Compositions and methods for enhancing cancer chemotherapy

Abstract

To provide safe and effective compositions and methods for enhancing the efficacy and / or reducing the side effects of cancer chemotherapy and increasing the sensitivity of chemotherapy-resistant cancer cells.SOLUTION: Nutritional supplements comprising fish oil and selenium have been found to provide various activities that are beneficial in treating cancer and related conditions. The supplement provides a synergistic effect in reducing cancer cell growth when used in combination with a chemotherapeutic agent and can reduce growth in drug resistant cancer cells when used in combination with a chemotherapeutic agent to which the cells are resistant. Effects in reducing angiogenesis, reducing metastasis, decreasing the number of circulating cancer cells, and altering AXL signaling have also been found. The use of the supplement was found to reduce the wasting associated with cachexia and decrease circulating cytokines associated with inflammation. The overall effect was found to prolong survival in a clinical study.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 62 / 519,093, filed June 13, 2017, U.S. Provisional Patent Application No. 62 / 519,096, filed June 13, 2017, U.S. Provisional Patent Application No. 62 / 595,002, filed December 5, 2017, and U.S. Provisional Patent Application No. 62 / 670,275, filed May 11, 2018, and all other cited external materials are incorporated herein by reference in their entirety. In the event that the definition or use of a term in a reference incorporated by reference is inconsistent with or contrary to the definition of the term provided herein, the definition of the term provided herein shall control.

[0002] (Technical field) The field of the invention is cancer chemotherapy, particularly cancer chemotherapy in combination with nutritional supplements. [Background technology]

[0003] The background discussion includes information that may be useful in understanding the present invention. None of the information provided herein is admitted to be prior art or related to the invention claimed herein, or that any publication specifically or implicitly mentioned is prior art.

[0004] While chemotherapy protocols used in the treatment of cancer can clearly benefit patients, they may be ineffective or less effective in some cancers, particularly recurrent or refractory tumors. Furthermore, chemotherapy is associated with significant side effects, including nausea, weight loss, hair loss, immunosuppression, and dermatitis.

[0005] Attempts have also been made to enhance the effectiveness of chemotherapy. Some studies suggest that fish oil intake can improve chemotherapy outcomes, while others suggest that fish oil can inhibit them (Daenen et al., JAMA Oncol (2015) 1(3):350-358). Formulation of chemotherapeutic agents as nanoparticles has also been attempted (Xu et al., Coll. Surf. B: Biointerfaces (2006) 48(1):50-57). However, it is unclear whether all chemotherapeutic agents are suitable for such modification. Co-delivery of chemotherapeutic agents with siRNA designed to prevent multidrug resistance has also been investigated. However, such siRNAs are sequence-specific and may not be suitable for some tumors.

[0006] The reduction of the side effects of chemotherapy is generally aimed at alleviating symptoms.For example, antiemetic drugs can be used to reduce nausea, along with improving diet and avoiding certain foods and eating small meals frequently.Unfortunately, this approach is not always effective.In some cases, chemotherapeutic agents are selected to reduce toxicity in order to reduce side effects, but these agents may also reduce the effectiveness against tumor cells.

[0007] Attempts have also been made to improve the sensitivity of cancer cells that are resistant to chemotherapeutic agents. For example, Kelley, U.S. Pat. No. 7,906,554 (Patent Document 1), describes the use of isoflavones in improving the efficacy of chemotherapeutic agents. The only data presented is for a compound known to have significant anticancer activity alone (dehydroequol), but the effect does not appear to be clearly demonstrated. Kim and Kim, U.S. Pat. No. 8,346,3943 (Patent Document 2), describe the role of glucosamine in sensitizing cancer cells to chemotherapeutic agents. The use of glucosamine has been described. However, it is unclear whether the concentrations of glucosamine found to be effective (1 mM or higher) can be achieved in vivo. U.S. Patent No. 9,095,602 to Gleave et al. describes the use of antisense DNA to suppress TRPM-2 gene activity to enhance chemosensitivity in refractory cancers. However, such an approach requires a means to safely and selectively deliver the antisense DNA to tumors. In addition, it is unclear whether such specific gene suppression will be effective across a variety of cancer cell types and chemotherapy mechanisms. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] U.S. Patent No. 7,906,554 [Patent Document 2] U.S. Patent No. 83463943 [Patent Document 3] U.S. Patent No. 9,095,602 [Non-patent literature]

[0009] [Non-Patent Document 1] Daenen et al, JAMA Oncol (2015)l(3):350-358 [Non-patent document 2] Xu et al,Coll.Surf.B:Biointerfaces(2006)48(l):50-57 Summary of the Invention [Problem to be solved by the invention]

[0010] Thus, there remains a need for safe and effective compositions and methods for enhancing the efficacy and / or reducing the side effects of cancer chemotherapy and increasing the sensitivity of chemotherapy-resistant cancer cells. [Means for solving the problem]

[0011] The present subject matter provides compositions and methods that can improve cancer treatment using supplements containing fish oil and selenium, either alone or in combination with chemotherapeutic agents. Specific formulations of preferred dietary supplements containing fish oil and selenium are provided that have been proven to be effective and palatable.

[0012] The use of supplements containing fish oil and selenium has been found to reduce proliferation in different tumor cell lines, and synergistically when used in combination with various chemotherapeutic agents. Increases in biochemical markers associated with apoptosis and oxidative stress, along with shifts in cell cycle phase distribution, have also been observed in cancer cells treated in this way. Similarly, reductions in tumor size have been observed in animal models of various tumor types upon treatment with such supplements, either alone or in combination with chemotherapeutic agents. In addition to reducing tumor size, the use of supplements containing fish oil and selenium (either alone or in combination with chemotherapeutic agents) has been found to reduce the number and size of tumor-associated blood vessels, demonstrating antiangiogenic effects. Certain chemotherapeutic agents have also been found to have previously undocumented antiangiogenic effects when used alone. In addition to reducing tumor size, the use of such supplements (either alone or in combination with chemotherapeutic agents) has been found to reduce indicators of tumor progression and the incidence of metastasis. Similarly, in clinical settings, the use of supplements containing fish oil and selenium has been found to reduce the number of circulating tumor cells.

[0013] Surprisingly, the stem cell properties of cancer stem cells and sphere cells were reduced by the use of a supplement containing fish oil and selenium. AXL signaling was also altered. Such a supplement could prevent drug-resistant tumor cells from undergoing chemotherapy to which they are resistant. Treatment with a supplement containing fish oil and selenium has been found to alter the expression of biochemical markers associated with the immune response to cancer cells in both cancer cells and cells of the immune system, indicating that such supplements can complement or enhance immunotherapy.

[0014] Supplements containing fish oil and selenium have also been found to be effective in reducing the wasting associated with cachexia, even beyond their caloric contribution. Despite concurrent chemotherapy, the mass of fat, muscle, and various organ tissues is improved or maintained. Biochemical markers associated with tissues affected by cachexia also increase with the use of such supplements. Blood levels of various cytokines, many of which are associated with inflammation, are also normalized with the use of such supplements.

[0015] Overall, the use of supplements containing fish oil and selenium, especially in combination with chemotherapy, provides antitumor benefits and reduces disease-related cachexia and inflammation, and this combination of benefits has been found to improve survival in clinical studies of cancer patients.

[0016] That is, the present invention relates to the following.

[0017] A composition for the treatment of cancer, the composition comprising: a pharmaceutical formulation containing a compound used in the treatment of said cancer and effective in inhibiting tumor cell replication; fish oil; and selenium; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, and docetaxel. the pharmaceutical formulation comprises a plurality of chemotherapeutic compounds; the plurality of chemotherapeutic compounds are selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel; the selenium is provided as a peptide or amino acid prepared from selenium yeast; The selenium and the fish oil are provided as nutritional supplements. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0018] 1. A method for increasing the effectiveness of a cancer chemotherapeutic agent, comprising administering a nutritional supplement comprising fish oil and a selenium peptide in combination with the cancer chemotherapeutic agent, wherein the cancer chemotherapeutic agent comprises one or more pharmaceutical compounds; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, and docetaxel. the pharmaceutical formulation comprises a plurality of chemotherapeutic compounds; the plurality of chemotherapeutic compounds are selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel; The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises the ingredients as shown in Table 1. The nutritional supplement is formulated according to the amounts shown in Table 1.

[0019] 1. A method of altering cell cycle distribution of tumor cells, comprising administering to a first population of said tumor cells a nutritional supplement comprising fish oil and a selenium peptide in combination with a cancer chemotherapeutic agent to effect an alteration in a portion of said tumor cells in said cell cycle phase; Preferably, the alteration comprises an increase in the fraction of the tumor cells in S phase compared to the second population of tumor cells treated with the cancer chemotherapeutic agent alone. the alteration comprises an increase in the fraction of the tumor cells in the subGI phase compared to a second population of tumor cells treated with the cancer chemotherapeutic agent alone. the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel; The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises the ingredients as shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0020] 1. A method of increasing the sensitivity of resistant tumor cells to a cancer chemotherapeutic agent, comprising contacting the resistant tumor cells with a nutritional supplement comprising selenium, fish oil, or a combination of selenium and fish oil, wherein the cancer chemotherapeutic agent comprises one or more pharmaceutical compounds; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel. the pharmaceutical formulation comprises a plurality of chemotherapeutic compounds; The plurality of chemotherapeutic compounds are selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel. The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises the ingredients as shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0021] 1. A method for reducing the cachectic effect of a cancer chemotherapeutic agent, comprising providing a nutritional supplement comprising selenium, fish oil, or a combination of selenium and fish oil to an individual undergoing chemotherapy using said cancer chemotherapeutic agent, wherein said cancer chemotherapeutic agent comprises one or more pharmaceutical compounds; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel. the pharmaceutical formulation comprises a plurality of chemotherapeutic compounds; the plurality of chemotherapeutic compounds are selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, and docetaxel; The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0022] 1. A composition for the treatment of cancer, comprising fish oil and selenium, wherein the fish oil and selenium are provided in amounts effective to provide a synergistic effect on a cancer treatment modality, the synergistic effect exceeding the additive effect of the fish oil or the selenium when provided individually in the corresponding amounts; Preferably, the cancer treatment modality is selected from the group consisting of inhibiting cancer cell replication, reducing metastasis, increasing the effectiveness of chemotherapeutic drugs, sensitizing resistant cancer cells to chemotherapeutic drugs, and reducing the cachectic side effects of chemotherapeutic drugs. The selenium is provided as a peptide or amino acid prepared from selenium yeast. The composition comprises at least three components as shown in Table 1, The composition is formulated according to the amounts shown in Table 1.

[0023] 1. A method of reducing stem cell characteristics of cancer stem cells, comprising contacting the cancer stem cells with a composition comprising selenium, fish oil, or a combination of selenium and fish oil; Preferably, the selenium yeast is in the form of a peptide or amino acid derived from selenium yeast; p-mTOR or vimentin expression is decreased in the cancer stem cells; Beclin-1 is increased in the cancer stem cells. The contacting is achieved by administering to a patient having cancer a composition comprising a plurality of compounds as set forth in Table 1; The compounds are provided in a formulation containing the amounts of the compounds as shown in Table 1.

[0024] 1. A method for reducing proliferation of cancer stem cells, comprising contacting the cancer stem cells with a composition comprising selenium, fish oil, or a combination of selenium and fish oil; Preferably, the selenium is in the form of a peptide or amino acid derived from selenium yeast. GRP78 expression is decreased in said cancer stem cells. CHOP expression is increased in the cancer stem cells. The contacting is achieved by administering to a patient having cancer a composition comprising a plurality of compounds as set forth in Table 1; The compounds are provided in a formulation containing the amounts of the compounds as shown in Table 1.

[0025] A method of treating cancer, comprising administering, at a first time, a nutritional supplement comprising fish oil and selenium. and administering a chemotherapeutic agent at a second time point, wherein the first time point precedes the second time point by a pretreatment interval; Preferably, the pretreatment interval is about 72 hours. The nutritional supplement comprises the ingredients in Table 1.

[0026] 1. A method of reducing angiogenesis in a tumor, the method comprising administering a nutritional supplement comprising fish oil and selenium; Preferably, the method further comprises administering a chemotherapeutic agent. The nutritional supplement comprises the ingredients in Table 1.

[0027] 1. A method of reducing the ability to respond to oxidative stress in a tumor, the method comprising administering a nutritional supplement comprising fish oil and selenium; Preferably, the method further comprises administering a chemotherapeutic agent. The nutritional supplement comprises the ingredients in Table 1.

[0028] 1. A method of reducing circulating tumor cells in a patient having a tumor, the method comprising administering a nutritional supplement comprising fish oil and selenium; Preferably, the method further comprises administering a chemotherapeutic agent. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The patient's first circulating tumor cell content prior to administration of the nutritional supplement is at least 10 times greater than the patient's second circulating tumor cell content after administration of the nutritional supplement.

[0029] 1. A method of enhancing tumor-specific immunotherapy, comprising administering a nutritional supplement comprising fish oil and selenium; Preferably, the method further comprises administering a chemotherapeutic agent. further comprising administering a tumor-specific immunotherapy. The nutritional supplement comprises at least three ingredients as shown in Table 1.

[0030] 1. A method of inducing apoptosis in tumors, comprising administering a nutritional supplement comprising fish oil and selenium; Preferably, the method further comprises administering a chemotherapeutic agent. The nutritional supplement comprises at least three ingredients as shown in Table 1.

[0031] A method for improving two-year survival in a tumor-bearing patient population, comprising administering fish oil and selenium. 10. A method of administering a nutritional supplement comprising: Preferably, the method further comprises administering a chemotherapeutic agent. The nutritional supplement comprises at least three ingredients as set forth in Table 1. the two-year survival rate of a tumor-bearing patient population receiving the nutritional supplement is at least twice the two-year survival rate of a tumor-bearing population receiving a generic supplement; the two-year survival rate of a tumor-bearing patient population receiving the nutritional supplement and having a BMI of less than 19 is at least five times longer than the two-year survival rate of a tumor-bearing population receiving a generic supplement and having a BMI of less than 19; The tumor-bearing patient population includes patients with head and / or neck tumors.

[0032] 1. A method of modulating AXL in tumor cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease the concentration of AXL in the tumor cells; Preferably, the tumor cells are drug-resistant tumor cells. The nutritional supplement comprises at least three ingredients as set forth in Table 1. further comprising administering a chemotherapeutic agent;

[0033] 1. A method of modulating heat shock proteins in tumor cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease the concentration of said heat shock proteins in said tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises at least three ingredients as set forth in Table 1. The method further comprises administering a chemotherapeutic agent.

[0034] 1. A method of modulating mTOR phosphorylation in tumor cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease mTOR phosphorylation in the tumor cells; Preferably, the tumor cells are drug-resistant tumor cells. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The method further comprises administering a chemotherapeutic agent.

[0035] 1. A method of modulating AXL and heat shock proteins in tumor cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease both the concentration of AXL and the concentration of heat shock proteins in the tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises at least three ingredients as set forth in Table 1. The method further comprises administering a chemotherapeutic agent.

[0036] 1. A method for modulating m-TOR phosphorylation and AXL in cancer cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease both the concentration of AXL and the mTOR phosphorylation in the tumor cells; Preferably, the tumor cells are drug-resistant tumor cells. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The method further comprises administering a chemotherapeutic agent.

[0037] A method for regulating mTOR phosphorylation and heat shock proteins in tumor cells. administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease both the levels of heat shock proteins and the phosphorylation of mTOR in the tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises at least three ingredients as set forth in Table 1. further comprising administering a chemotherapeutic agent;

[0038] 1. A method of modulating mTOR phosphorylation, AXL, and heat shock proteins in tumor cells, comprising administering a nutritional supplement comprising fish oil and selenium in an amount effective to decrease all of the AXL concentration, the heat shock protein concentration, and the mTOR phosphorylation in the tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises at least three ingredients as set forth in Table 1. The method further comprises administering a chemotherapeutic agent.

[0039] 1. A method of reducing VEGF in a tumor, comprising providing a nutritional supplement in an amount sufficient to reduce the concentration of VEGF in the tumor, wherein the nutritional supplement comprises selenium and fish oil; Preferably, the nutritional supplement comprises at least three ingredients as shown in Table 1: The method includes the further step of providing a chemotherapeutic agent.

[0040] 1. A method of reducing HIF-α in a tumor, comprising providing a chemotherapeutic agent in an amount effective to reduce the concentration of HIF-α in the tumor; Preferably, the chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. providing a nutritional supplement comprising selenium and fish oil. The nutritional supplement comprises at least three ingredients as shown in Table 1.

[0041] 1. A method for reducing HIF-α in a tumor, comprising providing a nutritional supplement in an amount sufficient to reduce the concentration of HIF-α in the tumor, wherein the nutritional supplement comprises selenium yeast and fish oil; Preferably, the nutritional supplement comprises at least three ingredients as shown in Table 1: The method includes the further step of providing a chemotherapeutic agent.

[0042] 1. A method of reducing CD31 in a tumor, comprising providing a chemotherapeutic agent in an amount effective to reduce the concentration of CD31 in the tumor; Preferably, the chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. providing a nutritional supplement comprising selenium and fish oil. The nutritional supplement comprises at least three ingredients as shown in Table 1.

[0043] 1. A method of reducing CD31 in a tumor, comprising providing a nutritional supplement in an amount sufficient to reduce the concentration of CD31 in the tumor, wherein the nutritional supplement comprises selenium and fish oil; Preferably, the nutritional supplement is a supplement shown in Table 1. The method includes the further step of providing a chemotherapeutic agent.

[0044] 1. A method of reducing PD-Ll expression in cancer cells, comprising treating said cancer cells with a chemotherapeutic agent. wherein the chemotherapeutic agent is provided in an amount sufficient to reduce PD-Ll expression in the cancer cells; Preferably, the method further comprises contacting the tumor cells with a nutritional supplement comprising fish oil and selenium, wherein the supplement is provided in an amount sufficient to reduce the PD-Ll expression in the cancer cells. The selenium is provided as selenium yeast. The nutritional supplement comprises at least three ingredients from Table 1. The at least three ingredients are provided in the amounts shown in Table 1.

[0045] 1. A method of reducing PD-Ll expression in cancer cells, comprising contacting said cancer cells with a nutritional supplement comprising fish oil and selenium, wherein said supplement is provided in an amount sufficient to reduce said PD-Ll expression in said cancer cells; Preferably, the method further comprises contacting the tumor cells with a nutritional supplement comprising fish oil and selenium, wherein the supplement is provided in an amount sufficient to reduce the PD-Ll expression in the cancer cells. The selenium is provided as selenium yeast. The nutritional supplement comprises at least three ingredients from Table 1. The at least three ingredients are provided in the amounts shown in Table 1.

[0046] 1. Use of a chemotherapeutic compound and a nutritional supplement in the manufacture of a medicament effective for treating cancer, wherein the chemotherapeutic compound is a compound utilized in the treatment of cancer and effective in inhibiting tumor cell replication, and the nutritional supplement comprises fish oil and selenium; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, and docetaxel. the chemotherapeutic compounds are utilized in the treatment of the cancer and comprise a plurality of chemotherapeutic compounds effective in inhibiting the replication of the tumor cells; the plurality of chemotherapeutic compounds are selected from the group consisting of two or more of Iressa, Tarceva, Alimta, cisplatin, Taxol, Adriamycin, Avastin, and docetaxel; The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises three or more ingredients shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0047] 1. Use of a nutritional supplement in the manufacture of a medicament effective to increase the effectiveness of a cancer chemotherapeutic agent, said nutritional supplement comprising fish oil and selenium, said nutritional supplement provided in an amount sufficient to increase the effectiveness of said cancer chemotherapeutic agent, said cancer chemotherapeutic agent comprising one or more pharmaceutical compounds; Preferably, the one or more pharmaceutical compounds are selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, and docetaxel. The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises three or more ingredients as set forth in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0048] 1. Use of a nutritional supplement in the manufacture of a medicament effective when used in combination with a chemotherapeutic agent to alter cell cycle distribution of a plurality of tumor cells, wherein the nutritional supplement comprises fish oil and selenium, and the nutritional supplement is provided in an amount effective when used in combination with the chemotherapeutic agent to cause an alteration in cell cycle distribution of the plurality of tumor cells; Preferably, said modification is a first proportion of tumor cells in S phase following said cancer chemotherapy. an increase in a second proportion of tumor cells in the plurality compared to a corresponding second proportion of tumor cells in the plurality treated with the agent alone; the alteration comprises an increase in a third proportion of the plurality of tumor cells in the subGI phase compared to a corresponding third proportion of the plurality of tumor cells treated with the cancer chemotherapeutic agent alone. the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel; The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises three or more ingredients shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0049] 1. Use of a nutritional supplement in the manufacture of a medicament effective for increasing the sensitivity of tumor cells resistant to a cancer chemotherapeutic agent, wherein the nutritional supplement comprises selenium, fish oil, or a combination of selenium and fish oil, and the cancer chemotherapeutic agent comprises one or more pharmaceutical compounds; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel. The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises three or more ingredients shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0050] 1. Use of a nutritional supplement in the manufacture of a medicament effective for reducing the cachectic effect of a cancer chemotherapeutic agent, wherein the nutritional supplement comprises selenium, fish oil, or a combination of selenium and fish oil, and the cancer chemotherapeutic agent comprises one or more pharmaceutical compounds; Preferably, the pharmaceutical compound is selected from the group consisting of Iressa, Tarceva, Alimta, cisplatin, taxol, adriamycin, avastin, and docetaxel. The selenium is provided as a peptide or amino acid prepared from selenium yeast. The nutritional supplement comprises three or more ingredients shown in Table 1. The nutritional supplements are formulated according to the amounts shown in Table 1.

[0051] 1. Use of a nutritional supplement in the manufacture of a medicament effective for modifying stem cell characteristics of cancer stem cells, wherein the nutritional supplement comprises selenium, fish oil, or a combination of selenium and fish oil; Preferably, the selenium is in the form of a peptide or amino acid derived from selenium yeast. the stem cell characteristic is p-mTOR expression or vimentin expression, and the p-mTOR expression or the vimentin expression is decreased in the cancer stem cells; the stem cell characteristic is Beclin-1 expression, and the Beclin-1 expression is increased in the cancer stem cells. The nutritional supplement comprises three or more compounds as shown in Table 1.

[0052] 1. Use of a nutritional supplement in the manufacture of a medicament effective for reducing the proliferation of cancer stem cells, wherein the nutritional supplement comprises selenium, fish oil, or a combination of selenium and fish oil; Preferably, the selenium is in the form of a peptide or amino acid derived from selenium yeast. the agent is effective to reduce GRP78 expression in the cancer stem cells. the agent is effective to increase CHOP expression in the cancer stem cells. The nutritional supplement comprises three or more ingredients as set forth in Table 1. The three or more ingredients are provided in the amounts set forth in Table 1.

[0053] 1. Use of a nutritional supplement in the manufacture of a medicament effective for reducing angiogenesis in a tumor, said nutritional supplement comprising fish oil and selenium; Preferably, the agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing angiogenesis in the tumor. The nutritional supplement comprises three or more ingredients as shown in Table 1.

[0054] 1. Use of a nutritional supplement in the manufacture of a medicament effective for reducing the ability of a tumor to respond to oxidative stress, wherein the nutritional supplement comprises fish oil and selenium; Preferably, the agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing the ability of the tumor to respond to the oxidative stress. The nutritional supplement comprises three or more ingredients as shown in Table 1.

[0055] 1. Use of a nutritional supplement in the manufacture of a medicament effective to reduce circulating tumor cells in a patient having a tumor, said nutritional supplement comprising fish oil and selenium; Preferably, the agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing the circulating tumor cells in the patient. The nutritional supplement comprises three or more ingredients shown in Table 1. The agent is effective to reduce a first circulating tumor cell content in the patient before administration of the nutritional supplement to a second circulating tumor cell content in the patient after administration of the nutritional supplement, wherein the second circulating tumor cell content is 10% or less of the first circulating tumor cell content.

[0056] 1. Use of a nutritional supplement in the manufacture of a medicament effective for enhancing tumor-specific immunotherapy, wherein the nutritional supplement comprises fish oil and selenium; Preferably, said agent provides a synergistic effect in enhancing said tumor-specific immunotherapy when used in combination with a chemotherapeutic agent. The nutritional supplement comprises three or more ingredients as shown in Table 1.

[0057] 1. Use of a nutritional supplement in the manufacture of a medicament effective for inducing apoptosis in tumors, said nutritional supplement comprising fish oil and selenium; Preferably, said agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in inducing apoptosis in said tumor. The nutritional supplement comprises three or more ingredients listed in Table 1.

[0058] 1. Use of a nutritional supplement in the manufacture of a medicament effective to improve two-year survival in a tumor-bearing patient population, said nutritional supplement comprising fish oil and selenium; Preferably, the agent provides a synergistic effect when used in combination with a chemotherapeutic agent in improving the two-year survival rate in a tumor-bearing patient population. The nutritional supplement comprises three or more ingredients as set forth in Table 1. the agent is effective to improve the two-year survival rate of a tumor-bearing patient population receiving the nutritional supplement by at least two times the two-year survival rate of a tumor-bearing population receiving a generic supplement; the agent is effective to improve the two-year survival rate of a population of patients receiving the nutritional supplement and bearing tumors with a BMI of less than 19 to be at least five times longer than the two-year survival rate of a population receiving a generic supplement and bearing tumors with a BMI of less than 19; The tumor-bearing patient population includes patients with head and / or neck tumors.

[0059] 1. Use of a nutritional supplement in the manufacture of a medicament effective to modulate AXL in tumor cells, said nutritional supplement comprising fish oil and selenium in amounts effective to reduce the concentration of AXL in said tumor cells; Preferably, the tumor cells are drug-resistant tumor cells. The nutritional supplement comprises three or more ingredients as set forth in Table 1. The agent provides a synergistic effect in reducing the concentration of AXL in the tumor cells when used in combination with a chemotherapeutic agent, and further comprises the step of administering the chemotherapeutic agent.

[0060] 1. Use of a nutritional supplement in the manufacture of a medicament effective for modulating heat shock protein expression in tumor cells, said nutritional supplement comprising fish oil and selenium in amounts effective to decrease the concentration of said heat shock protein in said tumor cells; The heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises three or more ingredients as set forth in Table 1. The agent provides a synergistic effect in reducing HSP90 expression in the tumor cells when used in combination with a chemotherapeutic agent, further comprising administering the chemotherapeutic agent.

[0061] 1. Use of a nutritional supplement in the manufacture of a medicament effective to modulate mTOR phosphorylation in tumor cells, said nutritional supplement comprising fish oil and selenium in amounts effective to decrease said mTOR phosphorylation in said tumor cells; the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises three or more ingredients as set forth in Table 1. The agent provides a synergistic effect in reducing mTOR phosphorylation in the tumor cells when used in combination with a chemotherapeutic agent, further comprising administering the chemotherapeutic agent.

[0062] 1. Use of a nutritional supplement in the manufacture of a medicament effective for modulating both AXL and heat shock proteins in tumor cells, wherein the nutritional supplement comprises fish oil and selenium in amounts effective to reduce both the concentration of AXL and the concentration of heat shock proteins in the tumor cells; The heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises three or more ingredients as set forth in Table 1. The agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing the expression of the heat shock protein and the AXL in the tumor cells, and further comprises the step of administering the chemotherapeutic agent.

[0063] 1. Use of a nutritional supplement in the manufacture of a medicament effective for modulating m-TOR phosphorylation and AXL in tumor cells, wherein the nutritional supplement comprises fish oil and selenium in amounts effective to decrease both the concentration of AXL and the mTOR phosphorylation in the tumor cells; Preferably, the tumor cells are drug-resistant tumor cells. The nutritional supplement comprises three or more ingredients as set forth in Table 1. The agent, when used in combination with a chemotherapeutic drug, provides a synergistic effect in reducing the mTOR phosphorylation and the AXL expression in the tumor cells, and further comprises a step of administering the chemotherapeutic drug.

[0064] 1. Use of a nutritional supplement in the manufacture of a medicament effective for modulating mTOR phosphorylation and heat shock proteins in tumor cells, said nutritional supplement comprising fish oil and selenium in amounts effective to decrease both the concentration of said heat shock proteins and said mTOR phosphorylation in said tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises three or more ingredients shown in Table 1. The method further comprises administering the chemotherapeutic agent, wherein the agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing the mTOR phosphorylation and the expression of heat shock proteins in the tumor cells.

[0065] Use of a nutritional supplement in the manufacture of a medicament effective for modulating mTOR phosphorylation, AXL expression, and heat shock protein expression in tumor cells, wherein the nutritional supplement comprises fish oil and selenium in amounts effective to decrease all of the AXL concentration, the heat shock protein concentration, and the mTOR phosphorylation in the tumor cells; Preferably, the heat shock protein is HSP90. the tumor cells are drug-resistant tumor cells; The nutritional supplement comprises three or more ingredients shown in Table 1. The agent, when used in combination with a chemotherapeutic agent, provides a synergistic effect in reducing mTOR phosphorylation, the heat shock protein expression, and AXL expression in the tumor cells.

[0066] 1. Use of a nutritional supplement in the manufacture of a medicament effective to reduce vascular growth factor (VEGF) in a tumor, said nutritional supplement comprising selenium and fish oil, provided in an amount sufficient to reduce the concentration of VEGF in said tumor; The nutritional supplement comprises three or more ingredients as set forth in Table 1. The method includes the further step of providing a chemotherapy agent.

[0067] 1. Use of a chemotherapeutic agent in the preparation of a medicament effective to reduce HIF-α in a tumor, wherein the chemotherapeutic agent is provided in an amount effective to reduce the concentration of HIF-α in the tumor; Preferably, the chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. The medicament further comprises a nutritional supplement comprising selenium and fish oil. The nutritional supplement comprises at least three ingredients as shown in Table 1.

[0068] 1. Use of a nutritional supplement in the manufacture of a medicament effective to reduce HIF-α in a tumor, said nutritional supplement comprising fish oil and selenium, said nutritional supplement provided in an amount sufficient to reduce the concentration of HIF-α in said tumor; Preferably, the nutritional supplement comprises at least three ingredients as shown in Table 1.

[0069] 1. Use of a chemotherapeutic agent in the preparation of a medicament effective to reduce CD31 in a tumor, said medicament comprising said chemotherapeutic agent in an amount effective to reduce the concentration of said CD31 in said tumor; Preferably, the chemotherapeutic agent is selected from the group consisting of taxol, adriamycin, and avastin. The medicament further comprises a nutritional supplement comprising selenium and fish oil. The nutritional supplement comprises three or more ingredients as shown in Table 1.

[0070] 1. Use of a nutritional supplement in the manufacture of a medicament effective to reduce CD31 in a tumor, said nutritional supplement comprising selenium and fish oil in amounts sufficient to reduce the concentration of CD31 in said tumor; Preferably, the nutritional supplement comprises three or more ingredients shown in Table 1.

[0071] 1. Use of a chemotherapeutic agent in the preparation of a medicament effective in reducing PD-L1 expression in cancer cells, wherein the medicament comprises the chemotherapeutic agent in an amount sufficient to reduce the PD-L1 expression in the cancer cells; Preferably, the medicament further comprises a nutritional supplement comprising fish oil and selenium. The selenium is provided as selenium yeast. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The at least three ingredients are provided in the amounts shown in Table 1.

[0072] 1. Use of a nutritional supplement in the manufacture of a medicament effective for reducing PD-L1 expression in cancer cells, wherein the nutritional supplement comprises fish oil and selenium, and the supplement is provided in an amount sufficient to reduce the PD-L1 expression in the cancer cells; Preferably, the selenium is provided as selenium yeast. The nutritional supplement comprises at least three ingredients as set forth in Table 1. The at least three ingredients are provided in the amounts shown in Table 1.

[0073] Various objects, features, aspects and advantages of the present subject matter will become more apparent from the following detailed description of preferred embodiments. [Brief explanation of the drawings]

[0074] [Figure 1] Figure 1 is a histogram of the effect of treatment with Iressa (3 μM-9 μM) in combination with PBS or various supplements on the proliferation of A549 lung tumor cells. [Figure 2] Figure 2 is a histogram of the effect of treatment with Iressa (0.125 μM to 1 μM) in combination with PBS or various supplements on the proliferation of A549 lung tumor cells. [Figure 3] Figure 3 is a histogram of the effect of treatment with Iressa (0.5 μM to 4 μM) in combination with PBS or various supplements on the proliferation of A549 lung tumor cells. [Figure 4] Figure 4 is a histogram of the effect of treatment with PBS or Tarceva (5 μM to 20 μM) in combination with various supplements on the proliferation of A549 lung tumor cells. [Figure 5] Figure 5 is a histogram of the effect of treatment with PBS or Tarceva (0.625 µM to 5 µM) in combination with various supplements on the proliferation of A549 lung tumor cells. [Figure 6] Figure 6 is a histogram of the effect of treatment with PBS or Alimta (0.125 μM to 1 μM) in combination with various supplements on the proliferation of A549 lung tumor cells. [Figure 7] FIG. 7 is a histogram of the effect of treatment with PBS or cisplatin (0.125 μg / mL to 1 μg / mL) in combination with various supplements on the proliferation of A549 lung tumor cells. [Figure 8]FIG. 8 is a histogram of the effect of treatment with docetaxel (1.25 μM to 5 μM) in combination with PBS or various supplements on the proliferation of MDA-MB-231 breast cancer cells. [Figure 9] FIG. 9 is a histogram of the effect of treatment with PBS or cisplatin (1.25 μg / mL to 10 μg / mL) in combination with various supplements on the proliferation of MDA-MB-231 breast cancer cells. [Figure 10] FIG. 10 shows the effect of a nutritional supplement containing selenium, a supplement containing fish oil, and a nutritional supplement containing both fish oil and selenium on the growth of A549 cells in culture. [Figure 11] FIG. 11 is a graph showing the effect of a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on the expression of Ki-67. [Figure 12] FIG. 12 shows the initial body weight of mice used in a nutritional supplement study containing Tarceva and fish oil and selenium. [Figure 13] FIG. 13 is a histogram showing tumor weights in mice treated with Tarceva, a nutritional supplement containing fish oil and selenium, or a combination thereof. [Figure 14] FIG. 14 is a histogram of the effects of a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent in mice implanted with human breast cancer cells. [Figure 15] FIG. 15 is a graphical representation of the effect of combined treatment with a nutritional supplement containing fish oil and selenium and chemotherapy on tumor volume and burden. [Figure 16] FIG. 16 is a histogram and photograph of the effect of treatment with Iressa and Iressa in combination with a nutritional supplement containing fish oil and selenium on the weight and weight distribution of tumors resulting from injection of lung cancer cells. [Figure 17]FIG. 17 is a graph of the effect on tumor weight of adding a nutritional supplement containing fish oil and selenium to a taxol and adriamycin treatment regime in an in vivo model of breast cancer. [Figure 18] FIG. 18 is a graph of the effect on tumor volume of adding a nutritional supplement containing fish oil and selenium to a taxol and adriamycin treatment regime in an in vivo model of breast cancer. [Figure 19] FIG. 19 is a histogram of the effect on tumor volume of treatment with a nutritional supplement containing fish oil and selenium and Avastin. [Figure 20] FIG. 20 is a histogram of the effect on tumor volume of treatment with a nutritional supplement containing fish oil and selenium and taxol. [Figure 21] FIG. 21 is a graph of the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on tumor volume and tumor weight. [Figure 22] FIG. 22 is a photograph and histogram of the effect on tumor weight distribution and weight gain of a prior art nutritional supplement and a nutritional supplement containing fish oil and selenium in combination with chemotherapy. [Figure 23] FIG. 23 is a schematic diagram of a typical study protocol for evaluating the effects of treatment with a nutritional supplement containing fish oil and selenium in combination with taxol or adriamycin. [Figure 24] FIG. 24 is a photograph showing tumors and associated blood vessels in mice from the untreated control group. [Figure 25] Figure 25 is a photograph showing a tumor and associated blood vessels from a mouse treated with a nutritional supplement. [Figure 26] FIG. 26 is a photograph showing tumors and associated blood vessels from mice treated with taxol. [Figure 27]FIG. 27 is a photograph showing tumors and associated blood vessels from mice treated with a combination of taxol and a nutritional supplement containing fish oil and selenium. [Figure 28] FIG. 28 is a comparative photograph showing tumors and associated blood vessels from mice treated with either taxol or a combination of taxol and a nutritional supplement containing fish oil and selenium. [Figure 29] FIG. 29 is a photograph showing tumors and associated blood vessels from mice treated with adriamycin. [Figure 30] FIG. 30 is a photograph showing tumors and associated blood vessels from mice treated with a combination of adriamycin and a nutritional supplement containing fish oil and selenium. [Figure 31] FIG. 31 is a comparative photograph showing tumors and associated blood vessels from mice treated with a combination of adriamycin or a combination of adriamycin and a nutritional supplement containing fish oil and selenium. [Figure 32] Figure 32 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of oxidative stress (i.e., HIF-α) and angiogenesis (i.e., VEGF) markers in an in vivo model of human cancer. Actin is included as a control. [Figure 33] Figure 33 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of angiogenesis (i.e., CD31) and metastasis (i.e., MMP-9) markers in an in vivo model of human cancer. Actin is included as a control. [Figure 34] FIG. 34 is a distribution of cell cycle phases of A549 tumor cells treated with chemotherapeutic agents (Alimta, 1 μM or Tarceva, 5 μM) and a nutritional supplement containing fish oil and selenium. [Figure 35]Figure 35 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of various apoptotic markers in an in vivo model of breast cancer. Actin is included as a control. [Figure 36] Figure 36 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of Bcl-2, p-Bcl-2, and caspase 3 apoptosis markers in an in vivo model of breast cancer. Actin is included as a control. [Figure 37] Figure 37 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of the apoptosis marker caspase 8 in an in vivo model of breast cancer. Actin is included as a control. [Figure 38] Figure 38 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on the expression of cytochrome C in an in vivo model of breast cancer. Actin is included as a control. [Figure 39] FIG. 39 is a photograph of a cell culture plate showing apoptosis in A549 lung cancer cells treated with a supplement containing fish oil and selenium. [Figure 40] Figure 40 is a trypan blue staining of A549 lung cancer cells treated with a supplement containing fish oil and selenium. [Figure 41] FIG. 41 is a histogram of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma concentrations of tumor MDA in an in vivo model of breast cancer. [Figure 42] FIG. 42 is a histogram of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor MDA levels in an in vivo model of breast cancer. [Figure 43]FIG. 43 is a histogram of the effect of treatment with Iressa (0.625 μM to 5 μM) in combination with PBS or various supplements on the proliferation of A549 tumorsphere (i.e., stem) cells. [Figure 44] FIG. 44 is a histogram of the effect of treatment with PBS or Tarceva (1.25 μM to 10 μM) in combination with various supplements on the proliferation of A549 tumorsphere (ie, stem) cells. [Figure 45] FIG. 45 is a histogram of the effect of treatment with docetaxel (1.25 μM to 5 μM) in combination with PBS or various supplements on the proliferation of MDA-MB-231 tumorsphere (ie, stem) cells. [Figure 46] FIG. 46 is a histogram of the effect of treatment with PBS or cisplatin (1.25 μg / mL to 10 μg / mL) in combination with various supplements on the proliferation of MDA-MB-231 tumorsphere (i.e., stem) cells. [Figure 47] FIG. 47 shows the cell cycle phase distribution of A549 tumor sphere cells (i.e., stem cells) treated with chemotherapeutic agents (Alimta, 1 μM or Tarceva, 5 μM) and a nutritional supplement containing fish oil and selenium. [Figure 48] Figure 48 is a Western blot showing the expression of various proteins in parental and sphere (i.e., stem) A549 cells. [Figure 49] Figure 49 is a Western blot showing elevated expression of stem cell-associated proteins in A549 sphere (ie, stem) cells. [Figure 50] FIG. 50 is a Western blot showing the reduction of stem cell properties in A549 sphere (ie, stem) cells treated with a nutritional supplement containing fish oil and selenium. [Figure 51] FIG. 51 is a Western blot showing the reduction of stem cell properties in A549 sphere (ie, stem) cells treated with a nutritional supplement containing fish oil and selenium. [Figure 52]FIG. 52 is a Western blot showing the reduction of stem cell properties in A549 sphere (ie, stem) cells treated with a nutritional supplement containing fish oil and selenium. [Figure 53] Figure 53 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on cancer stem cell markers in an in vivo model of breast cancer. Actin is included as a control. [Figure 54] FIG. 54 is a photomicrograph of Iressa-resistant cancer cells treated with a nutritional supplement containing fish oil and selenium and / or Iressa. [Figure 55] Figure 55 is a histogram of the response of HCC827 and resistant HCC827Gr cell lines to Iressa. [Figure 56] Figure 56 is a Western blot showing the differences in expression of various markers between HCC827 and resistant HCC287Gr cells, which exhibit stem cell properties. [Figure 57] FIG. 57 is a histogram showing sensitization of resistant HCC827Gr cells to Iressa up to 1 μM upon combined treatment with a nutritional supplement containing fish oil and selenium. [Figure 58] FIG. 58 is a histogram showing sensitization of resistant HCC827Gr cells to Iressa up to 4 μM upon combined treatment with a nutritional supplement containing fish oil and selenium. [Figure 59] Figure 59. Inhibition of colony formation by resistant HCC827Gr cells by treatment with Iressa in combination with a nutritional supplement containing fish oil and selenium. [Figure 60] FIG. 60 is a histogram showing inhibition of colony formation by resistant HCC827Gr cells by treatment with Iressa in combination with a nutritional supplement containing fish oil and selenium. [Figure 61]FIG. 61 is a frequency distribution of cell cycle phases for treatment of Iressa-resistant HCC827GR cells with Iressa, a nutritional supplement containing fish oil and selenium, and the combined treatment. [Figure 62] FIG. 62 is a frequency distribution of cell cycle phases in Iressa-resistant HCC827GR cells after pretreatment with a nutritional supplement of the present inventive concepts, with or without the addition of Iressa. [Figure 63] FIG. 63 shows the frequency of lung cancer metastasis in an animal model treated with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent. [Figure 64] Figure 64 is a photomicrograph showing the results of immunohistochemical staining for cells bearing tumor markers in transplanted mice treated with Tarceva, a nutritional supplement containing fish oil and selenium, and a combination of Tarceva and the supplement. [Figure 65] Figure 65 is a graph of the effect of a nutritional supplement containing fish oil and selenium on metastatic tumor burden when used in combination with taxol or adriamycin in an in vivo breast cancer model. [Figure 66] Figure 66. Frequency of metastases in tumor-bearing mice treated with nutritional supplements containing fish oil and selenium and / or multiple chemotherapeutic agents. [Figure 67] Figure 67 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor expression of MMP-9 in an in vivo model of breast cancer. Actin is included as a control. [Figure 68] FIG. 68 shows the reduction in circulating tumor cells in clinical patients treated with a nutritional supplement containing fish oil and selenium. [Figure 69] Figure 69 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on the immune checkpoint proteins CD279 and CD274 in an in vivo model of breast cancer. Actin is included as a control. [Figure 70]Figure 70 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on primary tumor PD-L1 and T cell PD-1 in an in vivo model of human cancer. Actin is included as a control. [Figure 71] Figure 71 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on metastatic tumor (i.e., breast) PD-L1 and T cell PD-1 in an in vivo model of metastatic human cancer. Actin is included as a control. [Figure 72] Figure 72 is a histogram and western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on tumor PD-L1 and T cell PD-1 in an in vivo model of human cancer. Actin is included as a control. [Figure 73] FIG. 73 is a graph showing the CD4 / CD8 ratios of breast cancer patients receiving nutritional supplements containing fish oil and selenium. [Figure 74] FIG. 74 is a graph showing the CD4 / CD8 ratios of lymphoma patients receiving nutritional supplements containing fish oil and selenium. [Figure 75] FIG. 75 is a histogram showing the relative loss of adipose tissue and muscle mass in tumor-implanted mice treated with a nutritional supplement containing fish oil and selenium and / or chemotherapeutic agents. [Figure 76] FIG. 76 is a schematic representation of the effect of oncogene products on energy expenditure. [Figure 77] FIG. 77 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and combined treatment with such a supplement and a chemotherapeutic agent on PTHrP and IL-6 expression in tumors. [Figure 78] Figure 78 shows a typical experimental design for characterizing the effects of combination therapy using a nutritional supplement containing fish oil and selenium with various chemotherapeutic drugs in tumor-bearing mice. [Figure 79]Figure 79 is a Western blot, histogram, and photomicrograph showing the expression of autophagy-related protein expression in tumor-bearing mice treated with omega-3 fatty acids. [Figure 80] FIG. 80 is a histogram and photomicrographs showing the preservation of adipose tissue in tumor-bearing mice treated with a nutritional supplement containing fish oil and selenium and / or chemotherapeutic agents. [Figure 81] FIG. 81 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on the preservation of white and brown adipose tissue in tumor-bearing mice. [Figure 82] FIG. 82 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on the preservation of gastrocnemius muscle mass in tumor-bearing mice. [Figure 83] FIG. 83 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on the preservation of gastrocnemius muscle mass and total skeletal muscle mass in tumor-bearing mice. [Figure 84] FIG. 84 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on brain and liver weights in tumor-bearing mice. [Figure 85] FIG. 85 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on heart and spleen weights in tumor-bearing mice. [Figure 86] FIG. 86 is a histogram showing the effect of treatment with a nutritional supplement containing fish oil and selenium and / or a chemotherapeutic agent on lung weight and metastasis-free mammary gland weight in tumor-bearing mice. [Figure 87]Figure 87 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on muscle mitochondrial and inflammatory (i.e., IL6) markers in an in vivo model of breast cancer. GAPDH is included as a control. [Figure 88] Figure 88 is a Western blot showing the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on complex III and complex IV mitochondrial complex proteins in an in vivo model of human cancer. GAPDH is included as a control. [Figure 89] Figure 89 shows the results of an rtPCR and Western blot study of IL-6, PTHrP, UCP-1, ATGL, and HSL expression in animal models of human cancer. [Figure 90] Figure 90 is a graph showing the reduction of cachexia in head and neck cancer patients using a nutritional supplement containing fish oil and selenium. The left panel shows the mean change in body weight of patients treated with a nutritional supplement containing fish oil and selenium (squares) and a generic supplement (triangles). The right panel shows the mean change in body weight of patients with a BMI of 19 or less treated with a nutritional supplement containing fish oil and selenium (squares), the mean change in body weight of patients with a BMI > 19 treated with a nutritional supplement containing fish oil and selenium (open circles), patients with a BMI of 19 or less treated with a generic supplement (triangles), and patients with a BMI > 19 treated with a generic supplement (filled circles). [Figure 91] Figure 91 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on food intake in an in vivo model of human cancer. [Figure 92] Figure 92 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma levels of VEGF in an in vivo model of breast cancer. [Figure 93]Figure 93 is a histogram of the effect of a nutritional supplement containing fish oil and selenium in combination with the drugs taxol, adriamycin, or avastin on plasma concentrations of VEGF in an in vivo model of breast cancer. [Figure 94] Figure 94 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma levels of TNF-α in an in vivo model of breast cancer. [Figure 95] Figure 95 is a histogram of the effect of a nutritional supplement containing fish oil and selenium in combination with the drugs taxol, adriamycin, or avastin on plasma concentrations of TNF-α and IL6 in an in vivo model of breast cancer. [Figure 96] FIG. 96 is a graph of the effect of a nutritional supplement containing fish oil and selenium, alone and in combination with taxol, adriamycin, or avastin, on plasma concentrations of IL-1-β and IL-10 in an in vivo model of human cancer. [Figure 97] FIG. 97 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma concentrations of IL-1-β in an in vivo model of breast cancer. [Figure 98] Figure 98 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma levels of IL6 in an in vivo model of breast cancer. [Figure 99] Figure 99 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma levels of IL2 in an in vivo model of breast cancer. [Figure 100] Figure 100 is a histogram of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma concentrations of IFN-γ and IL-2 in an in vivo model of breast cancer. [Figure 101]Figure 101 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma concentrations of IL10 in an in vivo model of breast cancer. [Figure 102] Figure 102 is a graph of the effect of a nutritional supplement containing fish oil and selenium in combination with a chemotherapeutic agent on plasma concentrations of IFN-γ in an in vivo model of breast cancer. [Figure 103] Figure 103 is a schematic diagram of AXL signaling and factors that influence tumor progression. [Figure 104] Figure 104 is a Western blot of AXL and p-MET expression in HCC827 and HCC827GR (Iressa-resistant) cells. [Figure 105] Figure 105 is a Western blot of the combined effect of selenium (Nutrient A) and fish oil (Nutrient B) on AXL expression in drug (Iressa)-resistant HCC827GR cells. GAPDH serves as a control. [Figure 106] Figure 106 is a schematic diagram of the pathway of AXL protein expression, providing various points for regulation of AXL expression. [Figure 107] Figure 107 is a Western blot of HSP90 and AXL expression in drug (Iressa) resistant HCC827GR cells and non-resistant parental HCC827 cells. GAPDH serves as a control. [Figure 108] Figure 108 is a Western blot showing the decrease in AXL and HSP90 expression in drug (Iressa) resistant tumor cells using a combination of selenium (Nutrient A) and fish oil (Nutrient B). [Figure 109] Figure 109 is a schematic diagram of the mTOR pathway. [Figure 110] Figure 110 is a Western blot showing a decrease in AXL, HSP90, and p-mTOR expression in drug (Iressa) resistant tumor cells using a combination of selenium (Nutrient A) and fish oil (Nutrient B). [Figure 111]Figure 111 is a Western blot showing the effect of treatment with a nutritional supplement containing fish oil and selenium, alone and in combination with a chemotherapy compound, on heat shock protein (HSP90) in tumor cells from an in vivo model of human cancer. [Figure 112] Figure 112 is a Western blot showing the effect of treatment with a nutritional supplement containing fish oil and selenium, alone and in combination with a chemotherapeutic compound, on p-AXL in tumor cells from an in vivo model of human cancer. [Figure 113] Figure 113 is a graph of two-year survival rates for head and neck cancer patients using a nutritional supplement containing fish oil and selenium and a generic prior art supplement. DETAILED DESCRIPTION OF THE INVENTION

[0075] The following description includes information that may be useful in understanding the present invention. No admission is made that any of the information presented herein is prior art or relevant to the present invention, or that any publication specifically or implicitly cited is prior art.

[0076] The subject of the present invention provides compositions and methods for using nutritional supplements (e.g., a supplement containing fish oil, selenium in the form of selenium yeast, chromium, and a specific plant-derived substance (NutraWell) and / or a supplement containing selenium and fish oil) in combination with chemotherapy.Unexpectedly, the combination therapy using chemotherapeutic agents and supplements provides significant synergistic effects in reducing tumor size and / or cell proliferation.In addition, the side effects of radiation therapy are alleviated, and the expression of genes related to apoptosis is regulated in tumor cells.Unexpectedly, the supplements of the present invention have also been found to increase the sensitivity of resistant cancer cells to chemotherapeutic agents.

[0077] It will be appreciated that the disclosed technology provides many advantageous technical effects, including enhancing the effectiveness of current chemotherapy protocols used in the treatment of cancer, and in the treatment of cancers that are resistant to chemotherapy, while reducing the side effects associated with these approaches.

[0078] The following discussion provides many example embodiments of the inventive subject matter. While each embodiment represents a single combination of inventive elements, it is understood that the inventive subject matter includes all possible combinations of the disclosed elements. Thus, if one embodiment includes elements A, B, and C and a second embodiment includes elements B and D, it is understood that the inventive subject matter also includes other remaining combinations of A, B, C, or D, even if not explicitly disclosed.

[0079] In some embodiments, numbers expressing properties such as amounts of ingredients, concentrations, and reaction conditions used to describe and claim certain embodiments of the present invention should be understood to be modified in some instances by the term "about." Accordingly, in some embodiments, the numerical parameters set forth in the specification and appended claims are approximations that may vary depending on the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the present invention are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as practicable. The numerical values ​​presented in some embodiments of the present invention may contain certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0080] While some of the findings described below are directed to the use of fish oil and selenium sources, Applicant noted that the nutritional supplement formulation presented in Table 1 (i.e., NutraWell) incorporates fish oil and selenium. The selenium is preferably provided as selenium yeast or components thereof (e.g., peptides and / or amino acids prepared from selenium yeast). Thus, the effects found in the fish oil and selenium yeast studies can be extended to the use of this nutritional supplement. NutraWell has been found to have a high level of tolerability and unexpected beneficial antitumor activity in combination with conventional therapy. As shown below, such nutritional supplements also demonstrate significant beneficial effects when used in combination with one or more chemotherapeutic agents.

[0081] [Table 1-1]

[0082] [Table 1-2]

[0083] [Table 1-3]

[0084] The compositions shown in Table 1 contain, in addition to selenium and fish oil, ingredients with various physiological and biochemical effects, including anti-inflammatory activity, blood glucose lowering, cholesterol lowering, and anti-tumor activity. In a preferred embodiment, the nutritional supplement contains at least three ingredients shown in Table 1. Such additional ingredients provide supplementation of essential vitamins, minerals, and amino acids at high levels. Other ingredients (e.g., enzymes, lecithin) function to aid in the digestion and absorption of the composition components when ingested. The combination of these complementary activities provides a synergistic effect beyond the simple additive effects of the individual components. It should be understood that the compositions shown in Table 1 also contain certain flavoring agents (e.g., brown sugar, honey, vanilla flavor, and masking agents) that function to improve palatability and acceptability. Certain ingredients (e.g., honey, brown sugar, milk, rice protein, casein) can provide both flavor and caloric energy. The inventors have found that combinations of the aforementioned flavoring agents are effective in providing compliance with the consumption of the nutritional supplement in effective amounts. In some embodiments, such flavoring agents can be omitted without adversely affecting the effectiveness of the nutritional supplement.

[0085] The ingredients shown in Table 1 can be provided as a single formulation (e.g., as a pill, tablet, capsule, powder, liquid, suspension, etc.) or can be divided into different formulations (e.g., as a pill, tablet, capsule, powder, liquid, suspension, or combinations thereof). The amounts shown in Table 1 are exemplary and are typical daily dosages provided to an adult of normal height and otherwise normal health. These amounts can be adjusted to account for differences in weight, gender, medical condition, etc. For example, a relatively small patient weighing 40 kilograms or less may benefit from a dosage provided at or below the lower end of the provided range, while a relatively large patient weighing 100 kilograms or more may require a dosage provided at the upper end (or higher) of the indicated range. In some embodiments, such a daily amount can be distributed as multiple doses throughout the day. In some such embodiments, the composition of each such distributed dose can be identical. In other embodiments, the composition of such distributed doses can vary, provided that the sum of such doses provides the required supplementation.

[0086] Tumor cell proliferation Surprisingly, the present inventors have found that treating various cancer cells with a combination of fish oil and selenium yeast has a direct effect on cell proliferation, especially when used in combination with cancer chemotherapy. Examples of the effects of combined treatment of tumor cells with chemotherapy and a nutritional supplement containing fish oil and selenium peptides can be seen in Figures 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. These figures show the effects of treatment of various cancer cell lines with chemotherapy and fish oil supplements, selenium peptide supplements, and supplements created by combining fish oil and the selenium peptides. Figures 1, 2, and 3 show the effects of treatment with Iressa in combination with PBS or various supplements on the proliferation of A549 lung tumor cells. As shown, the use of a supplement containing both fish oil and selenium reduces A549 cell proliferation, and the combination also provides an unexpected synergistic effect in reducing A549 cell proliferation when used in combination with Iressa. Similar effects are seen in Figures 4 and 5 when such a nutritional supplement is used in combination with Tarceva to treat A549 cancer cells. The effect of a nutritional supplement containing both fish oil and selenium in combination with Alimta on the proliferation of A549 cancer cells is shown in Figure 6, and the effect of such a nutritional supplement in combination with cisplatin on the proliferation of A549 cancer cells is shown in Figure 7. The synergistic effect of the combination of a nutritional supplement containing fish oil and selenium and a chemotherapeutic agent is noticeable in many instances, especially at lower drug concentrations.

[0087] The inventors surprisingly found that combination therapy using a chemotherapeutic agent and a fish oil / selenium peptide supplement also provided similar effects in an unrelated triple-negative breast cancer cell line (MDA-MB-231). Figure 8 shows typical results on cell proliferation after 72 hours of combined treatment of these cells with docetaxel and a fish oil / selenium supplement. While both the chemotherapeutic agent and the supplement containing fish oil and selenium reduced proliferation, their combined use produced a surprising synergistic effect. Similar effects were observed when MDA-MB-231 cells were treated with a nutritional supplement containing fish oil, selenium, a combination of fish oil and selenium, and cisplatin. Surprisingly, a synergistic effect in reducing proliferation was observed when both the selenium-containing supplement and the selenium / fish oil supplement were used in combination with cisplatin.

[0088] The effect of combining a nutritional supplement containing fish oil and selenium with a chemotherapy drug was also identified in an in vivo study, in which the Ki-61 proliferation marker was identified in mammary tumors of a mouse model by immunocytochemistry. A typical result of such a study is shown in Figure 11, where Ki-61 cells are indicated by arrows. As shown, treatment with the fish oil / selenium supplement ("N") reduced the number of positive cells (indicating a reduction in the proliferation index), similar to treatment with chemotherapy drugs (taxol, adriamycin). Treatment with a combination of both the nutritional supplement and the chemotherapy drug resulted in an even greater reduction in the proliferation index.

[0089] Overall, the inventors have found that a nutritional supplement containing fish oil and selenium (e.g., in the form of selenium yeast or yeast products) can reduce proliferation in a variety of tumor cell types and can provide synergistic effects when used in combination with a variety of different chemotherapeutic agents, effects seen both in cell culture and in in vivo animal models of human disease.

[0090] Tumor size The present inventors have also found that a nutritional supplement containing fish oil and selenium reduces tumor size in an in vivo animal model of human disease, and surprisingly, does so synergistically when used in combination with a chemotherapeutic drug. This article provides an example of a typical testing protocol for such a nutritional supplement ("Nutrawell") and the chemotherapy drug Tarceva. Figure 12 shows the initial body weights of mice used in such studies, which can be used to calculate tumor weight. Figure 13 shows the results of the study, tumor weight (top panel) and tumor weight distribution (bottom panel). As shown, 28 days after tumor implantation, mice were sacrificed and the tumors were characterized. All untreated mice developed large (>5g) tumors. Mice treated with either Tarceva or NutraWell still exhibited a significant number of such large tumors. However, mice treated with a combination of Tarceva and a fish oil / selenium nutritional supplement exhibited no large tumors and had a significant proportion of small (<3g) tumors.

[0091] Similar results were seen in mice implanted with cells derived from human breast cancer and treated with a fish oil / selenium nutritional supplement in combination with chemotherapy drugs commonly used to treat breast cancer (e.g., Taxol, Adriamycin, Avastin, etc.). Figure 14 shows an exemplary treatment protocol (top panel) and the results of a tumor volume study (bottom panel) performed on control (untreated) mice, mice treated with a fish oil / selenium nutritional supplement, and mice treated with various chemotherapy drugs with or without combined treatment with a fish oil / selenium nutritional supplement. Tumor volumes were recorded at two different time periods to demonstrate the effect on growth. Untreated (control) mice grew in tumor volume by approximately 120 mm over a 4-day period. 3As shown, the chemotherapeutic drugs Taxol, Adiamycin, and / or Avastin used in combination with the fish oil / selenium nutritional supplement exhibit a significantly reduced tumor growth rate compared to the use of these chemotherapeutic drugs alone. It should also be understood that the fish oil / selenium nutritional supplement alone reduced tumor growth.

[0092] Surprisingly, combination therapy with a fish oil / selenium nutritional supplement was found to synergistically enhance the effects of the chemotherapy agents. As shown in Figure 15, tumor volume in implanted mice (top panel) was not significantly affected by treatment with Alimta and cisplatin, indicating resistance to this combination chemotherapy. The addition of Tarceva to this chemotherapy regimen resulted in only a slight improvement in terms of tumor volume reduction. However, when Alimta and cisplatin were combined with the fish oil / selenium nutritional, a significant reduction was observed, and an even greater reduction was seen when NutraWell was used in combination with all three chemotherapy agents. Similar results were seen for tumor weight (bottom panel).

[0093] The effect of combined treatment with a nutritional supplement containing fish oil and selenium and the chemotherapy drug Iressa on tumor weight in mice implanted with lung cancer cells can be seen in Figure 16. As shown, treatment with Iressa alone only slightly reduced tumor weight (top left panel), while combined treatment with Iressa and a nutritional supplement containing fish oil and selenium reduced tumor weight by more than 50% compared to untreated control mice. The weight distribution (top right panel) also shows a slight reduction in tumor weight upon treatment with the chemotherapy drug (most of the mice have tumors >6 grams), but combined treatment with a nutritional supplement containing fish oil and selenium resulted in all tumors weighing less than 6 grams, and many weighing less than 3 grams. Representative photographs of tumors obtained from mice in the control and treatment groups are shown in the bottom panel of Figure 16.

[0094] Further in vivo studies were carried out using the protocol shown in Table 2.

[0095] [Table 2]

[0096] Figures 17 and 18 show representative data for tumor weight and volume (respectively) in test groups of mice implanted with breast cancer cells, as described in Table 2. As shown, the addition of a nutritional supplement containing fish oil and selenium to a treatment regime utilizing taxol or adriamycin resulted in a significant reduction in tumor weight compared to the use of these chemotherapeutic agents alone. Similar studies were performed to determine the effect of a nutritional supplement of the present invention in combination with Avastin or taxol. A nutritional supplement containing fish oil and selenium was provided to mice implanted with tumor cells at 0.4 g twice daily, and Avastin or taxol was provided intraperitoneally at 5 mg / kg every four days. The effect on tumor volume in mice treated with a nutritional supplement containing fish oil and selenium and Avastin can be seen in Figure 19. The effect on tumor volume in mice treated with a nutritional supplement containing fish oil and selenium and taxol can be seen in Figure 20. Improvement in tumor volume reduction is seen with combined treatment with a nutritional supplement containing both Avastin and taxol. Similar results are shown in FIG. 21, showing the effect of combined treatment with a nutritional supplement containing fish oil and selenium and taxol, adriamycin, or avastin on tumor volume (left panel) and tumor weight (right panel).

[0097] To determine whether the results of in vivo studies were influenced by the general nutritional status of the animal subjects receiving nutritional supplements, we compared the results of combined treatment with a nutritional supplement containing fish oil and selenium ("TNuF") and equivalent calorie supplementation with a commercially available nutritional supplement (Prosure™, "P"). As shown in Figure 22 (top panel), combined treatment with a chemotherapeutic agent and Prosure™ resulted in a slight reduction in apparent tumor size compared to treatment with the chemotherapeutic agent alone. However, combined treatment with a nutritional supplement containing fish oil and selenium and the chemotherapeutic agent reduced the apparent tumor size by approximately 50% compared to subjects treated with the chemotherapeutic agent alone. A similar effect was seen in tumor weight distribution (see 20180449S4, bottom panel). It is clear that the improvements in reducing tumor size and weight are not solely the result of improvements in nutritional status.

[0098] Neovascularization / inhibition of angiogenesis In many cases, the growth of solid tumors is promoted by angiogenesis and / or neovascularization, leading to the development of blood vessels within the tumor mass.The present inventors have also found that a nutritional supplement containing fish oil and selenium can reduce angiogenesis / neovascularization in tumors, both when used alone and when combined with chemotherapy drugs.Unexpectedly, the present inventors have also noted that some chemotherapy drugs alone reduce angiogenesis / neovascularization.The present inventors believe that such a reduction in neovascularization induces oxidative stress in tumors and / or makes tumors treated in this way more sensitive to oxidative stress.

[0099] The present inventors have demonstrated that the use of a nutritional supplement containing fish oil and selenium in combination with chemotherapeutic agents (e.g., Taxol, Adriamycin, Avastin) can improve in vivo breast cancer survival. We found that the compound has a direct and observable effect on tumor angiogenesis in a mouse model. In an exemplary study, mice were treated as shown in the diagram in Figure 23. Figure 24 provides a photograph of a typical tumor and associated blood vessels in a mouse from an untreated control group. Several relatively large-diameter blood vessels can be seen associated with the tumor. Figure 25 provides a similar photograph taken of a mouse from a group treated with the nutritional supplement alone. The number of tumor-associated blood vessels, as well as their diameter, are reduced compared to those in the untreated control group. Figures 26 and 27 provide photographs of typical tumors taken from mice treated with taxol and taxol in combination with a nutritional supplement containing fish oil and selenium ("N"), respectively. Surprisingly, treatment with taxol alone reduced the number of tumor-associated blood vessels and their diameter, which is not a known effect of treatment with this chemotherapy drug (see Figure 26). As shown in Figure 27, combined treatment with taxol and a nutritional supplement containing fish oil and selenium resulted in a dramatic reduction in tumor-associated blood vessels. Similar results were seen in the study shown in FIG.

[0100] Figures 29 and 30 provide photographs of typical tumors taken from mice in groups treated with adriamycin and adriamycin in combination with a nutritional supplement containing fish oil and selenium, respectively, in a similar study. Similar results were seen in a separate but similar study on the effects of a nutritional supplement containing fish oil and selenium in combination with either adriamycin or Avastin, as shown in Figure 31. In all cases, treatment with a nutritional supplement containing fish oil and selenium significantly reduced the tumor angiogenesis and tumor volume compared to similar treatments without the nutritional supplement. It is clear that nutritional supplements can reduce the angiogenesis of solid tumors and can enhance the reduction in solid tumor angiogenesis seen with treatment with conventional chemotherapeutic agents.

[0101] The effects of treatment with a fish oil and selenium-containing supplement and chemotherapeutic agents on angiogenesis / neovascularization can also be observed at the molecular level. VEGF and CD31 are markers associated with tumor angiogenesis via the HIF-α pathway. Figure 32 shows Western blots of HIF-α and VEGF in tumors treated with a fish oil and selenium-containing nutritional supplement ("N"), taxol, adriamycin, Avastin, and nutritional supplements in combination with these chemotherapeutic agents. As shown, the nutritional supplements reduce HIF-α levels, while the chemotherapeutic agents have relatively little effect. A similar reduction is seen in VEGF for both the fish oil and selenium-containing nutritional supplement and, to a lesser extent, taxol and adriamycin. In all cases, the combination of the nutritional supplement and chemotherapeutic agents results in a dramatic reduction in HIF-α and VEGF. Figure 33 shows the results of a similar study characterizing CD31. A nutritional supplement containing fish oil and selenium resulted in a dramatic decrease in CD31, and a similar decrease was seen after treatment with Adriamycin and Avastin. An even greater decrease in CD31 was seen after combined therapy with the nutritional supplement and the chemotherapeutic agent.

[0102] Effect on cell cycle phase distribution The inventors have found that combination therapy with a nutritional supplement containing fish oil and selenium and chemotherapy drugs also significantly affects the cell cycle phase distribution of cancer cells both in vivo and in vitro. As shown in Figure 34, A549 lung cancer cells show a slight shift to sub-GI when treated with a supplement containing fish oil and selenium, and a moderate shift to sub-GI when treated with Alimta. Combined treatment with the nutritional supplement and Alimta shows a larger additive shift to sub-GI, indicating a synergistic effect. Treatment with Tarceva alone is significantly higher than untreated control cells. Although it has little effect on cell cycle distribution compared to selenium, when used in combination with a nutritional supplement containing fish oil and selenium, a significant shift to sub-GI is observed. Results of a similar study using the chemotherapy drug Tarceva are shown in 8FIG3B. The results are summarized in Table 3.

[0103] [Table 3]

[0104] It is clear that the treatment of fish oil and selenium supplements in combination with various chemotherapeutic drugs can shift the cell cycle distribution of cancer cells to sub-GI phase.It should be understood that sub-GI phase is related to apoptosis, and therefore the combined treatment of chemotherapeutic drugs and fish oil and selenium supplements can be useful in inducing apoptosis and / or apoptotic events in cancer cells.

[0105] Apoptosis / autophagy As described above, cell cycle phase distribution studies suggest that combination therapy using a supplement containing fish oil and selenium in combination with a chemotherapeutic drug can induce apoptosis and / or apoptotic events in cancer cells. The present inventors have found that a nutritional supplement containing fish oil and selenium is effective in enhancing apoptosis in tumors in an in vivo model of breast cancer. For example, Figure 35 shows the effect of such a nutritional supplement, the chemotherapeutic agents Taxol, Avastin, or Adriamycin, and the use of these chemotherapeutic agents in combination with the nutritional supplement on the expression of apoptosis markers (specifically, VEGF, p53, and HIF-α) in advanced tumors in mice injected with breast cancer cells. As shown, the use of the nutritional supplement increases the expression of p53 while decreasing the expression of VEGF and HIF-α. This effect is observed when the nutritional supplement is used alone or in combination with a chemotherapeutic drug, with the combination providing an enhanced effect.

[0106] The results of a similar study targeting Bcl-2, p-Bcl-2, and caspase 3 are shown in Figure 36, and the results of a similar study targeting caspase 8 are shown in Figure 37. As shown, the use of a nutritional supplement containing fish oil and selenium, both alone and in combination with chemotherapy drugs, reduces Bcl-2 expression and increases p-Bcl-2 and caspase 3 expression. The use of a nutritional supplement containing fish oil and selenium, both alone and in combination with chemotherapy drugs, reduces caspase 8 expression. Figure 38 shows the effects of a nutritional supplement containing fish oil and selenium ("N"), Taxol, Adriamycin, and Avastin on Bcl-2 expression and caspase 3 expression. Figure 1 shows the effects of treatment with selenium and combination treatments on cytochrome C. As shown, treatment with the supplements results in a slight increase in cytochrome C, while little or no increase is seen with treatment with taxol or adriamycin. These effects are synergistically enhanced in some cases when a nutritional supplement containing fish oil and selenium is used in combination with a chemotherapy drug.

[0107] In addition to biochemical markers of apoptosis, apoptotic effects can be directly observed. For example, as shown in Figure 39, the growth of A549 cells in culture is slightly affected by exposure to a selenium-containing supplement or a fish oil-containing supplement, but dramatically affected by a supplement containing both fish oil and selenium. Similarly, as shown in Figure 40, trypan blue staining of A549 cells in culture shows only a slight effect in inducing cell death when exposed to a selenium-containing supplement or a fish oil-containing supplement, but a dramatic increase in cell death when exposed to a supplement containing both fish oil and selenium. While either selenium or fish oil has some effect, their combined effects are clearly greater than additive (i.e., synergistic).

[0108] Oxidative stress / hypoxia The inventors have found that nutritional supplements of the present invention are effective in enhancing tumor oxidative stress (e.g., inhibiting the tumor's ability to respond to oxidative stress) in an in vivo model of breast cancer. For example, Figure 41 shows the effects of a nutritional supplement containing fish oil and selenium, the chemotherapeutic agents Taxol or Adriamycin, and the use of these chemotherapeutic agents in combination with the nutritional supplement on the levels of malondialdehyde (MDA) in tumors of mice injected with breast cancer cells. As shown, the use of the nutritional supplement (either alone or in combination with the chemotherapeutic agent) is effective in increasing the relative concentration of MDA, indicating enhanced oxidative stress in the tumor. A similar effect is seen with Avastin (see Figure 42). A synergistic effect is also seen (e.g., when the nutritional supplement is used in combination with Adriamycin). As shown in Figure 32, this is consistent with the results for HIF-α, a marker of hypoxia. It should be understood that such an effect is complementary to the reduction in neovascularization discussed above and shown in Figures 24 and 25.

[0109] Sphere / Stem Cell Properties The present inventor has also found that nutritional supplements containing fish oil and selenium, combined with chemotherapy drugs, can be effective in reducing the stem cell properties of cancer cells.The occurrence of cancer cells that exhibit such properties is thought to be involved in both metastasis and the occurrence of resistance to chemotherapy drugs.It should be understood that the sphere cells that form in cell culture at least partially replicate the occurrence of cancer stem cells in vivo.

[0110] As shown in Figure 43, A549 sphere cells are relatively resistant to treatment with Iressa, but show reduced proliferation in the presence of a supplement containing fish oil and selenium. When Iressa and the supplement are used in combination, the effect is enhanced and may be synergistic (e.g., at higher Iressa concentrations). A549 lung cancer sphere cells are similarly resistant to Tarceva, and a synergistic reduction in proliferation is observed when the cells are treated with a combination of Tarceva (e.g., 10 μM) and a supplement containing fish oil and selenium (see Figure 44).

[0111] Similar effects are observed in MDA-MB-231 breast cancer sphere cells. As shown in Figure 45, these cells exhibit a small decrease in proliferation when exposed to a supplement containing fish oil and selenium, and are moderately sensitive to docetaxel, while the cells are not sensitive to docetaxel. When cells and the supplements are used in combination, a synergistic effect is observed, resulting in a significant reduction in proliferation. MDA-MB-231 breast cancer sphere cells are relatively resistant to both cisplatin and a supplement containing fish oil and selenium, but when cisplatin and such a supplement are used in combination, a significant reduction in proliferation is observed (see Figure 46).

[0112] The effects of cisplatin, selenium, fish oil, a combination of selenium and fish oil, and supplements combined with cisplatin on cell cycle phase distribution are shown in Figure 3C. As shown, selenium and fish oil alone produce a very small shift toward sub-GI, while the combination of selenium and fish oil produces a relatively large shift. Cisplatin also produces a small shift toward sub-GI, which is enhanced when used in combination with a supplement containing fish oil and selenium. The results are listed in Table 4.

[0113] [Table 4]

[0114] Similar results are seen for A549 lung cancer sphere cells treated with Alimta, Tarceva, and these chemotherapy agents in combination with a supplement containing fish oil and selenium, as shown in Figure 47. Results from these studies are summarized in Table 5.

[0115] [Table 5]

[0116] It should be understood that sub-Gl is related to apoptosis.Therefore, the present inventor believes that the use of a supplement containing fish oil and selenium in combination with chemotherapeutic agents can be effective in inducing apoptosis in cancer cells or stem cells, even if such cells are resistant to said chemotherapeutic agents.

[0117] The inventors have also combined a supplement containing fish oil and selenium with chemotherapy drugs. We also found that the use of IL-16 can alter the expression of biochemical markers associated with cancer spheres or stem cells, matching the levels associated with non-stem cancer cells. As shown in Figure 48, the expression of various biochemical markers (particularly CD133, C-myc, IL-6, and CD44) differs between parental A549 lung cancer cells and sphere cells derived from the same cell line. Figure 49 shows the expression levels of stem cell-associated markers (i.e., "stemness") in such A549 sphere cells, particularly E-cadherin (an epithelial marker) and vimentin (a mesenchymal marker), compared to parental A549 cells.

[0118] As shown in Figure 50, the use of a supplement containing fish oil and selenium alters the levels of various biochemical markers in A549 sphere cells, particularly p-AMPKα, p-mTOR, and vimentin. It should be noted that p-mTOR and vimentin are elevated in A549 sphere cells and decreased to levels similar to those of parental A549 cells upon exposure to a supplement containing both fish oil and selenium, indicating a decrease in stem cell characteristics. The results of a similar study conducted on A549 sphere cells are shown in Figure 51, which shows an increase in LC3-I and LC3II upon exposure to a supplement containing both fish oil and selenium. A similar study also shows a decrease in GRP78, N-cadherin, and β-catenin (see Figure 52).

[0119] While the aforementioned study focused on A549 sphere cells, similar results were obtained from in vivo studies of animal models of human breast cancer. Figure 53 shows the results of a study characterizing CD29 and CD24 expression in tumors from animals treated with a nutritional supplement containing fish oil and selenium ("N"), taxol, adriamycin, Avastin, and these chemotherapeutic agents in combination with the nutritional supplement. As shown, treatment with the nutritional supplement containing fish oil and selenium resulted in a modest reduction in CD29 expression, while treatment with the chemotherapeutic agents had little effect. However, combination therapy was found to potently reduce CD29 levels, indicating that such a combination provides a synergistic effect in reducing tumor stem cell characteristics in vivo. While CD24 expression was more responsive to treatment with the nutritional supplement containing fish oil and selenium, a synergistic effect in reducing CD24 levels was evident when used in combination with chemotherapeutic agents (particularly Avastin).

[0120] Overall, the inventors have found that the use of a nutritional supplement comprising fish oil and selenium can reduce stem cell properties and / or "stemness" in cancer cells, and can synergistically reduce them when used in combination with chemotherapeutic drugs.

[0121] Reversal of drug resistance As mentioned above, the development of stem cell characteristics (e.g., "stemness") in cancer cells is associated with the development of drug resistance. Surprisingly, the present inventors have found that a nutritional supplement containing fish oil and selenium is effective in sensitizing drug-resistant cancer cells to chemotherapeutic drugs to which they are resistant. For example, HCC827Gr cells are human lung cancer-derived cells that have developed resistance to chemotherapeutic drugs such as Iressa. The results of exposing such cells in culture to Iressa in the presence and absence of selenium and fish oil supplements are shown in Figure 54. As shown in Figure 54, exposure to Iressa does not significantly reduce the number of viable, resistant HCC827Gr cells, but when Iressa is used in combination with selenium and fish oil supplements, a significant reduction in the number of viable cells is observed.

[0122] This sensitizing effect is also found when characterizing cell proliferation. As shown in Figure 55, the parental HCC827 lung cancer cell line (left panel) is sensitive to Iressa, with proliferation reduced by more than 80% at 0.125 μM Iressa. In contrast, drug-resistant HCC827Gr lung cancer cells (right panel) maintain proliferation at nearly 80% of control values, even in the presence of 1 μM Iressa. The drug-sensitive parental cell line and the drug-resistant cell line continue to proliferate. Differences between the drug-sensitive parental cell line and the drug-resistant cell line are also evident in the expression of several biochemical markers. As shown in Figure 56, differences in gene expression (shown as mRNA) of various stem cell, metabolic, and epithelial-mesenchymal transition (EMT) genes are evident between the sensitive parental HCC827 cells and the resistant HCC287Gr cells.

[0123] Figures 57 and 58 show the effects of combined treatment of resistant HCC827Gr cells with Iressa together with selenium, fish oil, or selenium and fish oil-containing supplements. As shown, in the absence of supplementation, Iressa results in a maximum reduction of only 20% in cell proliferation. Treatment with a supplement containing both fish oil and selenium results in a moderate reduction in proliferation (approximately 50%). However, treatment with a supplement containing both selenium and fish oil reduces cell proliferation to a level similar to that seen in the sensitive HCC827 parent cell line (i.e., approximately 20% of the untreated control). It is clear that the use of a supplement containing fish oil and selenium is effective in sensitizing drug-resistant cancer cells to chemotherapy drugs, including drugs to which they are resistant.

[0124] As shown in Figures 59 and 60, a similar increase in colony formation reduction was observed upon treatment with a supplement containing selenium and fish oil. Significant colony formation by HCC827Gr cells was observed in the presence of 0.5 μM Iressa, but was significantly reduced in cells treated with a combination of selenium and fish oil. In suppressing colony formation by these resistant cancer cells, a synergistic effect between Iressa (which does not prevent increased colony formation by resistant HCC827Gr cells) and the selenium and fish oil supplement is evident, as colony formation was reduced to a level lower than that of cells treated with the selenium and fish oil supplement alone.

[0125] The inventors also found that treatment of such resistant cancer cells with fish oil and selenium yeast altered the cell cycle phase distribution of these cells, as shown in Figure 61. The numerical results are summarized in Table 6. As shown, treatment with Iressa alone had little or no effect on the cell cycle phase distribution of resistant HCC827GR cells. However, treatment of such cells with a combination of fish oil and selenium yeast strongly shifted the cell cycle distribution to the S and SubG1 phases. Similar results were observed with combined treatment with Iressa and the nutritional supplement, with a significant increase in the percentage of cells in SubG1 and a concomitant decrease in cells in the G2 / M phase. This indicates that a nutritional supplement containing fish oil and selenium yeast has an unexpected synergistic effect on cell cycle distribution in such chemotherapy-resistant cells.

[0126] [Table 6]

[0127] The present inventors have surprisingly found that pretreatment of chemotherapy-resistant cells (e.g., HCC827GR) with a nutritional supplement containing fish oil and selenium yeast can have the effect of sensitizing these resistant cells to subsequent exposure to the chemotherapeutic agent to which they are resistant. XFIG4 was detected by pretreatment of the cells with fish oil and selenium yeast for 72 hours prior to exposing them to Iressa. 1 shows the results of pre-treating Iressa-resistant HCC827GR cells with a nutritional supplement containing 100mg Iressa-resistant HCC827GR cells. As shown, such pre-treated cells show a significant decrease in cell proliferation compared to cells pre-treated with PBS.

[0128] Such pretreatment with a nutritional supplement containing fish oil and selenium yeast also affects cell cycle phase distribution compared to untreated cells, as shown in Figure 62. The numerical results of such studies are summarized in Table 7.

[0129] [Table 7]

[0130] As shown, pretreatment with a nutritional supplement containing fish oil and selenium yeast shifts the cell cycle phase distribution of these Iressa-resistant cells to a significantly higher percentage of Sub-G1 and S-phase cells. Application of Iressa to such pretreated cells provides an even higher percentage of cells in the Sub-G1 phase. It should be understood that the sub-G1 phase is associated with apoptosis.

[0131] Reduced progression / metastasis The present inventors have found that supplements containing fish oil and selenium can effectively reduce tumor metastasis and / or progression, especially when used in combination with chemotherapy. For example, the present inventors have found that using a nutritional supplement containing a combination of fish oil and selenium in conjunction with chemotherapy can reduce metastasis from the primary tumor site. As shown in Figure 63, untreated mice exhibit extensive metastasis of tumor cells from the lung cancer cell implantation site. The metastatic sites are indicated by arrows in the upper panel of Figure 63, and the findings are listed in the lower panel. This is reduced to some extent by treatment with Tarceva ("TT") or the fish oil and selenium supplement ("TN"), but the extent of lymph node metastasis is most pronounced using the combination of the two ("TTN"). Without wishing to be bound by theory, the present inventors believe this may be due to sensitization of cancer stem cells to the effects of Tarceva by the fish oil and selenium supplement and / or by a reduction in stem cell properties that promote metastasis. This reduction in metastasis can also be seen at the microscopic level in the reduction in the number of cells identified by immunohistochemistry as expressing EGFR and VEGF, characteristics of implanted tumor cells. As shown in Figure 64, the number of such cells identified in lung tissue sections from mice treated with both a fish oil and selenium supplement and a chemotherapeutic agent is dramatically reduced. Similar results were observed for the use of a fish oil and selenium nutritional supplement in reducing metastatic tumor burden in the mammary glands of test animals implanted with breast cancer cells, as shown in Figure 65. As shown, the use of the fish oil and selenium nutritional supplement (N) or taxol resulted in a moderate reduction in mammary tumor burden, while a dramatic reduction was observed when taxol and the supplement were used in combination. Adriamycin was found to be more effective than taxol in reducing metastatic tumor burden, but combination therapy with a fish oil and selenium nutritional supplement resulted in an even greater reduction in metastasis.

[0132] A nutritional supplement containing fish oil and selenium was used in combination with multiple chemotherapy regimens. A reduction in metastasis was also observed when using a nutritional supplement containing fish oil and selenium and a chemotherapy drug. Combination therapy using a nutritional supplement containing fish oil and selenium and a chemotherapy drug can also reduce the incidence of metastasis compared to the use of chemotherapy drugs alone. As shown in Figure 66, lung metastasis occurred in all tumor-bearing mice that did not receive treatment, and liver metastasis occurred in 40% of such mice. This was reduced in tumor-bearing mice receiving combination chemotherapy using multiple drugs (AC = Adriamycin / Cisplatin, Tarc = Tarceva), but lung metastasis still occurred in 80% of such mice. However, combination therapy using a nutritional supplement containing fish oil and selenium ("Nutra") reduced the incidence of lung metastasis by 40% compared to untreated control mice.

[0133] In addition to observing a decrease in metastatic tumors and tumor cells, a decrease in biochemical markers associated with tumor metastasis and progression was also observed with treatment using a nutritional supplement containing fish oil and selenium, which was enhanced by combined therapy with chemotherapy. As shown in Figure 67, use of the nutritional supplement significantly reduced the expression of MMP-9, a tumor-associated protein associated with tumor progression in breast cancer cells. This is particularly seen when used in combination with chemotherapy (e.g., taxol ("tax"), adriamycin ("adyri"), and avastin). Loss of MMP-9 is associated with a decrease in tumor malignancy.

[0134] Surprisingly, the inventors have also found that the use of a nutritional supplement containing fish oil and selenium can reduce the number of circulating cancer cells in human patients. Data from a patient treated with a nutritional supplement containing fish oil and selenium for 16 weeks is shown in Figure 68. The top panel shows results from week 1, with a circulating tumor cell "CTC" count of 187. The bottom panel shows results from week 16, with a CTC count of 9 (essentially background). The inventors believe that treatment with a nutritional supplement containing fish oil and selenium can provide a dramatic reduction in circulating tumor cells, e.g., reduce apparent CTC counts to essentially background or normal levels (or below), thereby reducing the chance and / or extent of metastasis. The inventors believe that this effect can be enhanced by combination therapy with one or more chemotherapeutic agents.

[0135] Immune checkpoint / immunotherapy While chemotherapy agents have long been used to treat cancer, immunological approaches have recently been investigated. Often, such approaches target the modulation of immune checkpoint molecules, providing activation of components of the immune system and / or inhibition of components that reduce the immune response to tumor cells. Surprisingly, the present inventors have found that a nutritional supplement containing fish oil and selenium can modify the expression of such immune checkpoint molecules. For example, as shown in Figure 69, the present inventors have found that a nutritional supplement containing fish oil and selenium is effective in reducing immune checkpoint proteins (e.g., CD279, CD274), and can do so when used in combination with conventional chemotherapy drugs (e.g., taxol ("tax"), adriamycin ("adyri"), and avastin).

[0136] In animal models treated with a nutritional supplement containing fish oil and selenium, either alone or in combination with various chemotherapeutic agents (taxol, adriamycin, avastin), the modulation of PD-L1 in tumor cells and PD-1 in T cells is shown in Figure 70 for lung cancer cells and in Figure 71 for breast cancer. As shown, tumor-associated PD-L1 is reduced to some extent by treatment with the nutritional supplement, but each of the chemotherapeutic agents is dramatically reduced by their use in combination (showing a synergistic effect). PD-1 expression in T cells in the same animals can also be modulated by treatment with a supplement containing fish oil and selenium, an effect that is comparable to the combined use of chemotherapeutic agents. The results of a similar study using a nutritional supplement containing fish oil and selenium ("N") in combination with Iressa ("I") or Tarceva ("T") are shown in Figure 72. PD-1 expression in T cells (left panel) was only slightly reduced by treatment with Iressa or Tarceva (TI and TT, respectively) but returned to near-normal levels by combination therapy with the nutritional supplements (TIN and TTN, respectively). Conversely, PD-L1 expression in tumor cells was essentially unaffected by chemotherapy alone but was reduced by combination therapy with a nutritional supplement containing fish oil and selenium. Overall, this suggests that a combination of a supplement containing fish oil and selenium, a chemotherapy agent, or such a supplement and one or more chemotherapy agents may enhance or improve the outcomes of immunotherapy in patients with cancer.

[0137] Clinical data also demonstrate improved immune function in cancer patients receiving nutritional supplements containing fish oil and selenium. For example, as shown in Figures 73 and 74, the CD4 / CD8 ratios in breast cancer and lymphoma patients so treated are improved compared to patients receiving a placebo.

[0138] Overall, the inventors believe this indicates that nutritional supplements containing fish oil and selenium can enhance immunotherapeutic approaches to cancer treatment in a complementary, additive, or synergistic manner. The inventors believe this indicates that nutritional supplements containing fish oil and selenium can enhance immunotherapeutic approaches to cancer treatment in a complementary, additive, or synergistic manner.

[0139] Weakness / cachexis Individuals with cancer often lose weight due to loss of adipose tissue and muscle wasting (cachexia). This weight loss can result in serious health problems in addition to those resulting directly from tumor growth and progression. The inventors have found that a nutritional supplement containing fish oil and selenium can reduce the symptoms of cachexia, especially when used in combination with one or more chemotherapeutic agents. Tumor-bearing mice also experience cachexia with or without treatment with chemotherapeutic agents. Using such an animal model, the inventors unexpectedly found that treatment with a nutritional supplement containing fish oil and selenium can reduce these effects. As shown in Figure 75, loss of white adipose tissue (WAT), brown adipose tissue (BAT), and muscle mass (gastrocnemius) is consistent with the cachexia observed in tumor-bearing mice and is only slightly reduced by chemotherapy with Tarceva. Treatment with a nutritional supplement containing fish oil and selenium ("Nutrawell") improves the retention of adipose tissue and muscle, regardless of whether or not concurrent treatment with the chemotherapy is also used.

[0140] As shown in Figure 76, tumor cells can express gene products (e.g., PTHrP and IL-6) that can lead to excessive cellular energy expenditure and wasting. Some of these gene products (e.g., IL-6) are also associated with inflammation. These processes are thought to be associated with the development of cachexia. As shown in Figure 77, treatment with chemotherapeutic agents such as Tarceva can actually increase the expression of such gene products. Surprisingly, treatment with a nutritional supplement containing fish oil and selenium ("Nutrawell") or chemotherapy-combined treatment reduces the expression of such genes. Such a reduction in excess energy expenditure can beneficially help reduce inflammation and wasting, and can also provide more energy for recovery and immune response against tumors.

[0141] In addition to the burden provided by the tumor, chemotherapy itself presents a variety of unpleasant and potentially dangerous These cachectic effects are associated with a number of potential adverse effects. Surprisingly, the present inventors have found that providing a nutritional supplement containing fish oil and selenium as a combination therapy with chemotherapy drugs can reduce or eliminate these cachectic effects. An example of a typical study utilizing human tumor cells implanted in mice and a nutritional supplement containing fish oil and selenium ("Nutrawell") is shown in Figure 78. As shown, some groups of mice were treated with multiple chemotherapy drugs. As shown in Figure 79, treatment with omega-3 fatty acids (e.g., Tω3, Tω6), such as those found in fish oil, inhibits the expression of proteins associated with autophagy (e.g., LC3B-I and / or LC3B-III) in tumor-bearing mice. Such a decrease in autophagy may be associated with the preservation of body weight and tissue in tumor-bearing mice, particularly during chemotherapy. As shown in Figures 80 and 81, treatment of tumor-bearing mice with chemotherapy drugs reduces adipose tissue weight beyond the effect of the tumor alone. However, the combination of such chemotherapy with a nutritional supplement containing fish oil and selenium ("Nutra") results in dramatic improvements in adipose tissue mass.

[0142] As shown in Figure 82, combination therapy with a nutritional supplement containing fish oil and selenium can also reduce muscle wasting (characterized by gastrocnemius muscle mass in this case). Similar data was obtained for both gastrocnemius muscle weight and total skeletal muscle weight (see Figure 83). As shown, combination therapy with a nutritional supplement containing fish oil and selenium ("N") and chemotherapy drugs (Alimta / Cisplatin or AC, Tarceva or T) results in improved muscle mass preservation compared to that seen with chemotherapy drug treatment without the addition of a supplement. Similar results were seen in evaluations of the effects of a nutritional supplement containing fish oil and selenium ("N") with different chemotherapy drugs on the weights of various organs, including the brain (Figure 84, left panel), liver (Figure 84, right panel), heart (Figure 85, left panel), spleen (Figure 85, right panel), lung (Figure 86, left panel), and metastasis-free mammary gland (Figure 86, right panel).

[0143] The reduction in cachexia symptoms associated with treatment using a nutritional supplement containing fish oil and selenium in combination with chemotherapy drugs is also evident at the molecular level. The inventors have found that the use of such nutritional supplements, whether used alone or in combination with chemotherapy drugs (e.g., taxol, adriamycin, avastin), is effective in preventing or reversing changes in the expression of specific biochemical markers associated with muscle weakness and inflammation. This is also evident at the molecular level, as shown in Figure 87. Figure 87 shows the expression of muscle-related proteins (UCP3, UCP2) found in mitochondria and inflammation-related cytokines (IL6) in the cytosol. Figure 88 shows the expression of another set of mitochondrial muscle markers, complex IV and complex III. The use of a nutritional supplement containing fish oil and selenium, whether used alone or in combination with taxol, adriamycin, or avastin, resulted in a significant increase in mitochondrial proteins. Conversely, inflammation in muscle tissue was reduced, as indicated by a decrease in IL-6, whether used alone or in combination with taxol, adriamycin, or avastin.

[0144] Treatment with a nutritional supplement containing fish oil and selenium, with or without combined chemotherapy, was also found to regulate the expression of biochemical markers related to adipose tissue and metabolism. As shown in Figure 89 (lower left panel), the expression of UCP-1 and ATGL is increased in tumor tissue (T) compared to non-tumor tissue (NT). This is somewhat reduced by treatment with taxol (TT) and treatment with a nutritional supplement containing fish oil and selenium (TN), but a significant (i.e., synergistic) decrease in expression is seen in the combined treatment (TTN). In Western blots, hormones along with UCP-1 Similar results are evident for sensitive lipase (HLS) and adipose tissue triglyceride lipase (ATGL) (Figure 89, right panel).

[0145] Surprisingly, treatment with a nutritional supplement containing fish oil and selenium was also found to reduce the expression of PTHrP in tumor-bearing animals (TN, see the upper left panel of Figure 89). This effect was synergistically enhanced when a chemotherapy drug (Taxol) (TT), which actually increases expression when used alone, was used in combination therapy (TTN). This indicates that supplements containing fish oil and selenium may be useful in treating mineral imbalances in cancer patients.

[0146] Improvement of cachexia in head and neck cancer patients has also been observed in clinical studies when using a nutritional supplement containing fish oil and selenium. As shown in Figure 90, such a nutritional supplement is effective in both maintaining weight (left panel) and, in some cases (e.g., with a BMI of <19), increasing weight (right panel). Similar results are seen in animal models, as shown in Figure 22 (left panel). It should be understood that these effects are not due to a simple increase in calorie intake, as treatment with a similar calorie content of conventional nutritional supplements did not produce this effect.

[0147] The presence of tumors and the use of chemotherapy are often associated with nausea and loss of appetite, which may contribute to cachexia. Surprisingly, the inventors have found that the use of a nutritional supplement containing fish oil and selenium can increase food intake in animal models, despite the additional calories provided by the supplement itself (see Figure 91).

[0148] Cytokine expression The presence of tumors is often associated with inflammation, a process mediated at least in part by cytokines. Cytokines may also be involved in other tumor-related processes, such as angiogenesis. Surprisingly, the inventors have discovered that a nutritional supplement containing fish oil and selenium can modulate the levels of various cytokines, particularly when used in combination with chemotherapy drugs. As shown in Figure 92, the use of such a nutritional supplement in combination with chemotherapy drugs (in this example, taxol, or "tax," and adriamycin, or "adria") in an in vivo model of breast cancer was found to reduce plasma VEGF concentrations, indicating that it can provide anti-inflammatory and anti-angiogenic effects. Similar results are seen with Avastin (see Figure 93). While the effect is seen with the supplement alone, combination therapy with such a supplement enhances the VEGF-lowering effect of the chemotherapy drug.

[0149] As shown in Figures 94 and 95 (left panels), a similar effect is seen in the reduction of TNF-α, which is associated with tumor angiogenesis, growth, and metastasis in in vivo breast cancer models. A similar effect is seen in the reduction of IL-1β (see also Figure 96, left panel), which is associated with tumor angiogenesis, growth, and metastasis. The effect of a nutritional supplement containing a combination of fish oil and selenium, both alone and in combination with chemotherapy, on plasma IL-1β in an in vivo model of breast cancer is shown in Figure 97. A similar study (see Figures 98 and 95, right panels) showed a reduction in plasma IL-6, which is associated with tumor angiogenesis, protection from tumor oxidative stress, and anti-apoptotic effects. Plasma concentrations of IL-10 were similarly reduced (in some cases, to undetectable levels) in such in vivo tumor models treated with a supplement containing fish oil and selenium, both alone and in combination with chemotherapy, despite elevated concentrations when such drugs were used alone (N) (Figure 96, right panel).

[0150] Conversely, the present inventors have demonstrated that the use of a nutritional supplement containing fish oil and selenium, both alone and We have found that the plasma concentrations of some cytokines can be increased both in combination with chemotherapy and in combination with chemotherapy drugs. For example, as shown in Figures 99 and 100 (right panels), the plasma concentration of IL2 is increased in an in vivo model of breast cancer. It should be understood that IL2 is considered an immunotherapeutic agent for the treatment of some cancers. Similar results are seen for IL10, another immune-activating cytokine, in an in vivo breast cancer model (see Figure 101). The plasma concentration of IFN-γ, another immune-activating cytokine, is also observed in such studies, as shown in Figures 102 and 100 (left panels).

[0151] Overall, the present inventor believes that the nutritional supplement comprising fish oil and selenium can be useful both as a monotherapy and in combination with one or more chemotherapeutic drugs to reduce the plasma concentration of cytokines associated with inflammation and tumor growth / progression.It should be understood that even if such chemotherapeutic drugs increase the plasma concentration of such cytokines when used in the absence of such supplements, such reduction can be brought about, and this reduction is enhanced (shows synergistic effect) by combined therapy.Similarly, the use of such supplements can increase the plasma concentration of immune-activating cytokines both as a monotherapy and in combination with one or more chemotherapeutic drugs.

[0152] AXL signaling As shown in Figure 103, AXL signaling in tumors is associated with a wide variety of downstream effects that promote tumor progression. As shown, AXL signaling within the tumor microenvironment can promote immunosuppression, the development of a cancer stem cell phenotype, resistance to various anticancer therapies (including anticancer drugs), tumor cell proliferation, resistance to apoptosis and / or autophagy, epithelial-to-mesenchymal transition, and metastasis. Thus, controlling AXL signaling (e.g., reverting the cells to a state more closely resembling that of normal cells) can provide a therapeutic target for the treatment of cancer, particularly drug-resistant cancers.

[0153] In some embodiments of the present invention, the present inventors have found that a nutritional supplement containing fish oil and selenium is particularly effective in modifying AXL signaling in cells, particularly in drug-resistant tumor cells. As shown in Figure 104, drug (Iressa)-resistant HCC827GR cells express high levels of both AXL and p-MET protein compared to sensitive parent HCC827 cells. Surprisingly, the present inventors have found that the combination of selenium yeast (nutrient A) and fish oil (nutrient B) dramatically reduces AXL expression in drug-resistant HCC827GR (see Figure 105). It should be noted that treatment with 1 μM Iressa is ineffective.

[0154] The inventors further investigated the mechanism by which the aforementioned nutritional supplements can regulate AXL protein concentration in cells. Figure 106 illustrates the pathway of AXL expression, various points of which can be targeted to regulate AXL expression. As shown, AXL depends on heat shock protein 90 (HSP90) for proper folding. Improper protein folding can lead to an increased rate of degradation. As shown in Figure 107, both AXL and HSP90 protein levels are significantly elevated in drug (Iressa)-resistant HCC827GR tumor cells compared to non-resistant parental HCC827 cells.

[0155] Surprisingly, the present inventors found that a supplement containing a combination of selenium yeast (nutrient A) and fish oil (nutrient B) reduced both AXL and HSP90 in drug (Iressa)-resistant HCC827GR cells (see Figure 108). A significant synergistic effect was also found when the combined nutritional supplement was used in combination with Iressa.

[0156] The mTOR pathway is also involved in drug resistance in tumor cells. In response to growth factors, mTOR can be phosphorylated, affecting cell growth and proliferation. Surprisingly, the present inventors have found that the use of a supplement that combines selenium yeast (nutrient A) and fish oil (nutrient B) can reduce the level of p-mTOR in drug (Iressa)-resistant HCC827GR cells both in the absence and presence of the drug (see Figure 110). Even more surprisingly, the combined nutritional supplement can reverse the elevated p-mTOR level in drug-resistant cells treated with chemotherapeutic agents.

[0157] The effects of nutritional supplements containing fish oil and selenium were also studied using an in vivo animal model of triple-negative human breast cancer, both alone and in combination with chemotherapy agents. The results are shown in Figures 111 and 112. Animals were treated with nutritional supplements containing fish oil and selenium (e.g., as prepared in Table 1), taxol ("tax"), adriamycin ("adyri"), avastin, or a combination of the nutritional supplements and individual chemotherapy agents. As shown in Figure 111, HSP90 expression (as determined by Western blot) in untreated tumor samples was high. Surprisingly, this was reduced to some extent by treatment with only the nutritional supplement containing fish oil and selenium. Treatment with taxol had minimal effect on HSP90 expression, but a significant reduction was observed when taxol was used in combination with the nutritional supplement containing fish oil and selenium. Similar results were seen with adriamycin treatment and adriamycin combination treatment with such supplements. Although HSP90 expression in such tumors is reduced to some extent by treatment with Avastin, combined treatment with Avastin and a nutritional supplement containing fish oil and selenium results in a dramatic reduction in HSP90 expression, demonstrating a synergistic effect.

[0158] Figure 112 shows the results of a similar study characterizing tumor p-AXL content. As shown, p-AXL is evident in untreated tumors and is surprisingly reduced following treatment with only a nutritional supplement containing fish oil and selenium. Treatment with taxol alone reduces p-AXL expression, which is reduced to almost undetectable levels by combined therapy with taxol and a nutritional supplement containing fish oil and selenium (showing a synergistic effect). Treatment with adriamycin or avastin alone resulted in little or no apparent reduction in p-AXL expression, while combined therapy with each of these drugs and such supplements resulted in a significant reduction in expression. Overall, it is clear that the use of nutritional supplements containing fish oil and selenium can be effective in modifying AXL signaling (e.g., reducing p-AXL and HSP90 expression), and that such supplements can be used in combination with various chemotherapeutic agents to achieve synergistic effects in modifying such AXL signaling.

[0159] survival As mentioned above, the use of a nutritional supplement containing fish oil and selenium has many beneficial effects on the treatment of cancer, including reducing tumor cell proliferation, tumor size and angiogenesis, resistance to chemotherapy, and metastasis. These effects are synergistically enhanced by combination therapy with one or more chemotherapeutic agents. In addition, such supplements can reduce or reverse the symptoms of cancer-associated cachexia, as well as alter plasma cytokine concentrations. Such benefits directly address many quality of life issues for individuals living with cancer. In addition, the inventors have discovered that the use of a nutritional supplement containing fish oil and selenium can also provide direct benefits on survival. As shown in Figure 113, the use of such a nutritional supplement ("Nutraceuticals") can also provide direct benefits on survival. Cancer patients receiving a "generic formula" ("fish oil") have a two-year survival rate that is more than double that of similarly treated patients receiving the calorie equivalent of a conventional supplement ("generic formula"). When studying patients with a BMI of less than 19, the fish oil and The use of a nutritional supplement containing fish oil and selenium results in a 500% increase in two-year survival rate compared to generic prescriptions. The inventors believe that the use of a nutritional supplement containing fish oil and selenium can provide a very significant improvement in the survival of cancer patients, especially those with a low BMI.

[0160] It will be apparent to those skilled in the art that many more modifications besides those already described are possible without departing from the inventive concepts herein. The inventive subject matter, therefore, is not to be limited except by the spirit of the appended claims. Moreover, in interpreting both the specification and the claims, all terms should be interpreted in the broadest possible manner consistent with the context. In particular, the terms "comprises" and "comprising" should be interpreted to refer to elements, components, or steps in an inclusive manner, meaning that a referenced element, component, or step may be present, utilized, or combined with other applications, components, or steps not specifically referenced. When the specification and claims refer to at least one selected from the group consisting of A, B, C, ..., and N, the context should be interpreted as requiring only one element from the group, not A+N, or B+N, etc.

Claims

[Claim 1] 1. A composition for the treatment of cancer, comprising: a pharmaceutical formulation for use in treating said cancer, said pharmaceutical formulation comprising a compound effective in inhibiting tumor cell replication; Fish oil and Selenium and A composition comprising:

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