Phosphate derivatives and their uses

Abstract

To provide a compound for treating cancer or inhibiting cancer metastasis.SOLUTION: The present invention provides a compound with general formula (I), or a tautomer, mesomer, racemate, enantiomer, and diastereoisomer thereof, or a mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof (where L represents an alkyl group or the like, Y and Z each represent N, O, S, a single bond or the like, D represents a phosphonic acid group or the like, E represents O or a methylene group, R1 and R2 each represent hydrogen, halogen, an alkyl group or the like, n1 represents an integer of 0-8, n2 represents an integer of 1-6, and n3 represents an integer of 1-3).SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to the field of medical technology, in particular to phosphate derivatives and their use for inhibiting tumor growth and metastasis. [Background technology]

[0002] The microenvironmental characteristics of solid tumors include low extracellular pH, low oxygen concentration, and high glucose uptake. These three factors are closely related and cooperate with each other to promote tumor initiation, growth, invasion, and metastasis. There are two pathways for glucose metabolism: mitochondrial oxidative phosphorylation and glycolysis. Normal cells are oxidatively phosphorylated by aerobic cycling mitochondria, while cancer cells are metabolized by glycolysis (anaerobic and aerobic glycolysis). Research has shown that tumor cells have extremely high glucose uptake, and PET (positron emission tomography) uses this principle to detect 2- 18 Imaging was performed using F-2-deoxyglucose. Malignant tumors grow rapidly and consume large amounts of glucose, resulting in the accumulation of radiopharmaceuticals and imaging. Cancer cells grow very rapidly, and if they grow too quickly, they often become anoxic. Therefore, they stop aerobic oxidation, which requires mitochondria, and energy is provided by anaerobic glycolysis of glucose. After glucose is metabolized to pyruvate, it is not aerobically oxidized through the mitochondrial tricarboxylic acid cycle but is converted to lactate, which is then excreted from the cell by lactate dehydrogenase (LDH). Hypoxia also naturally impairs glycolysis, and the end product of glycolysis is lactate. Abnormal cancer cell metabolism creates an acidic environment in tumor tissue. Cancer cells thrive better in this acidic environment. Lactate produced by glycolysis significantly promotes cancer cell aggressiveness against normal cells and promotes their own adaptability and regulation. A hypoxic-acidic microenvironment mediates tumor resistance to conventional chemotherapy, and radiation therapy also plays a crucial role in immunotherapy resistance.

[0003] The pH (pHe) of the peripheral microenvironment of malignant tumors is 6.5-6.9, and can reach 6.2 in the core, while that of normal cells is 7.3-7.5. The pH of normal human saliva is 6.5-7.4, but in cancer patients it can be as low as 4.5-5.7.

[0004] The acidic peripheral microenvironment of malignant tumors is primarily generated by the following mechanisms: the Wargurg effect leads to lactate aggregation, and hypoxia drives the function of carbonic anhydrase and proton transporters, which, combined, lead to a decrease in the pH of the tumor cell's external environment. After tumor cells absorb large amounts of glucose, the large amounts of lactate generated by highly efficient glycolysis can be transported into the tumor microenvironment by monocarboxylate transport pumps (MCTs). Intracellular CO2 is released into the extracellular space by diffusion, and hypoxia in tumor tissue cells induces the activation of hypoxia-inducible factor 1 (HIF-1), which induces high expression of CA-IX in tumor cells and catalyzes the reaction of CO2 with HO to form carbonic acid. Therefore, glycolysis and hypoxia are the main pathways for the acidic microenvironment of tumor cells (Modern Biomedical Progress 2014, Vol. 14, No. 19, 3775-3777).

[0005] There are two types of lactic acid: L-lactic acid and D-lactic acid. Most of the lactic acid produced by cancer cells is D-lactic acid. When the concentration of D-lactic acid in the cells is too high and the concentration of methylglyoxal is too low, cancer forms. The following is believed: Under normal circumstances, the intestinal mucosa has a barrier function against D-lactic acid, and large amounts of D-lactic acid are excreted in the feces, so it generally does not cause cancer; however, only in cases of intestinal mucosa damage, intestinal surgery, or enteritis can the intestinal mucosa's permeability increase, allowing D-lactic acid to enter the bloodstream. Blood D-lactic acid is the first step in the development of cancer, resulting in over 90% of cancer patients suffering from prolonged gastrointestinal discomfort, prolonged constipation, and prolonged enteritis. Because cell membranes have a barrier function against D-lactic acid, blood D-lactic acid does not necessarily cause cancer. However, when cells are inflamed, abscesses, ulcers, undergone surgery, or trauma, the barrier function of the cell membrane is lost, allowing D-lactic acid to enter the cells, which is the second step in the development of cancer. When the amount of D-lactic acid that enters the cells reaches a certain level, insulin is activated, which then activates glyoxalase, which then converts methylglyoxal into D-lactic acid, resulting in the formation of cancer. This is the cancer trilogy. The vicious cycle that forms here is called an autocatalytic reaction in chemistry, and no one can control it. Autocatalytic reactions have one characteristic: as long as the raw materials are supplied and glucose is continuously ingested, the reaction rate will become faster and faster and will not automatically stop. As a result, the D-lactic acid concentration is very high and the methylglyoxal concentration is very low, and the imbalance between the two is the root cause of cancer.

[0006] Tumor cells themselves adapt to the acidic microenvironment primarily through mechanisms such as autophagy and upregulation of transport systems. Continuous stimulation by various carcinogens and growth factors significantly disrupts hydrogen ion dynamics within tumor cells, leading to abnormal cellular pH balance. To avoid the toxicity of the acidic microenvironment, tumor cells export hydrogen ions, ultimately creating an acidic extracellular environment and an alkaline intracellular environment. Tumor cells achieve this by regulating plasma membrane hydrogen ion-related transport proteins, such as the sodium / hydrogen exchanger (NHE), Na / K ATPase, vesicular H-ATPase (V-ATPase), H / C1 total transporter, and monocarboxylate transporter (MCT). Tumor cells have other mechanisms for adapting to the acidic microenvironment, such as upregulating the expression of genes such as VEGF, promoting tumor angiogenesis; releasing cathepsin B and other proteolytic enzymes, causing extracellular matrix degradation and abnormal immune function; evading the host immune response and immunotherapy, making tumor cells less sensitive to radiation therapy, and developing drug resistance in tumor cells, thereby avoiding drug damage caused by chemotherapy.

[0007] The acidic microenvironment is a powerful weapon for tumor cells to achieve their invasiveness and metastasis. Tumor cells themselves generate proliferation signals, undergo apoptosis, immortalization, invasion, metastasis, immune evasion, and new blood vessel generation, all of which are achieved through mechanisms that induce an acidic environment. All areas of tumor research, from pathogenesis to treatment, are closely related to the tumor's acidic microenvironment.

[0008] The acidic tumor microenvironment affects T cell function. A hypoxia-induced tumor acidic microenvironment is a powerful obstacle to T cell function. Severe hypoxia and lactic acid strongly inhibit T cell activation, proliferation, and cytotoxicity. IL-2-induced T cell proliferation ceases when the extracellular pH is 6.7, but the extracellular pH of hypoxic tumor cells can be as low as 5.8–6.5. At this time, lymphocytes actually enter a state of apoptosis, while tumor cells are resistant and can continue to proliferate. In vitro experiments have shown that an acidic pH can reduce the secretion of IFN-γ and TNF-α by T cells, indicating that an acidic microenvironment can broadly block the production of proinflammatory cytokines. Furthermore, certain acid receptor families can convert changes in extracellular acidity into intracellular signals. The acidic microenvironment can promote the expression of G proteins, T cell inhibitory receptors, and T cell death-associated gene 8, the latter of which mediates the translation and expression of c-myc in lymphocytes.

[0009] The acidic microenvironment influences tumor aggressiveness. The acidic microenvironment outside tumor cells stimulates cell proliferation, activates transcription factors, enhances target gene expression, and promotes tumor formation; tumor cells can use lactic acid to maintain their own acidic microenvironment to promote metastasis; it induces extracellular matrix remodeling, increases tumor invasiveness and metastasis, slows cellular metabolism of carcinogens, and inhibits cellular repair of carcinogen-induced DNA damage.

[0010] The acidic tumor microenvironment affects tumor cells in the following ways: It alters their biological activity; its active growth makes tumor tissue highly susceptible to relative blood perfusion, leading to the accumulation of abnormal acid metabolites and the upregulation of genes such as VEGF, promoting the formation of tumor angiogenesis and providing a favorable environment for tumor metastasis; it induces abnormal immune function, including in other components of the immune system such as DCs, MDSCs, and macrophages; it allows cells to evade the host immune response and immunotherapy; for example, lactic acidosis is a strong negative prognostic indicator for sepsis; it reduces tumor cell sensitivity to radiation therapy, induces drug resistance in tumor cells, and avoids drug damage from chemotherapy.

[0011] Damage caused by products of the tumor acidic microenvironment to normal tissues adjacent to cancer mainly results in: necrosis and apoptosis of peripheral normal cells; and degradation of the extracellular matrix through the release of cathepsin B and other proteolytic enzymes.

[0012] The pH gradient formed by the difference between intracellular and extracellular pH, high pressure in tissue fluids, and low oxygen tension hinder the function of many chemotherapy drugs. A weakly acidic environment (pH = 6–7) is optimal for mAbs. An excessively acidic microenvironment can cause mAb degradation and ultimately reduce its activity, so the extracellular acidic environment within solid tumor tissues may reduce the therapeutic efficacy of mAbs.

[0013] Based on the above-mentioned important role of the acidic microenvironment in tumors, researchers have conducted extensive research on the relationship between the tumor microenvironment and anti-cancer effects. Nuclear magnetic resonance spectroscopy analysis showed that oral or intrasplenic injection of NaHCO3 in mice with metastatic breast cancer selectively increased the extracellular pH of tumors and reduced the formation of lymph node and liver metastases. Proton pump inhibitors (PPIs), primarily pantoprazole, lansoprazole, and omeprazole, are the first-line drugs for the treatment of peptic ulcers. They exert their acid-inhibiting effects by inhibiting the activity of H+ / K+-ATPase. In recent years, the anti-tumor effects of PPIs have gradually been recognized, making them a hot topic in anti-tumor therapy. In vitro and in vivo experiments have demonstrated that PPI-sensitized tumor cell lines are more sensitive to chemotherapy drugs such as cisplatin, 5-fluorouracil, and vinblastine, and omeprazole can induce cisplatin-induced sensitivity in human solid tumors inoculated into nude mice. The mechanism of PPIs is primarily through the inhibition of V-ATPase activity by the drug, which increases extracellular and intracellular pH in organelles such as lysosomes, thereby increasing intracellular drug concentrations and reversing the sensitivity of cells with high expression of multidrug resistance (MDR) genes to antitumor drugs. The ability of PPIs to inhibit tumor cells is directly related to the acidity level of the medium; PPIs can induce selective cytotoxicity in B-cell tumors by activating a large number of reactive oxygen species (ROS) and disrupting the structure of lysosomal membranes, leading to caspase-dependent apoptosis. The use of PPIs also enhances chemotherapy drug sensitivity and inhibits the growth of human xenograft tumors in animal models, without causing systemic reactions. Downregulation of NHE1 also significantly suppresses glioma cell invasion and metastasis. All of these findings suggest that targeted intervention of NHE1 expression can regulate the pH inside and outside tumor cells, altering their biological activity and thus hindering tumor growth and metastasis.

[0014] Tumor metastasis is the underlying cause of clinical treatment failure for malignant tumors. More than 80% of cancer deaths are due to tumor metastasis. Lactic acid accumulation in tumors not only reflects tumor malignancy but is also closely related to its distant metastasis. Tumor metastasis refers to the invasion of tumor cells from the primary site into lymphatic vessels, blood vessels, and body cavities, migrating to other locations and continuing to proliferate, forming tumors of the same type as the primary tumor. This process is called metastasis. Benign tumors do not metastasize, but only malignant tumors do. Common metastatic pathways are as follows: lymphatic metastasis: malignant tumors of epithelial tissues primarily metastasize via lymphatic vessels; hematogenous metastasis: all types of malignant tumors, including sarcoma, kidney cancer, liver cancer, thyroid follicular cancer, and choriocarcinoma, can occur; and implantation metastasis: commonly seen in cancers of abdominal organs. Lactic acid enters endothelial cells via MCT1, causing the degradation and phosphorylation of IκBα, stimulating the autoimmune nuclear factor κB / interleukin-8 pathway, leading to cell migration and angiogenesis. Research has shown that in mouse xenograft models of human colorectal and breast cancer, lactic acid released from tumor cells can be transported via MCT4 and further stimulate interleukin-8-dependent tumor angiogenesis and tumor growth, thereby achieving tumor metastasis. Lactic acid also mediates the body's immune response to cancer cells, aiding their spread. Research has shown that lactic acid is not only present but is involved in every stage of cancer growth.

[0015] Professor Sun Xueying of Harbin Medical University pointed out that immunotherapy, including immunological checkpoint blockade monoclonal antibodies and antigen receptor T cell immunotherapy, has made great progress and is being applied to the clinical treatment of various malignant tumors. However, the tumor-specific microenvironment influences the efficacy of immunotherapy. A detailed understanding of the effects of the tumor's hypoxic and acidic microenvironment on immunotherapy and research into the mechanisms by which cancer cells evade immune surveillance and attack will help explore new ideas and methods to improve the efficacy of tumor immunotherapy (World Chinese Journal of Digestology 2017 July 28;25(21):1934-1944).

[0016] Tumors contain large amounts of lactic acid, which dissociates into lactate anions and hydrogen ions, two "helpers" for cancer cells, allowing them to determine the amount they "consume" based on the amount of "food" they consume. Assuming a lack of glucose, as long as either of these two factors is removed, cancer cells will die rapidly. They found that using a base such as sodium bicarbonate (baking soda) to remove hydrogen ions from the tumor eliminates the synergistic effect of lactic acid and hydrogen ions, rapidly and effectively killing tumor cells lacking or lacking glucose. "Conventional transcatheter arterial chemoembolization (cTACE) involves blocking the tumor's "food channels" and using sodium bicarbonate to remove hydrogen ions from the tumor. This not only denies the tumor "food," but also forces it to rapidly consume more, resulting in rapid "starvation." "The encouraging results of the clinical study showed that 37 patients were treated with cTACE, 18 of whom responded well; and 40 patients were treated with TILA-TACE, 40 of whom responded well.

[0017] Robert Gillies of the H. Lee Moffitt Cancer Research Center in Florida, USA, has conducted extensive research on buffers with buffering properties in the acidic tumor microenvironment, such as sodium bicarbonate, imidazole and its derivatives, Tris, and lysine, and has concluded that these buffers can effectively inhibit tumor growth and metastasis in mice. Among these, sodium bicarbonate was also used in clinical trials conducted by the University of Arizona.

[0018] Many studies have shown that increasing the pH of the tumor microenvironment can effectively inhibit tumor growth and metastasis. Therefore, the acidic microenvironment of solid tumors is a very promising target for cancer therapy, especially for metastasis.

[0019] Therefore, there is an urgent need in the field of biomedical technology to effectively tailor regulators of the acidic microenvironment of tumors, especially novel small molecule compounds with higher pH values. Summary of the Invention [Problem to be solved by the invention]

[0020] The present invention solves the technical problem of the current lack of small molecule compounds that can effectively increase the pH value of the acidic tumor microenvironment, and provides novel phosphate derivatives with monodentate or polydentate alkaline groups with a pH of >8.0. By adjusting the acidity of the tumor microenvironment, the proliferation and metastasis of cancer cells can be effectively inhibited, thereby achieving better efficacy in clinical cancer treatment. [Means for solving the problem]

[0021] In order to solve the above technical problems, the present invention is implemented through the following technical solutions. In one aspect of the present invention, there is provided a compound having the general formula (I), or a tautomeric, meso-, racemic, enantiomeric, diastereomeric, or mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, JPEG0007732678000002.jpg38170L is a C1-C10 alkyl group, cycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkyl heteroaryl group, alkenyl group, alkynyl group, straight or branched chain of 3-15 carbon atoms containing N, O or S atoms, straight or branched chain of 1-15 carbon atoms containing N, O or S atoms, straight or branched chain of repeating units, tandem bisaryl group, tandem bisheteroaryl group, tandem aryl group group and heteroaryl group, N or O or S or JPEG0007732678000003.jpg16170 and are selected from bisaryl groups and bisheteroaryl groups connected via a 16170, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group may each independently be substituted with one or more substituents selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and is preferably a phenyl group or a naphthyl group having one or more substituents; JPEG0007732678000004.jpg97170JPEG0007732678000005.jpg150170JPEG0007732678000006.jpg45170R 1 and R 2 are each independently hydrogen, halogen, alkyl group, cycloalkyl group, hydroxyalkylhydrocarbyl group, alkoxyalkyl group, alkoxycycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, alkenyl group, alkynyl group, amino group, hydroxy group, sulfhydryl group, carboxy group, alkoxy group, cycloalkoxy group, haloalkyl group, cyano group, thioalkyl group, sulfo group, sulfone group, sulfoxide group, phosphate group, alkyl phosphonic acid group, aryl phosphate group, aryl phosphonic acid group, group), wherein the alkyl, cycloalkyl, heterocyclyl, aryl or heteroaryl groups can each be independently substituted with one or more substituents selected from hydroxy, halogen, alkyl, alkoxy, cycloalkyl, heterocyclyl, aryl and heteroaryl groups, or can be empty; R 3 and R 4are each independently hydrogen, an alkyl group, a cycloalkyl group, a cycloalkylalkyl group, an alkylcycloalkyl group, an aryl group, an arylalkyl group, an alkylaryl group, a heteroaryl group, a heteroarylalkyl group, an alkylheteroaryl group, a heterocyclyl group, a heterocyclyl alkyl group, an alkyl heterocyclyl group, an alkenyl alkyl group, an alkynyl alkyl group, and wherein the alkyl group, cycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkylheteroaryl group, heterocyclyl group, heterocyclylalkyl group, alkylheterocyclyl group, alkenylalkyl group, and alkynylalkyl group are optionally unsubstituted or substituted with one or more substituents, each of which independently is selected from the group consisting of alkyl group, cycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkylheteroaryl group, heterocyclyl group, heterocyclylalkyl group, alkylheterocyclyl group, alkenyl group, alkynyl group, amino group, hydroxy group, sulfhydryl group, carboxy group, alkoxy group, cycloalkoxy group, haloalkyl group, alkoxycarbonyl group, acyloxy group, amide group, urea group, alkylsulfonyl group, and alkylsulfonyl group. group), aromatic sulfonyl group, haloalkyl group, halogen, cyano group, nitro group, nitroso group, thiocyano group, isothiocyano group, thioalkyl group, sulfo group, phosphate group, phosphonic acid group, alkyl phosphate group, alkyl phosphonic acid group, aryl phosphate groupphosphate group), arylphosphonic acid group, or R 3 and R 4 together with the N atom to which they are attached form a heterocyclyl group which is monocyclic, bicyclic or tricyclic, or a fused, bridged or spirocyclic ring, said heterocyclyl group containing at least one N atom or containing one or two or three heteroatoms arbitrarily selected from N, S and O, said heterocyclyl group being unsubstituted or substituted with one or more substituents which are each independently selected from alkyl groups, cycloalkyl groups, cycloalkylalkyl groups, alkylcycloalkyl groups, aryl groups, arylalkyl groups, alkylaryl groups, heteroaryl groups, heteroarylalkyl groups, alkylheteroaryl groups, heterocyclyl groups, a heterocyclylalkyl group, an alkylheterocyclyl group, an alkenyl group, an alkynyl group, an amino group, a hydroxy group, a sulfhydryl group, a carboxy group, an alkoxy group, a cycloalkoxy group, a haloaryl group, an alkoxycarbonyl group, an acyloxy group, an amido group, a urea group, an alkylsulfonyl group, an aromatic sulfonyl group, a haloalkyl group, a halogen atom, a cyano group, a nitro group, a nitroso group, a thiocyano group, an isothiocyano group, a thioalkyl group, a sulfo group, a phosphate group, a phosphonic acid group, an alkylphosphate group, an alkylphosphonic acid group, an arylphosphate group, or an arylphosphonic acid group, A 1 , A 2 are each independently selected from H, Li, Na, K, Cs and their corresponding cations, or A 1 , A 2 together form Ca, Mg, Al, Sc, Ti, Cr, Co, Fe, Ni, Cu, Zn, Cd, Hg and their respective corresponding cations, E is an O atom or C(R 1 R 2 ) and R 5 , R 6are each independently selected from a hydrogen atom, a halogen, an alkyl group, an alkoxyalkyl group, a cycloalkyl group, an alkoxycycloalkyl group, a hydroxyalkyl group, a hydroxycycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group can each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; or R 5 , R 6 can form a 3-8 membered ring, said ring can contain 1-2 O, N, and / or S heteroatoms; R 7 , R 8 are each independently selected from a hydrogen atom, an alkyl group, a hydroxyalkyl group, a cycloalkyl group, an alkoxyalkyl group, an alkoxycycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group may each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; R 9are selected from a hydrogen atom, a halogen, an alkylhydrocarbyl group, an alkoxyalkyl group, a cycloalkyl group, an alkoxycycloalkyl group, a hydroxyalkyl group, a hydroxycycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, and an acyl group, and the alkyl group, alkoxyalkyl group, cycloalkyl group, alkoxycycloalkyl group, hydroxyalkyl group, hydroxycycloalkyl group, heterocyclyl group, aryl group, heteroaryl group, and acyl group can each be independently substituted with one or more substituents selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; R 10 is selected from a hydrogen atom, a halogen, an alkylhydrocarbyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, or is empty, and said alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl group can each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; R 12 are selected from a hydrogen atom, an alkylhydrocarbyl group, an alkoxyalkyl group, a cycloalkyl group, an alkoxycycloalkyl group, a hydroxyalkyl group, a hydroxycycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, a heterocyclylalkyl group, an arylalkyl group and a heteroarylalkyl group, and the alkylhydrocarbyl group, alkoxyalkyl group, cycloalkyl group, alkoxycycloalkyl group, hydroxyalkyl group, hydroxycycloalkyl group, heterocyclyl group, aryl group, heteroaryl group, heterocyclylalkyl group, arylalkyl group and heteroarylalkyl group are each independently optionally substituted with one or more substituents selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group and a heteroaryl group; n1 is selected from 1, 2, 3, 4, 5, 6, 7, and 8; n2 is selected from 1, 2, 3, 4, 5, and 6; n3 is selected from 1, 2, and 3; n4 is selected from 0, 1, 2, 3, and 4; n5 is selected from 0, 1, 2, and 3, n6 is selected from 1, 2, and 3.

[0022] Preferably, the compound can be a compound of formula (II) or a tautomer, meso-isomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, JPEG0007732678000007.jpg37170L is a C1-C10 alkyl group, cycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkylheteroaryl group, alkenyl group, alkynyl group, linear or branched chain of 3-15 carbon atoms containing N, O or S atoms, linear or branched chain of 1-15 carbon atoms containing N, O or S atoms, tandem bisaryl group, tandem bisheteroaryl group, tandem aryl and heteroaryl group, N, O or S or JPEG0007732678000008.jpg16170, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group may each independently be substituted with one or more substituents selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and is preferably a phenyl group or a naphthyl group having one or more substituents; R 1 and R 2are each independently selected from hydrogen, halogen, alkyl groups, cycloalkyl groups, hydroxyalkyl hydrocarbyl groups, alkoxyalkyl groups, alkoxycycloalkyl groups, cycloalkylalkyl groups, alkylcycloalkyl groups, alkenyl groups, alkynyl groups, amino groups, hydroxy groups, sulfhydryl groups, carboxy groups, alkoxy groups, cycloalkoxy groups, haloalkyl groups, cyano groups, thioalkyl groups, sulfo groups, sulfone groups, sulfoxide groups, phosphate groups, alkylphosphonic acid groups, arylphosphate groups, and arylphosphonic acid groups, and the alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl groups can each be substituted with one or more substituents independently selected from hydroxy groups, halogen, alkyl groups, alkoxy groups, cycloalkyl groups, heterocyclyl groups, aryl groups, and heteroaryl groups; JPEG0007732678000009.jpg54170 trimethylolmethyl group and dimethylolalkylmethyl group, A 1 , A 2 are each independently selected from H, Li, Na, K, Cs and their corresponding cations, or A 1 , A 2 together form Ca, Mg, Al, Sc, Ti, Cr, Co, Fe, Ni, Cu, Zn, Cd, Hg and their respective corresponding cations, E is C(R 1 R 2 ) and R 5 , R 6are each independently selected from a hydrogen atom, a halogen, an alkyl group, an alkoxyalkyl group, a cycloalkyl group, an alkoxycycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group can each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; or R 5 , R 6 can form a 3-8 membered ring, said ring can contain 1-2 O, N, and / or S heteroatoms; R 7 , R 8 are each independently selected from a hydrogen atom, an alkyl group, a hydroxyalkyl group, a cycloalkyl group, an alkoxyalkyl group, an alkoxycycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group may each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; R 9 is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group may each be independently substituted with one or more substituents selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; R 10is selected from a hydrogen atom, a halogen, an alkyl group, a cycloalkyl group, a heterocyclyl group, an aryl group, a heteroaryl group, or is empty, and the alkyl group, cycloalkyl group, heterocyclyl group, aryl group, or heteroaryl group can each be substituted with one or more substituents independently selected from a hydroxy group, a halogen, an alkyl group, an alkoxy group, a cycloalkyl group, a heterocyclyl group, an aryl group, and a heteroaryl group; n1 is selected from 0, 1, 2, 3, 4, 5, 6, 7, and 8; n2 is selected from 1, 2, 3, 4, 5, and 6; n3 is selected from 1, 2, and 3; n4 is selected from 0, 1, 2, 3, and 4; n5 is selected from 0, 1, 2, and 3, n6 is selected from 1, 2, and 3.

[0023] Preferably, the compound can be a compound of formula (II) or a tautomer, meso-isomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, JPEG0007732678000010.jpg63170R 3 and R 4are each independently selected from hydrogen, an alkyl group, a cycloalkyl group, a cycloalkylalkyl group, an alkylcycloalkyl group, an aryl group, an arylalkyl group, an alkylaryl group, a heteroaryl group, a heteroarylalkyl group, an alkylheteroaryl group, a heterocyclyl group, a heterocyclylalkyl group, an alkylheterocyclyl group, an alkenyl group, and an alkynyl group, wherein said alkyl group, cycloalkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkylheteroaryl group, a heterocyclyl group, a heterocyclylalkyl group, an alkylheterocyclyl group, an alkenyl group, and an alkynyl group are optionally unsubstituted or substituted with one or more substituents, wherein said substituents are each independently selected from alkyl groups, cycloalkyl groups, cycloalkyl groups, alkylcycloalkyl groups, aryl groups, arylalkyl groups, alkylaryl groups, heteroaryl groups, heteroarylalkyl groups, alkylheteroaryl groups, heterocyclyl groups, heterocyclylalkyl groups, alkylheterocyclyl groups, alkenyl groups, and alkynyl groups. alkyl group, cycloalkylalkyl group, alkylcycloalkyl group, aryl group, arylalkyl group, alkylaryl group, heteroaryl group, heteroarylalkyl group, alkylheteroaryl group, heterocyclyl group, heterocyclylalkyl group, alkylheterocyclyl group, alkenyl group, alkynyl group, amino group, hydroxy group, sulfhydryl group, carboxy group, alkoxy group, cycloalkoxy group, haloaryl group, alkoxycarbonyl group, acyloxy group, amido group, urea group, alkylsulfonyl group, aromatic sulfonyl group, haloalkyl group, halogen, cyano group, nitro group, nitroso group, thiocyano group, isothiocyano group, thioalkyl group, sulfo group, phosphate group, phosphonic acid group, alkylphosphate group, alkylphosphonic acid group, arylphosphate group, arylphosphonic acid group, or R 3 and R 4together with the N atom to which they are attached form a heterocyclyl group which is monocyclic, bicyclic or tricyclic, or a fused, bridged or spirocyclic ring, said heterocyclyl group containing at least one N atom or containing one or two or three heteroatoms arbitrarily selected from N, S and O, said heterocyclyl group being unsubstituted or substituted with one or more substituents which are each independently selected from alkyl groups, cycloalkyl groups, cycloalkylalkyl groups, alkylcycloalkyl groups, aryl groups, arylalkyl groups, alkylaryl groups, heteroaryl groups, heteroarylalkyl groups, alkylheteroaryl groups, heterocyclyl groups, The substituents are selected from heterocyclylalkyl groups, alkylheterocyclyl groups, alkenyl groups, alkynyl groups, amino groups, hydroxy groups, sulfhydryl groups, carboxy groups, alkoxy groups, cycloalkoxy groups, haloaryl groups, alkoxycarbonyl groups, acyloxy groups, amido groups, urea groups, alkylsulfonyl groups, aromatic sulfonyl groups, haloalkyl groups, halogens, cyano groups, nitro groups, nitroso groups, thiocyano groups, isothiocyano groups, thioalkyl groups, sulfo groups, phosphate groups, phosphonic acid groups, alkylphosphate groups, alkylphosphonic acid groups, arylphosphate groups, and arylphosphonic acid groups, either alone or in any combination.

[0024] Preferably, the compound can be a compound of formula (III), or a tautomer, meso-isomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, JPEG0007732678000011.jpg39170 R 11is selected from hydrogen, halogen, C1-C6 alkyl groups, C3-C8 cycloalkyl groups, hydroxyalkylhydrocarbyl groups, alkoxyalkyl groups, alkoxycycloalkyl groups, cycloalkylalkyl groups, alkylcycloalkyl groups, alkenyl groups, alkynyl groups, amino groups, hydroxy groups, alkoxy groups, cycloalkoxy groups, haloalkyl groups, cyano groups, thioalkyl groups, sulfo groups, sulfone groups, sulfoxide groups, aryl groups, and heteroaryl groups, and the alkyl groups, cycloalkyl groups, heterocyclyl groups, aryl groups, and heteroaryl groups can each be independently substituted with one or more substituents selected from hydroxy groups, halogen groups, alkyl groups, alkoxy groups, cycloalkyl groups, heterocyclyl groups, aryl groups, and heteroaryl groups.

[0025] Preferably, the compound can be a compound of formula (IV) or a tautomer, meso-isomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, JPEG0007732678000012.jpg89170R 11 is selected from hydrogen, halogen, C1-C6 alkyl groups, C3-C8 cycloalkyl groups, hydroxyalkylhydrocarbyl groups, alkoxyalkyl groups, alkoxycycloalkyl groups, cycloalkylalkyl groups, alkylcycloalkyl groups, alkenyl groups, alkynyl groups, amino groups, hydroxy groups, alkoxy groups, cycloalkoxy groups, haloalkyl groups, cyano groups, thioalkyl groups, sulfo groups, sulfone groups, sulfoxide groups, aryl groups, or heteroaryl groups, and said alkyl groups, cycloalkyl groups, heterocyclyl groups, aryl groups, or heteroaryl groups are each independently substituted with one or more substituents selected from hydroxy groups, halogen groups, alkyl groups, alkoxy groups, cycloalkyl groups, heterocyclyl groups, aryl groups, and heteroaryl groups.

[0026] Preferably, the compounds of general formula (I) include compounds of the following specific structures: JPEG0007732678000013.jpg74170JPEG0007732678000014.jpg227170JPEG0007732678000015.jpg236170JPEG0007732678000016.jpg237170 JPEG0007732678000017.jpg248170JPEG0007732678000018.jpg225170JPEG0007732678000019.jpg253170JPEG0007732678000020.jpg59170

[0027] In another aspect of the present invention, there is provided a pharmaceutical composition comprising a therapeutically effective amount of a compound represented by the general formula (I) or a tautomer, meso-isomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, and one or more pharmaceutically acceptable carriers, diluents, or excipients.

[0028] The above-mentioned acceptable carriers are non-toxic, can aid administration, and do not adversely affect the therapeutic effects of the compound. Such carriers can be any solid excipient, liquid excipient, semi-solid excipient, or gas excipient in aerosol compositions that are commonly available to those skilled in the art. Solid pharmaceutical excipients include starch, cellulose, talc, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, magnesium stearate, sodium stearate, stearyl glyceryl ester, sodium chloride, anhydrous skim milk, etc. Liquid and semi-solid excipients can be selected from glycerin, propylene glycol, water, ethanol, and oils, including those derived from petroleum, animal, vegetable, or synthetic oils, such as peanut oil, soybean oil, mineral oil, and sesame oil. Preferred liquid carriers, particularly injectable solutions, include water, saline, aqueous glucose solution, and glycol. Other auxiliary agents, such as flavoring agents and sweeteners, can also be added to the composition.

[0029] The compound of the present invention is administered in a therapeutically effective amount, and the administration method can be oral, systemic (for example, through the skin, nasally, or via suppository), or parenteral (for example, intramuscular, intravenous, or subcutaneous).The preferred administration method is oral or intravenous administration, which can be adjusted according to the severity of the disease.

[0030] The actual amount of a compound of the invention administered (i.e., active ingredient) will depend on many factors, including the severity of the disease being treated, the age and relative health of the subject being treated, the potency of the compound used, the route and form of administration, and other factors.

[0031] Various dosage forms of the pharmaceutical composition of the present invention can be prepared according to conventional methods in pharmaceutical technology.For example, compound is mixed with one or more carriers, and then prepared into desired dosage forms such as tablets, pills, capsules, semisolids, powders, sustained-release dosage forms, solutions, suspensions, formulations, gas aerosols, etc.

[0032] In another aspect of the present invention, there is further provided the use of the compound represented by said general formula (I) or its tautomer, meso form, racemate, enantiomer, diastereomer, or mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, or said pharmaceutical composition in a drug for preventing and / or treating a disease caused and mediated by an acidic microenvironment.

[0033] The cancers include breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumors, ovarian tumors, peritoneal tumors, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, mastoid kidney tumors, head and neck tumors, leukemia, lymphoma, myeloma, non-small cell lung cancer, head and neck cancer, uterine cancer, and testicular cancer. , fallopian tube cancer, endometrial cancer, vaginal cancer, vulvar cancer, anal cancer, esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, urethral cancer, penile cancer, transitional cell carcinoma, ureteral cancer, renal cell carcinoma, renal pelvis cancer, Hodgkin's disease, non-Hodgkin's lymphoma, soft tissue sarcoma, childhood solid tumors, lymphocytic lymphoma, central nervous system (CNS) tumors, primary central nervous system lymphoma, tumor angiogenesis, spinal cord tumors, brain stem glioma, pituitary adenoma, melanoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, chronic or acute leukemia, and / or combinations of the various cancers.

[0034] In another aspect of the present invention, there is further provided the use of said compound or a tautomer, meso form, racemate, enantiomer, diastereomer, or mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, or said pharmaceutical composition in the preparation of a medicament for inhibiting cancer metastasis.

[0035] In another aspect of the present invention, there is further provided the use of said compound or a tautomer, meso form, racemate, enantiomer, diastereomer, or mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, or a pharmaceutical composition according to claim 7 in the preparation of a medicament for the prevention and / or treatment of a disease caused by acidosis.

[0036] In another aspect of the present invention, there is further provided the use of said compound or a tautomer, meso form, racemate, enantiomer, diastereomer, or mixture form thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, in combination with at least one other anti-cancer agent in the preparation of a medicament for treating cancer or inhibiting cancer metastasis. [Effects of the Invention]

[0037] The compounds of the present invention have significant antitumor activity, and experiments have confirmed that these compounds have inhibitory effects on the proliferation of various cancer cells, making them suitable for the treatment of various cancers. In particular, they have good therapeutic effects on kidney cancer, liver cancer, prostate cancer, stomach cancer, ovarian cancer, and colon cancer, and have a very significant inhibitory effect on cancer metastasis caused by in situ cancer.

[0038] The phosphate derivatives of the present invention can be used as regulators of the acidic tumor microenvironment in combination with clinical anticancer drugs, and can effectively inhibit cell proliferation and metastasis. They have good therapeutic effects on various cancers, especially kidney cancer, liver cancer, prostate cancer, gastric cancer, ovarian cancer, colon cancer, etc., with few toxic side effects, and have very broad application prospects. [Brief explanation of the drawings]

[0039] [Figure 1] FIG. 2 shows the results of tumor inhibition of the compound of Example 7 against a liver cancer cell Hep3B2.1-7-Luc orthotopic xenograft tumor model in Example 23 of the present invention. [Figure 2] FIG. 2 is a drug-time curve diagram after intraperitoneal administration of the compound of Example 7 to BALB / c nude mice in Example 23 of the present invention. [Figure 3] 1 is a histogram of the drug concentration distribution in each tissue and organ 24 hours after intraperitoneal administration of the compound of Example 7 to a BALB / c nude mouse in Example 23 of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0040] The invention will now be described in more detail with reference to the drawings and specific embodiments.

[0041] Example 1: Synthesis of sodium (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene)) bis(methylene)diphosphonate JPEG0007732678000021.jpg54170

[0042] Step (1): Synthesis of diethyl 4-(bromomethyl)benzylphosphonate. 53.0 g (0.201 mol) of 1,4-bis(bromomethyl)benzene and 33.4 g (0.201 mol) of triethyl phosphite were placed in a 250 mL single-neck round-bottom flask and heated to 90°C in an oil bath with magnetic stirring for 3 hours. The reaction mixture was cooled to solidify, and 50 mL of dichloromethane was added to the solid. The resulting well-dispersed suspension was poured into 100 mL of petroleum ether, stirred overnight, filtered, and the filtrate was collected. The residue was washed with 100 mL of a 1:10 mixture of dichloromethane and petroleum ether. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 25.0 g of colorless liquid diethyl 4-(bromomethyl)benzylphosphonate. The yield was 38.7%. 12 H 18 BrO3P, MS(ES+) m / z: 321.0(M+H) + .

[0043] Step (2): Synthesis of ethyl (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(methylene)diphosphonate. 25.0 g (77.9 mmol) of diethyl 4-(bromomethyl)benzylphosphonate, 4.29 g (35.4 mmol) of tris(hydroxymethyl)aminomethane, 12.2 g (88.5 mmol) of anhydrous potassium carbonate, and 150 mL of acetonitrile were placed in a 500 mL single-neck round-bottom flask and heated to 60 °C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 100 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 16.1 g of a colorless liquid, ethyl (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(methylene)diphosphonate. The yield was 75.6%. 28 H 45 NO9P2, MS(ES+) m / z: 602.2(M+H) + .

[0044] Step (3): Synthesis of sodium (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(methylene)diphosphonate: 16.1 g (26.8 mmol) of ethyl (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(methylene)diphosphonate and 150 mL of concentrated hydrochloric acid were placed in a 500 mL single-neck round-bottom flask, heated to 105°C in an oil bath, and magnetically stirred for 42 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and freeze-dried. The solid was dissolved in pure water, and 2N aqueous sodium hydroxide solution was added. Recrystallization from ethanol yielded 16.0 g of a white solid of sodium (4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(methylene)diphosphonate crystalline hydrate. The yield was 76.9%. 1 H NMR(500MHz,D2O)δ7.15(s,4H),3.90(s,2H),3.61(s,3H),2.72(d,J = 19.8 Hz,2H);C 20 H 25 NNa4O9P2 xH2O,MS(ES+)m / z:490.2(M+H) + .

[0045] Example 2: Synthesis of sodium(2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonate JPEG0007732678000022.jpg67170

[0046] Step (1): Synthesis of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonate tetramethyl ester. 24.8 g (0.200 mol) of dimethyl methylphosphonate and 400 mL of dry tetrahydrofuran were placed in a 2000 mL three-necked round-bottom flask. With magnetic stirring, 88 mL of n-butyllithium n-hexane solution (0.22 mol, 2.5 M) was slowly added dropwise in a dry ice acetone bath. The addition time was 50 minutes. Stirring was continued at this temperature for 1 hour. Within 30 minutes, a solution of 35.7 g (0.100 mol) of 1,3,5-tris(bromomethyl)benzene in 200 mL of dry tetrahydrofuran was added dropwise at -78 °C, and stirring was continued for 1 hour. A major proportion of the product was detected by LCMS. The reaction was immediately quenched with potassium dihydrogen phosphate solution (1M, 100 ml) and left at room temperature overnight. The mixture was separated, and the aqueous phase was extracted four times with 50 ml of a 3:1 mixture of chloroform and isopropanol. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spun dry. The residue was purified by column chromatography to give 11.7 g of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester as a yellow oil. The yield was 26.4%. 15 H 25 BrO6P2, MS(ES+)m / z:443.0(M+H) + .

[0047] Step (2): 2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonate tetramethyl ester Synthesis of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonic acid tetramethyl ester. 290 mg (0.653 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 137 mg (1.31 mmol) of diethanolamine, 90 mg (0.653 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL one-neck round-bottom flask and heated in a 40°C oil bath for 16 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 240 mg of colorless liquid 2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonic acid tetramethyl ester. The yield was 78.6%. 19 H 35 NO8P2, MS(ES+) m / z: 468.2(M+H) + .

[0048] Step (3): Synthesis of sodium(2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonate). 240 mg (0.513 mmol) of 2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonate. Tetramethylphosphonate and 5 ml of concentrated hydrochloric acid were placed in a 50 ml round-bottom flask and heated to 105°C in an oil bath under reflux for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid. This solid was purified by resin column chromatography and freeze-dried. The resulting solid was dissolved in pure water, added with 2N aqueous sodium hydroxide, and recrystallized from ethanol to yield 250 mg of a white solid: sodium 2,2'-(5-(((bis(2-hydroxyethyl)amino)methyl)-1,3-phenylene)bis(2,1-ethanediyl)diphosphonate hydrate. The yield was 85.8%. 1 C 15 H 23 NNa4O8P2 xH2O,MS(ES+)m / z:412.1(M+H) + .

[0049] Example 3: Synthesis of sodium(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(4,1-butandyl)diphosphonate JPEG0007732678000023.jpg52170

[0050] Step (1): Synthesis of tert-butyl 1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylcarbamate. 12.1 g (0.100 mol) of tris(hydroxymethyl)aminomethane and 100 ml of tetrahydrofuran were placed in a 500 ml single-neck round-bottom flask. A solution of 21.8 g (0.100 mol) of di-tert-butyl dicarbonate in 80 ml of tetrahydrofuran was added dropwise over 30 minutes in a water bath at 15 °C with magnetic stirring. The reaction mixture was stirred at 15 °C for 16 hours and then rotary evaporated to obtain 22.1 g of viscous liquid 1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl tert-butylcarbamate. The yield was 99%. CH 19 NO5, MS(ES+) m / z: 244.0(M+Na) + .

[0051] Step (2): Synthesis of tert-butyl 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylcarbamate. 22.1 g (0.100 mol) of 1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl tert-butylcarbamate and 200 mL of dichloromethane were placed in a 1000 mL three-necked round-bottom flask, and 18.6 g (0.32 mol) of crushed potassium hydroxide and 53.0 g (0.31 mol) of benzyl bromide were added with magnetic stirring in an ice-water bath at 0°C. The reaction mixture was stirred at 0-15°C for 16 hours, filtered to remove inorganic salts, washed with 200 ml of water three times, dried over anhydrous sodium sulfate, and rotary evaporated to obtain a crude product, which was then subjected to silica gel column chromatography to obtain 35.0 g of viscous liquid tert-butyl 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylcarbamate. The yield was 71.2%. 30 H 37 NO5, MS(ES+) m / z: 514.3 (M+Na)+ .

[0052] Step (3): Synthesis of 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-amine. 26.0 g (52.9 mmol) of tert-butyl 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylcarbamate and 120 mL of dichloromethane were placed in a 500 mL three-neck round-bottom flask, and 40 mL of trifluoroacetic acid was added with magnetic stirring in an ice-water bath at 0°C. The reaction mixture was stirred at 0-15°C for 16 hours. The reaction mixture was rotary evaporated to obtain a crude product, which was washed with petroleum ether, then adjusted to pH 9 with 200 ml of water and 5% sodium bicarbonate, extracted twice with 200 ml of dichloromethane, washed with 200 ml of water, washed with 200 ml of saturated saline, dried over anhydrous sodium sulfate, and spin-dried to obtain 14.0 g of viscous liquid 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-amine. The yield was 67.5%. 30 H 37 NO5, MS(ES+) m / z: 392.2(M+H) + .

[0053] Step (4): Synthesis of 4-(1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylamino)butylphosphonate diethyl ester and 4,4'-(1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylamino)butylphosphonate tetraethyl ester. 782 mg (2.00 mmol) of 1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-amine, 915 mg (4.00 mmol) of 4-chlorobutylphosphonic acid diethyl ester, 552 mg (4.0 mmol) of anhydrous potassium carbonate, and 20 ml of acetonitrile were placed in a 100 ml single-necked round-bottom flask and magnetically stirred in an oil bath at 80°C for 30 hours. The reaction mixture was filtered and rotary evaporated to obtain the crude product, which was purified by column chromatography to obtain 608 mg of a viscous liquid, 4,4'-(1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylazanediyl)bis(4,1-butanediyl)diphosphonic acid tetraethyl ester, in a 39.2% yield. 41 H 63 NO9P2, MS(ES+) m / z:776.4(M+H) + At the same time, 249 mg of viscous liquid 4-(1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylamino)butylphosphonic acid diethyl ester was obtained in a yield of 21.3%. 33 H 46 NO6P, MS(ES+) m / z:584.3(M+H) + .

[0054] Step (5): Synthesis of sodium 4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(4,1-butanediyl)diphosphonate. 608 mg (0.784 mmol) of 4,4'-(1,3-bis(benzyloxy)-2-(benzyloxymethyl)propan-2-ylazanediyl)bis(4,1-butanediyl)diphosphonic acid tetraethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 100 mL one-neck round-bottom flask and refluxed in an oil bath at 105 °C with magnetic stirring for 18 hours. The reaction mixture was rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 279 mg of a white solid of sodium 4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(4,1-butanediyl)diphosphonate crystalline hydrate. The yield was 62.3%. 12 H 25 NNa4O9P2 xH2O,MS(ES+)m / z:394.1(M+H) + .

[0055] Example 4: Synthesis of sodium 2,2'-(4,4'-(2-hydroxyethyl azandyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000024.jpg51170

[0056] Step (1): Synthesis of dimethyl 4-(bromomethyl)phenethylphosphonate. 12.4 g (0.100 mol) of dimethyl methylphosphonate and 200 ml of dry tetrahydrofuran were placed in a 1000 ml three-necked round-bottom flask. Under magnetic stirring, 40 ml of n-butyllithium n-hexane solution (0.10 mol, 2.5 M) was slowly added dropwise in a dry ice acetone bath at -78 °C. The addition time was 30 min. Stirring was continued at this temperature for 1 h. Within 30 min, 26.4 g (0.100 mol) of 1,4-bis(bromomethyl)benzene in 100 ml of dry tetrahydrofuran was added dropwise at -78 °C, and stirring was continued for 1 h. A major proportion of the product was detected by LCMS. The reaction was immediately quenched with potassium dihydrogen phosphate solution (1 M, 50 ml) and allowed to stand overnight at room temperature. The mixture was separated, and the aqueous phase was extracted twice with 50 ml of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spun dry. The residue was purified by column chromatography to give 11.8 g of dimethyl 4-(bromomethyl)phenethylphosphonate as a yellow oil. The yield was 38.4%. 11 H 16 BrO3P, MS(ES+) m / z: 307.0(M+H) + .

[0057] Step (2): Synthesis of 2,2'-(4,4'-(2-hydroxyethylazandyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate tetramethyl ester. 675 mg (2.2 mmol) of dimethyl 4-(bromomethyl)phenethylphosphonate, 61 mg (1.0 mmol) of ethanolamine, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL one-neck round-bottom flask and heated to 60 °C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 330 mg of a colorless liquid, 2,2'-(4,4'-(2-hydroxyethylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 64.0%. 24 H 37 NO7P2, MS(ES+) m / z: 514.2(M+H) + .

[0058] Step (3): Synthesis of sodium 2,2'-(4,4'-(2-hydroxyethylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate. 328 mg (0.638 mmol) of 2,2'-(4,4'-(2-hydroxyethylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and freeze-dried. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 200 mg of a white solid of sodium 2,2'-(4,4'-(2-hydroxyethylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 46.1%. 20 H 25 NNa4O7P2 xH2O, MS(ES+)m / z:458.1(M+H) + .

[0059] Example 5 Synthesis of sodium 2,2',2",2'"-(5,5'-(2-hydroxyethylazandiyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate JPEG0007732678000025.jpg83170

[0060] Step (1): Synthesis of octamethyl 2,2',2",2'"-(5,5'-(2-hydroxyethylazandiyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate. A 100 mL single-neck round-bottom flask was charged with 972 mg (2.2 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 61 mg (1.00 mmol) of ethanolamine, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 20 mL of acetonitrile, and the mixture was heated to 60°C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled and filtered, the filtrate was collected, and the residue was washed twice with 20 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 550 mg of a colorless liquid: octamethyl 2,2',2",2'"-(5,5'-(2-hydroxyethylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate. The yield was 70%. 32 H 55 NO 13 P4, MS(ES+)m / z:786.3(M+H) + .

[0061] Step (2): Synthesis of sodium 2,2',2",2'"-(5,5'-(2-hydroxyethylazandiyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate. 550 mg (0.70 mmol) of 2,2',octamethyl 2",2'"-(5,5'-(2-hydroxyethylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate and 10 ml of concentrated hydrochloric acid were placed in a 50 ml one-neck round-bottom flask and heated to 105 °C in an oil bath with magnetic stirring for 42 hours. The reaction solution was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and freeze-dried. The solid was dissolved in pure water, alkalized with 2 N aqueous sodium hydroxide, and recrystallized from ethanol to obtain 390 mg of a white solid, sodium 2,2',2",2'"-(5,5'-(2-hydroxyethylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate crystalline hydrate. The yield was 58%. 24 H 31 NNaO 13 P4 xH2O, MS(ES+)m / z:674.1(M+H) + .

[0062] Example 6: Synthesis of sodium (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl-azandyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate JPEG0007732678000026.jpg62170

[0063] Step (1): Synthesis of (5-(bromomethyl)-1,3-phenylene)bis(methylene)diphosphonic acid tetraethyl ester. 7.14 g (20.0 mol) of 1,3,5-tris(bromomethyl)benzene and 6.68 g (40.2 mol) of triethyl phosphite were placed in a 50 mL single-neck round-bottom flask and heated to 90 °C in an oil bath with magnetic stirring for 3 hours. The reaction mixture was cooled and solidified. 30 mL of dichloromethane was added to the solid. The resulting suspension was poured into 30 mL of petroleum ether, stirred overnight, filtered, and the filtrate was collected. The residue was washed with 30 mL of a 1:5 dichloromethane:petroleum ether mixture. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 2.45 g of a pale liquid, (5-(bromomethyl)-1,3-phenylene)bis(methylene)diphosphonic acid tetraethyl ester. The yield was 26%. 17 H 29 BrO6P2, MS(ES+) m / z: 493.0(M+Na) + .

[0064] Step (2): Synthesis of (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate octaethyl ester. 1.04 g (2.2 mmol) of (5-(bromomethyl)-1,3-phenylene)bis(methylene)diphosphonic acid tetraethyl ester, 121 mg (1.0 mmol) of tris(hydroxymethyl)aminomethane, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 15 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60 °C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 360 ​​mg of a colorless liquid of (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate octaethyl ester. The yield was 40%. 38 H 67 NO 15 P4, MS(ES+)m / z:902.4(M+H) + .

[0065] Step (3): Synthesis of sodium (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate. 360 mg (0.40 mmol) of (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate octaethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105°C in an oil bath, and magnetically stirred for 42 hours. The reaction mixture was cooled and rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 210 mg of a white solid of sodium (5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(methylene)tetraphosphonate crystalline hydrate. The yield was 57%. 22 H 27 NNaO 15 P4 xH2O, MS(ES+)m / z:678.1(M+H) + .

[0066] Example 7 Synthesis of sodium 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazandiyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate JPEG0007732678000027.jpg33170

[0067] Step (1): Synthesis of 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate. 972 mg (2.2 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 121 mg (1.0 mmol) of tris(hydroxymethyl)aminomethane, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60°C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 465 mg of a colorless liquid of 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate. The yield was 55%. 34 H 59 NO 15 P4, MS(ES+)m / z:846.3(M+H) + .

[0068] Step (2): Synthesis of sodium 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate. 465 mg (0.55 mmol) of 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 42 hours. The reaction mixture was cooled and rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 360 mg of a white solid of sodium 2,2',2",2'"-(5,5'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate crystalline hydrate. The yield was 65%. 26 H 35 NNaO 15 P4 xH2O, MS(ES+)m / z:606.2(M+H) + .

[0069] Example 8 Synthesis of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000028.jpg33170

[0070] Step (1): Synthesis of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 663 mg (1.5 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 363 mg (3.0 mmol) of tris(hydroxymethyl)aminomethane, 207 mg (1.5 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60°C in an oil bath with magnetic stirring for 16 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 493 mg of a colorless liquid of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 68%. 19 H 35 NO9P2, MS(ES+) m / z: 484.2(M+H) + .

[0071] Step (2): Synthesis of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate. 493 mg (1.02 mmol) of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105°C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 349 mg of a white solid of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 60%. 15 H 23 NNa4O9P2 xH2O, MS(ES+)m / z:428.1(M+H) + .

[0072] Example 9 Synthesis of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000029.jpg35170

[0073] Step (1): Synthesis of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 450 mg (0.93 mmol) of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester, 284 mg (2.0 mmol) of methyl iodide, 276 mg (2.0 mmol) of anhydrous potassium carbonate, and 4 mL of acetonitrile were placed in a 10 mL microwave tube and heated to 60 °C in an oil bath with magnetic stirring for 2 hours. The reaction mixture was cooled, filtered, the filtrate was collected, and the residue was washed twice with 10 mL of dichloromethane. The filtrates were combined and rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 300 mg of a colorless liquid of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 64%. 20 H 37 NO9P2, MS(ES+) m / z: 498.2(M+H) + .

[0074] Step (2): Synthesis of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate. 300 mg (0.60 mmol) of 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 170 mg of a white solid of sodium 2,2'-(5-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylmethylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 54%. 16 H 25 NNa4O9P2 xH2O, MS(ES+)m / z:442.1(M+H) + .

[0075] Example 10: Sodium 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazandiyl)bis(methylene)bis(benzenealkyl-5,5 3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate JPEG0007732678000030.jpg70170

[0076] Step (1): Synthesis of 2-butoxymethyl-2-nitro-1,3-propanediol. 15.1 g of tris(hydroxymethyl)nitromethane (0.100 mol) was placed in a 250 ml single-neck round-bottom flask and 50 ml of 50% aqueous potassium hydroxide solution was added dropwise with magnetic stirring at 0 °C in an ice-water bath. After 30 minutes of magnetic stirring at 0 °C, 6.85 g (50 mmol) of bromobutane was added dropwise. The mixture was then magnetically stirred at 0-15 °C for 16 hours. With cooling in an ice-water bath, the reaction mixture was poured into 200 ml of saturated aqueous ammonium chloride solution, saturated with NaCl, and extracted continuously with dichloromethane-isopropanol (10:1) until most of the product was extracted. The organic phases were combined and rotary evaporated to obtain a viscous liquid, which was purified by column chromatography to obtain 4.14 g of pale yellow liquid 2-butoxymethyl-2-nitro-1,3-propanediol. The yield was 40%. 17 NO5, MS(ES+) m / z: 230.1(M+Na) + .

[0077] Step (2): Synthesis of 2-amino-2-butoxymethyl-1,3-propanediol. 4.14 g (40 mmol) of 2-butoxymethyl-2-nitro-1,3-propanediol, 800 mg of Raney nickel, 4 ml of concentrated aqueous ammonia, and 80 ml of methanol were placed in a 250 ml single-neck round-bottom flask and hydrogen was added at 30°C for 16 hours. After filtration, the residue was washed twice with 30 ml of methanol. The combined organic phases were rotary evaporated to obtain 3.3 g of a white solid, 2-amino-2-butoxymethyl-1,3-propanediol. The yield was 92%. CH 19 NO3, MS(ES+) m / z: 178.1(M+H) + .

[0078] Step (3): Synthesis of 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate. 972 mg (2.2 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 178 mg (1.0 mmol) of 2-amino-2-butoxymethyl-1,3-propanediol, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL one-neck round-bottom flask, heated to 60 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 ml of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 380 mg of a colorless liquid: 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate. The yield was 42%. 38 H 67 NO 15 P4, MS(ES+) m / z: 902.3(M+H) + .

[0079] Step (4): Synthesis of sodium 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate. 380 mg (0.42 mmol) of 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)octamethyltetraphosphonate and 10 ml of concentrated hydrochloric acid were placed in a 50 ml one-neck round-bottom flask, heated to 105°C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and freeze-dried. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 180 mg of a white solid of sodium 2,2',2",2'"-(5,5'-(1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(benzenealkyl-5,3,1-triyl))tetra(ethane-2,1-diyl)tetraphosphonate crystalline hydrate. The yield was 41%. 30 H 43 NNaO 15 P4 xH2O, MS(ES+)m / z:790.2(M+H) + .

[0080] Example 11: Synthesis of sodium 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000031.jpg31170

[0081] Step (1): Synthesis of 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 662 mg (1.5 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(2,1-ethanediyl)phosphonic acid tetramethyl ester, 533 mg (3.0 mmol) of 2-amino-2-butoxymethyl-1,3-propanediol, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60°C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 567 mg of a colorless liquid of 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 70%. 23 H 43 NO9P2, MS(ES+) m / z:540.2(M+H) + .

[0082] Step (2): Synthesis of sodium 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate. 220 mg (0.407 mmol) of 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 160 mg of a white solid of sodium 2,2'-(5-((1-butoxy-3-hydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-1,2-phenylene)bis(ethane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 62%. 19 H 31 NNa4O9P2 xH2O, MS(ES+)m / z:484.2(M+H) + .

[0083] Example 12: Synthesis of sodium (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl-azunediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonate JPEG0007732678000032.jpg49170

[0084] Step (1): Synthesis of methyl 5-(bromomethyl)picolinate. 7.56 g (50.0 mmol) of methyl 5-(methyl)picolinate, 9.79 g (55 mmol) of N-bromosuccinimide, and 100 mL of carbon tetrachloride were placed in a 500 mL single-neck round-bottom flask, heated to 90 °C in an oil bath, and magnetically stirred for 10 minutes. 650 mg (2.5 mmol) of dibenzoyl peroxide was added. The reaction mixture was refluxed for 2 hours. After cooling, 100 mL of dichloromethane was added, washed with 100 mL of water, washed twice with 100 mL of 5% sodium bicarbonate, washed with 100 mL of water, dried, filtered, and spin-dried to obtain the crude product. Purification by silica gel column chromatography yielded 8.05 g of viscous liquid methyl 5-(bromomethyl)picolinate. The yield was 70%. C8H8BrNO2, MS (ES+) m / z: 252 (M+Na) + .

[0085] Step (2): Synthesis of methyl 5-(diethoxyphosphoryl)methyl)picolinate. 8.05 g (35.0 mol) of methyl 5-(bromomethyl)picolinate and 8.35 g (50 mmol) of triethyl phosphite were placed in a 100 mL one-neck round-bottom flask, heated to 90°C in an oil bath, and magnetically stirred for 3 hours. The reaction mixture was cooled and rotary evaporated to give a viscous liquid. This was purified by column chromatography to give 4.02 g of methyl 5-(diethoxyphosphoryl)methyl)picolinate as a viscous liquid. The yield was 40%. ;C 12 H 18 NO5P, MS(ES+) m / z: 288.1(M+H) + .

[0086] Step (3): Synthesis of diethyl (6-(hydroxymethyl)pyridin-3-yl)methylphosphonate. 3.75 g (13.1 mmol) of methyl 5-((diethoxyphosphoryl)methyl)picolinate and 50 ml of anhydrous tetrahydrofuran were placed in a 250 ml three-necked round-bottom flask. 15.6 ml (15.6 mmol) of 1 M lithium aluminum hydride was added dropwise to the mixture in an ice-water bath with magnetic stirring. The reaction mixture was allowed to react at 0°C for 2 hours. Under water cooling, 10 g of solid sodium sulfate decahydrate was added to quench the reaction. The mixture was filtered, and the residue was washed twice with 50 ml of tetrahydrofuran. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spin-dried to obtain the crude product. Purification by column chromatography yielded 2.59 g of a viscous liquid diethyl (6-(hydroxymethyl)pyridin-3-yl)methylphosphonate in a 76% yield. 11 H 18 NO4P, MS(ES+) m / z: 260.1(M+H) + .

[0087] Step (4): Synthesis of diethyl (6-(bromomethyl)pyridin-3-yl)methylphosphonate. 2.25 g (8.7 mmol) of diethyl (6-(hydroxymethyl)pyridin-3-yl)methylphosphonate and 50 mL of dichloromethane were placed in a 250 mL single-neck round-bottom flask. 4.72 g (17.4 mmol) of phosphorus tribromide was added with magnetic stirring in an ice-water bath at 0 °C. The reaction mixture was stirred at 0 °C for 3 hours. The mixture was spin-dried, added to 30 g of ice, adjusted to neutral with 5% sodium bicarbonate, and extracted three times with 60 mL of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spin-dried at low temperature to obtain a viscous liquid. Washing with a petroleum ether-dichloromethane mixture yielded 1.6 g of diethyl (6-(bromomethyl)pyridin-3-yl)methylphosphonate as a viscous liquid. The yield was 57%. C 11 H 17 BrNO3P, MS(ES+)m / z:322.0(M+H) + .

[0088] Step (5): Synthesis of (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonic acid ethyl ester. 1.5 g (4.66 mmol) of (6-(bromomethyl)pyridin-3-yl)methylphosphonic acid diethyl ester, 257 mg (2.12 mmol) of tris(hydroxymethyl)aminomethane, 645 mg (4.66 mmol) of anhydrous potassium carbonate, and 15 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 40°C in an oil bath with magnetic stirring for 16 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 30 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 320 mg of a colorless liquid of ethyl (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonate. The yield was 24%. 26 H 43 N3O9P2, MS(ES+)m / z:604.2(M+H) + .

[0089] Step (6): Synthesis of sodium (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonate. 300 mg (0.50 mmol) of ethyl (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonate and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 16 hours. The reaction mixture was cooled and rotary evaporated to give a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 50 mg of a white solid of sodium (6,6'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(6,3-pyridylidene))bis(methylene)diphosphonate crystalline hydrate. The yield was 15%. 18 H 23 N3Na4O9P2 xH2O, MS(ES+)m / z:492.1(M+H) + .

[0090] Example 13: Synthesis of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazandiyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate JPEG0007732678000033.jpg48170

[0091] Step (1): Synthesis of diethyl 1-(4-(bromomethyl)phenyl)propan-2-ylphosphonate. 1.66 g (10.0 mol) of diethyl diethylphosphonate and 40 mL of dry tetrahydrofuran were placed in a 250 mL three-necked round-bottom flask. With magnetic stirring, 4.0 mL of n-butyllithium n-hexane solution (10.0 mmol, 2.5 M) was slowly added dropwise in a dry ice acetone bath. The addition time was 2 minutes. Stirring was continued at −45°C for 30 minutes. At −78°C, 2.64 g (10.0 mol) of 1,4-bis(bromomethyl)benzene in 10 mL of dry tetrahydrofuran was added dropwise, and stirring was continued at −45°C for 1 hour. The reaction was immediately quenched with potassium dihydrogen phosphate solution (1 M, 40 mL) and allowed to stand overnight at room temperature. The mixture was separated, and the aqueous phase was extracted three times with 40 mL of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spun dry. The residue was purified by column chromatography to give 1.2 g of diethyl 1-(4-(bromomethyl)phenyl)propan-2-ylphosphonate as a yellow oil. The yield was 34%. 14 H 22 BrO3P, MS(ES+)m / z:349.0(M+H) + .

[0092] Step (2): Synthesis of ethyl 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate. 1.2 g (3.45 mmol) of diethyl 1-(4-(bromomethyl)phenyl)propan-2-ylphosphonate, 190 mg (1.57 mmol) of tris(hydroxymethyl)aminomethane, 476 mg (3.45 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60 °C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The filtrates were combined and rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 672 mg of a colorless liquid of ethyl 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate. The yield was 65%. 32 H 53 NO9P2, MS(ES+) m / z: 658.3(M+H) + .

[0093] Step (3): Synthesis of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate. 672 mg (1.02 mmol) of ethyl 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 514 mg of a white solid of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(propane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 72%. 24 H 33 NNa4O9P2 xH2O, MS(ES+)m / z:546.2(M+H) + .

[0094] Example 14: Synthesis of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropane-2-ylazandyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000034.jpg20170

[0095] Step (1): Synthesis of 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate methyl ester. 676 mg (2.2 mmol) of dimethyl 4-(bromomethyl)phenethylphosphonate, 121 mg (1.00 mmol) of tris(hydroxymethyl)aminomethane, 304 mg (2.2 mmol) of anhydrous potassium carbonate, and 10 mL of acetonitrile were placed in a 50 mL single-neck round-bottom flask and heated to 60°C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 10 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 380 mg of a colorless liquid of 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 68%. 26 H 41 NO8P2, MS(ES+) m / z: 558.2(M+H) + .

[0096] Step (2): Synthesis of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate. 380 mg (0.68 mmol) of 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask, heated to 105 °C in an oil bath, and magnetically stirred for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 337 mg of a white solid of sodium 1,1'-(4,4'-(1,3-dihydroxy-2-methylpropan-2-ylazanediyl)bis(methylene)bis(4,1-phenylene))bis(ethane-2,1-diyl)diphosphonate crystalline hydrate. The yield was 74%. 22 H 29 NNa4O8P2 xH2O, MS(ES+)m / z:502.2(M+H) + .

[0097] Example 15: Synthesis of sodium 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonate JPEG0007732678000035.jpg47170

[0098] Step (1): Synthesis of methyl 3-bromo-2-(bromomethyl)propionate. 12.3 g (50.0 mol) of 3-bromo-2-(bromomethyl)propionic acid and 6.9 g (50 mmol) of anhydrous potassium carbonate were placed in a 250 mL one-neck round-bottom flask, and 10.65 g (75 mmol) of methyl iodide was added dropwise. The mixture was stirred magnetically at 20 °C for 3 hours. The reaction mixture was filtered, the residue was washed with 50 mL of dichloromethane, and the combined organic phases were rotary evaporated to give 11.7 g of viscous liquid methyl 3-bromo-2-(bromomethyl)propionate. The yield was 90%. C5H8Br2O2, MS (ES+) m / z: 281.0 (M+Na). + .

[0099] Step (2): Synthesis of methyl 3-(diethoxyphosphoryl)-2-((diethoxyphosphoryl)methyl)propionate. 11.0 g (42.5 mol) of methyl 3-bromo-2-(bromomethyl)propionate and 21.2 g (127 mmol) of triethyl phosphite were placed in a 100 mL one-neck round-bottom flask, heated to 120°C in an oil bath, and magnetically stirred for 3 hours. The reaction mixture was cooled and rotary evaporated to obtain a viscous liquid. This was purified by column chromatography to obtain 7.96 g of viscous liquid methyl 3-(diethoxyphosphoryl)-2-((diethoxyphosphoryl)methyl)propionate. The yield was 50%. 13 H 28 O8P2, MS(ES+)m / z:375.1(M+H) + .

[0100] Step (3): Synthesis of 2-(hydroxymethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. 7.5 g (20.0 mmol) of methyl 3-(diethoxyphosphoryl)-2-((diethoxyphosphoryl)methyl)propionate and 100 mL of anhydrous tetrahydrofuran were placed in a 500 mL three-necked round-bottom flask. 24 mL (24 mmol) of 1 M lithium aluminum hydride was added dropwise in an ice-water bath with magnetic stirring. The reaction mixture was allowed to react at 0°C for 2 hours. Under water cooling, 15 g of solid sodium sulfate decahydrate was added to quench the reaction. The mixture was filtered, and the residue was washed twice with 50 mL of tetrahydrofuran. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spin-dried to obtain the crude product. Purification by column chromatography yielded 4.50 g of viscous liquid 2-(hydroxymethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. The yield was 65%. 12 H 28 O7P2, MS(ES+)m / z:347.1(M+H) + .

[0101] Step (4): Synthesis of 2-(bromomethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. 4.3 g (12.4 mmol) of 2-(hydroxymethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester and 80 ml of dichloromethane were placed in a 250 ml single-neck round-bottom flask. 5.04 g (18.6 mmol) of phosphorus tribromide was added with magnetic stirring in an ice-water bath at 0°C. The reaction mixture was stirred at 0°C for 3 hours. The mixture was spin-dried, 40 g of ice was added, the pH was adjusted to neutral with 5% sodium bicarbonate, and extracted three times with 80 ml of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by column chromatography to obtain 3.1 g of viscous liquid 2-(bromomethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. The yield was 61%. 12 H 27 BrO6P2, MS(ES+)m / z:409.0(M+H) + .

[0102] Step (5): Synthesis of 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. 614 mg (1.50 mmol) of 2-(bromomethyl)propane-1,3-diyldiphosphonic acid tetraethyl ester, 363 mg (3.00 mmol) of tris(hydroxymethyl)aminomethane, 207 mg (1.5 mmol) of anhydrous potassium carbonate, and 30 mL of acetonitrile were placed in a 100 mL single-neck round-bottom flask and heated to 80°C in an oil bath with magnetic stirring for 16 hours. The reaction mixture was cooled and filtered. The filtrate was collected, and the residue was washed twice with 30 mL of dichloromethane. The filtrates were combined and rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 276 mg of a colorless liquid of 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonic acid tetraethyl ester. The yield was 41%. 16 H 37 NO9P2, MS(ES+) m / z: 450.2(M+H) + .

[0103] Step (6): Synthesis of sodium 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonate. 276 mg (0.61 mmol) of 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonic acid tetraethyl ester and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask and heated to 105 °C in an oil bath with magnetic stirring for 16 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to obtain 220 mg of a white solid, crystalline sodium 2-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)propane-1,3-diyldiphosphonate hydrate. The yield was 72%. 17NNa4O9P2 xH2O, MS(ES+)m / z:338.1(M+H) + .

[0104] Example 16: Synthesis of 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-sodium trimethylphosphonate JPEG0007732678000036.jpg56170

[0105] Step (1): Synthesis of ethyl 1-(bromomethyl)methane-1,1,1-trimethylphosphonate. 7.76 g (20.0 mol) of tetrabromopentopentanol and 13.3 g (80.0 mmol) of triethyl phosphite were placed in a 100 mL one-neck round-bottom flask, heated to 120°C in an oil bath, and magnetically stirred for 4 hours. The reaction mixture was cooled and rotary evaporated to obtain a viscous liquid. This was purified by column chromatography to obtain 5.60 g of ethyl 1-(bromomethyl)methane-1,1,1-trimethylphosphonate as a viscous liquid. The yield was 50%. 17 H 38 BrO9P3, MS(ES+)m / z:559.1(M+H) + .

[0106] Step (2): Synthesis of ethyl 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-trimethylphosphonate. 2.7 g (4.83 mmol) of ethyl 1-(bromomethyl)methane-1,1,1-trimethylphosphonate, 726 mg (6.0 mmol) of tris(hydroxymethyl)aminomethane, 690 mg (5.0 mmol) of anhydrous potassium carbonate, and 80 mL of acetonitrile were placed in a 250 mL single-neck round-bottom flask and heated to 80°C in an oil bath with magnetic stirring for 16 hours. The reaction mixture was cooled and filtered. The filtrate was collected, and the residue was washed twice with 60 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 520 mg of a colorless liquid of ethyl 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-trimethylphosphonate. The yield was 18%. 21 H 48 NO 12 P3, MS(ES+)m / z:600.2(M+H) + .

[0107] Step (3): Synthesis of 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-trimethylphosphonate. 500 mg (0.83 mmol) of ethyl 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-trimethylphosphonate and 10 mL of concentrated hydrochloric acid were placed in a 50 mL one-neck round-bottom flask and heated to 105 °C in an oil bath with magnetic stirring for 24 hours. The reaction mixture was cooled and rotary evaporated to obtain a solid, which was purified by resin column chromatography and lyophilized. The solid was dissolved in pure water, alkalized with 2 N aqueous sodium hydroxide, and recrystallized from ethanol to obtain 210 mg of a white solid, 1-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)methane-1,1,1-trimethylphosphonate crystalline hydrate. The yield was 39%. 18 NNa6O 12 P3 xH2O, MS(ES+)m / z:432.1(M+H)+ .

[0108] Example 17: Synthesis of sodium 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000037.jpg78170

[0109] Step (1): Synthesis of dimethyl 5-benzyloxyisophthalate. 21.02 g (100 mol) of dimethyl 5-hydroxyisophthalate and 100 ml of dry N,N-dimethylformamide were placed in a 500 ml three-necked round-bottom flask. With magnetic stirring, 20.7 g (150 mmol) of anhydrous potassium carbonate and 25.7 g (150 mmol) of benzyl bromide were added. The mixture was stirred overnight at room temperature. The reaction mixture was poured into 500 ml of ice water and extracted twice with 200 ml of ethyl acetate. The combined organic phases were washed with 200 ml of water and 200 ml of saturated brine, dried over anhydrous sodium sulfate, filtered, and spin-dried to obtain the crude product. The crude product was purified by column chromatography to yield 27 g of oily dimethyl 5-benzyloxyisophthalate. The yield was 90%. 17 H 16 O5, MS(ES+)m / z:323.1(M+Na) + .

[0110] Step (2): Synthesis of (5-(benzyloxy)-1,3-phenylene)dimethanol. 27 g (90.0 mmol) of dimethyl 5-benzyloxyisophthalate and 200 ml of anhydrous tetrahydrofuran were placed in a 1000 ml three-necked round-bottom flask, and 198 ml of 1 M lithium aluminum hydride in tetrahydrofuran was slowly added dropwise in an ice-water bath at 0°C. The reaction mixture was magnetically stirred at 0-20°C for 2 hours. The reaction mixture was cooled in an ice-water bath, and 50 g of sodium sulfate decahydrate was added in batches to quench the reaction. The mixture was filtered, the filtrate was collected, and the residue was washed twice with 200 ml of dichloromethane. The filtrates were combined, dried, and rotary evaporated to obtain 20.2 g of a viscous liquid (5-(benzyloxy)-1,3-phenylene)dimethanol. The yield was 92%. 15 H 16 O3, MS(ES+) m / z: 267.1(M+Na) + .

[0111] Step (3): Synthesis of 1-(benzyloxy)-3,5-bis(bromomethyl)benzene. 20.2 g (82.7 mmol) of (5-(benzyloxy)-1,3-phenylene)dimethanol and 200 ml of anhydrous dichloromethane were placed in a 1000 ml three-necked round-bottom flask, and 67.2 g (248 mmol) of phosphorus tribromide was slowly added dropwise in an ice-water bath at 0°C. The reaction solution was magnetically stirred at 0-20°C for 2 hours. The reaction solution was poured onto 500 g of ice and extracted twice with 200 ml of dichloromethane. The combined organic phases were washed with 200 ml of water, dried, and rotary evaporated to obtain 27.5 g of a pale yellow solid, 1-(benzyloxy)-3,5-bis(bromomethyl)benzene. The yield was 89%. 15 H 14 Br2O, MS(ES+) m / z: 391.0 (M+Na) + .

[0112] Step (4): Synthesis of 2,2'-(5-(benzyloxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 23.06 g (186 mol) of dimethyl methylphosphonate and 250 mL of dry tetrahydrofuran were placed in a 1000 mL three-necked round-bottom flask. With magnetic stirring, 74.4 mL of n-butyllithium n-hexane solution (186 mmol, 2.5 M) was slowly added dropwise over 30 minutes in a dry ice acetone bath. Stirring was continued at -78 °C for 30 minutes. At -78 °C, 27.5 g (74.3 mol) of 1-(benzyloxy)-3,5-bis(bromomethyl)benzene in 100 mL of dry tetrahydrofuran was added dropwise, and stirring was continued at -78 °C for 2 hours. The reaction was immediately quenched with potassium dihydrogen phosphate solution (1 M, 100 mL) and allowed to stand overnight at room temperature. The mixture was separated, and the aqueous phase was extracted three times with 200 ml of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and spun dry. The residue was purified by column chromatography to obtain 25.4 g of 2,2'-(5-(benzyloxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester as a colorless oil. The yield was 75%. 21 H 30 O7P2, MS(ES+)m / z:479.2(M+Na) + .

[0113] Step (5): Synthesis of 2,2'-(5-hydroxy-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 25.4 g (55.7 mol) of 2,2'-(5-(benzyloxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester, 1 g of 10% palladium-carbon, and 250 ml of ethanol were placed in a 1000 ml three-necked round-bottom flask. H2 was added at 20°C with magnetic stirring and the reaction was carried out for 18 hours. After filtration and spin drying, 19.4 g of yellow solid 2,2'-(5-hydroxy-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester was obtained. The yield was 95%. 14 H 24O7P2, MS(ES+)m / z:389.1(M+Na) + .

[0114] Step (6): Synthesis of 2,2'-(5-(2-(2-bromoethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 1.83 g (5.0 mol) of 2,2'-(5-hydroxy-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester, 2.32 g (10.0 mmol) of 1-bromo-2-(2-ethoxy)ethane, 1.38 g (10.0 mmol) of anhydrous potassium carbonate, and 20 mL of anhydrous N,N-dimethylformamide were placed in a 100 mL single-neck round-bottom flask and reacted at 60 °C for 18 hours with magnetic stirring. The reaction mixture was poured into 120 mL of ice water and extracted twice with 120 mL of ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by column chromatography to obtain 1.47 g of viscous liquid 2,2'-(5-(2-(2-bromoethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 57%. 18 H 31 BrO8P2, MS(ES+)m / z:517.1(M+H) + .

[0115] Step (7): Synthesis of 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 1.35 g (2.6 mol) of 2,2'-(5-(2-(2-bromoethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester, 726 mg (6.0 mmol) of tris(hydroxymethyl)aminomethane, 414 mg (3.0 mmol) of anhydrous potassium carbonate, and 80 mL of acetonitrile were placed in a 250 mL one-neck round-bottom flask, heated to 80 °C in an oil bath, and magnetically stirred for 16 hours. The reaction mixture was cooled, filtered, the filtrate collected, and the residue washed twice with 60 ml of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 914 mg of a colorless liquid: 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 63%. 22 H 41 NO 11 P2, MS(ES+)m / z:558.2(M+H) + .

[0116] Step (8): Synthesis of sodium 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 860 mg (1.54 mmol) of 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask and magnetically stirred at 0°C in an ice-water bath. Under nitrogen protection, 3 mL of trimethylbromosilane was added dropwise. The reaction mixture was slowly warmed to 20°C within 1 hour. Stirring was continued for 6 hours. Rotary evaporation gave a viscous liquid, to which 10 ml of dichloromethane and 5 ml of methanol were added, followed by stirring at 20°C for 30 minutes, spin-drying, purification by resin column chromatography, and freeze-drying. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide, and recrystallized from ethanol to give 236 mg of a white solid, crystalline sodium 2,2'-(5-(2-(2-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)ethoxy)ethoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate hydrate. The yield was 24%. 18 H 29 NNaO 11 P2 xH2O, MS(ES+)m / z:502.2(M+H) + .

[0117] Example 18: Synthesis of sodium 2,2'-(5-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000038.jpg61170

[0118] Step (1): Synthesis of 2-methyl-4,4-dihydroxymethyl-4,5-dihydrooxazole. 5.0 g (41.3 mmol) of tris(hydroxymethyl)aminomethane, 50 ml of N,N-dimethylformamide, 9.1 ml of triethyl orthoacetate, and 7.2 ml of N,N-dimethylformamide were placed in a 250 ml single-neck round-bottom flask and stirred at 120°C for 8 hours. The solvent was spun off, and 100 ml of petroleum ether was added to precipitate the solid. The solid was collected to obtain 4.31 g of a pale yellow solid, 2-methyl-4,4-dihydroxymethyl-4,5-dihydrooxazole. The yield was 72%. CH 11 NO3, MS(ES+) m / z: 146.1(M+H) + .

[0119] Step (2): Synthesis of 2,2'-(5-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. 576 mg (2.2 mmol) of triphenylphosphine and 40 mL of dry toluene were placed in a 250 mL three-necked round-bottom flask, and 383 mg (2.2 mmol) of diethyl azodicarboxylate was slowly added dropwise in an ice-water bath at 0°C. After stirring for 30 minutes in the ice-water bath, 732 mg (2.0 mmol) of 2,2'-(5-hydroxy-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 436 mg (3.0 mmol) of 2-methyl-4,4-dihydroxymethyl-4,5-dihydrooxazole were added. The reaction mixture was heated to reflux for 16 hours. The toluene was spun off and the crude product was purified by column chromatography to give 325 mg of a viscous liquid of 2,2'-(5-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester. The yield was 33%. 20 H 33 NO9P2, MS(ES+) m / z: 494.2(M+H) + .

[0120] Step (3): Synthesis of sodium 2,2'-(5-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 325 mg (0.67 mmol) of 2,2'-(5-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask and magnetically stirred at 0 °C in an ice-water bath. Under nitrogen protection, 3 mL of trimethylbromosilane was added dropwise. The reaction mixture was slowly warmed to 20 °C within 1 hour. Stirring was continued for 6 hours. The resulting mixture was rotary evaporated to a viscous liquid, to which 2 ml of absolute ethanol and 2 ml of 6N aqueous hydrochloric acid were added, and the mixture was heated to 70°C in an oil bath with magnetic stirring for 1 hour. The reaction mixture was cooled and rotary evaporated to a viscous liquid. 10 ml of dichloromethane and 5 ml of methanol were added, and the mixture was stirred at 20°C for 30 minutes. The mixture was then spin-dried, purified by resin column chromatography, and lyophilized. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide, and recrystallized from ethanol to yield 82 mg of a white solid crystalline hydrate of sodium 2,2'-(5-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. The yield was 22%. 14 H 21 NNa4O9P2 xH2O, MS(ES+)m / z:414.1(M+H) + .

[0121] Example 19: Synthesis of sodium 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000039.jpg99170

[0122] Step (1): Synthesis of (3,5-bis(2-(dimethoxyphosphoryl)ethyl)phenyl)(2,4,6-trimethylphenyl)iodonium trifluoromethanesulfonate. 759 mg (4.4 mmol) of m-chloroperoxybenzoic acid and 40 mL of anhydrous dichloromethane were placed in a 50 mL dry, single-necked round-bottom flask, and 1.90 g (4.0 mmol) of 2,2'-(5-iodo-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester and 5.29 g (4.4 mmol) of mesitylene were added. In an ice-water bath, 1.02 g (6.8 mmol) of trifluoromethanesulfonic acid was slowly added dropwise. The reaction mixture was stirred at 20°C for 2 hours. The dichloromethane was spun off at low temperature, 100 ml of ether was added, filtered, and the solid was collected to give 2.74 g of a yellow solid, (3,5-bis(2-(dimethoxyphosphoryl)ethyl)phenyl)(2,4,6-trimethylphenyl)iodonium trifluoromethanesulfonate. The yield was 92%. 24 H 34 F3IO9P2S, MS(ES+)m / z:595.1(M+) + .

[0123] Step (2): Synthesis of 2,2'-(5-(4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 1.30 g (1.75 mmol) of (3,5-bis(2-(dimethoxyphosphoryl)ethyl)phenyl)(2,4,6-trimethylphenyl)iodonium trifluoromethanesulfonate (yellow solid) and 20 mL of dry dichloromethane were placed in a 100 mL three-necked round-bottom flask. At 20 °C, 282 mg (2.27 mmol) of 4-(hydroxymethyl)phenol and 677 mg (5.25 mmol) of N,N-dimethylformamide (10 mL) were added dropwise. The reaction mixture was stirred at 20 °C for 16 hours. The dichloromethane was spun off and the crude product was purified by column chromatography to give 620 mg of a viscous liquid of 2,2'-(5-(4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 75%. 21 H 30 O8P2, MS(ES+)m / z:473.1(M+H) + .

[0124] Step (3): Synthesis of 2,2'-(5-(4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 620 mg (1.31 mmol) of 2,2'-(5-(4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of anhydrous dichloromethane were placed in a 50 mL three-necked round-bottom flask, and 710 mg (2.62 mmol) of phosphorus tribromide was added dropwise slowly over an ice-water bath at 0°C. The reaction mixture was magnetically stirred at 0-20°C for 3 hours. The reaction mixture was poured onto 50 g of ice and extracted twice with 50 ml of dichloromethane. The combined organic phases were washed with 100 ml of water, dried, and rotary evaporated to give 652 mg of viscous liquid 2,2'-(5-(4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 93%. 21 H 29BrO7P2, MS(ES+)m / z:535.1(M+H) + .

[0125] Step (4): Synthesis of 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 652 mg (1.22 mol) of 2,2'-(5-(4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester, 484 mg (4.0 mmol) of tris(hydroxymethyl)aminomethane, 207 mg (1.5 mmol) of anhydrous potassium carbonate, and 20 mL of N,N-dimethylformamide were placed in a 50 mL one-neck round-bottom flask and magnetically stirred at 20 °C for 16 hours. The reaction mixture was filtered, the filtrate was collected, and the residue was washed twice with 60 mL of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 506 mg of a colorless liquid of 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 72%. 25 H 39 NO 10 P2, MS(ES+)m / z:576.2(M+H) + .

[0126] Step (5): Synthesis of sodium 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 506 mg (0.878 mmol) of 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask and magnetically stirred at 0°C in an ice-water bath. Under nitrogen protection, 3 mL of trimethylbromosilane was added dropwise. The reaction mixture was slowly warmed to 20°C within 1 hour. Stirring was continued for 6 hours. Rotary evaporation gave a viscous liquid, to which 10 ml of dichloromethane and 5 ml of methanol were added, followed by stirring at 20°C for 30 minutes, spin-drying, purification by resin column chromatography, and freeze-drying. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to give 215 mg of a white solid crystalline hydrate of sodium 2,2'-(5-(4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. The yield was 37%. 21 H 27 NNaO 10 P2 xH2O, MS(ES+)m / z:520.1(M+H) + .

[0127] Example 20: Synthesis of sodium 2,2'-(4'-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000040.jpg65170

[0128] Step (1): Synthesis of 2,2'-(4'-hydroxydiphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester. 954 mg (2.0 mmol) of 2,2'-(5-iodo-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonic acid tetramethyl ester, 146 mg (0.20 mmol) of [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, 424 mg (4.0 mmol) of anhydrous sodium carbonate, 10 mL of acetonitrile, and 1 mL of water were placed in a 50 mL single-neck round-bottom flask and stirred at 20 °C for 5 minutes. 413 mg (3.2 mmol) of N,N-dimethylformamide and 404 mg (3.2 mmol) of 4-hydroxythiophenol were added. The reaction mixture was stirred at 100 °C for 8 hours. The solvent was spun off and the crude product was purified by column chromatography to give 567 mg of a viscous liquid of 2,2'-(4'-hydroxydiphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 64%. 20 H 28 O7P2S, MS(ES+)m / z:443.1(M+H) + .

[0129] Step (2): Synthesis of 2,2'-(4'-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester. 369 mg (1.41 mmol) of triphenylphosphine and 30 mL of dry toluene were placed in a 250 mL three-necked round-bottom flask, and 245 mg (1.41 mmol) of diethyl azodicarboxylate was slowly added dropwise in an ice-water bath at 0°C. After stirring for 30 minutes in the ice-water bath, 567 mg (1.28 mmol) of 2,2'-(4'-hydroxydiphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester and 279 mg (1.92 mmol) of 2-methyl-4,4-dihydroxymethyl-4,5-dihydrooxazole were added. The reaction mixture was heated to reflux for 16 hours. The toluene was spun off and the crude product was purified by column chromatography to give 299 mg of a viscous liquid of 2,2'-(4'-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 41%. 26 H 37 NO9P2, MS(ES+) m / z:570.2(M+H) + .

[0130] Step (3): Synthesis of sodium 2,2'-(4'-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate. 299 mg (0.525 mmol) of 2,2'-(4'-((4-(hydroxymethyl)-2-methyl-4,5-dihydrooxazol-4-yl)methoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask. 3 mL of trimethylbromosilane was added dropwise at 0°C in an ice-water bath under magnetic stirring and nitrogen protection. The reaction mixture was slowly warmed to 20°C within 1 hour. Stirring was continued for 6 hours. The resulting mixture was rotary evaporated to a viscous liquid, to which 2 ml of absolute ethanol and 2 ml of 6N aqueous hydrochloric acid were added, and the mixture was heated to 70°C in an oil bath with magnetic stirring for 1 hour. The reaction mixture was cooled and rotary evaporated to a viscous liquid. 10 ml of dichloromethane and 5 ml of methanol were added, and the mixture was stirred at 20°C for 30 minutes. The mixture was then spin-dried, purified by resin column chromatography, and freeze-dried. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide, and recrystallized from ethanol to yield 142 mg of a white solid, crystalline sodium 2,2'-(4'-(2-amino-3-hydroxy-2-(hydroxymethyl)propoxy)diphenyl-3,5-diyl)bis(ethane-2,1-diyl)diphosphonate hydrate. The yield was 43%. 20 H 25 NNa4O9P2 xH2O, MS(ES+)m / z:490.1(M+H) + .

[0131] Example 21: Synthesis of sodium 2,2'-(5-(2,6-bis(2-(bishydroxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000041.jpg112170

[0132] Step (1): Synthesis of methyl 4-(benzyloxy)-3,5-dibromobenzoate. 6.2 g (20.0 mmol) of methyl 4-hydroxy-3,5-dibromobenzoate, 4.14 g (30.0 mmol) of anhydrous potassium carbonate, and 30 ml of anhydrous N,N-dimethylformamide were placed in a 100 ml single-neck round-bottom flask. The reaction mixture was stirred at 20°C for 16 hours. The reaction mixture was poured into 200 ml of ice water and extracted twice with 100 ml of ethyl acetate. The combined organic phases were washed with 200 ml of water and 200 ml of saturated brine, dried over anhydrous sodium sulfate, and the solvent was spun off. The crude product was purified by column chromatography to obtain 6.78 g of viscous liquid methyl 4-(benzyloxy)-3,5-dibromobenzoate. The yield was 85%. 15 H 12 Br2O3, MS(ES+)m / z:421.0(M+Na) + .

[0133] Step (2): Synthesis of methyl 4-(benzyloxy)-3,5-bis(2-(bisethoxyphosphoryl)vinyl)benzoate. 6.5 g (16.3 mmol) of methyl 4-(benzyloxy)-3,5-dibromobenzoate, 8.03 g (48.9 mmol) of diethyl vinylphosphonate, 6.59 g (65.2 mmol) of triethylamine, 496 mg (1.63 mmol) of tris(o-methylphenyl)phosphine, 183 mg (0.82 mmol) of palladium acetate, and 100 mL of acetonitrile were placed in a 250 mL single-neck round-bottom flask and magnetically stirred at 80 °C under nitrogen protection for 16 hours. After rotary evaporation, the crude product was purified by column chromatography to give 6.18 g of a viscous liquid of methyl 4-(benzyloxy)-3,5-bis(2-(bisethoxyphosphoryl)vinyl)benzoate. The yield was 67%. 27 H 36 O9P2, MS(ES+)m / z:566.2(M+H) + .

[0134] Step (3): Synthesis of methyl 4-hydroxy-3,5-bis(2-(bisethoxyphosphoryl)ethyl)benzoate. 3.0 g (5.30 mmol) of methyl 4-(benzyloxy)-3,5-bis(2-(bisethoxyphosphoryl)vinyl)benzoate, 300 mg of 10% palladium on carbon, and 30 ml of ethanol were placed in a 100 ml single-neck round-bottom flask. H2 was added with magnetic stirring at 20 °C and the reaction was allowed to proceed for 16 hours. After filtration and rotary evaporation, 2.29 g of a pale red solid, methyl 4-hydroxy-3,5-bis(2-(bisethoxyphosphoryl)ethyl)benzoate, was obtained. The yield was 90%. 20 H 34 O9P2, MS(ES+)m / z:481.2(M+H) + .

[0135] Step (4): Synthesis of ethyl 2,2'-(2-hydroxy-5-(hydroxymethyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 2.7 g (4.37 mmol) of methyl 4-hydroxy-3,5-bis(2-(bisethoxyphosphoryl)ethyl)benzoate and 30 mL of anhydrous tetrahydrofuran were placed in a 100 mL three-necked round-bottom flask. 8.7 mL (8.7 mmol) of 1 M lithium aluminum hydride was slowly added dropwise at 0°C with magnetic stirring. The reaction was allowed to proceed at 0-20°C for 2 hours. 5 g of sodium sulfate decahydrate was added to quench the reaction, followed by filtration. The residue was washed twice with 30 mL of dichloromethane. The combined organic phase was then rotary evaporated. The crude product was purified by column chromatography to obtain 1.34 g of a pale red solid, ethyl 2,2'-(2-hydroxy-5-(hydroxymethyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. The yield was 68%. 19 H 34 O8P2, MS(ES+)m / z:453.2(M+H) + .

[0136] Step (5): Synthesis of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 1.64 g (2.21 mmol) of the yellow solid (3,5-bis(2-(dimethoxyphosphoryl)ethyl)phenyl)(2,4,6-trimethylphenyl)iodonium trifluoromethanesulfonate and 20 ml of dry dichloromethane were placed in a 100 ml three-necked round-bottom flask, and at 20°C, 1.20 g (2.65 mmol) of ethyl 2,2'-(2-hydroxy-5-(hydroxymethyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate and 685 mg (5.3 mmol) of N,N-dimethylformamide in 10 ml of dichloromethane were added dropwise. The reaction mixture was stirred at 20°C for 16 hours. The dichloromethane was spun off and the crude product was purified by column chromatography to give 815 mg of a viscous liquid of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 46%. 33 H 56 O 14 P4, MS(ES+) m / z:801.3(M+H) + .

[0137] Step (6): Synthesis of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 815 mg (1.02 mmol) of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(hydroxymethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of anhydrous dichloromethane were placed in a 50 mL three-necked round-bottom flask, and 551 mg (2.03 mmol) of phosphorus tribromide was slowly added dropwise to the mixture in an ice-water bath at 0°C. The reaction mixture was magnetically stirred at 0-20°C for 3 hours. The reaction mixture was poured onto 50 g of ice and extracted twice with 50 ml of dichloromethane. The combined organic phases were washed with 100 ml of water, dried, and rotary evaporated to give 764 mg of viscous liquid 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 87%. 33 H 55 BrO 13 P4, MS(ES+)m / z:863.2(M+H) + .

[0138] Step (7): Synthesis of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 764 mg (0.884 mol) of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-(bromomethyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester, 424 mg (3.5 mmol) of tris(hydroxymethyl)aminomethane, 138 mg (1.0 mmol) of anhydrous potassium carbonate, and 10 mL of N,N-dimethylformamide were placed in a 50 mL one-neck round-bottom flask and magnetically stirred at 20 °C for 16 hours. The reaction mixture was filtered, the filtrate was collected, and the residue was washed twice with 60 ml of dichloromethane. The combined filtrate was rotary evaporated to give a viscous liquid, which was purified by column chromatography to give 543 mg of a colorless liquid, 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 68%. 37 H 65 NO 16 P4, MS(ES+)m / z:904.3(M+H) + .

[0139] Step (8): Synthesis of sodium 2,2'-(5-(2,6-bis(2-(bishydroxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 500 mg (0.553 mmol) of 2,2'-(5-(2,6-bis(2-(bisethoxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask, and 3 mL of trimethylbromosilane was added dropwise at 0 °C in an ice-water bath under nitrogen protection with magnetic stirring. The reaction mixture was slowly warmed to 20°C within 1 hour. Stirring was continued for 16 hours. Rotary evaporation gave a viscous liquid, to which 10 ml of dichloromethane and 5 ml of methanol were added, followed by stirring at 20°C for 30 minutes. The liquid was then spin-dried, purified by resin column chromatography, and freeze-dried. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide, and recrystallized from ethanol to give 97 mg of a white solid, crystalline sodium 2,2'-(5-(2,6-bis(2-(bishydroxyphosphoryl)ethyl)-4-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)phenoxy)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate hydrate. The yield was 18%. 25 H 33 NNaO 16 P4 xH2O, MS(ES+)m / z:736.1(M+H) + .

[0140] Example 22: Synthesis of sodium 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bishydroxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate JPEG0007732678000042.jpg103170

[0141] Step (1): Synthesis of methyl 3-bromo-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. 6.82 g (20.0 mmol) of methyl 3-bromo-5-iodobenzoate, 3.28 g (20.0 mmol) of diethyl vinylphosphonate, 4.04 g (40.0 mmol) of triethylamine, 608 mg (2.0 mmol) of tris(o-methylphenyl)phosphine, 223 mg (1.0 mmol) of palladium acetate, and 100 ml of acetonitrile were placed in a 250 ml one-neck round-bottom flask and magnetically stirred at 80 °C under nitrogen protection for 4 hours. After rotary evaporation, the crude product was purified by column chromatography to obtain 2.41 g of viscous liquid methyl 3-bromo-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. The yield was 32%. 14 H 18 BrO5P, MS(ES+)m / z:399.0(M+Na) + .

[0142] Step (2): Synthesis of methyl 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. 2.2 g (5.83 mmol) of methyl 3-bromo-5-(2-(bisethoxyphosphoryl)vinyl)benzoate, 2.22 g (8.75 mmol) of bis(pinacolato)diboron, 1.72 g (17.5 mmol) of potassium acetate, 213 mg (0.29 mmol) of [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, and 40 mL of dioxane were placed in a 100 mL single-neck round-bottom flask and magnetically stirred at 85 °C under nitrogen protection for 6 hours. After rotary evaporation, the crude product was purified by column chromatography to give 1.86 g of methyl 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. The yield was 75%. 20 H 30 BO7P, MS(ES+)m / z:447.2(M+Na) + .

[0143] Step (3): Synthesis of methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. 1.7 g (4.01 mmol) of methyl 3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate, 1.95 g (4.4 mmol) of 2,2'-(5-(bromomethyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester, 1.11 g (8.02 mmol) of potassium carbonate, 293 mg (0.40 mmol) of [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, and 30 mL of ethylene glycol dimethyl ether were placed in a 100 mL one-neck round-bottom flask and magnetically stirred at 80 °C under nitrogen protection for 18 hours. After rotary evaporation, the crude product was purified by column chromatography to give 1.80 g of methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate. The yield was 68%. 29 H 43 O 11 P3, MS(ES+)m / z:683.2(M+Na) + .

[0144] Step (4): Synthesis of methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzoate. 1.8 g (2.72 mmol) of methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)vinyl)benzoate, 180 mg of 10% palladium on carbon, and 30 ml of ethanol were placed in a 100 ml single-neck round-bottom flask. H2 was added with magnetic stirring at 20°C and the reaction was carried out for 16 hours. After filtration and rotary evaporation, 1.67 g of a viscous liquid, methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzoate, was obtained in a 93% yield. 29 H 43 O 11P3, MS(ES+)m / z:685.2(M+H) + .

[0145] Step (5): Synthesis of 2,2'-(5-(3-(hydroxymethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 1.67 g (2.53 mmol) of methyl 3-(3,5-bis(2-(bismethoxyphosphoryl)ethyl)benzyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzoate and 30 mL of anhydrous tetrahydrofuran were placed in a 100 mL three-necked round-bottom flask. 5.1 mL (5.1 mmol) of 1 M lithium aluminum hydride was slowly added dropwise at 0°C with magnetic stirring. The reaction was allowed to proceed at 0-20°C for 2 hours. 5 g of sodium sulfate decahydrate was added to quench the reaction, followed by filtration. The residue was washed twice with 30 mL of dichloromethane. The combined organic phases were then rotary evaporated. The crude product was purified by column chromatography to obtain 1.22 g of a viscous liquid of 2,2'-(5-(3-(hydroxymethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 76%. 28 H 45 O 10 P3, MS(ES+)m / z:635.2(M+H) + .

[0146] Step (6): Synthesis of 2,2'-(5-(3-(bromomethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 1.22g (1.92mmol) of 2,2'-(5-(3-(hydroxymethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10ml of anhydrous dichloromethane were placed in a 50ml three-necked round-bottom flask, and 1.04g (3.84mmol) of phosphorus tribromide was added dropwise slowly over an ice-water bath at 0°C. The reaction mixture was magnetically stirred at 0-20°C for 3 hours. The reaction mixture was poured onto 50 g of ice and extracted twice with 50 ml of dichloromethane. The combined organic phases were washed with 100 ml of water, dried, and rotary evaporated to give 1.14 g of a viscous liquid, 2,2'-(5-(3-(bromomethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 85%. 28 H 44 BrO9P3, MS(ES+)m / z:697.2(M+H) + .

[0147] Step (7): Synthesis of 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. 1.14 g (1.63 mol) of 2,2'-(5-(3-(bromomethyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester, 594 mg (4.90 mmol) of tris(hydroxymethyl)aminomethane, 225 mg (1.63 mmol) of anhydrous potassium carbonate, and 15 mL of N,N-dimethylformamide were placed in a 50 mL single-neck round-bottom flask and magnetically stirred at 20 °C for 16 hours. The reaction mixture was filtered, the filtrate was collected, and the residue was washed twice with 60 ml of dichloromethane. The combined filtrate was rotary evaporated to obtain a viscous liquid, which was purified by column chromatography to obtain 842 mg of a colorless liquid of 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester. The yield was 70%. 32 H 54 NO 12 P3, MS(ES+)m / z:738.3(M+H) + .

[0148] Step (8): Synthesis of sodium 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bishydroxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate. 800 mg (1.08 mmol) of 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bisethoxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate methyl ester and 10 mL of dry dichloromethane were placed in a 50 mL one-neck round-bottom flask. 3 mL of trimethylbromosilane was added dropwise under nitrogen protection with magnetic stirring at 0 °C in an ice-water bath. The reaction mixture was slowly warmed to 20 °C within 1 hour. Stirring was continued for 16 hours. Rotary evaporation gave a viscous liquid, to which 10 ml of dichloromethane and 5 ml of methanol were added, followed by stirring at 20°C for 30 minutes, spin-drying, purification by resin column chromatography, and freeze-drying. The solid was dissolved in pure water, alkalized with 2N aqueous sodium hydroxide solution, and recrystallized from ethanol to give 221 mg of a white solid, crystalline sodium 2,2'-(5-(3-((1,3-dihydroxy-2-(hydroxymethyl)propan-2-ylamino)methyl)-5-(2-(bishydroxyphosphoryl)ethyl)benzyl)-1,3-phenylene)bis(ethane-2,1-diyl)diphosphonate hydrate. The yield was 25%. 24 H 32 NNa6O 12 P3 xH2O, MS(ES+)m / z:626.2(M+H) + .

[0149] Example 23 Pharmacokinetic study of the compounds of the present invention in a liver cancer cell Hep3B2.1-7-Luc orthotopic xenograft tumor model 1. Cell culture: Hep3B2.1-7-Luc cells were cultured in vitro as adherent cells in EMEM medium supplemented with 1.5 g / L sodium bicarbonate, 10% heat-inactivated fetal bovine serum, 100 U / ml penicillin, and 100 μg / ml streptomycin at 37°C and 5% CO2. They were subcultured twice a week. When the cells were in the exponential growth phase, they were harvested, counted, and inoculated.

[0150] 2. Animals: BALB / c nude mice, female, 6-8 weeks old, weighing 18-22 grams. Tumor inoculation size: 1.25 x 10 6 A suspension of Hep3B2.1-7-luc cells was added to 50% Matrigel and inoculated in situ into the left hepatic lobule of each mouse, which was then sutured. Seven days after cell inoculation, the mice were imaged using an IVIS Lumina XR small animal imager, and mice with appropriate signal intensity (10 per group) were selected for pharmacokinetic studies.

[0151] 3.Animal image processing process: 1) An appropriate amount of D-luciferin was weighed out, and the solution was adjusted to a concentration of 15 mg / ml with DPBS. The solution was filtered and sterilized using a 0.2 μm filter, and stored in a dark place at −20° C.

[0152] 2) Using a 25 x 5 / 8" syringe, one mouse was injected with 150 mg luciferin / kg, based on 10 ul / g of mouse body weight (e.g., 20 g mouse, 0.2 ml).

[0153] 3) After 10-12 minutes, the animals were anesthetized with isoflurane.

[0154] 4) The mice were placed in the imaging system cartridge with their ventral sides facing up for tumor cell detection.

[0155] 5) The exposure time ranged from 1 minute to 1 second, and the imaging results were expressed in photons / second by the imaging software. Bioimaging was performed once a week, with the final imaging before the end of the experiment.

[0156] 4. Experimental endpoint: The experimental endpoint was to investigate whether tumor growth could be inhibited, delayed, or cured. To monitor tumor growth, mice were imaged once a week via the IVIS Lumina XR small animal imager. The tumor signal was determined by the number of photons of exposure per second.

[0157] 5. Data analysis: Taking the data of the compound of Example 7 as an example, the results are shown in Figure 1. The compound of Example 7 has a significant effect of inhibiting tumor growth.

[0158] Example 7 The results of the pharmacokinetic experiment of the compound are shown in Figures 2 and 3 and Table 1.

[0159] JPEG0007732678000043.jpg52170

[0160] From Figure 3, it can be seen that 24 hours after the compound of Example 7 of the present invention was intraperitoneally administered to BALB / c nude mice, the drug compound was mainly distributed in the tissues and organs of the mice, such as the liver, kidney, lung, intestine, spleen, and prostate, and the compound of the present invention is particularly suitable for the treatment of cancers such as liver cancer, kidney cancer, prostate cancer, lung cancer, and colon cancer.

[0161] The above examples are merely illustrative of the embodiments of the present invention, and although the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make some modifications and improvements without departing from the concept of the present invention, and all of these fall within the scope of protection of the present invention. Therefore, the patent protection scope of the present invention should be subject to the scope of the attached claims.

Claims

1. A pharmaceutical composition comprising a therapeutically effective amount of a compound represented by any of the following formulae, or a tautomer, meso form, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents, or excipients, wherein the pharmaceutical composition is administered orally, intravenously, or intraperitoneally.

2. 10. Use of the pharmaceutical composition of claim 1 in the preparation of a medicament for preventing and / or treating cancer.

3. The cancers include breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumor, ovarian tumor, peritoneal tumor, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, mastoid kidney tumor, head and neck tumor, leukemia, lymphoma, myeloma, non-small cell lung cancer, head and neck cancer, uterine cancer, and testicular cancer. , fallopian tube cancer, endometrial cancer, vaginal cancer, vulvar cancer, anal cancer, esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal cancer, urethral cancer, penile cancer, transitional cell carcinoma, ureteral cancer, renal cell carcinoma, renal pelvic cancer, Hodgkin's disease, non-Hodgkin's lymphoma, soft tissue sarcoma, childhood solid tumors, lymphocytic lymphoma, central nervous system (CNS) tumors, primary central nervous system lymphoma, tumor angiogenesis, spinal cord tumors, brainstem glioma, pituitary adenoma, melanoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, chronic or acute leukemia, and / or combinations of the various cancers.

3. The use according to claim 2.

4. Use of the pharmaceutical composition according to any one of claims 1 to 3 in the preparation of a medicament for inhibiting cancer metastasis.

5. 10. Use of the pharmaceutical composition of claim 1 in the preparation of a medicament for the prevention and / or treatment of diseases caused by acidosis.

6. 10. Use of the pharmaceutical composition according to any one of claims 1 to 4 in combination with at least one other anti-cancer agent in the preparation of a medicament for treating cancer or inhibiting cancer metastasis.

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