin, and anticancer drug ansamitocin P-3 [55].According to the different aromatic cores, ansamycins can be divided into benzene series and naphthalene series [56].Hygrocins are a subclassification of naphthalenic ansamycins, whose amides are five-membered or seven-membered nitrogen heterocycles [57].After knocking out the gene gdmAI responsible for the synthesis of the high-yield ansamycin analogue geldanamycin in Streptomyces sp.LZ35, hygrocins C−F (73-76, Figure 9) were produced and characterized [58].In 2015, strain LZ35 was genetically modified by overexpression of hgc1, a LAL-type activator, and was found to produce three hygrocin derivatives, hygrocins H-J (77-79, Figure 9

Ansamycins
Ansamycins are a class of important macrocyclic lactam alkaloids obtained mainly from actinomycetes, of which the more representative include the anti-tuberculosis drug laofomycin, Hsp90 inhibitor geldamycin, and anticancer drug ansamitocin P-3 [55].According to the different aromatic cores, ansamycins can be divided into benzene series and naphthalene series [56].Hygrocins are a subclassification of naphthalenic ansamycins, whose amides are five-membered or seven-membered nitrogen heterocycles [57].After knocking out the gene gdmAI responsible for the synthesis of the high-yield ansamycin analogue geldanamycin in Streptomyces sp.LZ35, hygrocins C−F (73-76, Figure 9) were produced and characterized [58].In 2015, strain LZ35 was genetically modified by overexpression of hgc1, a LAL-type activator, and was found to produce three hygrocin derivatives, hygrocins H-J (77-79, Figure 9 Three ansamycin derivatives with unprecedented skeletons, ansalactams B-D (89-91, Figure 9), were isolated from marine sediment-derived Streptomycetes sp.CNH189 [61].Antibacterial activity tests showed that compounds 89-91 had weak inhibitory activity against MRSA, with MIC values of 31.2, 31.2, and 62.5 µg/mL, respectively.A chemical study of the strain Streptomyces sp.KFD18 produced four ansamycin analogues named divergolides T-W (92-95, Figure 9) [62].All these substances showed various degrees of cytotoxic activity against cancer cell lines SGC-7901, K562, Hela and A549; compounds with hydroxylation of C-7 and R configuration of C-2 exhibited weak cytotoxic effect.

Oxazoles and Thiazoles
Oxazole and thiazole rings exhibit a wide range of pharmacological activities, such as antiproliferative, anti-inflammatory and antimicrobial activity, by binding non-covalently to many enzyme and receptor targets [69,70].Most of these alkaloids have been mentioned in other chapters, and this chapter summarizes the remaining five compounds 109-113 (Figure 11).Under LC-MS/MS molecular networking guidance, five siderophores containing oxazole or thiazole rings were isolated from S. diastaticus NBU2966 collected from marine sponge Axinellida sp.[71].However, none of these metabolites demonstrated activity against S. aureus, MRSA, B. subtilis, and P. aeruginosa.

Oxazoles and Thiazoles
Oxazole and thiazole rings exhibit a wide range of pharmacological activities, such as antiproliferative, anti-inflammatory and antimicrobial activity, by binding non-covalently to many enzyme and receptor targets [69,70].Most of these alkaloids have been mentioned in other chapters, and this chapter summarizes the remaining five compounds 109-113 (Figure 11).Under LC-MS/MS molecular networking guidance, five siderophores containing oxazole or thiazole rings were isolated from S. diastaticus NBU2966 collected from marine sponge Axinellida sp.[71].However, none of these metabolites demonstrated activity against S. aureus, MRSA, B. subtilis, and P. aeruginosa.

Oxazoles and Thiazoles
Oxazole and thiazole rings exhibit a wide range of pharmacological activities, such as antiproliferative, anti-inflammatory and antimicrobial activity, by binding non-covalently to many enzyme and receptor targets [69,70].Most of these alkaloids have been mentioned in other chapters, and this chapter summarizes the remaining five compounds 109-113 (Figure 11).Under LC-MS/MS molecular networking guidance, five siderophores containing oxazole or thiazole rings were isolated from S. diastaticus NBU2966 collected from marine sponge Axinellida sp.[71].However, none of these metabolites demonstrated activity against S. aureus, MRSA, B. subtilis, and P. aeruginosa.

Pyridines
Pyridine alkaloids are a class of important skeletons for maintaining the pharmacological activity of drugs [72].This chapter summarizes pyridine and its derivatives of marine Streptomyces sp., including pyridone, benzopyridine (quinoline), and the saturated variant, piperidine.

Quinolines
Quinolines have been designed as important skeletons in drug structures for more than two centuries [77].Classic examples are quinine, the first effective antimalarial drug in history, and the broad-spectrum antibiotic fluoroquinolone [78].This subsection provides an overview of quinoline alkaloids of marine Streptomyces sp.origin and their de-

Pyrazines
Owing to nitrogen atoms acting as hydrogen bonding acceptors and the structure being conducive to nucleophilic reaction, pyrazine is commonly used as a classical pharmacophore [86].Pyrazine derivatives have been reported for applications as antitumor drugs, diuretics, anti-inflammatory and anti-infective drugs [87].Griseusrazin A (158, Figure 15) was isolated from a strain S. griseus subsp.griseus 09-0144 and activated the expression of heme oxygenase 1 which inhibits the upstream NF-κB pathway [88].Therefore, it could downregulate the expression of related enzymes inducible nitric oxide synthase (iNOS) and COX-2 at the transcriptional level as well as the production of inflammatory mediators NO and PGE2.Compounds 159-161 (Figure 15

Pyrazines
Owing to nitrogen atoms acting as hydrogen bonding acceptors and the structure being conducive to nucleophilic reaction, pyrazine is commonly used as a classical pharmacophore [86].Pyrazine derivatives have been reported for applications as antitumor drugs, diuretics, anti-inflammatory and anti-infective drugs [87].Griseusrazin A (158, Figure 15) was isolated from a strain S. griseus subsp.griseus 09-0144 and activated the expression of heme oxygenase 1 which inhibits the upstream NF-κB pathway [88].Therefore, it could downregulate the expression of related enzymes inducible nitric oxide synthase (iNOS) and COX-2 at the transcriptional level as well as the production of inflammatory mediators NO and PGE 2 .Compounds 159-161 (Figure 15 [91].Compound 166 was inactive, which indicated that the carbonyl was the key active moiety.

Diketopiperazines
Piperazine alkaloids of biological origin are most commonly of the 2,5-diketopiperazines (2,5-DKPs) type, with a cyclodipeptide structure formed by condensation of two amino acids [92].These simple dipeptides reported for multiple biological activities have a flexible skeleton with multiple chiral centers and four hydrogen bonding sites [93].

Diketopiperazines
Piperazine alkaloids of biological origin are most commonly of the 2,5-diketopiperazines (2,5-DKPs) type, with a cyclodipeptide structure formed by condensation of two amino acids [92].These simple dipeptides reported for multiple biological activities have a flexible skeleton with multiple chiral centers and four hydrogen bonding sites [93].

Phenazines and Phenoxazines
To the best of our knowledge, phenazines and phenoxazines are mainly derived from SMs of Streptomyces and Pseudomonas isolated from soil or marine habitats [99].Most of these alkaloids are characterized by promising biological activities such as antibacterial, antiviral, antitumor and antiparasitic effects [100].Six antitumor phenoxazines venezuelines A-E (178-182, Figure 17) and maroxazinone (183, Figure 17) were isolated from two sediments-derived Streptomyces [101,102].Compound 179 showed moderate antitumor activity against five cancer cell lines with IC50 values of 5.74-9.67µM and weak activity against human hepatoma cell Bel 7042 (IC50 >10 µM).Notably, the cytotoxicity of this compound may be explained by significant upregulation of the orphan nuclear receptor Nur77 (apoptosis-associated) expression.183 showed moderate antiproliferative activity against MCF7, HEPG2 and HCT116 cell lines with IC50 values of 4.32, 2.90 and 8.51 µg/mL, respectively.Cytotoxic activity of phenoxazinones was stronger than phenoxazines and the

Phenazines and Phenoxazines
To the best of our knowledge, phenazines and phenoxazines are mainly derived from SMs of Streptomyces and Pseudomonas isolated from soil or marine habitats [99].Most of these alkaloids are characterized by promising biological activities such as antibacterial, antiviral, antitumor and antiparasitic effects [100].Six antitumor phenoxazines venezuelines A-E (178-182, Figure 17) and maroxazinone (183, Figure 17) were isolated from two sediments-derived Streptomyces [101,102].Compound 179 showed moderate antitumor activity against five cancer cell lines with IC 50 values of 5.74-9.67µM and weak activity against human hepatoma cell Bel 7042 (IC 50 >10 µM).Notably, the cytotoxicity of this compound may be explained by significant upregulation of the orphan nuclear receptor Nur77 (apoptosis-associated) expression.183 showed moderate antiproliferative activity against MCF7, HEPG2 and HCT116 cell lines with IC 50 values of 4.32, 2.90 and 8.51 µg/mL, respectively.Cytotoxic activity of phenoxazinones was stronger than phenoxazines and the increase of substituents weakened the activity.Sponges can host microorganisms colonization due to their porous structure, therefore the metabolites of sponge symbiotic microorganisms are important sources of marine natural products [103].Streptomyces sp.HB202 was isolated from the sponge Halichondria panicea and yielded three phenazine alkaloids streptophenazines I-K (184-186, Figure 17) [104].These compounds inhibited the activity of inflammatory response associated enzyme phosphodiesterase (PDE 4B) with IC 50 values ranging from 11.6 to 12.2 µM.In addition, compound 186 had antibacterial activity against B. subtilis and S. epidermidis (IC 50 = 21.6 ± 6.8 µM and 14.5 ± 2.0 µM, respectively).

Indolizidines
Indolizidine were reported to have broad biological activity, such as antitumor activity, anti-infective system disease activity, and anti-inflammatory activity [108].Most indolizidines are obtained from plants and animals, rarely from microbial sources [109].
Eight indolizidine alkaloids cyclizidines B-I (194-201, Figure 18) were detected in the EtOAc extracts of the strain Streptomyces sp.HNA39 [110].Cyclizidine C (195) showed the most promising activity against PC-3 and HCT-116 cancer cell lines, with IC 50 values of 0.52 ± 0.03 µM and 8.3 ± 0.1 µM, respectively.Moreover, compounds 195, 198, 200 and 201 exhibited moderate inhibitory activities against protein kinase ROCK2.In another report, a low-yielding indolizidine named cyclizidine J (202, Figure 18) was detected in strain HNA39 [111].This compound had an uncommon chlorine atom substitution at the C-8 position.However, 202 lacked inhibitory activity against cancer cell line PC-3 and protein kinase.A stress culture of marine hydrothermal vent actinomycetes with heavy metal ions can activate silent biosynthetic pathways [112].After the addition of 100 µmol/L Ni 2+ to the medium of metal-resistant Streptomyces sp.WU20, a cyclizidine analogue (203, Figure 18) that was absent before addition was purified [113].The authors hypothesized that the ring opening of the five-membered ring in the structure of alkaloid 203 was due to the inhibition of normal biosynthesis by heavy metal stress.Compound 203 was bacteriostatic against B. subtilis with MIC of around 32 µg/mL.Chemical analysis of symbiotic strain Streptomyces sp.HZP-2216E from fresh sea lettuce Ulva pertusa led to the discovery of an indolizinium alkaloid, streptopertusacin A (204, Figure 18), which had a 40 mg/mL MIC value against MRSA [114].The antitumor activity of alkaloids was enhanced when C-8 was replaced by a hydroxyl group, but the cytotoxicity was lost after the aromatization of the indolizidine core.

Pyrrolizidines
Pyrrolizidines are mainly derived from plants as toxic components of chemical defense [115].Bacterial-derived pyrrolizidines have been reported less frequently, with a total of 12 species and about 60 compounds of this class identified as of 2021 [116].These Pyrrolizidines are commonly biosynthesized by multidomain NRPS gene clusters and are post-modified as well by flavine adenosine dinucleotide (FAD)-dependent monooxygenases [117].
Bohemamine is a rare pyrrolizidine subtype derived only from actinomycetes [118].The fermentation broth of strain S. spinoverrucosus SNB-048 purified two bohemamine-type pyrrolizidines named spithioneines A and B (205 and 206, Figure 18) with rare ergothioneine moiety [119].In the same year, six derivatives of bohemamines D−I (207-212, Figure 18) were again isolated from strain SNB-048 [120].Unfortunately, none of the compounds showed significant activity.Tracing the cytotoxic activity of S. spinoverrucosus SNB-032 metabolites led to the isolation of an analogue, 5-Br-bohemamine C (213, Figure 18), as well as three dimeric bohemamines dibohemamines A-C (214-216, Figure 18) [121].The authors confirmed that the dimer formation was a non-enzymatic Baylis-Hillman addition reaction of monomeric compounds using formaldehyde in the medium.Compounds 215 and 216 exhibited potent cytotoxicity against an NSCLC cell line A549 with IC 50 values of 0.140 and 0.145 µM, respectively.In addition, compound 216 showed moderate activity against an HCC1171 cell line (IC 50 = 1.2 µM).Bohemamine-type pyrrolizidines exhibited significant cytotoxic activity due to the polymerization of dimer.
position.However, 202 lacked inhibitory activity against cancer cell line PC-3 and protein kinase.A stress culture of marine hydrothermal vent actinomycetes with heavy metal ions can activate silent biosynthetic pathways [112].After the addition of 100 µmol/L Ni 2+ to the medium of metal-resistant Streptomyces sp.WU20, a cyclizidine analogue (203, Figure 18) that was absent before addition was purified [113].The authors hypothesized that the ring opening of the five-membered ring in the structure of alkaloid 203 was due to the inhibition of normal biosynthesis by heavy metal stress.Compound 203 was bacteriostatic against B. subtilis with MIC of around 32 µg/mL.Chemical analysis of symbiotic strain Streptomyces sp.HZP-2216E from fresh sea lettuce Ulva pertusa led to the discovery of an indolizinium alkaloid, streptopertusacin A (204, Figure 18), which had a 40 mg/mL MIC value against MRSA [114].The antitumor activity of alkaloids was enhanced when C-8 was replaced by a hydroxyl group, but the cytotoxicity was lost after the aromatization of the indolizidine core.19) [122].Compounds 217 and 219 displayed positive activity against HCC366, A549 and HCC44 cell lines (IC 50 = 2.2-8.4 µM).For compound 218, the authors hypothesized that the reason for its inactivity was the inability of the structure to pass through the cell membrane.Antimycin is an antibiotic with antibacterial, insecticidal and anticancer activity, consisting of a rare nine-membered dilactone core [123].Antimycins E-H (220-223, Figure 19) were isolated from Streptomyces sp.THS-55, and showed extremely significant cytotoxic activity against the HeLa cell line (IC 50 < 0.1 µM) by downregulating the levels of E6/E7 oncoproteins [124].The potency was dependent on the long-chain substituent of R 2 and the acyl group of R 3 .In addition, neoantimycins A and B (224 and 225, Figure 19) were isolated from S. antibioticus and exhibited weak cytotoxic activity against the SF-268 cancer cell line [125].Bagremycin is a phenol ester formed from p-hydroxystyrene and p-hydroxybenzoic acid with antimicrobial activity [126].Bagremycins C and D (226 and 227, Figure 19) were isolated in 2017 from Streptomyces sp.Q22 [127].The following year, bagremycins F and G (228 and 229, Figure 19) were purified from coastal mud-sourced Streptomyces sp.ZZ745 [128].Compound 226 inhibited the G 0 /G 1 cell cycle in four glioma cells (U87MG, U251, SHG44 and C6) with IC 50 values of 2.2 to 6.4 µM.Furthermore, compounds 228 and 229 showed 41.8 and 67.1 µM MIC values against E. coli.One N-acetyl macrolide analogue N-acetylborrelidin B (230, Figure 19) was detected by a strain S. mutabilis MII with stronger activity against Staphylococcus warneri (18 mm zone of inhibition) [129].

Macrolactams
Macrolactams are a class of macrocyclic compounds in which amide units are integrated into a polyketide skeleton above twelve carbons [130].These compounds often contain an azacyclic core skeleton or azacyclic substituent modifications that result in alkalinity [131].

Macrolactams
Macrolactams are a class of macrocyclic compounds in which amide units are integrated into a polyketide skeleton above twelve carbons [130].These compounds often contain an azacyclic core skeleton or azacyclic substituent modifications that result in alkalinity [131].

Conclusions and Future Perspectives
Microorganisms in special habitats have cryptic and extraordinary potential for biosynthesizing unique SMs with diverse biological properties.Marine Streptomyces as an excellent producer of therapeutic agents has become a global hotspot in natural product research and continue to play a paramount role in the production of new alkaloids for drug discovery.This review comprehensively summarizes as many as 261 new alkaloids discovered in marine Streptomyces in the past decade (January 2013-June 2023).Among these metabolites, 199 compounds have promising therapeutic effects.For instance, compounds 38, 41, 42, 97, 98, 142 and 193 exhibited excellent antimicrobial activity, and 114, 188 and 220-239 displayed remarkable cytotoxic effects.In addition, compounds 5-16 showed significant selective inhibition on protein kinases PKC, ROCK2 and BTK.This work will pave the way for further development of marine Streptomyces-derived alkaloids.
Although strategies for the isolation of marine microbial metabolites have been revolutionized over the past decades, there are still difficulties to overcome.Traditional means of separating natural products are subject to randomization.The efficient isolation of novel natural products and removal of inactive known compounds have been thorny issues hindering the development of natural medicinal chemistry.The annotation of BGCs, as well as LC-MS/MS-based metabolite structure prediction methods, will remain hot research topics in this field in the future.BGCs of marine microorganisms often have low or no expression under routine laboratory culture conditions.BGC activation techniques, such as OSMAC strategy, strain co-culture, ribosome engineering, heterologous expression of gene clusters and overexpression/knockout of regulatory genes and ribosome engineering, provide methodological references to break this bottleneck.In addition to the discovery of these therapeutic agents, it is crucial to solve the problem of compound supply.Currently, in addition to the total synthesis route design of natural products, the modification of industrial production strains using genetic engineering approaches and optimization of microbial fermentation and extraction, as well as purification processes at various levels, should be employed in the preparation of these substances.

Conclusions and Future Perspectives
Microorganisms in special habitats have cryptic and extraordinary potential for biosynthesizing unique SMs with diverse biological properties.Marine Streptomyces as an excellent producer of therapeutic agents has become a global hotspot in natural product research and continue to play a paramount role in the production of new alkaloids for drug discovery.This review comprehensively summarizes as many as 261 new alkaloids discovered in marine Streptomyces in the past decade (January 2013-June 2023).Among these metabolites, 199 compounds have promising therapeutic effects.For instance, compounds 38, 41, 42, 97, 98, 142 and 193 exhibited excellent antimicrobial activity, and 114, 188 and 220-239 displayed remarkable cytotoxic effects.In addition, compounds 5-16 showed significant selective inhibition on protein kinases PKC, ROCK2 and BTK.This work will pave the way for further development of marine Streptomyces-derived alkaloids.
Although strategies for the isolation of marine microbial metabolites have been revolutionized over the past decades, there are still difficulties to overcome.Traditional means of separating natural products are subject to randomization.The efficient isolation of novel natural products and removal of inactive known compounds have been thorny issues hindering the development of natural medicinal chemistry.The annotation of BGCs, as well as LC-MS/MS-based metabolite structure prediction methods, will remain hot research topics in this field in the future.BGCs of marine microorganisms often have low or no expression under routine laboratory culture conditions.BGC activation techniques, such as OSMAC strategy, strain co-culture, ribosome engineering, heterologous expression of gene clusters and overexpression/knockout of regulatory genes and ribosome engineering, provide methodological references to break this bottleneck.In addition to the discovery of these therapeutic agents, it is crucial to solve the problem of compound supply.Currently, in addition to the total synthesis route design of natural products, the modification of industrial production strains using genetic engineering approaches and optimization of microbial fermentation and extraction, as well as purification processes at various levels, should be employed in the preparation of these substances.

Mar. Drugs 2024, 22 , x 2 of 30 Figure 1 .
Figure 1.Source and statistics of new compounds from Streptomyces over the past three decades.

Figure 1 .
Figure 1.Source and statistics of new compounds from Streptomyces over the past three decades.

Figure 1 .
Figure 1.Source and statistics of new compounds from Streptomyces over the past three decades.

Figure 2 .
Figure 2. (a) Structural classes of marine Streptomyces-derived alkaloids reported in January 2013-June 2023; (b) sources of marine Streptomyces strains; and (c) number of new alkaloids discovered between January 2013-June 2023.

Figure 2 .
Figure 2. (a) Structural classes of marine Streptomyces-derived alkaloids reported in January 2013-June 2023; (b) sources of marine Streptomyces strains; and (c) number of new alkaloids discovered between January 2013-June 2023.
Mar. Drugs 2024, 22, x 5 of 30 confirmed that spmH functioned as halogenase and acted in early biosynthesis using L-Trp as a substrate.Compounds 32 and 33 showed various degrees of cytotoxicity against four cancer cell lines (SF-268, MCF-7, HepG2 and A549) and the presence or absence of chlorine atoms had no significant effect on the cytotoxic activity.
Mar. Drugs 2024, 22, x 6 of 30 an important functional group to maintain activity.Overexpression of the orf2011 gene encoding the LuxR family regulator in the Streptomyces sp.HK-18 resulted in the production of two members of the xiamycins family containing an N-O bone linked aliphatic chain structure named lipoxiamycins A and B (43 and 44, Figure 5) as well as increased production of xiamycins dimers dixiamycins A and C (45 and 46, Figure 5) [35].An antiinflammatory assay showed that compounds 43 and 45 could significantly inhibit the production of lipopolysaccharide-induced NO with IC50 values of 9.89 ± 0.92 and 4.12 ± 0.22 µM, respectively.