Selective Oxidative Dearomatization of Angular Tetracyclic Phenols by Controlled Irradiation under Air: Synthesis of an Angucyclinone-Type Double Peroxide with Anticancer Properties

Article abstract of DOI:10.1021/acs.orglett.8b02515

Angular tetracyclic p-peroxyquinols, p-quinols, and a pentacyclic double peroxide, showing anticancer properties, were synthesized from the corresponding phenols by an environmentally friendly solvent- and wavelength-controlled irradiation under air in th

Full text of DOI:10.1021/acs.orglett.8b02515

Letter
pubs.acs.org/OrgLett
Cite This: Org. Lett. XXXX, XXX, XXX−XXX
Selective Oxidative Dearomatization of Angular Tetracyclic Phenols
by Controlled Irradiation under Air: Synthesis of an Angucyclinone-
Type Double Peroxide with Anticancer Properties
†
‡
‡
,†,§
́
María J. Cabrera-Afonso, Silvia R. Lucena, Angeles Juarranz, Antonio Urbano,*
and M. Carmen Carren
o*,†,§
̃
†
́
́
Departamento de Química Organica, Universidad Autonoma de Madrid (UAM), Cantoblanco, 28049-Madrid, Spain
‡Departamento de Biología, UAM, Cantoblanco, 28049-Madrid, Spain
§Institute for Advanced Research in Chemical Sciences (IAdChem), UAM, Cantoblanco, 28049-Madrid, Spain
S
* Supporting Information
ABSTRACT: Angular tetracyclic p-peroxyquinols, p-quinols,
and a pentacyclic double peroxide, showing anticancer
properties, were synthesized from the corresponding phenols
by an environmentally friendly solvent- and wavelength-
controlled irradiation under air in the absence of an external
photosensitizer.
As part of our continuing efforts toward the synthesis of
ngucyclines and their aglucones, named angucyclinones,
Aare a group of natural quinones of poliketide origin, which
exhibit a wide range of biological properties.1 Common
features of their structure are a benz[a]anthracene ABCD
angular tetracyclic skeleton with a methyl group at C3 and
oxygen functionalities at C1, C7, C8, and C12. Due to biological
activity and challenging structures, extensive studies on their
synthesis1,2 have been reported. Among them, there is a
subgroup incorporating oxygenated substituents at the angular
4a and 12b positions such as Aquayamycin (Scheme 1). The
presence of hydroxyls at the junction of the A and B rings still
represents a significant synthetic challenge.3 Only a few
synthetic model studies,4 together with several total
syntheses,5,6 have been reported so far.
angucyclinones,7 we have reported a novel synthetic approach
to tricyclic models of derivatives with angular hydroxyls based
on the oxidative dearomatization mediated by the system
Oxone/NaHCO3/CH3CN, as a source of singlet oxygen, of an
angular tricyclic phenol such as 1 (Scheme 1a).8,9 This key
step allowed selective access to p-peroxyquinol 2, a common
precursor of six differently substituted oxygenated tricyclic core
models of natural angucyclinones. These results led us to
consider a similar route to tetracyclic models of angucyclinones
starting from adequately substituted angular tetracyclic phenols
such as 3 (Scheme 1b).
Herein, we report the oxidative dearomatization of
tetracyclic phenols 3 into p-peroxyquinols 4 using our previous
methodology with Oxone. We also describe a new environ-
mentally friendly photooxidation process without any added
photosensitizer to achieve the same oxidative dearomatization
process and the unprecedented and selective formation of a
pentacyclic double peroxide 5 from 3a by judicious choice of
the solvent and the wavelength of the light source. Biosynthetic
studies carried out by Rohr on Aquayamycin10 by culturing the
Streptomyces precursor under a 18O2 atmosphere, revealed
incorporation of the heavy isotope at the C12b position. The
oxygenated groups introduced at C12b in our angucyclinone-
type derivatives 4 and 5 were thus incorporated in a similar
manner from molecular oxygen. Considering that peroxides
can be exploited for therapeutic benefits against cancer,11 the
Scheme 1. Oxidative Dearomatization toward Tricyclic and
Tetracyclic Models of Angucyclinones
Received: August 6, 2018
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.8b02515
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
cytotoxicity of 5 was tested in selected cancer cell lines. Other
angucyclinones12 have demonstrated antitumoral activity,
producing apoptosis in various cancer cell lines (MDA-MB-
231, A549, and HT29).
When tetracyclic phenol 3a (see Supporting Information
(SI) for the synthesis) was submitted to the oxidative
dearomatization under our previous conditions (Oxone/
NaHCO3/CH3CN-H2O),8,9 no evolution occurred. By chang-
ing the solvent to acetone and working in the dark (phenol 3a
was shown to be photosensitive), a 50:50 mixture of p-
peroxyquinol 4a and p-quinol 6a resulted (Scheme 2), which
after treatment with NaI allowed us to obtain p-quinol 6a, with
the angular OH at C12b (76% overall yield from 3a).
in a highly stereoselective manner. The structure of 5 was
confirmed by X-ray diffraction (CCDC 1832265), evidencing
that both oxygen molecules had been incorporated from the
same face.
The double oxidative dearomatization of rings B and C of 3a
upon irradiation with sunlight (Scheme 3) contrasted with that
obtained with visible light (Scheme 2), where only ring B of
phenol 3a was oxidized. We reasoned that such a difference
could be due to the range of UV wavelength provided by
sunlight. Then, we decided to evaluate other light sources with
maximum emissions at the near-UV to achieve the double
oxidative dearomatization process. Thus, irradiation of phenol
3a with blue LEDs (λmax 450 nm) in CHCl3 under air
furnished, in only 2 h, the double peroxide 5 in an improved
47% yield (Scheme 3). A similar result was obtained upon
irradiation under air for 3a with a UV lamp (λmax 370 nm) in
CHCl3 for 6 h, which gave rise to derivative 5, in 39% yield.
Surprisingly, when phenol 3a was irradiated with blue LEDs
under air using acetone instead of CHCl3, the p-peroxyquinol
4a was formed in only 15 min (70% yield, Scheme 3). This
result, as well as the formation of p-peroxyquinol 4a in CHCl3
by irradiation with visible light, suggested that p-peroxyquinol
4a could be the intermediate in the double oxidative
dearomatization of phenol 3a leading to double peroxide 5.
This was confirmed by irradiation of p-peroxyquinol 4a in
CHCl3, with blue LEDs under air affording compound 5 in
only 20 min and quantitative yield. Further treatment of p-
peroxyquinol 4a with NaI in THF gave p-quinol 6a in 76%
yield.
Scheme 2. Oxidative Dearomatization of Phenol 3a with
Oxone/NaHCO3/Acetone or Irradiation with Visible Light
The observation that phenol 3a was photosensitive and
evolved into mixtures of products upon exposure to light (see
SI) encouraged us to investigate this photochemical process in
depth. Thus, the exposition of a CHCl3 solution of phenol 3a
to irradiation by laboratory fluorescent light under air for 24 h
gave rise to an unseparable 75:25 mixture of 4a and 6a, whose
reduction with NaI afforded tetracyclic p-quinol 6a (44%
isolated yield from 3a, Scheme 2). When a household lamp
was used as the light source under identical conditions, a
slightly lower 34% isolated yield of p-quinol 6a was obtained.
More interestingly, when phenol 3a was exposed to sunlight,
in CHCl3 under air, a new oxidized product, the pentacyclic
double peroxide 5, was obtained after 6 h, in 31% isolated yield
(Scheme 3). In this case, a double oxidative dearomatization at
rings B and C of 3a had taken place, with incorporation of two
molecules of oxygen and formation of three stereogenic centers
The system Oxone/NaHCO3/CH3CN had been shown to
provide singlet oxygen (1O2) which, upon reaction with p-alkyl
phenols, led to p-peroxyquinols through the corresponding
endoperoxides.9 We reasoned that p-peroxyquinol 4a and
double peroxide 5 could be also formed by reaction with O2
generated by aerobic irradiation of phenol 3a, which could act
as a self-sensitizer.
1
To gain insight into the reaction mechanism, we carried out
1
several studies (see SI). The presence of O2 in the reactions
effected under our photochemical conditions was confirmed by
a trapping experiment with 9,10-dimethylanthracene, which
afforded the corresponding endoperoxide.13 The role of 1O2 as
the oxidant in the synthesis of peroxide 5 from 3a was
supported by the increased reaction rate (15 min vs 2 h)
observed when irradiation with blue LEDs of 3a was
performed in a deuterated solvent (see SI), which is known
14
1
1
to increase the lifetime of O2. The presence of O2 in the
formation of 4a from 3a was also supported by addition of the
1O2 quencher 1,4-diazabicyclo[2.2.2]octane (DABCO),15
which greatly reduced the product yield (a 64% of starting
phenol 3a remained unchanged, see SI). We also checked the
influence of DABCO and the radical scavenger TEMPO in the
formation of 5 from 4a. When p-peroxyquinol 4a was
irradiated in CHCl3 in the presence of DABCO or TEMPO,
the starting hydroperoxide 4a remained unchanged (see SI).
These results suggested that 1O2 could be the oxidant
responsible for the formation of the p-peroxyquinol 4a which
could later evolve into double peroxide 5 by a radical pathway,
Scheme 3. Oxidative Dearomatization of 3a by a Solvent-
and Wavelength-Controlled Irradiation under Air
1
with O2 having an essential role.
1
Generation of O2 under our conditions could only occur if
3a was acting as phothosensitizer, after excitation by light. UV
spectra of 3a indicated bands at λ 330−450 nm, almost
overlapping with the blue LEDs emission (λ 370−490 nm, see
SI). Accordingly, upon blue LED irradiation, phenol 3a could
B
DOI: 10.1021/acs.orglett.8b02515
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
itself act as a photosensitizer producing the photoexcited
1O2.16
Scheme 6. Aerobic Photooxidation of 7 and 3c
Based on these results, we can advance a possible
mechanism for the formation of 4a and 5 from 3a (Scheme
1
4). First, reaction of phenol 3a with O2, through a [4 + 2]
Scheme 4. Mechanistic Proposal for the Aerobic
Photooxidation of 3a into 4a and 5
more electron-rich C ring of 7, probably through the
endoperoxide intermediate 8, which could not be detected.
On the other hand, irradiation of the 1-oxo substituted
tetracyclic phenol 3c (see SI for the synthesis) under similar
conditions gave quinone 10 (6 h, 99% yield) resulting from the
oxidation of the more electron-rich p-dimethoxy substituted C
ring. When such irradiation was effected in acetone, the
endoperoxide 11 could be isolated in almost quantitative yield.
Formation of 10 in CHCl3 could likely be due to a radical
homolytic cleavage of the O−O bond on the undetected
endoperoxide 11.
The anticancer properties of peroxide 5 were tested in three
established human cell lines, larynx Hep-2, breast MDA-MB,
and cervix HeLa cells. First, we evaluated the cytotoxicity of
the new angucyclinone-type derivative 5 in these cell lines by
using two different conditions of treatment: long incubation
period (24 h) with a low concentration of drug (10−8 M) and a
short-term incubation (5 h) with higher drug concentration
(10−7 M, 5 × 10−7 M, 2.5 × 10−7 M, and 10−6 M). The final
concentration of acetone in the culture medium was always
lower than 5%.
cycloaddition, selectively occurred at the most electron-rich p-
alkyl substituted phenol B ring, leading to the corresponding
1,4-endoperoxide which evolved into p-peroxyquinol 4a. Once
4a was formed, a radical process favored by irradiation in the
chlorinated solvent and triggered by 1O217 through abstraction
of the hydrogen of the hydroperoxide 4a afforded the peroxy
radicals [4a]· and HOO·. Intramolecular attack of [4a]· to the
C12 position from the face containing the peroxy radical gave
the cyclic peroxide moiety. This attack triggers the movement
of electrons represented, favoring reaction at C7 with the
HOO· situated on the same face to give the bis-peroxide 5 and
explaining the exclusive formation of the diastereomer with the
cis relative configuration of both peroxides.18
When treating cells with 10−8 M for 24 h, only Hep-2 and
MDA-MB lines significantly decreased their survival compared
with the untreated control cells (see SI). The most sensible
line was MDA-MB, with a percentage of 25% of lethality.
When incubating cells for 5 h, the survival rate was dependent
on the drug concentration. No effects were seen for DMEM/
acetone 5%. The lower concentration of the drug tested (10−7
M) did not cause a relevant effect in any of the lines; the
induced toxicity was lower than 10%. However, the rest of the
tested concentrations were highly effective, inducing more than
90% of toxicity for all cell lines, with the exception of Hep-2
cells treated with concentrations of 2.5 × 10−7 M in which only
70% of cell death was detected. Thus, with long-term
incubation, the most sensitive cell line in terms of survival
was MDA-MB and Hep-2 was the most resistant (Figure 1a,b).
This drug demonstrates an extremely high effectivity at
short-term treatment from a concentration of 2.5 × 10−7 M,
being even more effective (in terms of lethality) than
Doxorubicin (Doxo), whose use is extended in patients with
cancer. Doxo presented an IC50 of 5 × 10−7 M in two
mammary cancer cell lines (MDA-MB-231 and MCF-7),19 and
it was even higher in other studies, presenting an IC50 of 1.84
× 10−5 M in MDA-MB-231,20 between 9.16 × 10−6 M and
5.46 × 10−6 M for Hep-2, HeLa, and MCF-7 cell lines.21 From
comparison of the efficiency of Doxo with four angucyclinones
of new synthesis employed for the treatment of the breast
MCF-7 cell line, again, peroxide 5 was observed to be more
lethal at lower concentrations, presenting these four com-
pounds with an IC50 between 3.4 × 10−7 M and 51.3 × 10−7
M.22
We also performed the oxidative dearomatization of 8-
methoxy-substituted phenol 3b (see SI for the synthesis) with
Oxone/K2CO3/acetone−H2O followed by reduction with NaI,
giving p-quinol 6b with a moderate 30% global yield for the
two steps (Scheme 5). Fortunately, irradiation of phenol 3b
Scheme 5. Oxidative Dearomatization of Phenol 3b with
Oxone or Photooxidation with Blue LEDs
with blue LEDs under air in acetone for 15 min furnished p-
peroxyquinol 4b in 81% yield. Further reduction of 4b with
NaI gave p-quinol 6b in 51% yield.
The interest of such photooxidation led us to extend the
study to other analogues (Scheme 6). Thus, irradiation of the
benzyl-protected derivative 7 (see SI for the synthesis) with
blue LEDs (450 nm) under air in CHCl3 gave quinone 9 (30
min, 48% yield), resulting from the exclusive oxidation of the
C
DOI: 10.1021/acs.orglett.8b02515
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
ASSOCIATED CONTENT
* Supporting Information
■
S
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.8b02515.
Synthetic procedures, characterization data, and copies
1
of H and 13C NMR spectra (PDF)
Accession Codes
CCDC 1832265 contains the supplementary crystallographic
data for this paper. These data can be obtained free of charge
via www.ccdc.cam.ac.uk/data_request/cif, or by emailing
data_request@ccdc.cam.ac.uk, or by contacting The Cam-
bridge Crystallographic Data Centre, 12 Union Road,
Cambridge CB2 1EZ, UK; fax: +44 1223 336033.
AUTHOR INFORMATION
Corresponding Authors
■
*E-mail: antonio.urbano@uam.es.
*E-mail: carmen.carrenno@uam.es.
ORCID
Figure 1. Effect of Peroxide 5 on larynx Hep-2, breast MDA-MB, and
cervix HeLa after a short incubation period with variable
concentrations of drug: (a) morphological changes induced in the
tumoral cells; (b) cell survival after treatments evaluated by the MTT
assays; (c) morphology of cell nuclei after treatments evaluated under
the fluorescence microscopy.
Antonio Urbano: 0000-0003-2563-1469
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
We thank MINECO (Grants CTQ2017-83309-P, CTQ2014-
53894-R, and FIS PI15/00974) for financial support.
■
We have also evaluated the changes induced in the cell and
nuclear morphology after treatments. Control HeLa and Hep-2
cells present a polygonal morphology, typical of keratinocytes,
while the features of the MDA-MB cells were more spindled.
These morphologies were maintained in cultures treated with
low drug concentrations and in control acetone (5%). In
incubations with drug concentrations of 2.5 × 10−7 M, cells
become rounded and detach from the substrate. Conversely,
with concentrations of 10−6 M, cells remained attached to the
well but exhibited an irregular shape. In this case, the 3D
structure of the cells seemed to be lost. Nuclear morphology
was evaluated 5 h after treatment with the highest
concentrations (5 × 10−7 M and 10−6 M). Whereas control
cells showed rounded nuclei and brilliant blue fluorescent
chromatin (after DAPI staining), treated cultures showed
nuclei with chromatin irregularly distributed forming highly
fluorescent discrete aggregates or with a very low or null blue
fluorescence, indicating a loss of the DNA content of the
nucleus (Figure 1c).
REFERENCES
■
(1) (a) Kharel, M. K.; Pahari, P.; Shepherd, M. D.; Tibrewal, N.;
Nybo, S. E.; Shaaban, K. A.; Rohr, J. Nat. Prod. Rep. 2012, 29, 264.
(b) Krohn, K.; Rohr, J. Top. Curr. Chem. 1997, 188, 127.
̃
(2) Carreno, M. C.; Urbano, A. Synlett 2005, 2005, 1.
(3) Baranczak, A.; Sulikowski, G. A. Angew. Chem., Int. Ed. 2009, 48,
6005.
(4) (a) Lebrasseur, N.; Fan, G.-J.; Oxoby, M.; Looney, M. A.;
Quideau, S. Tetrahedron 2005, 61, 1551. (b) Lebrasseur, N.; Fan, G.-
J.; Quideau, S. ARKIVOC 2004, 13, 5. (c) Kraus, G. A.; Wan, Z.
Tetrahedron Lett. 1997, 38, 6509. (d) Nicolas, T. E.; Franck, R. W. J.
Org. Chem. 1995, 60, 6904.
(5) (a) Krohn, K.; Frese, P. Tetrahedron Lett. 2001, 42, 681.
(b) Krohn, K.; Frese, P.; Florke, U. Chem. - Eur. J. 2000, 6, 3887.
(6) (a) Kusumi, S.; Nakayama, H.; Kobayashi, T.; Kuriki, H.;
Matsumoto, Y.; Takahashi, D.; Toshima, K. Chem. - Eur. J. 2016, 22,
18733. (b) Khatri, H. R.; Nguyen, H.; Dunaway, J. K.; Zhu, J. Chem. -
Eur. J. 2015, 21, 13553. (c) Matsumoto, T.; Yamaguchi, H.; Tanabe,
M.; Yasui, Y.; Suzuki, K. Tetrahedron Lett. 2000, 41, 8393.
(7) Carren
Eur. J. 2007, 13, 879 and references cited therein.
(8) Vila-Gisbert, S.; Urbano, A.; Carreno, M. C. Chem. Commun.
2013, 49, 3561.
̃
o, M. C.; Ribagorda, M.; Somoza, A.; Urbano, A. Chem. -
̃
In summary, we have discovered that the angular tetracyclic
phenols 3a−b can be directly transformed into the C12b
-
̃
́
́
(9) Carreno, M. C.; Gonzalez-Lopez, M.; Urbano, A. Angew. Chem.,
Int. Ed. 2006, 45, 2737−2740.
substituted hydroperoxides 4a−b by irradiation with blue
LEDs under air in acetone without the need of any added
photosensitizer. When irradiation of phenol 3a was effected in
CHCl3, a double oxidative dearomatization took place
incorporating two molecules of oxygen into the tetracyclic
structure of phenol 3a, in a very selective way, giving rise to the
double peroxide 5, whose anticancer properties were evaluated.
Our in vitro studies indicate the high ability of peroxide 5 to
induce lethality in different carcinoma cell lines.
(10) Udvarnoki, G.; Henkel, T.; Machinek, R.; Rohr, J. J. Org. Chem.
1992, 57, 1274.
(11) (a) Gorrini, C.; Harris, I. S.; Mak, T. W. Nat. Rev. Drug
Discovery 2013, 12, 931. (b) Raza, M. H.; Siraj, S.; Arshad, A.;
Waheed, U.; Aldakheel, F.; Alduraywish, S.; Arshad, M. J. Cancer Res.
Clin. Oncol. 2017, 143, 1789.
́
(12) Lombo, F.; Abdelfattah, M. S.; Brana, A. F.; Salas, J. A.; Rohr,
̃
́
J.; Mendez, C. ChemBioChem 2009, 10, 296.
(13) Kotani, H.; Ohkubo, K.; Fukuzumi, S. J. Am. Chem. Soc. 2004,
126, 15999.
D
DOI: 10.1021/acs.orglett.8b02515
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
(14) Hurst, J. R.; McDonald, J. D.; Schuster, G. B. J. Am. Chem. Soc.
1982, 104, 2065.
(15) Silverman, S. K.; Foote, C. S. J. Am. Chem. Soc. 1991, 113,
7672.
(16) Carney, J. M.; Hammer, R. J.; Hulce, M.; Lomas, C. M.;
Miyashiro, D. Tetrahedron Lett. 2011, 52, 352.
(17) Sun, J.-G.; Yang, H.; Li, P.; Zhang, B. Org. Lett. 2016, 18, 5114.
(18) As suggested by a reviewer, a second [4 + 2] cycloaddition with
1O2 at the C-ring of 4a, directed by the OOH, followed by cyclization
and opening of the endoperoxide formed, could be an alternative
mechanism for the second step.
(19) Sapio, L.; Sorvillo, L.; Illiano, M.; Chiosi, E.; Spina, A.;
Naviglio, S. Molecules 2015, 20, 15910.
(20) Li, Z. L.; Chen, C.; Yang, Y. C.; Wang, T.; Yang, X.; Yang, S.;
Liu, C. Int. J. Clin. Exp. Pathol. 2015, 8, 4378.
(21) Mohammed, M. M.; Ibrahim, N. A.; Awad, N. E.; Matloub, A.
A.; Mohamed-Ali, A. G.; Barakat, E. E.; Mohamed, A. E.; Colla, P. L.
Nat. Prod. Res. 2012, 26, 1565.
(22) Boonlarppradab, C.; Suriyachadkun, C.; Rachtawee, P.;
Choowong, W. J. Antibiot. 2013, 66, 305.
E
DOI: 10.1021/acs.orglett.8b02515
Org. Lett. XXXX, XXX, XXX−XXX