Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Jan 14;64(1):741-767.
doi: 10.1021/acs.jmedchem.0c01735. Epub 2021 Jan 5.

Synthesis, Tumor Specificity, and Photosensitizing Efficacy of Erlotinib-Conjugated Chlorins and Bacteriochlorins: Identification of a Highly Effective Candidate for Photodynamic Therapy of Cancer

Ravindra R Cheruku  1 , Joseph Cacaccio  1 , Farukh A Durrani  1 , Walter A Tabaczynski  1 , Ramona Watson  1 , Kevin Siters  2 , Joseph R Missert  1 , Erin C Tracy  3 , Mykhaylo Dukh  1 , Khurshid Guru  4 , Richard C Koya  5 , Pawel Kalinski  5 , Heinz Baumann  3 , Ravindra K Pandey  1
Affiliations

Synthesis, Tumor Specificity, and Photosensitizing Efficacy of Erlotinib-Conjugated Chlorins and Bacteriochlorins: Identification of a Highly Effective Candidate for Photodynamic Therapy of Cancer

Ravindra R Cheruku et al. J Med Chem. .

Abstract

Erlotinib was covalently linked to 3-(1'-hexyloxy)ethyl-3-devinylpyropheophorbide-a (HPPH) and structurally related chlorins and bacteriochlorins at different positions of the tetrapyrrole ring. The functional consequence of each modification was determined by quantifying the uptake and subcellular deposition of the erlotinib conjugates, cellular response to therapeutic light treatment in tissue cultures, and in eliminating of corresponding tumors grown as a xenograft in SCID mice. The experimental human cancer models the established cell lines UMUC3 (bladder), FaDu (hypopharynx), and primary cultures of head and neck tumor cells. The effectiveness of the compounds was compared to that of HPPH. Furthermore, specific functional contribution of the carboxylic acid side group at position 172 and the chiral methyl group at 3(1') to the overall activity of the chimeric compounds was assessed. Among the conjugates investigated, the PS 10 was identified as the most effective candidate for achieving tumor cell-specific accumulation and yielding improved long-term tumor control.

PubMed Disclaimer

Figures

Figure 1:
Figure 1:
Structures of the starting chlorins and bacteriochlorin used in present study
Figure 2
Figure 2
Uptake and retention by HNT1 cells (passage 116). Confluent cell cultures were incubated for 30 min in serum-free medium containing 3.2 μM of the indicated PS. The cellular level and subcellular distribution of PS fluorescence indicates the binding to and mode of uptake by the cells. Subsequent treatment of the cells for 5 h with serum and PS-containing medium determines the competing effect of serum proteins on uptake and steady-state accumulation of the PSs. Follow-up incubation for 15 h in serum-containing medium but without PS defines the rate of egress of PS from cells. PS fluorescence is recorded at 100X magnification by different-length exposure of the camera. Images allow digital quantification of PS fluorescence level.
Figure 3.
Figure 3.
Cell type specific retention of Erlotinib-conjugates. Five-day old co-culture of FaDu-49 and CFSE-stained HN-121 T-Fb were incubated with medium containing 10% FBS and 1.6 μM PS followed by a 48-h chase in medium without PS. Cell-associated fluorescence signals were recorded.
Figure 4
Figure 4
Five-day old cocultures of HN-143 T-EC with HN-166 T-Fb were incubated with medium containing 10% FBS and 3 mM indicated PS for 5 h and then chased for 40 h with medium without PS. Phase contrast images at 100X magnification (upper panel in each section) and corresponding fluorescent images of all cultures (500 ms exposure time) were taken at the indicated time points (lower point in each section).
Figure 5
Figure 5
PS-dose dependent photoreaction in HN-85-1-3 T-EC. Level of PS taken up was quantified by fluorescence prior to 665 nm light treatment (3 J/cm2). A, Immunoblot analysis of STAT3 and EGFR immediately after light treatment. B, Duplicate cultures were incubated for 24 h after PDT and then the percentage of surviving cells determined.
Figure 6:
Figure 6:
Compounds selected to determine the impact of structural requirements for tumor-uptake and in vivo PDT efficacy.
Figure 7:
Figure 7:
In vivo uptake in tumor, liverand skin (female SCID mice bearing UMUC3 tumor) of PS 10 at the dose of 0.47, 0.25 and 0.125 mmole/kg (A, B & C); PS 11 at a dose of 0. 47 mmole/kg (D); and PS 19 and HPPH at a dose of 0.47 mmol/kg (E & F).The comparative in vivo efficacy of PS 10 at variable drug dose (0.47, 0.25 and 0.125 mmol/kg), PS 19 and HPPH at a dose of 0.47 mole/kg is shown in Figure 7G. All tumors were exposed with the same light dose (665 nm, 135J/cm2,75 mW/cm2), at the time of maximal tumor uptake of the PS. For PS-uptake study (tumor, liver and skin), 3 female SCID mice bearing UMUC3 tumors were used (Figures 7A-7F). For determining in-vivo efficacy the number of female SCID mice used in each study are indicated in the Figure 7G). Statistical analysis; log rank (Mantel-Cox test), p-value:<0.0001.
Figure 8:
Figure 8:
In vivo uptake of PS 10 at the dose of 0.47, 0.25 and 0.125 μmole/kg and HPPH in tumor, liver and skin of SCID mice bearing FaDu tumors (3 mice/group), is shown in A, B, C and D respectively. The in vivo PDT efficacy (long-term tumor response) of PS 10 at variable doses: 0.47, 0.25 and 0.125 μmole/kg and HPPH at a dose of 0.47 μmole/kg is shown in Figure 8E. Tumors were exposed to light (665 nm, 135 J/cm2, 75 mW/ cm2) at 24h post injection and tumor-growth was measured daily for 60 days. For PS-uptake study (tumor, liver and skin), 3 female SCID mice bearing FaDu tumors were used (Figures 8A-8D). For determining in-vivo efficacy, the number of female SCID mice used in each study are indicated in the Figure 8E). Statistical analysis; log rank (Mantel-Cox test), p-value:<0.0001.
Figure 9:
Figure 9:
Structure of the PS-Erlotinib conjugates in which erlotinib moiety is attached either at meta- ( PS 10) or para- position (PS 16) of the pyropheophorbide-a.
Figure 10:
Figure 10:
In vivo uptake of PS 16 at a dose of 0.47μmole/kg in tumor, liver and skin of SCID mice bearing (A) UMUC3 and (B) FaDu tumors. The in vivo PDT efficacy (long-term tumor response) of PS 16 at a dose of 0.47 μmole/kg and PS 16 for UMUC 3 and FaDu tumors are shown Figures C and D respectively. The tumors were exposed to light (665 nm, 135J/cm2, 75 mW/cm2) at 24 h post-injection of the PS. A direct correlation between the PS uptake and tumor response (cure) was observed./kg. Statistical analysis log rank (Mantel-Cox test), (C) UMUC3, p-value: <0.0001 and (D) FaDu tumors, p-value: 0.0019. For PS-uptake study (tumor, liver and skin), 3 female SCID mice bearing either UMUC3 or FaDu tumors were used (Figures 9A & 9B). For determining in-vivo efficacy the number of female SCID mice used in each study are indicated in the Figure 9C & 9D.
Figure 11:
Figure 11:
In vivo uptake of PS 5 at the dose of 0.47μmol/kg in tumor, liver and skin of SCID mice bearing (A) UMUC3 tumors and (B) FaDu tumors. The in vivo PDT efficacy (long-term tumor response) of PS 5 at a dose of 0.47 μmole/kg for UMUC3 and FADU tumors are shown in (C) and (D) respectively. The tumors were exposed to light (665 nm, 135J/cm2, 75 mW/cm2) at 24 h post-injection of the PS. A direct correlation between the PS uptake and tumor response (cure) was observed. For PS-uptake study (tumor, liver and skin), 3 female SCID mice bearing either UMUC3 or FaDu tumors were used (Figures 11A & 11B). For determining in-vivo efficacy the number of female SCID mice used in each study are indicated in the Figure 9C & 9D. Statistical analysis log rank (Mantel-Cox test), (C): p-value: <0.0046 and (D): p-value: 0.0153.
Scheme 1:
Scheme 1:
Erlotinib moiety conjugated at position-17 of the hexyl ether analog of pyropheophorbide-a
Scheme 2:
Scheme 2:
Erlotinib moiety conjugated at position-3 of pyropheophorbide-a and bacteriopyro-pheophorbide-a
Scheme 3:
Scheme 3:
Erlotinib moiety conjugated either at p- position of the benzyloxy group having a chiral center at position 3(1’)- or at the m-position without a chiral center at position 3(1’) of the conjugate.
Scheme 4:
Scheme 4:
Erlotinib moiety conjugated at position 20 of methyl meso-pyropheophorbide a
Scheme 5:
Scheme 5:
The hexyl ether analog of pyropheophorbide-a (HPPH) conjugated with three Erlotinib moieties.
Scheme 6:
Scheme 6:
Erlotinib moiety conjugated to the fused imide ring system of purpurinimide and Bacteriopurpurinimide
Scheme 7:
Scheme 7:
Erlotinib moiety conjugated at position-13 of chlorin e6 with an amide linker
Scheme 8:
Scheme 8:
Erlotinib moiety conjugated at position-13 of chlorin e6 with an ethylene glycol linker
Scheme 9:
Scheme 9:
Erlotinib moiety conjugated at position-13 of chlorin by a longer ethylene glycol linker
Scheme 10:
Scheme 10:
Erlotinib moiety conjugated at position-15 of chlorin e6
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Juarranz A; Jaen P; Sanz-Rodriguez F; Cuevas J and Goncalez S Photodynamic therapy of cancer: Basic principles and applications, Clinical and Translational Oncology, 2008, 10, 148–154. - PubMed
    1. Kessel D Photodynamic therapy: A brief history, J. Clin. Med, 2019, 8, 1581, and references therein. - PMC - PubMed
    1. Hiramatsu R; Kawabata S; Tanaka H; Sakurai Y; Suzuki M; Ono K; Miyatake S-I; Kuroiwa T; Hao E and Vicente MGH, Tetrakis(p-carboranylthio-tetrafluorophenyl) chlorin (TPFC): Application for photodynamic therapy and boron neutron capture therapy. J. Pharmaceutical Sci, 2015, 104, 962–970 - PubMed
    1. Lo P-C; Rodriguez-Morgade MS; Pandey RK; Ng DKP; Torres T and Dumoulin F The unique features and promises of phthalocyanines as advanced photosensitizers for photodynamic therapy of cancer. Chem. Soc. Rev, 2020, 49, 1041–1056. - PubMed
    1. Shaker YM; Sweed AMK; Moylan C; Rogers L; Ryan AA; Pettidemange R and Senge MO, Current developments in using MESO-(TETRA) substituted porphyrins in PDT in HANDBOOK OF PHOTODYNAMIC THERAPY, Updates on Recent Applications of Porphyrin-Based Compound (Eds: Pandey RK; Kessel D and Dougherty TJ), World Scientific, New Jersey, 2016, pp 95–149.

Publication types

MeSH terms

LinkOut - more resources