Alkyl TPP compounds for mitochondrial targeting and anti-cancer therapy

Abstract

To provide novel therapeutic compounds that target and inhibit a mitochondrial function, and may serve as anti-cancer therapeutics.SOLUTION: Alkyl-triphenylphosphonium compounds having a saturated, linear alkyl chain of about 9 to about 18 carbons may be used as therapeutic agents to treat cancer. Demonstrative compound dodecyl-TPP (d-TPP) dose-dependently inhibits the propagation of breast cancer stem cells and targets the bulk of the adherent of cancer cells, by decreasing MCF-7 cell viability. Further, d-TPP potently inhibits a mitochondrial oxygen consumption rate, while simultaneously shifting cell metabolism toward a glycolytic pathway. This shift to a strict metabolic dependency on glycolysis may be used to eradicate the residual glycolytic CSC population, by using additional metabolic stressors. For example, d-TPP may be combined with a glycolytic inhibitor or an OXPHOS inhibitor, through co-administration or sequential administration, to treat cancer and eradicate CSCs.SELECTED DRAWING: Figure 2

Description

Technical Field

[0001] Related Applications This application claims the benefit of U.S. Provisional Patent Application No. 62 / 834,932, filed Apr. 16, 2019, and U.S. Provisional Patent Application No. 62 / 842,893, filed May 3, 2019, which are hereby incorporated by reference in their entirety.

[0002] This disclosure relates to inhibiting mitochondrial function and eradicating cancer, particularly cancer stem cells, using alkyltriphenylphosphonium compounds such as dodecyl TPP.

Background Art

[0003] Researchers are working on developing new anti-cancer treatments. Conventional cancer therapies (e.g., radiation therapy, alkylating agents such as cyclophosphamide, antimetabolites such as 5-fluorouracil) attempt to selectively detect and eradicate rapidly growing cancer cells by interfering with the cellular mechanisms involved in cell growth and DNA replication. Other cancer therapies use immunotherapies (e.g., monoclonal antibodies) that selectively bind to mutant tumor antigens on rapidly growing cancer cells. Unfortunately, after these therapies, tumors often recur at the same site or at one or more different sites, indicating that not all cancer cells have been eradicated. Recurrence may be due to insufficient chemotherapy dosing and / or the emergence of cancer clones that are resistant to the therapy. Therefore, new cancer treatment strategies are needed.

[0004] Advances in mutation analysis have enabled detailed studies of the genetic mutations that occur during cancer progression. Despite having knowledge of the genomic landscape, modern oncology still focuses on cancer. Identifying primary driver mutations across all subtypes was difficult. The harsh reality is that each patient's tumor is unique, and a single tumor can have multiple different clonal cells. This seems to be due to the possibility of containing vesicles. Therefore, what is needed is several different ones. This is a new approach that focuses on the commonalities between different types of cancer. Targeting metabolic differences between these two groups is a promising novel cancer treatment strategy. Analysis of transcriptional profiling data from the material reveals mitochondrial biosynthesis and / or mi An increase in over 95 mRNA transcripts related to tochondria translation was revealed. Of the 95 upregulated mRNAs, more than 35 are mitochondrial ribosomal tangent. Mitochondrial ribosomal protein (MRP) It codes for multiple mitri. Similarly, proteomics analysis of human breast cancer stem cells has shown that multiple mitri Significant excess of bosomal proteins and other proteins involved in mitochondrial biosynthesis. Overexpression was revealed.

[0005] Mitochondria are essential for meeting cellular requirements and adapting to the cellular microenvironment. Through division, elongation, and connection with one another, a tubular network or fragmented granules is formed. In this context, it is a very dynamic organelle. The balance between mitochondrial fusion and fission is, The morphology, abundance, function, and spatial distribution of mitochondria are determined, and therefore, ATP production, Mitochondrial-dependent processes such as itophagy, apoptosis, and calcium homeostasis. It influences many biological processes. And mitochondrial dynamics are Mitochondrial metabolism, respiration, and oxidative stress can be regulated. Therefore, fission activity and It is not surprising that an imbalance with fusion activity negatively impacts multiple pathological conditions, including cancer. Cancer cells often exhibit fragmented mitochondria, and their division is either enhanced or reduced. While mitochondrial dynamics are often associated with cancer, how do they influence tumorigenesis? A comprehensive understanding of the mechanisms by which these effects occur remains necessary.

[0006] In particular, when cancer cells move toward tumor growth and metastatic dissemination, the increased bioenergy of cancer cells To support the demands of biosynthesis, complete and enhanced metabolic function is necessary. However, mitochondrial-dependent metabolic pathways extract energy from multiple fuel sources. This provides a biochemical platform for cancer cells.

[0007] In recent years, energy metabolism and mitochondrial function have been identified as being involved in the maintenance and proliferation of CSCs. The above CSCs are associated with the dynamics of tumor initiation, metastatic spread, and anticancer These are characteristic cell subpopulations within tumor masses that contribute to resistance to therapy. For example, CSCs are A unique and distinctive increase in mitochondrial mass, as well as enhancement of mitochondrial biosynthesis, and This shows stronger activation of mitochondrial protein translation. These behaviors are related to mitochondrial protein translation. This suggests a strict dependence on the function. Consistent with these observations, the rise Mitochondrial metabolic function and OXPHOS were detected in CSCs of multiple types of tumors. Similarly, the fluorescent mitochondrial dye MitoTracker enriches CSCs and It is used in purification. This dye accumulates in CSCs and is scaffold-independent in vitro. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness. Enable the characterization of cancer cell subpopulations based on the degree of growth and high tumor-initiating ability in vivo. In addition, during asymmetric cell division, young mitochondria cluster in daughter cells that maintain a stem-like phenotype, while daughter cells involved in differentiation lose stemness and receive older mitochondria. This further supports the idea of selecting the most functionally intact, surviving, undamaged mitochondria to support stemness.

[0008] Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function. Based on these observations, a new pharmacological approach aimed at targeting mitochondria in CSCs has been proposed, and some of it has been well applied in preclinical and clinical studies. For example, doxycline, an antibiotic that reduces mitochondrial protein translation as an off-target effect, has been suggested for use in the clinical management of early breast cancer patients for its ability to selectively target CSCs. Therefore, what is needed is the identification of candidates for cancer therapeutic compounds that target and inhibit mitochondrial function.

[0009] However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these However, since the lack of mitochondrial function can be overcome by adopting adaptive mechanisms in the tumor mass, there are certain limitations to the use of anti-mitochondrial agents alone in cancer therapy. These adaptive mechanisms include, for example, the ability of CSCs to shift from oxidative metabolism to alternative energy pathways in the multi-directional process of metabolic plasticity driven by both endogenous and exogenous factors within and around tumor cells in the niche. In particular, in CSCs, such manipulation of metabolic flexibility can be advantageous from a therapeutic perspective. Therefore, what is needed is to understand these Either prevent the metabolic shift, or use the above shift to inhibit cancer cell proliferation. This is a treatment approach. [Overview of the project] [Problems that the invention aims to solve]

[0010] The purpose of this disclosure is to target and inhibit mitochondrial function and to function as an anticancer therapy. The purpose of this disclosure is to identify and describe novel therapeutic compounds that can be used. Another purpose of this disclosure is to identify and describe cancer cells To prevent the metabolic shift and / or to utilize the above metabolic shift to inhibit the proliferation of cancer cells, To identify and describe novel therapeutic approaches using the above-mentioned therapeutic compounds to eradicate CSCs. And so it is. [Means for solving the problem]

[0011] Increased mitochondrial metabolism is associated with tumor initiation, dissemination to secondary sites, and treatment resistance. One of the most distinctive features of cancer stem cells (CSCs) is Using mitochondrial disruption agents, the maintenance and propagation of CSCs are efficiently inhibited, and neoplasms are treated. This can lead to better control. Triphenylphosphonium (TPP)-based Tochondria target compounds accumulate in the mitochondria of living cells, and are small, generally non-existent. A toxic, biologically active molecule used in anticancer treatment to block the growth and proliferation of CSCs. It will become a candidate for law.

[0012] This specification describes novel alkyl truffles such as dodecyl TPP (d-TPP). Cancer It is a therapeutic agent. The A-TPP compound is a cation, and is usually a salt (e.g., d-TPP bromide salt). It should be understood that it is available as a pharmaceutical. Embodiments of the approach of the present invention are pharmaceutical A pharmaceutical composition containing an effective amount of d-TPP or other A-TPP compounds, and a method for treating cancer and promoting cancer growth. It inhibits tumor recurrence, eradicates CSCs, prevents or reduces the likelihood of tumor recurrence, and prevents metastasis. This may take the form of either doing so or reducing the possibility thereof. In some embodiments, d - A-TPP compounds such as TPP are used as the first metabolic inhibitor, and glycolysis and / or OXPHOS It may be used in combination with a second metabolic inhibitor that interferes with the first metabolic process. Such combinations may be used in anticancer By using it as a therapy, it can treat cancer, inhibit cancer growth, eradicate CSCs, and tumors. To prevent or reduce the likelihood of tumor recurrence, and to prevent or reduce the likelihood of metastasis. Cut.

[0013] This disclosure demonstrates that the inhibition of CSC propagation is more than twice as effective as that of other TPP derivative compounds described below. This relates to compositions and therapeutic strategies containing d-TPP, which is advantageous. It targets the majority of cancer cells and reduces their survival rate, but it does not affect normal fibroblasts. The impact is limited. In cancer cells, especially CSCs, the mitochondrial membrane potential is normal or It may be higher than healthy cells. Therefore, using a TPP-based strategy, "normal" Mitochondria and "malignant" mitochondria, primarily in healthy individuals and diseased states. They can be distinguished based on the unique chemophysical characteristics of their organelles. Notably, d- TPP therapy determined a shift in energy metabolism toward activation of the glycolytic pathway. This is due to d-T It may be promoted as a compensatory response to the anti-mitochondrial effects induced by PP. The possibility is extremely high. This metabolic shift leads to the severe resistance of cancer cells to glycolysis after d-TPP treatment. A close dependence was revealed. In CSC, manipulating such metabolic flexibility is considered a therapeutic approach. This can be advantageous from a specific point of view. For example, by synchronizing CSCs with specific metabolic dependencies... Blocking the CSC's ability to shift energy between multiple energy pathways will lead to the eradication of CSCs. This may represent a useful strategy for residual CSC. This involves the use of a second metabolic inhibitor (glycolysis or oxphos) to further starve the population. This includes a "two-hit" treatment strategy.

[0014] In an embodiment of the "two-hit" strategy of the present invention, the d-TPP and other similar policies are converted to A-TPP. The compound is used as the primary metabolic inhibitor to damage CSC mitochondria and induce glycolysis in cancer cells. This shifts the state to one where the cancer cells are deprived of their bioenergy source, leading to propagation and recurrence. And a second metabolic inhibitor (glycolysis or oxphos) that inhibits metastasis is used. It should be understood that metabolic inhibitors can be administered simultaneously or sequentially. For example, d- TPP may be administered first to induce a metabolic shift of CSCs toward glycolysis, followed by the second Metabolism inhibitors may be administered.

[0015] Regarding the metabolic and biological effects induced by d-TPP on cancer cells, particularly breast cancer cells... The following is a description of the process. Using a 3D tumor-like mass assay with MCF-7 cells, d-TPP was evaluated. The treatment used demonstrated dose-dependent inhibition of breast CSC propagation in the suspension. These results indicate that d-TPP reduces the survival rate of MCF-7 cells, thereby reducing cancer cells This indicates that it targets the "bulk" of the attached material. Analysis of metabolic flux using Xfe96 showed that d-TPP is related to mitochondrial oxygen consumption. This study demonstrates that it strongly inhibits the rate of glycolysis while simultaneously shifting cellular metabolism to the glycolytic pathway.

[0016] Following this metabolic shift, the dependence of CSCs on glycolysis is used to introduce additional metabolic stressors. This allows for the eradication of the remaining glycolytic CSC population. The effects of d-TPP and a second metabolic inhibitor are significant. The "two-hit" treatment strategy of the present invention was validated using empirical combinations. The second metabolic inhibitors included natural and rigid compounds, some of which were FDA approved. It is purported to be a glycolysis inhibitor (e.g., vitamin C, 2-deoxyglucose i.e., 2DG) Alternatively, OXPHOS inhibitors (e.g., doxycycline, niclosamide, berberine chloride) It is an inhibitor known to exhibit the following behavior. An embodiment of a "two-hit" therapeutic strategy. This is a concentration of d-TPP that is toxic only to cancer cells but not to normal cells. In this case, CSC transmission was effectively reduced. Toxicity was observed in normal human fibroblasts (hTER). Evaluation was performed using T-BJ1) and the chicken chorioalinal membrane assay.

[0017] The results disclosed herein indicate that d-TPP is associated with the desired effects in normal cells. Without inducing any significant off-target effects, it stops the propagation of CSCs and removes most cancer cells. This demonstrates that it targets the TPP system in cancer treatment. This opens the way to further exploring the body's potential. Furthermore, TPP compounds are "normal" The potential for distinguishing between mitochondria and "malignant" mitochondria needs to be investigated. This is because distinct biochemical and metabolic changes in these organelles are present in certain cells. This suggests that it may precede a normal or pathological phenotype. Finally, the above The data suggests that manipulating energy mechanisms is a useful tool for controlling the stem cell properties of cancer cells. Confirm that it is.

[0018] In the approach of the present invention, the pharmaceutical composition is converted into an A-TPP of a pharmaceutically effective amount of d-TPP, etc. A compound (including its pharmaceutically acceptable salt) and a pharmaceutically acceptable ki for the above compound. It may contain a carrier, diluent, or excipient. Some embodiments of the above pharmaceutical composition are This also includes a pharmaceutically effective amount of a second metabolic inhibitor compound such as a glycolysis inhibitor or oxphos inhibitor. Good. The second metabolic inhibitor compound described above is, in some embodiments, a separately pharmaceutically acceptable compound. It may be in a suitable carrier. Compounds obtained by the approach of the present invention can be used as anticancer therapy agents. It can be used. The pharmaceutically effective amount of compounds obtained by the present invention's approach is a known method in the art. It can be administered to subjects in stages. d-TPP is a second metabolic inhibitor in some embodiments. It can be administered simultaneously with the drug compound. Alternatively, d-TPP can be administered prior to the second metabolic inhibitor, It can be optionally administered before and together with the second metabolic inhibitor. By administering the compound, cancer can be treated, CSCs eradicated, and tumor recurrence prevented. It is possible to prevent or reduce the possibility of metastasis, or to prevent or reduce the possibility of metastasis. In some embodiments, cancer is treated by administering a pharmaceutically effective amount of d-TPP. It can be shifted to a glucose state. In some embodiments, a pharmaceutically effective amount of d-T Administering PP can enhance the efficacy of chemotherapy. Several implementation methods In this state, administering a pharmaceutically effective amount of d-TPP reduces tumor recurrence and metastasis, and improves drug tolerance. Treatment, prevention, and / or enablement of at least one of the following: sex and radiotherapy resistance. The possibility can be reduced. [Brief explanation of the drawing]

[0019] [Figure 1] Figure 1 shows the structure of d-TPP (structure A) and the structure of b-TPP (structure B). [Figure 2] Figure 2 shows the results of a tumor-like mass assay on MCF-7 cells exposed to various concentrations of d-TPP and b-TPP. [Figure 3] Figure 3 shows the results of a tumor-like mass assay on MDA-MB-231 cells exposed to various concentrations of d-TPP. [Figure 4A-4C] Figures 4A-4C compare the results of cell viability tests using various concentrations of d-TPP in MCF-7 breast cancer cells and h-TERT normal cells. Figure 4A shows the results after 24 hours, Figure 4B shows the results after 48 hours, and Figure 4C shows the results after 72 hours. [Figure 5A-5B] Figures 5A-5B show the oxygen consumption rates of MCF-7 cells treated with various concentrations of d-TPP. [Figure 6A-6B] Figures 6A-6B show the ECAR results for multiple d-TPP concentrations in the range of 50 nM to 500 nM, as well as for a vehicle-only control. [Figures 7A-7E] Figures 7A–7E show the results of tumor-like mass assays against 100 nM d-TPP combined with a second inhibitor compound, according to several embodiments of the approach of the present invention. [Figures 8A-8D] Figures 8A-8D show the results of the xCELLigence analysis. Figures 8A and 8B show the results for 48 hours of d-TPP treatment, while Figures 8C and 8D show the results for 72 hours of d-TPP treatment. [Figure 9] Figure 9 shows the tumor weight results obtained from the CAM assay for various treatment groups. [Figure 10]Figure 10 shows the results of metastatic invasion obtained from CAM assays for various treatment groups. [Figure 11] Figure 11 shows embryo survival results obtained from CAM assays for various treatment groups. [Figure 12] Figure 12 shows Kaplan-Meier curves illustrating survival rates obtained from CAM assays for various treatment groups. [Modes for carrying out the invention]

[0020] The following description illustrates embodiments of the approach of the present invention, and how the approach of the present invention can be put into practice. This will be explained in sufficient detail to demonstrate the approach of the present invention in these specific embodiments. As described by reference, it is understood that the approach of the present invention can be embodied in different forms. This description shall be interpreted as any attached claim being a specific embodiment described herein. It should not be interpreted as being limited to a specific state. Rather, these embodiments are thoroughly explained in this disclosure. This will be complete and fully convey the scope of the approach of the present invention to those skilled in the art. It is provided in this manner.

[0021] This description uses various terms that should be understandable to those with ordinary skills in the relevant technical field. It is being used. The following explanation is provided to avoid misunderstandings. The term "to treat" "treat", "treated", "treating" g) and "treatment" refer to the condition, disorder, or disease being treated. In particular, at least one symptom, reduction, or improvement related to or caused by cancer This includes, in certain embodiments, the treatment is related to or caused by the cancer being treated. The present invention reduces and / or improves at least one of the symptoms that occur. This includes, in some embodiments, the treatment involves a certain type of cancer in the host's body. This includes causing the death of Gori cells, for example CSCs, and this is for example these cells By depriving them of the energy generation mechanism, further propagation of cancer cells can be prevented, and / or This can be achieved by inhibiting CSC function. For example, treatment can target one or more cancers. This may include a reduction in symptoms or complete eradication of cancer.

[0022] The terms "cancer stem cells" and "CSCs" refer to cells that, when transplanted into an animal host, self-renew, differentiate, and This refers to a subgroup of cancer cells within a tumor that has the ability to form tumors. ("The majority (bulk)") Compared to cancer cells, CSCs have a larger mitochondrial mass and enhanced mitochondrial biosynthesis. Therefore, mitochondrial protein translation is more activated. In short, "circulating tumor cells" are primary tumor cells. These are cancer cells that flow from the circulatory system or lymphatic vessels and are carried throughout the body by blood circulation. Using ellSearch Circulating Tumor Cell Test This allows for the detection of circulating tumor cells.

[0023] When used herein, the phrase "pharmaceutically effective amount" "Effective amount" refers to the regulation, adjustment, and development of protein kinase activity. Or inhibition, for example, inhibition of protein kinase activity, or therapeutic effects such as cancer treatment. To achieve this, it is necessary to administer it to the host, or to the host's cells, tissues, or organs. This refers to the quantity. A physician or veterinarian with ordinary skills in the relevant technical field may obtain the necessary pharmaceutical composition. The efficacy can be easily determined and prescribed. For example, a physician or veterinarian can determine the amount of the active ingredient contained in the pharmaceutical composition. The dose of the compound of the invention is started at a level below the level required to obtain the desired therapeutic effect, and then... The dosage can be gradually increased until the desired effect is achieved.

[0024] As used herein, the phrase "active compound" This refers to the A-TPP compounds described herein, which include their pharmaceutically acceptable salts, or This may include isotopic analogs. The phrase "active compound" also refers to glycolysis inhibitors or OXPHO Embodiments that include a second inhibitor, such as an S inhibitor, may include a second inhibitor compound. One or more active compounds are known to a person with ordinary skill in the art, Please understand whichever approach is preferable for administering the substance to the subject. Also, regarding activity... The amount of compound and the timing of its administration depend on the individual subject being treated (among multiple factors). In particular, for example, age and weight, method of administration, pharmacokinetic properties of one or more specific active compounds, Furthermore, please understand that this may depend on the judgment of the prescribing physician. Therefore, the variability among subjects may vary. Depending on the circumstances, any dosage described herein is intended to serve as an initial guideline. The physician has done so, and the physician has done so to achieve the treatment that the physician considers appropriate for the subject. Therefore, the dosage of the compound can be titrated. Considering the desired degree of treatment, the physician will determine the age of the subject and It is important to balance various factors such as body weight, the presence of pre-existing diseases, and the presence of other diseases. It is possible. As will be explained in more detail below, pharmaceutical preparations are not limited to It can be prepared for any desired route of administration, including oral, intravenous, or aerosol administration. ru.

[0025] Heterogeneity between and within tumors is one of the main factors explaining tumor progression and therapy failure. Therefore, CSCs are hierarchical towards the generation of a complete repertoire of multiple cell types within the tumor mass. Due to its ability to produce offspring that are differentiated in a specific way, it generates the heterogeneity described above, and this leads to Subsequently, pharmacological strategies that identify and selectively target CSCs are expected to advance the treatment prospects. It has been implemented through several approaches, some of which are the most promising but also the most challenging. Various theories have been proposed to explain its origin. One is the so-called "metabolic stem cell" theory. According to the "bo-stemness" model, certain metabolic phenotypes determine the stem cell nature of a tumor. This model allows us to determine that specific metabolic dynamics differ between non-cancerous cells and differentiated cancer cells. This suggests that it may promote the acquisition of stem cell characteristics. Similarly, tumor metabolism may promote cancer It is included in the novel characteristics. Based on these observations, cancer metabolism is selected for CSCs. It's not surprising that this could be seen as an opportunity to target them.

[0026] CSCs across diverse cancer types showed increased OXPHOS activity compared to non-stem cell cancer populations. This includes unique characteristics such as increased mitochondrial biosynthesis and higher mitochondrial mass. It exhibits characteristic features. Supporting these findings, telomerase activity is high (hTERT h igh Therefore, cancer cells that possess high immortality are hTERT low The corresponding These findings specifically indicate increased mitochondrial mass compared to other materials. This suggests that the CSC population can be specifically inhibited using endothelial inhibitors, which means that A path to identifying chemical strategies aimed at selectively targeting mitochondria in CSCs. It opens.

[0027] Mitochondrial inhibition inhibits cancer recurrence and metastasis, and eradicates cancer cells, especially CSCs. This is an effective strategy for that purpose. For example, the antibiotic doxycycline has CSC activity It exhibits behavior as a mitochondrial inhibitor that impairs mitochondria. Long-term treatment with doxycycline is In response to mitochondrial dysfunction, it activates the glycolysis pathway of CSCs. Vitamin C, etc. By using glycolysis inhibitors in combination with doxycycline, CSCs can be completely eradicated. It can be completely eliminated. The combination of doxycycline and vitamin C is for the clinical management of cancer patients. A safe approach is demonstrated. Doxycycline, at concentrations that exhibit biological activity, is controlled. It has a very small range of side effects, and vitamin C has no potential side effects. This is well known. In addition to this, a meta-analysis of 21 published studies, Based on data from preclinical studies, oral administration of vitamin C is associated with an increased risk of lung and breast cancer. Furthermore, it has been demonstrated that it reduces overall mortality and disease-specific mortality. Oral administration of ascorbate over a week prior to vaccination may lead to lymphoma xenotransfer. Tumor progression was reduced in the graft model.

[0028] Clinical verification of these findings regarding doxycycline is planned by the inventors in the recent This is provided in a pilot study. In this study, patients with early-stage breast cancer were given a short-term (14-day) treatment. They were enrolled for preoperative doxycycline administration (inter-operative). In particular, a study comparing postoperative versus preoperative breast tumors. Analysis of samples showed that, compared to controls, the stem cells in patients treated with doxycycline were different. A selective decrease in the cellular markers CD44 and ALDH was observed, which indicates a mitochondrial target. Clinical evidence that CSC transmission can be effectively stopped using a transformation strategy It is being offered. Nevertheless, in breast cancer patients, doxycidenomin in combination with vitamin C Further research is needed to investigate the effects of cyclins.

[0029] Triphenylphosphonium (TPP) ) is an example of a lipophilic cation that targets cellular mitochondria. The TPP portion is therapeutic. It can covalently bind to compounds, allowing these bound therapeutic compounds to be delivered to mitochondria. The therapeutic compound was found to be present in higher concentrations in mitochondria than in other parts of the cell. The inventors have found that certain compounds containing the TPP portion inhibit mitochondrial function. This demonstrated that it reduces CSC propagation. Mitochondrial targeting signaling The TPP portion, which acts as a "cargo" therapeutic compound, is covalently bonded to it. It can be chemically bonded to molecules. The inherent properties of cargo molecules, and their bonding to the TPP structure, It deeply influences the accumulation of therapeutic compounds in the mitochondria of living cells, and subsequently, the overall life It may affect physical functions.

[0030] Therefore, covalent modification of compounds to TPP cations is used for probes and contrast agents in mitochondrial. This is a method of delivery to Doria. TPP + Cations transport small molecules to mitochondria. It is an extremely effective chemical "vehicle" for that purpose. Furthermore, TPP The chemical synthesis of thione can be easily achieved, and the degree of accumulation in mitochondria is positively charged TP It is elevated by the chemical attraction between the P cation and the negative membrane potential of the inner mitochondrial membrane.

[0031] Certain chemical derivatives of TPP also target mitochondria and exhibit mitochondrial inhibitory activity. Therefore, it is useful for targeting and eradicating cancer cells. (Published November 21, 2018) International Patent Application PCT / US201, which is requested and incorporated in its entirety by reference. Issue 8 / 062174 describes examples of TPP derivatives with anticancer activity. For example, this publication The researchers have found that the TPP derivative compound 2-butene-1,4-bis-TPP(b-TPP) is, We demonstrated that it impairs tochondria metabolic function, which leads to inhibition of breast CSC activity. .

[0032] This specification describes a saturated linear alkyl chain having approximately 9 to approximately 18 carbon atoms. A-TPP compounds:

[0033] [ka]

[0034] (where "x" is 8-17, preferably 9-16, more preferably 11-14) Novel therapeutic approaches using integrated metabolic strategies to eradicate CSCs by leveraging advantageous characteristics - This is the case. In one exemplary embodiment, x is 11, and d- is described in the following embodiment. TPP is obtained.

[0035] In a demonstrative embodiment of the present invention, compound d-TPP is used in the mitochondrial of CSC. It is used as a therapeutic agent that inhibits rhyncholytic function. It has approximately 9 to 18 carbon atoms in its alkyl chain. Other A-TPP compounds can also be used without departing from the approach of the present invention. Please understand this. d-TPP is a compound that transports the mitochondrial-targeted TPP portion. Mitochondrial inhibitory effects result in a dose-dependent and time-dependent reduction in cell viability.

[0036] These effects reduce the formation of 3D tumor-like masses, which are analyzed as readouts of CSC activity. Metabolic flux analysis shows that d-TPP is involved in mitochondrial basal respiration and ATP production. It has been shown to strongly inhibit [the process]. The combination of these effects is explained below. The reason for the decline in the functional capacity of CSCs, as observed in response to d-TPP administration. The rationale is provided. Breast cancer cells preferentially exhibit a glycolytic phenotype in response to d-TPP. This metabolic switch enables a subset of CSCs to exert the anti-mitochondrial effect of d-TPP. You will be able to deal with it.

[0037] In some embodiments, the approach of the present invention involves a second metabolic inhibitor with d-TPP ( By combining glycolysis or oxphos (a "two-hit" combination), the above dependency is targeted. This further starves the remaining CSC population. The d-TPP compound is the first As a primary metabolic inhibitor acting as a hit (specifically as a mitochondrial inhibitor) ) is used, followed by a second metabolic inhibitor (e.g., glycolysis) that acts as a second hit. Alternatively, an oxphos inhibitor may be used.

[0038] Despite the acquisition of this compensatory glycolytic behavior, d-TPP therapy involves glycolysis inhibitors and OXP By increasing the sensitivity of CSCs to the action of HOS inhibitors, CSCs are weakened. Therefore, these effects of d-TPP enable a variety of combination therapies. In this case, d-TPP is administered together with one or more such inhibitors, This offers a "two-hit" therapeutic approach to eradicate C. Demonstration of a second metabolic inhibitor. Typical examples include the glycolysis inhibitors vitamin C and 2-deoxy-D-glucose (2- DG), as well as OXPHOS inhibitors such as doxycycline, azithromycin, and niclorox. Examples include thamides and berberine chloride. For example, tetracycline, chlorotetracycline. The Terramycin family, including Crin, minocycline, and tigecycline, or erythro Erythromycin, including tyrosromycin, telithromycin, clarithromycin, and roxithromycin Elavacycline, a member of the Mycin family that is also FDA-approved. Recyclin and omadacycline can also be used without departing from the approach of the present invention. The functional verification of the approach of the present invention is provided below, but here, MCF-7 cells are used. By treating with d-TPP and a second metabolic inhibitor simultaneously, CSC activity was almost completely restored. A sense of loss was obtained.

[0039] As mentioned above, some derivatives of the TPP portion have the potential to inhibit cancer growth and metastasis. It also possesses power. Using the ATP depletion assay as a surrogate marker for mitochondrial dysfunction. The inventors filed an application on November 21, 2018, which is in its entirety aided by reference to this application. In the international patent application PCT / US2018 / 062174, several TPP derivatives were identified. For example, 2-butene-1,4-bis-TPP(b-TPP) It showed considerable efficacy in inhibiting CSC propagation (IC-50 to 500 nM). Mitocon The accumulation of TPP derivatives in Doria, as well as the mitochondrial inhibitory and anticancer effects of said compounds, It is heavily dependent on specific structures. What is needed is further TP with anticancer efficacy. It is a P derivative compound.

[0040] The present inventors have developed another TPP derivative having considerable mitochondrial inhibitory and anticancer efficacy. Therefore, dodecyl TPP was identified. Regarding the metabolic and biological properties of d-TPP compounds: This will be explained below. Furthermore, by efficiently targeting the flexible cellular energy mechanism of CSCs, cancer can be rooted. The following explains the exceptional combination strategy. The d-TPP structure is shown below. This is an organophosphorus containing a quaternary phosphate with three phenyl rings and a saturated 12-carbon alkyl chain. It is a cation.

[0041] [ka]

[0042] As a cation, d-TPP may be a salt such as a bromide salt. Figure 1 shows a bromide salt. Compare d-TPP (structure A) with 2-butene-1,4-bis-TPP (structure B). As can be seen, d-TPP has only one TPP moiety and is a hydrophobic long-chain alkyl group. It has [this characteristic]. On the other hand, B-TPP has two TPP portions linked by butene.

[0043] Both d-TPP and b-TPP inhibit CSC propagation, but d-TPP is more effective. It is extremely potent. The tumor-like mass formation assay uses multiple concentrations of d-TPP or b-T to increase the tumor-like mass formation assay. Estrogen receptor (ER) positive after treatment with PP The results were used as readouts for CSC propagation in MCF-7 breast cancer cells. Figure 2 shows a comparison of the CSC propagation inhibitory activity of d-TPP and b-TPP in the following location. As shown, treatment with d-TPP involves the two highest concentrations among those tested. It resulted in an 80% reduction in CSC propagation at 500 nM and 1 μM, and A slight but significant reduction (20%) in tumor-like mass formation is extremely important in relation to d-TPP. This has already been observed at low concentrations (50 nM and 100 nM). Compared to b-TPP, d-TPP was more than twice as potent in inhibiting tumor-like mass formation.

[0044] The inhibitory effect of d-TPP was also tested in the triple-negative breast cancer cell line MDA-MB-231. A similar inhibitory trend against CSC propagation was observed in the triple-negative breast cancer cell line MDA-M. Observed in B-231. Tumor-like mass formation aggregates in MDA-MB-231 cells. The results of (i) are shown in Figure 3. As can be seen, d-TPP was 50 nM, the highest value tested. It also showed inhibitory effects at low concentrations, and achieved nearly equivalent inhibition of 80% at 1 μM. I did it.

[0045] Through our past efforts, we have found that certain TPP derivatives can improve mitochondrial function in cancer cells. This can interfere with the metabolic activity of cancer cells, ultimately leading to a decrease in their survival rate. This demonstrated that it is possible. Interestingly, these effects on specific TPP compounds are related to cancer cells. It is selectively induced in cells, but not in normal (i.e., healthy) cells. Cancer cell survival This demonstrates the ability of d-TPP to reduce the rate of death but not reduce the cell viability of normal cells. Therefore, both MCF-7 breast cancer cells and normal human fibroblasts (hTERT-BJ1) The preservation rate was determined for various increasing concentrations of d-TPP (from 50 nM to 1 μM). The evaluation was conducted over a period of time (from 24 hours to 72 hours). The results are summarized in Figures 4A to 4C. Specifically, Figure 4A shows the results after 24 hours, and Figure 4B shows the results after 48 hours. 4C shows the results after 72 hours. These data indicate that d-TPP is beneficial for the survival of MCF-7 cells. This demonstrates that the rate decreases in a time-dependent and dose-dependent manner. High concentrations of the compound (e.g.) The effects (250 nM, 500 nM, 1 μM) are evident as early as 24 hours after treatment. The sea urchins showed an early decline in cell viability of approximately 30%. 72 hours after treatment, cell viability was... The rate reduction was nearly 80% at the above high concentrations (250 nM, 500 nM, 1 μM). Even at low concentrations (50 nM, 100 nM), there was a slight but significant difference (nearly 30%). Inhibitory effects were detected. On the other hand, the cell viability of hTERT-BJ1 cells was relatively high. The TPP concentration only decreased at low concentrations (250 nM, 500 nM, 1 μM), and not at low concentrations (50 nM, 1 The 00nM concentration showed no toxicity. These results suggest that d-TPP may not be particularly toxic when used at low concentrations. This demonstrates that it can selectively target cancer cells rather than normal cells.

[0046] Metabolic flux analysis shows that d-TPP blocks ATP production and leads to metabolic inflexibility. This indicates that it activates glycolysis. Some cancer cells use OXPHOS as an energy source. It has the ability to switch to glycolysis. It can flexibly shift fuel sources according to local availability. This ability, which appears to be inherent, is related to abnormal cell proliferation, survival, and remote sites. This is a necessary condition for seeding. It impairs the flexibility of the energy mechanism of cancer cells. Pharmacological and / or metabolic approaches aimed at this purpose may adversely affect tumor progression. It will become that.

[0047] To understand the effect of d-TPP therapy on the metabolism of cancer cells, the inventors of the present invention have found Sea Metabolic flux analysis was performed using horse XFe96. Figures 5A and 5B show various results. The oxygen consumption rate (oxygen co) of MCF-7 cells treated with d-TPP at a certain concentration. The results of the OCR (Observation Rate) are shown. Basal respiration, proton leak, Figure 5A shows the OCR (pmo) for ATP production-related respiration, maximal respiration, and respiratory reserve volume. Figure 5B shows the time course of l / min / SRB, and the OCR is shown as the magnification change relative to the vehicle. This shows that after treatment with 50 nM d-TPP, there was a dramatic decrease in OCR in MCF-7 cells. Observed: The decrease in OCR occurred as the d-TPP concentration increased from 50 nM to 500 nM. Therefore, it grew larger. Mitochondrial basal respiration is approximately 250 nM IC50 (CSC). The activity decreases at the same concentration of the drug required to halve it, and similarly, ATP levels also decrease depending on the dose. It was completely depleted.

[0048] Regarding glycolysis, a clearly opposite trend was observed, and MCF-7 cells treated with d-TPP... ECAR (extracellular acidification rate) in cells As shown by the analysis of extracellular acidification rate, it increased significantly in a dose-dependent manner. The AR results are shown in Figures 6A and 6B. In Figure 6A, ECAR (mpH / min / SRB) is 5 Regarding d-TPP concentrations in the range of 0 nM to 500 nM and the control (vehicle only), time course Figure 6B shows ECAR for glycolysis, glycolytic reserve, and glycolytic reserve capacity. This is shown as a magnification change relative to d-TP. As can be seen, glycolysis in MCF-7 cells is d-TP The levels increased in a dose-dependent manner with P treatment. In summary, these data suggest that d-TPP is mi MCF-7 cells that produce ATP from oxidative phosphorylation after impaired tochondria function. This indicates that it impairs ability. In order to cope with such a stressful metabolic environment, Cancer cells are forced to shift to a glycolytic phenotype. After this metabolic shift, cancer cells... It relies heavily on glucose to meet high energy demands. In this scenario, d-T The anti-mitochondrial effect induced by PP treatment is an absolute and inflexible reliance on glycolysis. It acts as a functional metabolic synchronizer for survival.

[0049] d-TPP inhibits mitochondrial ATP production, shifting CSCs into a glycolytic state. Ability opens the door to a variety of potential treatment strategies, especially therapeutic combinations. d-TPP can be combined with compounds that inhibit glycolysis or oxphos. This timing of combination therapy allows the effects of d-TPP to occur first, followed by glycolysis. It can produce an OXPHOS inhibitory effect. Alternatively, d-TPP can be used as the second It may be administered concurrently with metabolic inhibitor compounds. For example, a combination of d-TPP and vitamin C. The combination offers a metabolic "two-hit" strategy that targets the metabolic vulnerability of CSCs. d-TPP inhibits ATP production, shifting CSCs to a glycolytic profile. Vitamin C inhibits glycolysis, leaving CSCs in a state where they have no metabolic options. For example, 2- Other ingredients include deoxyglucose, doxycycline, niclosamide, and berberine chloride. The inhibitor may be used in combination with d-TPP.

[0050] To demonstrate the effectiveness of this two-hit therapy approach, a tumor-like mass formation assay and cancer The dose of a drug that is selectively toxic only to normal cells but not to normal cells. Further evaluation was performed using d-TPP at a concentration of 00 nM. Figures 7A to 7E show various results. Results of tumor-like mass assays for 100 nM d-TPP combined with inhibitory compounds. This shows the effects of vitamin C, a natural glycolysis inhibitor, and synthetic glycolysis inhibitors, starting from Figures 7A and 7B. The harmful agent 2-deoxy-glucose (2-DG) was included along with d-TPP. Figure 7A. As can be seen in 7B, vitamin C and 2-DG have an effect on CSC activity compared to d-TPP. The inhibitory effect was enhanced. In particular, the treatment using vitamin C was effective when used in combination with 100 nM d-TPP. In that case, only over 50% of CSCs were found at 250 μM and over 70% at 500 μM. It inhibited transmission. Separately, the IC50 of vitamin C is related to the transmission of MCF-7CSC. The concentration was 1 mM. As can be confirmed, d-TPP affects the sensitivity of CSC to vitamin C. It increased sexual performance by nearly four times.

[0051] The inhibitory activity of 2-DG was already observed at a concentration of 10 mM, which is the same as the inhibitory activity of d-TPP alone. This resulted in a twofold increase in activity. The inhibitory effect was even more dramatic at 20 mM, C SC activity was almost completely suppressed, and the residual tumor-like mass formation ability was reduced to less than 10%.

[0052] Next, Figures 7C-7E show two FDA-approved compounds combined with d-TPP. , namely doxycycline and niclosamide, as well as the natural compound berberine chloride. The results show that each of these compounds behaves as an OXPHOS inhibitor. This behavior is known to occur in cancer cells weakened by d-TPP treatment, due to additional metabolic activity. This is based on the understanding that stressors can help eradicate the remaining CSC population. Doxycycline is known to impair mitochondrial biosynthesis and function. As shown in Figure 7C, low doses of the antibiotic doxycycline (e.g., 10) A concentration of μM was sufficient to double the potency of d-TPP against CSC activity. It was enhanced in the presence of 30 μM doxycycline. As shown in Figure 7D, Niclosamide, an anti-tapeworm drug that inhibits OXPHOS, has d- The effectiveness of the TPP was increased by approximately twofold at 250 nM and by approximately threefold at 500 nM. (Figure) 7E is a natural compound and OXPHOS inhibitor chloride combined with d-TPP. The results for berberine are shown. Berberine chloride is used in Coptidis rhizoma (Co ptis chinensis Franch) and Phellodendri cor Extracted from tex (Phellodendron amurense Ruprecht) It is a major alkaloid known for its antimalarial, anti-inflammatory, and antibiotic activity. The effect of berberine chloride was already evident at 1 μM, resulting in a 60% reduction. At the highest concentration tested (10 μM), berberine chloride inhibited CSC formation by more than 80%. This increased the effectiveness of the d-TPP by approximately five times.

[0053] These results indicate that mitochondrial disruptors such as d-TPP disrupt the normal energy mechanisms of cells. Impairing certain functions casts a shadow over the possibility of CSCs using alternative fuels and metabolic pathways. This demonstrates that it has an impact. This effect affects the biology and propagation of CSCs, as well as cancer cells. It has adverse effects on survival rate and proliferation. Due to these adverse effects, CSCs require chemotherapy and light therapy. They become more vulnerable to other cancer therapies, including radiation therapy. CSCs become more sensitive to other chemotherapeutic agents, radiation, and phototherapy. Effective amount of d-TPP

[0054] Using the xCELLigence system, d - The pharmacokinetics of TPP's effects were analyzed. The xCELLigence system is small in size. By measuring the cell number-dependent electrical impedance, the health and behavior of cells can be assessed. Evaluate real-time label-free monitoring. Real-time cell analysis, 50n Regarding 48-hour and 72-hour treatments with d-TPP at concentrations of M, 100nM, and 250nM: The study was conducted using only vehicles. The results of the xCELLigence analysis were as follows: Figures 8A-8D show the results for 48 hours of d-TPP treatment. Figures 8C and 8D show the results for 72 hours of d-TPP treatment. Furthermore, the effect of d-TPP on MCF-7 cells is dose-dependent and time-dependent. The lowest dose tested (50 nM d-TPP) showed a tendency for cell counts to decrease after 72 hours of treatment. At higher doses (100 nM), the main cell proliferation inhibitory effect was detected. Toxicity was present at a d-TPP concentration of 250 nM.

[0055] Further evaluation of both efficacy and toxicity is being conducted by Innovation (Latronche, France). Using a CAM model available from [source], we developed a new cell line from the MDA-MB-231 cell line in chicken embryos. This CAM assay was performed on human breast tumors. This CAM assay determines whether d-TPP contributes to tumor development. It was shown to inhibit both growth and transition. For example, at a concentration of 25 μM, d-TPP The treatment led to a 40% inhibition of tumor growth and a 75% inhibition of metastasis. Regarding the eggs, fertilized eggs were incubated at 37.5°C and 50% relative humidity for 9 days. The graft is dropped onto each egg through a small hole drilled through the shell into the air chamber, above the CAM. 1 cm on the eggshell 2 The window was opened. This transplantation process was applied to each egg in the test group. This transplantation process is an invasive surgical procedure, so it was used within a few hours after tumor transplantation. A certain degree of death is expected. The data reported in this specification are as follows: At least 10 eggs were successfully transplanted per test group.

[0056] MDA-MB-231 tumor cell line was treated with 10% FBS and 1% penicillin / strep The cells were cultured in DMEM medium supplemented with tomycin. After a 9-day incubation period, The cells were detached with trypsin, washed in complete medium, and suspended in transplantation medium. 1.10 6 individual After adding the cell inoculation to the CAM of each egg, the eggs were randomly divided into multiple test groups. The tumor was detectable within one day of transplantation. The negative control group was 0.125% in PBS. This is DMSO, and d-TPP glucoglycerides are available in concentrations of 6.25 μM, 25.0 mM, and 62.5 mM. The group was evaluated. Each egg in the group was treated 8 times per day. 8 days of treatment Subsequently, tumor growth was quantitatively evaluated. The upper part of the CAM (containing the tumor) was removed and treated with PBS. After being washed with cirrhosis fluid, the tumor was directly transferred to PFA for 48 hours of fixation. Next, the tumor was placed in a normal C1. The tumor was carefully isolated from AM tissue and weighed. The average tumor weight (mg) is shown in Figure 9. Yes. For all quantitative data related to the evaluation of these chicken eggs, (post-hoc tests between groups) One-way ANOVA (including) is performed using dedicated computer software called Prism (registered The analysis was conducted using (Trademark) (GraphPad Software). All analyses were performed using Statistical differences between groups are indicated on the graph by the presence of asterisks, which have the following meanings. Visualized as follows: No asterisk means p > 0.05; * means 0.5 ≥ p > 0. 0 means 1; ** means 0.01≧p>0.001; *** means 0.001≧p It tastes good. As can be seen in Figure 9, d-TPP is moderate in dose compared to the negative control. Tumor growth was significantly inhibited at both 25.0 μM and high doses (62.5 μM). Consistent with the assessment, the tumor growth inhibitory effect of d-TPP was dose-dependent.

[0057] Metastatic invasion was also quantitatively evaluated. (1 cm of lower CAM) 2 Collect the part and group The number of metastatic cells was evaluated in eight samples (n=8). Genotherapy was performed using a commercially available kit. The DNA was extracted and analyzed by qPCR using a primer specific to the human Alu sequence. The calculation of Cq for each sample, the average Cq, and the relative amount of the transition for each group was performed by B. Directly managed with io-Rad(registered trademark) CFX Maestro software. Rea l-Time PCR Data Markup Language (RDML) data According to the standard (http: / / www.rdml.org), Cq is the curvature of the amplification curve. It is defined as the largest cycle (fractional PCR cycle). Cq is qP The CR curve is obtained during the exponential amplification phase, where it is a straight line. The basic results of qPCR are as follows: As expected: A lower Cq value indicates a higher initial replication count of the target gene. PCR efficiency is 100%. In that case, the difference of one cycle between the two reactions is compared to the reaction with the higher Cq value. A lower Cq value indicates that the gene replication in the response is twice as high. Figure 10 shows the lower CAM. The results of metastatic invasion measured by qPCR for Alu sequences in the following cases are shown. Negative control and In comparison, metastatic regression can be observed after treatment with moderate doses of d-TPP. For the treated group, statistical evaluation was limited to only the three surviving samples within the group. This was carried out using [a specific method / tool], which affected the statistical analysis.

[0058] The tolerance of embryos treated with d-TPP was also quantitatively evaluated. Embryo viability was assessed post-transfer and during treatment. We performed tests daily. The number of dead embryos was also counted on the final day to assess the embryotoxicity induced by the treatment. It was worthwhile. Figure 11 shows the viable and dead embryos in each group described herein. Figure 12 shows the percentages for the coupler (from transplantation day to retrieval day). This is the Nmayer curve. d-TPP did not show toxicity at moderate and low doses. This can be confirmed (however, as expected, some deaths occurred in the negative control group). However, the higher the dose, the greater the mortality rate. To establish a drug regimen, toxicity is important. The need for such analysis should be understood by anyone with the normal skills in the relevant technical field. Yes. However, the above results indicate that the tumor growth inhibitory and metastasis inhibitory effects of d-TPP are toxic. It has been demonstrated that it exists even at concentrations far below the threshold.

[0059] The data presented here further validates the use of the metabolic "two-hit" approach. Therefore, the above approach manipulates cancer metabolism to increase the metabolic flexibility of CSCs more efficiently. It is used to enable CSC suppression by changing to absolute metabolic inflexibility. Therefore, d-TPP has the potential to act as a safe driving factor for such metabolic shifts. This opens the door to further investigation into the role of this compound in cancer biology and energetics. However, it is used as an adduct to conventional chemotherapy, in combination with other metabolic inhibitors. Further research is needed to support the use of TPP compounds. The same applies to other A-TPP compounds having saturated linear alkyl chains containing 8 carbon atoms. Please understand that the expected effects are to be seen.

[0060] The active compound is A-T, which has a saturated linear alkyl chain containing approximately 9 to 18 carbon atoms. PP compounds, such as d-TPP, target cancer cells, including CSCs and circulating tumor cells. By doing so, cancer is treated and eradicated, tumor recurrence is prevented or its likelihood is reduced, and metastasis is prevented. It can be used as a therapeutic agent to prevent or reduce the likelihood of d-TP. P shifts cancer cells into a glycolytic state, which is counteracted by chemotherapy, radiation therapy, and phototherapy. To sensitize cancer cells and reduce their resistance to other drugs such as doxycycline. It can be used as an active compound.

[0061] Regarding active compounds, the second demonstrably effective inhibitory compound has taken on various forms in the field. It is available at [location]. In the case of A-TPP compounds such as d-TPP, the above active compound is solid or It can be administered orally as a liquid. In some embodiments, d-TPP is a solution, a suspension, or It can be administered intramuscularly, intravenously, or by inhalation as an emulsion. Several implementations Morphologically, d-TPP (to avoid any ambiguity, this includes its salts) is liposome It can be administered as a suspension by inhalation, intravenous, or intramuscular administration. In this case, the active compound or its salt may be, for example, about 0.001, 0.01, 0.1, or 0.5 m Chromometers ~ approximately 5, 10, 20 micrometers or more, and arbitrarily approximately 1 ~ approximately Multiple solid particles or droplets having any desired particle size of 2 micrometers. It can be in a certain state. The specific form of administration may change, and within the scope of this disclosure External parameters (e.g., manufacturing, transportation, storage, shelf life, etc.) are common in d-TPP. Please understand that these can sometimes be determinants of morphology and concentration.

[0062] The pharmaceutical composition of the present invention includes d-TPP (including its salts) as the active compound. This is contained in any pharmaceutically acceptable carrier. If a solution is preferred, water is water-soluble. It can be a carrier of choice for the compound or its salt. Regarding water solubility, glycerol Polyethylene glycol, propylene glycol, polyethylene glycol, or mixtures thereof A suitable vehicle may be used. Furthermore, a method to improve water solubility may be used with the approach of the present invention. It can be used without deviation. In the latter case, the organic vehicle may contain a considerable amount of water. Yes, it is possible. And in any case, the solution can be prepared by preferred means known in the art, e.g. For example, sterilization can be achieved by filtration through a 0.22 micrometer filter. After sterilization, The solution can be dispensed into appropriate containers such as depyrogenic glass vials. This dispensing is optional. It is performed using an aseptic method. Afterwards, the sterile closure device can be placed on the vial, and The contents of the vial can be freeze-dried as needed. (e.g., glycolysis inhibitors or oxphos inhibitors) Embodiments comprising a second inhibitor compound are available in the art. The form can be administered simultaneously. The approach of the present invention is, unless otherwise specified, a particular It is not intended to be limited to specific forms of administration.

[0063] In addition to one or more active compounds, the pharmaceutical formulations of the present invention approach are publicly available in the art. Other known additives may be included. For example, some embodiments include acids (e.g., hydrochloric acid) and bases. Alternatively, a buffer solution (e.g., sodium acetate, sodium borate, sodium citrate, gluconate) It may contain pH adjusters such as sodium phosphate, sodium lactate, and sodium phosphate. Some embodiments use methylparaben, propylparaben, benzyl alcohol, and It may contain antimicrobial preservatives. Antimicrobial preservatives are often designed to make the formulation suitable for multiple doses. It is included when placed in the vial. The pharmaceutical formulations described herein are known in the art. It can be freeze-dried using this technique.

[0064] In embodiments involving oral administration of the active compound, the pharmaceutical composition may be in the form of capsules, tablets, pills, or powders. It can take the form of a preparation, solution, suspension, etc. Sodium citrate, calcium carbonate, Tablets containing various excipients such as calcium phosphate, polyvinylpyrrolidone, sucrose, Starch (for example, potato or tapioca) combined with a binder such as gelatin or acacia It can be used with various disintegrants such as cadre starch and certain complex silicates. Lubricants such as magnesium thearate, sodium lauryl sulfate, and talc are used for tableting. It may be included as a target. Similar types of solid compositions can be used in soft and hard-filled gelatin capsules. It can be used as a filler inside. Related materials include lactose and high molecular weight polyethylene. Lenglycol is also included. Aqueous suspensions and / or elixirs are preferred for oral administration. The compounds of the subject matter of this disclosure can be used as various sweeteners, flavoring agents, colorants, emulsifiers, and / or suspending agents. , as well as water, ethanol, propylene glycol, glycerin, and various similar It can be combined with diluents such as the combination of d-TPP as a second inhibitor. In embodiments in which the compound is included, the second inhibitor compound is not limited to the form of a d-TPP compound. It can be administered in a separate form without being combined with another substance.

[0065] Further embodiments provided herein include the activation disclosed herein. Examples include liposome formulations of the compound. The technique for forming the liposome suspension is the said technique It is publicly known in the field. When the compound is a water-soluble salt, using conventional liposome technology, This can be incorporated into lipid vesicles. In such cases, due to the water solubility of the active compound, The active compound can be substantially contained within the hydrophilic center or core of the liposome. The lipid layer can have any of the conventional compositions, and may also contain cholesterol. It can be made to contain cholesterol, or to be made to be cholesterol-free. The active compound of interest is If it is not water-soluble, here too, conventional liposome formation technology is used to form the salt into liposomes. It can be substantially contained within a hydrophobic lipid bilayer that forms the structure. In this case, the liposomes produced are obtained using standard sonication and homogenization techniques. Similarly, the size can be reduced. The active compounds disclosed herein The liposome formulation containing the product can be freeze-dried to produce a freeze-dried product, which can then be processed with pharmaceutically acceptable water or other substances. The liposome suspension can be regenerated by reconstituting it with a suitable carrier.

[0066] With regard to pharmaceutical compositions, the pharmaceutically effective amount of the active compound described herein is determined by a healthcare professional. This is determined and depends on the patient's condition, size and age, as well as the delivery route. In a limited embodiment, a dosage of approximately 0.1 to approximately 200 mg / kg has therapeutic efficacy. The weight ratio is the ratio of the weight of the active compound, including when salt is used, to the body weight of the subject. Yes. In some embodiments, the dosage is up to approximately 1-5, 10, 20, 30, or 40. This can be used to provide the amount of the active compound necessary to deliver a μM serum concentration of the active compound. In some embodiments, the concentration is approximately 1 mg / kg to approximately 10 mg / kg, and in some other embodiments... In this state, a dosage of approximately 10 mg / kg to approximately 50 mg / kg can be used for oral administration. Typically, a dosage of approximately 0.5 mg / kg to 5 mg / kg is used for intramuscular injection. In some embodiments, the dosage is about 1 μmol for intravenous or oral administration. / kg ~ approximately 50 μmol / kg, or arbitrarily approximately 22 μmol / kg ~ approximately 33 μmol / kg It can be a compound in the form of g. The oral dosage form is, for example, 5 mg per tablet or other solid dosage form. Contains any appropriate amount of active material, including 50, 100, 200, or 500 mg. It is possible.

[0067] The following paragraphs describe the materials and methods used in the experiments and data generation described above. To be listed: d-TPP bromide salt, doxycycline, ascorbic acid, 2-deoxy-D- Glucose (2-DG), berberine chloride, and niclosamide are used in Sigma Aldric Supplied from h. Ascorbic acid and 2-deoxy-D-G are dissolved in the cell culture medium. All compounds except lucose (2-DG) were dissolved in DMSO.

[0068] With regard to the cell cultures referenced herein, MCF7 and MDA-MB-231 breast cancer Cells were obtained from ATCC. Human immortalized fibroblasts (hTERT-BJ1) are Clo Supplied by ntech, Inc. The cells were placed in a humid atmosphere containing 5% CO2 at 37°C. Under the air, 10% FBS (fetal bovine serum), 2 Dirubecco's modified Eagle supplemented with mM GlutaMAX and 1% Pen-Strep Dulbecco's modified Eagle's medium:D They were cultured in MEM.

[0069] The tumor-like mass assay was performed as follows: MCF-7 or MDA-MB-231 cells The single-cell suspension was enzymatically decomposed (1× trypsin EDTA, Sigma Aldrich The cells were then prepared using (25 gauge needle) and manual dissection. Next, the cells were ( 2-Hydroxyethyl methacrylate (PolyHEMA, Sigma Aldrich) Tumor-like mass medium (DMEM-F12 / B) in a culture dish coated with a non-adherent medium. 27 / 20-ng / ml EGF / PenStrep) with 500 cells / cm 2 at density Seeds were sown. Cells were grown for 5 days in a humidified incubator at 37°C, atmospheric pressure, and 5% (v (v) Maintained in carbon dioxide / air. After 5 days of incubation, spheres larger than 50 μm were placed in the eyepiece. The percentage of seeded cells that formed spheres was calculated by counting using the scale. This is called percentage tumor-like mass formation. The tumor-like mass assay is performed in triple replication, and this I repeated it three times independently.

[0070] Extracellular acidification rate (ECAR) and real-time oxygen consumption rate for metabolic flux analysis (OCR) assay, Seahorse Extracellular Flux(X The analysis was performed using an Fe-96 analyzer (Seahorse Bioscience). In short, 15,000 MCF-7 cells per well are used with XFe-96 The cells were seeded into well cell culture plates and incubated overnight to allow them to adhere. Then, the cells were 2 Treatment was performed over 4 hours with increasing concentrations of d-TPP (50 nM ÷ 500 nM). Control cells treated with vehicle alone (DMSO) were treated in parallel. Next, the cells were placed in a preheated X F assay medium (or for OCR measurement, 10 mM glucose, 1 mM pyruvate) XF assay medium supplemented with salts and 2 mM L-glutamine, with pH adjusted to 7.4. The cells were then washed. The cells were then incubated in a non-CO2 incubator at 37°C for 1 hour. The XF assay was maintained in 175 μL / well. During the incubation period, 80m 5 μL of M glucose, 9 μM of oligomycin, and (for ECAR measurement) 1 M 2-deoxyglucose, or (for OCR measurement) 10 μM oligomycin 9 μM FCCP, 10 μM rotenone, 10 μM antimycin A, XF assembly The protein was placed in a culture medium and inserted into the injection port of the XFe-96 sensor cartridge. After normalizing the measured values ​​by content, the dataset was analyzed using XFe-96 software. (SRB). All experiments were repeated three times independently.

[0071] Based on the measured cellular protein content, MCF-7 and hTERT-BJ1 cells In this context, the cell viability is measured using sulforhodamine B (SRB). The assay was performed over 24, 48, or 72 hours. d-TPP (50 nM ÷ 1 μM) After treatment with ), the cells were placed in a cold room and treated with 10% trichloroacetic acid (trichloro The samples were fixed with acetic acid (TCA) for 1 hour and dried overnight at room temperature. Next, the cells were incubated with SRB for 15 minutes, washed twice with 1% acetic acid, and at a minimum... It was then air-dried for one hour. Finally, the protein-binding dye was added in 10 mM Tris pH8. The sample was dissolved in solution 0.8 and read at 540 nm using a plate reader.

[0072] xCELLigence Real-time Cell Analysis The alysis (RTCA) system is manufactured by ACEA Biosciences Inc. The xCELLigence RTCA system was supplied with a cell index (Cell I Attached cells by measuring electrical impedance expressed as ndex (CI) value This provides a useful approach for real-time monitoring of the biological state of [the organism]. 50 00 MCF-7 cells were seeded in a 16-well plate (E plate). 24 hours after seeding After a while, the cells are transported by a vehicle or multiple concentrations of d-TPP (50 nM to 250 nM) that increase. The treatment was then continued for another 48 or 72 hours. RTCA is induced by cells. This is performed by measuring the value of the electrical impedance, and the above value is obtained over 96 hours. It was automatically recorded every 15 minutes. This approach allowed for the initiation and dynamics of cellular responses. This allows for the quantification of the substance. The experiment was conducted independently three times using four samples for each condition. It was returned.

[0073] Unless otherwise specified, the data disclosed herein are 3 per experiment. The mean ± mean standard obtained over three or more independent experiments involving more than one technical replication. It is expressed as a quasi-error (SEM). Statistical significance was measured using a t-test. p ≤ 0.0 A value of 5 was considered statistically significant.

[0074] The terminology used in the above description of embodiments of the approach of the present invention is not merely specific. This description is for illustrative purposes only and is not intended to be limiting. When used in the attached claims, the singular forms "a, an" and "above, the above" are used. The plural form "(the)" also includes the plural form unless the context explicitly indicates otherwise. The intention is to... The approach of the present invention is as follows: As will become clear from the analysis, it includes numerous alternatives, modifications, and equivalents.

[0075] The terms "first", "second", and "third" )", a)", b)", c) etc. are used herein to describe various approaches of the present invention. These terms may be used to describe elements, and the claims may be defined by these terms. It will be understood that these terms are not limited to the approach of the present invention. It is used solely to distinguish one element from another. Therefore, the first is described below. The first element does not depart from the teachings of the approach of the present invention and can also be referred to as an aspect of an element, and similarly can be referred to as the third element. Therefore, the terms "first", "second", "third", "a)", "b)", "c)", etc. are not intended to give an order or other hierarchical relationship to the associated elements, but are simply used for identification . The order of operations (or steps) is not limited to the order presented in the claims.

[0076] Unless otherwise defined, all terms (including technical and scientific terms) used in this specification have the same meaning as commonly understood by those skilled in the art. Further, terms defined as in a commonly used dictionary are to be interpreted as having a meaning consistent with the meaning of such terms in the context of the present application and the related art, and should not be interpreted in an idealized or overly formal sense unless clearly defined herein. It will be understood that all publications, patent applications, patents, and other reference materials mentioned in this specification are hereby incorporated by reference in their entirety. In case of conflict with the terms of the law, this specification shall prevail.

[0077] Also, as used in this specification, "and / or" means any and all possible combinations of one or more of the associated listed items, and the absence of a combination when interpreted as "or", and includes these.

[0078] ​Unless otherwise explicitly stated in the context, the appropriations of the present invention described herein It is specifically intended that the various features of the device can be used in any combination. Furthermore, the present invention approach, in some embodiments, with respect to demonstrative embodiments It is also taken into consideration that any of the described features or combinations of features may be excluded or omitted. .

[0079] When used herein, the transitional phrase "essentially consists of..." The material described is (essentially of) (and its grammatical variations). This is a step and does not substantially affect one or more basic novel features of the claims. things (those that do not materially affect t "The basic and novel characteristic(s)" It should be interpreted as encompassing. Therefore, when used herein, the term "essential" "to consist of..." is interpreted as equivalent to "to include, to be equipped with (comprising)." It must not be done.

[0080] For example, when referring to measurable values ​​such as quantity or concentration, the term used herein is used The word "about" means ±20%, ±10%, ±5%, ±1%, ± It is intended to include variations of 0.5%, or as small as ±0.1%. Measurable. The range provided herein with respect to a value is any other range and / or within that range. Individual values ​​may be included.

[0081] Having described specific embodiments of the approach of the present invention, the following claims are made: Without departing from the spirit or scope of the present invention, numerous obvious changes to these embodiments may be made. Because the form is possible, the attached claims are not specific details as described above. Please understand that this is not limited by that.

Claims

1. A pharmaceutical composition comprising a pharmaceutically effective amount of dodecyltriphenylphosphonium (d-TPP) or a pharmaceutically acceptable salt thereof, and a pharmaceutically effective amount of a second inhibitor compound, wherein the second inhibitor compound is (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A pharmaceutical composition containing the following:

2. The pharmaceutical composition according to claim 1, further comprising one of vitamin C and 2-deoxy-glucose.

3. The pharmaceutical composition according to claim 1, wherein the second inhibitory compound comprises one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline.

4. The pharmaceutical composition according to claim 1, wherein the second inhibitor compound comprises one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin.

5. The pharmaceutical composition according to claim 1, wherein the second inhibitor compound comprises niclosamide.

6. The pharmaceutical composition according to claim 1, wherein the second inhibitor compound comprises berberine chloride.

7. The pharmaceutical composition according to any one of claims 1 to 6, characterized in that it is used for at least one of the following: treatment of cancer, eradication of cancer stem cells, eradication of circulating tumor cells, prevention or reduction of tumor recurrence, prevention or reduction of metastasis, and sensitization of cancer to one of chemotherapy, radiotherapy, and phototherapy.

8. A composition for treating or preventing cancer, comprising a pharmaceutically effective amount of a dodecyltriphenylphosphonium (d-TPP) compound or a pharmaceutically acceptable salt thereof and a pharmaceutically effective amount of a second inhibitor compound, wherein the second inhibitor compound is (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A composition containing the following:

9. The composition according to claim 8, wherein the cancer is breast cancer.

10. The composition according to claim 8, further comprising vitamin C and one of 2-deoxy-glucose and doxycycline.

11. The composition according to claim 8, wherein the second inhibitor compound is one of doxycycline, niclosamide, and berberine chloride.

12. The composition according to claim 8, wherein the d-TPP compound or a pharmaceutically acceptable salt thereof is administered before the administration of the second inhibitor compound.

13. The composition according to claim 10, wherein the d-TPP compound or a pharmaceutically acceptable salt thereof is administered before the administration of vitamin C or 2-deoxyglucose.

14. A composition for treating or preventing one of tumor recurrence and metastasis of cancer, the composition comprising a pharmaceutically effective amount of a dodecyltriphenylphosphonium (d-TPP) compound or a pharmaceutically acceptable salt thereof and a pharmaceutically effective amount of a second inhibitor compound, the second inhibitor compound being (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A composition containing the following:

15. The composition according to claim 14, wherein the cancer is breast cancer.

16. The composition according to claim 14, further comprising one of vitamin C and 2-deoxy-glucose.

17. The composition according to claim 16, wherein the second inhibitor compound is one of doxycycline, niclosamide, and berberine chloride.

18. The composition according to claim 14, wherein the d-TPP compound or a pharmaceutically acceptable salt thereof is administered before the administration of the second inhibitor compound.

19. The composition according to claim 16, wherein the d-TPP compound or a pharmaceutically acceptable salt thereof is administered before the administration of vitamin C or 2-deoxy-glucose.

20. A composition for shifting cancer into a glycolytic state, the composition comprising a pharmaceutically effective amount of a dodecyltriphenylphosphonium (d-TPP) compound or a pharmaceutically acceptable salt thereof and a pharmaceutically effective amount of a second inhibitor compound, wherein the second inhibitor compound is (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A composition containing the following:

21. A composition for increasing the efficacy of cancer therapy, the composition comprising a pharmaceutically effective amount of dodecyltriphenylphosphonium (d-TPP) or a pharmaceutically acceptable salt thereof and a pharmaceutically effective amount of a second inhibitor compound, wherein the second inhibitor compound is (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A composition containing the following:

22. The composition according to claim 21, wherein the cancer therapy includes chemotherapy.

23. A composition for eradicating CSCs and circulating tumor cells in the body of a subject, wherein the composition is A pharmaceutically effective amount of dodecyltriphenylphosphonium (d-TPP) compound or a pharmaceutically acceptable salt thereof; and A pharmaceutically effective dose of the second inhibitor compound The second inhibitor compound includes, (a) one of doxycycline, tetracycline, chlortetracycline, minocycline, and tigecycline, (b) one of azithromycin, erythromycin, telithromycin, clarithromycin, and roxithromycin, (c) niclosamide, or (d) Berberine chloride A composition containing the following:

24. The composition according to claim 23, further comprising one of vitamin C and 2-deoxy-glucose.

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