阅读说明：本技术 焦脱镁叶绿酸轭合物及其在癌症治疗中以及作为荧光标记的用途 (Pyrophorbide conjugates and their use in cancer therapy and as fluorescent markers ) 是由 H·阿扎伊斯 P·科利内 N·德莱尔姆-费尔赖 O·莫拉莱斯 S·莫尔顿 C·弗罗绍 R 于 2018-07-20 设计创作，主要内容包括：本发明涉及式(I)化合物及其药学上可接受的盐。本发明还涉及所述式(I)化合物特别通过光动力疗法在治疗癌症中的用途。<Image he="548" wi="700" file="DDA0002416466230000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The present invention relates to compounds of formula (I) and pharmaceutically acceptable salts thereof. The invention also relates to the use of said compounds of formula (I) in the treatment of cancer, in particular by photodynamic therapy.)

1. A compound of formula (I):

2. the compound according to claim 1 for use as a medicament.

3. Use of a compound according to claim 1 for the treatment of cancer.

4. Use of a compound according to claim 3 for the treatment of cancer by photodynamic therapy.

5. Use of a compound according to claim 3 or 4 for reducing the risk of metastasis.

6. The compound for use according to claim 3 or 4, wherein the cancer is selected from ovarian cancer, lung cancer, kidney cancer, endometrial cancer, colorectal cancer or breast cancer, pancreatic cancer, brain cancer, gastric cancer, liver cancer, prostate cancer, testicular cancer, bladder cancer or head and neck cancer.

7. The compound for use according to claim 6, wherein the cancer is ovarian cancer.

8. Use of a compound according to any one of claims 2-7, wherein the compound is intended for intraperitoneal or intravenous administration.

9. A process for the preparation of a compound of formula (I),

characterized in that the process comprises the step of coupling a compound of formula (II):

10. process for the preparation of the compound of formula (I) according to claim 9, characterized in that it comprises the following steps:

(a) coupling a compound of formula (IV):

and N-Boc-2,2' - (ethylenedioxy) diethylamine, to obtain a compound of formula (III):

b) a step of deprotecting the compound of formula (III) obtained in step a) to obtain a compound of formula (II):

c) a step of coupling the compound of formula (II) obtained in step b) with folic acid to obtain a compound of formula (I):

11. use of a compound of formula (I) according to claim 1 as a fluorescent label.

12. A method for imaging in a subject comprising visualizing fluorescence emitted by a compound of formula (I) according to claim 1, said compound of formula (I) having been previously administered to said subject and activated by light source light.

Technical Field

The present invention relates to the field of photosensitizers, more particularly to their use in photodynamic therapy regimes for the treatment of cancer, in particular ovarian cancer.

Background

In france, ovarian cancer represents 4500 new cases per year. Poor prognosis of the disease is associated with delayed diagnosis, as most cases are diagnosed in stages III and IV of the international association of obstetrics and gynecology (FIGO), and survival rates decrease with progression of the disease. Ovarian cancer is the leading cause of death from gynecological cancers in the united states and western europe. In france, approximately 3500 deaths per year are attributed to this.

The disease often progresses to the appearance of metastases in the form of peritoneal carcinoma, which corresponds to the presence of numerous tumors on the surface of organs of the pelvic cavity. The peritoneum is a continuous serosa (formed by a monolayer of mesothelial cells) that covers the abdomen, pelvis, and internal organs, defining a virtual space of the peritoneal cavity. There is a distinction between the visceral peritoneum (which covers the exterior of the organ) and the parietal peritoneum (which covers the interior surface of the abdominal wall).

Current therapies are based, where possible, on chemotherapy surgery using platinum salts, in some cases also in combination with targeted therapies. It is well recognized that the absence of residual lesions after surgery is a major factor in good prognosis. Therefore, the ability of surgical treatment to eradicate all tumor implants is crucial. Systemic adjuvant chemotherapy or neoadjuvant chemotherapy has increased survival rates to five years, especially in the early stages: after three cycles of combined chemotherapy with carboplatin and paclitaxel, the survival rates in phases I and II were 81%. In the case of maximal cytopenia, and as a complement to macroscopically intact tumor reduction surgery, treatment strategies such as intraperitoneal chemotherapy and intraperitoneal photodynamic therapy can be envisaged. Despite extensive clinical trials, the benefits of intraperitoneal chemotherapy have not been demonstrated in this indication, and recommendation of this option outside of clinical trials is not suggested.

By virtue of photosensitizers that selectively target early lesions, as opposed to intraperitoneal chemotherapy with or without hyperthermia, and only targeting these lesions, it is possible to reduce the toxicity of the treatment, since the effect of light only occurs in the presence of photosensitizers within the tumor tissue. It may also make possible the treatment of microscopic metastases that are overlooked during surgery. Thus, targeting receptors overexpressed by ovarian tumor cells by means of more specific photosensitizers may enhance the efficacy of photodynamic therapy (PDT).

In this case, Schneider et al (Bioorganic)&Medicinal Chemistry,2005,13, 2799-. More specifically, the present invention is to provide a novel,

et al (Photodiagnosis and Photodynamics Therapy,2016,13,130-138) demonstrated in animal models that this photosensitizer is particularly specific for peritoneal metastasis of ovarian epithelial cancers, and therefore it was suggested to be used in the development of an intraperitoneal Photodynamic Therapy regimen that is not toxic to the patient. However, on the one hand, the photosensitizer exhibits very weak fluorescence, thus preventing its detection in the medical devices commonly used by gynecologists, and on the other hand, it exhibits low stability.

You et al (Bioorganic & Medicinal Chemistry,2015,23,1453-1462) also developed conjugates of pheophorbide-a, in particular photosensitizers comprising a pheophorbide-a unit conjugated to folic acid via a spacer of polyethylene glycol type (Pho-PEG-FA). In particular, You et al show that this photosensitizer (Pheo-PEG-FA) targets the folate receptor and, in view of its fluorescence, can be used in photodynamic therapy protocols for the treatment of cancers overexpressing the folate receptor. However, the photosensitizer exhibits relatively moderate fluorescence.

Therefore, there is a real need to develop new photosensitizers for photodynamic therapy. These photosensitizers must both have good therapeutic efficacy and selectively target lesions so as not to damage healthy tissue, and must also have sufficient fluorescence to visualize these lesions by medical devices.

Brief description of the invention

Faced with this problem, the present inventors have synthesized a novel photosensitizer comprising pyropheophorbide units (Pyro-PEG-FA) conjugated with folic acid via a polyethylene glycol type spacer by a method ensuring excellent yield. Analysis of the photosensitizer allows it to exhibit improved physicochemical properties compared to the structurally similar conjugates described above, and also has good photostability. The inventors have also shown that this novel photosensitizer can specifically target microscopic peritoneal metastases of ovarian epithelial cancer without compromising the quality of the anti-tumor effector immune response.

Accordingly, the present invention relates to compounds of formula (I):

the invention also relates to the use of the compounds of formula (I) as medicaments.

According to a particular embodiment of the invention, the compounds of formula (I) are used for the treatment of cancer, in particular by photodynamic therapy. In particular, the compounds of formula (I) of the present invention are useful for reducing the risk of metastasis. Preferably, the compounds of formula (I) are used for the treatment of a cancer selected from ovarian, lung, kidney, endometrial, colorectal or breast cancer, pancreatic cancer, brain cancer, gastric, liver, prostate, testicular, bladder or head and neck cancer. More preferably, the compounds of formula (I) are used for the treatment of ovarian cancer.

According to another particular embodiment, the compound of formula (I) is intended to be administered intraperitoneally or intravenously.

Another subject of the present invention relates to a process for the preparation of a compound of formula (I) as shown above, comprising the step of coupling a compound of formula (II):

another subject of the invention is also the use of the compounds of formula (I) as shown above as fluorescent labels.

Another subject of the invention also relates to a method for imaging in a subject, comprising visualizing the fluorescence emitted by a compound of formula (I) previously administered to said subject and photoactivated by a light source.

Drawings

FIG. 1: a method for the synthesis of Pyro-PEG-FA compounds of formula (I).

FIG. 2: time-dependent photodegradation (365nm,5 mW/cm) of Folic Acid (FA), Pyro-PEG-FA compound of formula (I) (Pyro-S-FA), Phoo-PEG-FA compound (Phoo-S-FA), and TPP-FA compound (P1-S-FA)²,[0.45mM]In DMSO).

FIG. 3A: the relative amounts of ATP in the ovarian tumor cells of SKOV-3(A) and OVCAR-3(B) subjected to PDT were measured.

FIG. 3B: effect of PDT on cytokine secretion by OVCAR-3 ovarian tumor cells.

FIG. 4A: photograph of laparoscopic images in immunocompetent rats that have developed peritoneal cancer after injection of Pyro-PEG-FA compound of formula (I).

FIG. 4B: the photograph of fig. 4A in white light.

Detailed Description

As illustrated and demonstrated by the inventors in the following examples, the present invention provides novel folic acid conjugated photosensitizers of formula (I):

i) can specifically target microscopic peritoneal metastasis of ovarian epithelial cancer,

ii) has a good therapeutic effect,

iii) sufficient fluorescence is provided by conventional medical equipment to visualize the lesion,

iv) the synthesis requires only a few steps, and

v) activating immune cell proliferation.

Pyro-PEG-FA compounds of formula (I)

Accordingly, the present invention relates to compounds of formula (I):

the compounds of formula (I) are of the type comprising a compound of the formulaA conjugated compound of pyropheophorbide-a unit (Pyro-PEG-FA) in which the spacer is conjugated with folic acid. Pyropheophorbide-a units provide the compounds of formula (I) with satisfactory fluorescence for visualizing lesions. The spacer of polyethylene glycol type comprises polyethylene glycol PEG₂Two monomers of (2). Folate units can specifically target the folate receptor. In the present specification, the compound of formula (I) is represented by the formula "Pyro-PEG-FA" or "Pyro-PEG₂-FA "represents.

The expression "pharmaceutically acceptable salts thereof" denotes salts of the compounds of the subject formula (I) having the desired biological activity. Pharmaceutically acceptable salts comprise salts of acidic or basic groups present in the particular compound. Pharmaceutically acceptable acid addition salts include, but are not limited to, the hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, acetate, lactate, salicylate, citrate, tartrate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisate, fumarate, gluconate, gulonate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate and p-toluenesulfonate salts, and palmitate salts (i.e., 1' -methylenebis (2-hydroxy-3-naphthoate)). Suitable base salts include, but are not limited to, aluminum, calcium, lithium, magnesium, potassium, sodium, zinc, and diethanolamine salts. In particular, a list of pharmaceutically acceptable salts is disclosed in the review by Berge et al (j.pharm.sci.1977,66(1), 1-19).

The compounds of formula (I) can be synthesized by a simple method comprising the step of coupling the compound of formula (II) with folic acid:

more particularly, the compound of formula (I) can be synthesized by a process comprising three steps starting from pyropheophorbide-a of formula (IV) sold by Boc sciences. According to a preferred embodiment, the compound of formula (I) is prepared by a process comprising the steps of:

(a) coupling a compound of formula (IV):

and N-Boc-2,2' - (ethylenedioxy) diethylamine, to obtain a compound of formula (III):

b) a step of deprotecting the compound of formula (III) obtained in step a) to obtain a compound of formula (II):

c) a step of coupling the compound of formula (II) obtained in step b) with folic acid to obtain a compound of formula (I):

this simple process, which requires only three steps, makes it possible to obtain the compound of formula (I) in very satisfactory yields, which are particularly higher than those obtained by the process using the compound of You et al (Pheo-PEG-FA). Therefore, the method is more advantageous and suitable for industrial applications.

Applications of

The compounds of formula (I) as described above may be used as medicaments, preferably for the treatment of cancer.

For the purposes of the present invention, the term "treatment" means amelioration, prevention, or reversal of a disease or disorder in the case of cancer. The term "treating cancer" is intended to equivalently mean reducing the number of metastases and/or reducing the risk of developing metastases. In the case of the treatment of metastases, the aim is in particular to reduce the recurrence rate or recurrence rate of the patient in the development of cancer. The term "treating cancer" is also intended to refer to inhibiting cancer cell proliferation.

Without wishing to be bound by a particular mechanism of action, the compounds of formula (I) may activate the immune system, thereby helping to combat cancer. Therefore, the compounds of formula (I) according to the invention are proposed for the treatment of cancer by immunotherapy, in particular by activating the immune system after photodynamic therapy.

More particularly, the compounds of formula (I) are useful for the treatment of cancer by photodynamic therapy.

Photodynamic therapy, commonly abbreviated PDT, consists in bringing pathological tissue in contact with a light-activatable molecule, called photosensitizer or photosensitizing agent, in this case a compound of formula (I) according to the invention, and then in irradiating the tissue with light having a wavelength (color) suitable for the activation characteristics of the photosensitizer. After the molecules are activated by light and react with oxygen in the tissue, highly toxic substances are locally produced to finally induce destruction of the cancer lesion. The main advantage of PDT is its selectivity. In fact, the light used is not harmful by itself; photosensitizers without light are non-toxic. To initiate the reaction, a combination of light, photosensitizer and oxygen is necessary. Thus, by optimizing the photosensitizer concentration and the dose of light, cells can be selectively destroyed.

The present invention therefore also relates to the use of a compound of formula (I) as described above for the treatment of cancer, wherein an effective dose of a compound of formula (I) is contacted with cancer cells and/or metastases, which are subsequently exposed to a light source.

The present invention also relates to a method for treating cancer comprising the steps of: contacting a cancer cell with an effective amount of a compound of formula (I) as described above, and exposing the cancer cell to a light source.

The term "effective dose" is intended to mean a dose sufficient to obtain satisfactory fluorescence and thus the desired therapeutic effect. The skilled person will be able to adjust the dosage depending on the severity and type of cancer to be treated. However, an effective dose of the compound of formula (I) may be from 0.1mg/kg to 100mg/kg, from 0.1mg/kg to 50mg/kg, from 0.1mg/kg to 10mg/kg, preferably from 0.1mg/kg to 5mg/kg, even more preferably from 0.1mg/kg to 3 mg/kg.

For the compounds of formula (I) according to the invention, photoactivation of the photosensitizer can be obtained by any type of light source known to the person skilled in the art. In particular, the photoactivation can be carried out by artificial light (lamps, lasers) under radiation, such as ultraviolet or infrared type radiation, or by visible or natural light. The skilled person will select a suitable irradiation pattern by means of light means placed in the body cavity according to the type of cancer to be treated. For example, for liver cancer, the optical device is placed in the peritoneal cavity. For ovarian cancer, the light device is placed in the pelvic cavity; and so on. As non-limiting examples of optical devices, mention may be made of optical balloons (Photodiognosis and Photodynamic Therapy,2016,16,23-26) as described by Munck et al, or light-emitting textiles (LEF) as described by Guyon et al (Journal of biological Optics,2012,17 (3).

The wavelength of light used is selected to obtain the most effective photosensitizer effect. In particular, wavelengths of 300-800nm, preferably 400-700nm, and more preferably about 668nm are used.

Usually at 1-200J/cm²Preferably 1 to 150J/cm²And even more preferably about 150 joules/cm²The radiation is applied at a dose of (a). On the cells, generally 1-10 joules/cm are applied²The dosage of (a).

Preferably, a light intensity of 1-150mW/cm is used²，1-100mW/cm²，30-70mW/cm²Preferably about 50mW/cm²The light source of (1). On the cells, the light intensity is 1-10mW/cm²Or more preferably about 5mW/cm²The light source of (1).

Photoactivation of the photosensitizer and exposure of the cancer cells to the light source may be carried out for 1 minute to 3 hours, 10 minutes to 2 hours, 30 minutes to 90 minutes, preferably 1 hour or 60 minutes.

The invention described herein is preferably carried out in humans.

The compounds of formula (I) have a folate unit and can therefore be used for the treatment of any type of cancer expressing a folate receptor by photodynamic therapy. Assaraf et al describe a list of these folate receptor expressing cancers (Drug resistance update, 2014,17,89-95) and include, inter alia, ovarian, lung, kidney, endometrial, colorectal and breast cancers, pancreatic, brain, gastric, liver, prostate, testicular and bladder cancers or head and neck cancers.

According to a preferred embodiment of the invention, the cancer is selected from ovarian, lung, kidney, endometrial, colorectal and breast cancer, pancreatic, brain, stomach, liver, prostate, testicular and bladder cancer or head and neck cancer. According to an even more preferred embodiment, the cancer is ovarian cancer.

Depending on the type of cancer to be treated, the compounds of formula (I) may be administered to the patient by any type of route known to those skilled in the art. For example, the intraperitoneal route or the intravenous route can be used for the treatment of ovarian cancer. According to a preferred embodiment of the invention, the compounds of formula (I) are intended for intraperitoneal or intravenous administration, preferably intravenous administration.

The compounds of formula (I) can be used as fluorescent labels in view of their improved absorption and fluorescence properties. For example, compounds of formula (I) as fluorescent markers are useful in the treatment of cancer by fluorescence-guided surgery and/or in imaging or diagnostic methods.

Thus, methods of imaging or diagnosing cancer in a subject are described, comprising administering to the subject and irradiating or photoactivating a compound of formula (I), and visualizing the emitted fluorescence. This approach allows visualization of folate receptors that are overexpressed in cancer cells. Thus, the imaging makes it possible to delineate cancer regions in appropriate circumstances and also to guide ablation-type surgery.

Thus, a subject of the present invention is a method for imaging in a subject, comprising visualizing the fluorescence emitted by a compound of formula (I) previously administered to said subject and photoactivated by a light source.

In view of their same properties, the compounds of formula (I) may also be used as photosensitizers in cosmetic and dermatological applications, such as photon rejuvenation, the treatment of common acne or skin aging.

The invention therefore also relates to the non-therapeutic use of the compounds of formula (I) as defined above as photosensitizers.

Other aspects and advantages of the present invention will become apparent from a reading of the following examples, which are to be regarded as illustrative rather than restrictive.

Detailed Description