Application of isothioblue injection as fluorescent tracer agent

Abstract

The invention discloses an application of an isosulfide blue injection in preparation of a fluorescent tracer agent, discloses a new application of isosulfide blue as the fluorescent tracer agent for the first time, and through clinical verification, the application retains dye tracing of the isosulfide blue and also uses a fluorescent tracing function of the isosulfide blue. During lymphatic vessel tracing, medical staff can conveniently find sentinel lymph nodes quickly and conveniently, the application range of the isosulfur blue is expanded, and the diversity of tracing selection of clinical medical staff is increased.

Description

Technical Field

[0001] The invention relates to the technical field of medicine, in particular to application of isosulfan blue injection as a fluorescent tracer. Background Art

[0002] The metastasis of cancer cells to lymph nodes through lymphatic vessels is the main way for breast cancer to spread locally. The axillary lymph node staging determined by surgical removal of axillary lymph nodes is an important basis for breast cancer prognosis and adjuvant treatment decisions. Traditional axillary lymph node dissection can determine whether the axillary lymph nodes are metastatic and the number of metastatic lymph nodes, but complications such as upper limb edema and nerve damage caused by the dissection will affect the patient's quality of life after surgery. Axillary lymph node dissection can allow patients with positive lymph nodes to obtain good local control of the disease, but it will be unnecessary overtreatment for patients with negative axillary lymph nodes, and it also brings the risk of complications such as upper limb edema that affect the quality of life after surgery. Sentinel lymph node biopsy (abbreviated as biopsy) is relatively minimally invasive and has fewer complications. It can determine whether the axillary lymph nodes are metastatic, and then decide to exempt axillary lymph node dissection for patients with negative biopsies. For patients with breast cancer with negative axillary lymph nodes, there is no significant difference in the local control rate and long-term survival rate between axillary dissection and surgery without dissection. Sentinel lymph node biopsy has now become the first choice for axillary staging in patients with early breast cancer with negative axillary lymph nodes, allowing many early breast cancer patients to avoid complications such as upper limb edema.

[0003] Sentinel lymph nodes are defined as the first lymph nodes to metastasize to the tumor. In actual operation, the tracer is first injected into the breast, and the first lymph node is removed according to the guidance of the tracer, and the metastasis is determined by pathological diagnosis. The key to ensuring the success rate and accuracy of sentinel lymph node biopsy lies in the tracer technology.

[0004] The current methods of lymphatic tracing mainly include dye method, radionuclide tracing method and fluorescent dye method. Although the radionuclide tracing method has high accuracy, it has certain radioactivity when used. The fluorescent dye method mainly uses indocyanine green. By injecting indocyanine green, the fluorescent signals of lymphatic vessels and lymph nodes can be clearly observed, thereby guiding the surgical sampling of lymph nodes. In particular, with the help of fluorescence detection of the surgical field, the missed detection of sentinel lymph nodes can be avoided to a large extent. The dye method mainly uses isosulfan blue, patent blue, nanocarbon, methylene blue, mitoxantrone, etc.

[0005] Isosulfan blue is a safe and highly sensitive dye. After subcutaneous injection, it can be selectively absorbed by lymphatic vessels and quickly enter the lymphatic system. It has low diffusion in soft tissues and is simple, fast, and intuitive. It is currently the only FDA-approved sentinel lymph node tracer with an accuracy rate of more than 97%. However, isosulfan blue is currently used as a dye method in clinical applications. During surgery, relevant tissues need to be dissected and observed to find sentinel lymph nodes along the tracing route, resulting in difficult surgery, blood easily contaminating the blue-dyed area during surgery, and tracing is not obvious. The lymphatic vessels and lymph nodes stained with isosulfan blue are light in color, and coupled with tissue coverage, it is easy for sentinel lymph nodes to be difficult to find and missed. The existing technology for the application of isosulfan blue is mainly the dye method, and there has been no related report on the application of isosulfan blue in fluorescent tracers. Summary of the invention

[0006] In view of the above-mentioned prior art, the present invention has discovered a new use of isosulfan blue through research, that is, its use in the preparation of fluorescent tracers.

[0007] In order to achieve the above object, the present invention is implemented through the following technical solutions: Through clinical trials, the present invention has discovered for the first time that isosulfan blue can not only be used as a dye tracer, but can also produce fluorescence characteristics under incident light irradiation, so that it can be used as a fluorescent tracer, so that it has both dye and fluorescence characteristics, performs well in lymphatic vessel tracing, and improves tracing efficiency. Therefore, isosulfan blue can be used as an effective reagent for fluorescent tracing and can be used to prepare lymphatic tracers.

[0008] The isosulfan blue injection produces fluorescence spectral characteristics after combining with serum proteins, and produces fluorescence for tracing under the irradiation of incident light of a specific wavelength. The wavelength range of the incident light is 620nm-690nm, and the isosulfan blue injection produces fluorescence with a wavelength of 690nm-790nm.

[0009] The molecular formula of the isosulfan blue injection is C 27 H 31 N2O6S2Na.

[0010] In specific application, the isosulfan blue injection is applied to the lymphatic vessel fluorescent tracer, and the isosulfan blue injection is subcutaneously injected near the area where lymphatic tracing is required, and then the tracing area is irradiated with incident light, and the isosulfan blue injection produces fluorescent properties to visualize the lymphatic vessels.

[0011] Compared with the prior art, the beneficial effects of the present invention are: The present invention discloses for the first time a new use of isosulfan blue as a fluorescent tracer. After clinical verification, this use retains the dye tracing of isosulfan blue, and uses its fluorescent tracing function at the same time, making up for its shortcoming of light staining to the naked eye. It greatly facilitates medical personnel to quickly and conveniently find sentinel lymph nodes during lymphatic tracing, improves tracing efficiency, and avoids missed detection of sentinel lymph nodes. The present invention expands the application scope of isosulfan blue, increases the diversity of tracing options for clinical medical staff, and uses a tracer with two properties for lymphatic tracing to improve tracing efficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Attached Figure 1 is a schematic diagram of the relevant lymph nodes in Example 1, Figure 1 (a) shows the removed lymph node tissue, and (b) shows the fluorescence imaging effect.

[0013] Attached Figure 2 is a schematic diagram related to Example 2, Figure 2 (a) is a schematic diagram of the injection site, (b) is a schematic diagram of the lymphatic vessels under fluorescent observation, (c) is a schematic diagram when marking with a streaking pen, and (d) is a schematic diagram of the lymphatic vessel marking position with a streaking pen.

[0014] Attached Figure 3 is a schematic diagram related to Example 2, Figure 3 (a) is a schematic diagram of the blue-stained lymphatic vessel observed under fluorescence and the corresponding body surface markers, and (b) is a schematic diagram of the blue-stained lymph node under fluorescence observation.

[0015] Attached Figure 4 is a schematic diagram related to Example 3, Figure 4 (a) is a schematic diagram of body surface marking based on fluorescent tracer, and (b) is a schematic diagram of fluorescent verification of lymph nodes.

[0016] Attached Figure 5 is a schematic diagram related to Example 4, Figure 5 (a) is a schematic diagram of lymphatic vessel development, (b) is a schematic diagram of body surface marking, (c) is a schematic diagram of in vitro specimens, and (d) is a schematic diagram of fluorescence development results. DETAILED DESCRIPTION

[0017] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, it should be understood that after reading the content described in the present invention, those skilled in the art may make various changes or modifications to the present invention, and these equivalent forms fall within the scope limited by the application equally.

[0018] The instruments, reagents, materials, etc. involved in the following embodiments, unless otherwise specified, are all conventional instruments, reagents, materials, etc. in the prior art and can be obtained through regular commercial channels. The experimental methods, detection methods, etc. involved in the following embodiments, unless otherwise specified, are all conventional experimental methods, detection methods, etc. in the prior art.

[0019] Example 1: Discovery of a new use of isosulfan blue injection as a fluorescent tracer Isosulfan blue has been used clinically in the United States for more than 40 years since it was launched in 1982. It is the first and only staining dye approved by the US FDA for lymphangiography, showing physiological inertness and no pharmacological effects.

[0020] Isosulfan blue is a safe and highly sensitive dye. After subcutaneous injection, it can be selectively absorbed by lymphatic vessels and quickly enter the lymphatic system. It has low diffusion in soft tissues and is simple, fast, and intuitive. It is currently the only FDA-approved sentinel lymph node tracer with an accuracy rate of more than 97%. The main side effect in clinical applications is allergy (0.25-2%), which generally returns to normal within 60 minutes. The tattoo-like blue staining on the skin can be eliminated in a short period of time. The mechanism of action is based on the physiological lymphatic system circulation. After subcutaneous injection, isosulfan blue injection binds to serum proteins, the lymphatic circulatory system recycles proteins, and the lymphatic vessels are displayed by blue dye.

[0021] The current clinical application of isosulfan blue injection and the manufacturer's drug instructions confirm that its known properties are simply as a tracer for the dye method.

[0022] like Figure 1 As shown in (a), the removed lymph node tissue, the applicant dripped an appropriate amount of isosulfan blue injection on part of the lymph node tissue, and irradiated the lymph tissue with incident light of an excitation wavelength of 660nm, and imaged the lymph tissue using a fluorescent camera. The imaging results are shown in Figure 1 As shown in (b), the imaging results show that the lymphatic tissue dripped with isosulfan blue injection has fluorescence spectrum characteristics, and its fluorescence phenomenon is clearly observed in the imaging. Therefore, the applicant has discovered for the first time a new use of isosulfan blue injection, which can be used as a fluorescent tracer, which is not discovered by current clinicians and pharmaceutical manufacturers. The newly discovered fluorescent property has achieved unexpected technical effects.

[0023] Example 2: Clinical application of isosulfan blue injection as a fluorescent tracer 1 Patient A underwent modified radical mastectomy for left breast cancer, with the tumor located in the upper outer quadrant; 13:15 Inject 1ml of isosulfan blue injection solution into the skin at 3, 6, 9, 12 points around the areola and on the surface of the mass. Figure 2 As shown in (a).

[0024] At 13:17, the lymphatic vessels were observed under fluorescence and marked on the body surface. Figure 2 As shown in (b), under fluorescent observation, the areola 9 and 12 injection points give rise to lymphatic vessels above the breast. Figure 2 Middle (c) shows the lymphatic vessels heading toward the axilla from the injection point on the surface of the tumor under fluorescent observation, which are marked with a streaking pen. Figure 2 Middle (d) shows the position of the lymphatic vessel marked with a streaking pen.

[0025] Figure 3 In (a), the tissue was dissected according to the marked position. The blue-stained lymphatic vessels were confirmed by both naked eye and fluorescent observation, which were consistent with the marked position on the body surface.

[0026] At 13:44, we continued to find two blue-stained sentinel lymph nodes along the blue-stained lymphatic vessels and verified them by fluorescence. Figure 3 Middle (b) shows blue-stained lymph nodes under fluorescence observation.

[0027] Analysis: Through the above clinical use, it can be found that isosulfan blue injection can be used as a fluorescent tracer in clinical practice. Compared with the traditional dye tracing method, it saves an average of 30 minutes from injection to finding the sentinel lymph node. The fluorescent tracing and its own dye tracing achieve mutual reference, allowing medical staff to quickly and accurately find the sentinel lymph node.

[0028] Example 3: Clinical application of isosulfan blue injection as a fluorescent tracer 2 Patient B, underwent right breast lumpectomy + modified radical mastectomy for right breast cancer, the mass was located below At 9:27, 1 ml of 5-fold diluted isosulfan blue injection was injected at the four points on the areola; 9:30 Observe the lymphatic vessels under fluorescence and mark them on the body surface. Figure 4 As shown in (a).

[0029] The lymphatic vessels originate from the 9 o'clock injection point on the areola and go towards the axilla.

[0030] According to the fluorescence tracing, a slightly blue-stained lymph node was dissected out, which was confirmed to be a sentinel lymph node by fluorescence verification. The other lymph nodes showed no fluorescence or blue staining. Figure 4 (b) shows the fluorescently verified lymph node.

[0031] Example 4: Clinical application of isosulfan blue injection as a fluorescent tracer Patient C, who underwent modified radical mastectomy for right breast cancer, had a tumor located on the outside At 8:54, 1 ml of 10-fold diluted isosulfan blue injection was injected at the four points on the areola; At 8:55, the lymphatic vessels were observed under fluorescence and marked on the body surface; Figure 5(a) shows the lymphatic vessels. Figure 5 (b) shows body surface markers; The lymphatic vessels originate from the 9 o'clock and 12 o'clock injection points on the areola and seem to lead to the axilla; At 9:51, the specimen was removed from the body, and a slightly blue-stained lymph node was dissected out. After fluorescence verification, it was found to be a sentinel lymph node. Two slightly blue-stained lymph nodes were dissected out behind the sentinel lymph node and marked as post-sentinel lymph nodes. Group I lymph nodes showed fluorescence imaging, while the other lymph nodes showed no blue staining or fluorescence. Figure 5 (c) in the middle is an in vitro specimen. Figure 5 Middle (d) is the fluorescence development result.

[0032] Analysis: According to current data, 1 ml of the original solution, 5-fold dilution and 10-fold dilution of isosulfan blue injection can all be used to observe fluorescent development of surface lymphatic vessels within 1-5 minutes. It has also been clinically proven that fluorescent tracers based on isosulfan blue injection can help medical staff find sentinel lymph nodes accurately and quickly during lymphatic tracing.

Claims

1. Application of isosulfan blue injection as a fluorescent tracer.

2. The use according to claim 1, characterized in that: The isosulfan blue injection produces fluorescence spectrum characteristics after combining with serum protein, and produces fluorescence under the irradiation of incident light of a specific wavelength, so it can be used as a fluorescent tracer.

3. The use according to claim 2, characterized in that: The incident light has a wavelength range of 620nm-690nm, and the isosulfan blue injection generates fluorescence with a wavelength of 690nm-790nm.

4. The use according to claim 1, characterized in that: The molecular formula of the isosulfan blue injection is C 27 H 31 N2O6S2Na.

5. The use according to any one of claims 1 to 4, characterized in that: The isosulfan blue injection is applied as a lymphatic fluorescent tracer.

6. The use according to claim 5, characterized in that: In specific applications, isosulfan blue injection is injected subcutaneously at 1-3 points near the area where lymphatic tracing is required, and then the tracing area is irradiated with incident light. The isosulfan blue injection produces fluorescent properties to visualize the lymphatic vessels.

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