KR20050020755A - Non-invasive diagnostic imaging technology for mitochondria using radiolabeled lipophilic salts - Google Patents

Abstract

The present invention uses a series of ¹⁸ F-labeled lipophilic phosphonium cations (PhCs) for the non-invasive assessment of ΔΨm, lipophilic ammonium cation analogues of PhCs and PET or SPECT to image the mitochondrial-related pathological state of patients And use it to detect.

Description

Non-invasive DIAGNOSTIC IMAGING TECHNOLOGY FOR MITOCHONDRIA USING RADIOLABELED LIPOPHILIC SALTS}

This application is based on US Provisional Application No. 60 / 354,563, filed February 6, 2002. Their teachings are incorporated herein by reference.

The present invention was made in support of Grant No. CA92871 of the National Institutes of Health. The United States Government has certain rights in the invention.

The present invention provides novel radiolabeled lipophilic salts, in particular radiolabeled lipophilic phosphonium and ammonium salts, from which mitochondrial surface potentials (ΔΨm) can be measured. The present invention also includes pharmaceutical compositions comprising such radiolabeled salts. Additionally, the present invention provides an imaging method for identifying tissues or cells in which mitochondria exhibit abnormal activity levels by locally placing radiolabeled lipophilic salts into dysfunctional mitochondria.

The invention also provides a non-invasive method for early and sensitive detection of tumor response of chemotherapeutic agents.

The invention also provides a method comprising the administration of a high energy radiolabeled lipophilic salt to a patient, particularly to a patient suffering from a disease or disorder associated with mitochondrial dysfunction.

The measurement of mitochondrial membrane potential (ΔΨm) provides the single most comprehensive reflection of mitochondrial bio-energetic function, primarily because it directly depends on the proper integration of the various metabolic pathways that collect in the mitochondria.

In addition to aging, many diseases, including cancer, apoptotic vascular disease, liver disease, degenerative and autoimmune disorders, are associated with mitochondrial dysfunction, and new pathological conditions associated with mitochondria are being identified each year.

Changes in ΔΨm include suppressed apoptosis (eg, over 100 diseases directly caused by mitochondrial dysfunction, such as DNA mutations and oxidative stress (eg, various types of myopathy). Cancer) or increased apoptosis (eg HIV, degenerative disease).

There is a SPECT image searcher labeled with a technetium center that can accumulate in the mitochondria, and a technetium labeled searcher has been used in mitochondrial based imaging techniques. There are also a number of commercially available imaging probes that detect a given pathology using imaging agents such as [ ^{99 m} Tc] MIBI, FDG.

[ ¹⁸ F] FDG detects malignant disorders due to increased glucose metabolism. In addition, as mentioned above, [ ¹⁸ F] FDG is unable to differentiate neoplasms from inflammation. [ ¹⁸ F] FDG is the most effective imaging explorer, but it does not distinguish between neoplasia and inflammation, and often poses a challenge in diagnosis.

Inflammation (eg, tuberculosis) in certain organs is a pathological condition that often occurs among patients with suspected malignant disorders. For example, more than 10% of the lungs indicated as hot spots by [ ¹⁸ F] FDG PET are inflammatory processes, not neoplasms, as revealed by surgery. In other words, about 10% of lung patients noted in [ ¹⁸ F] FDG PET would have to perform unnecessary chest surgery for diseases (inflammatory) that would otherwise be treated with non-surgical, low cost and pathological approaches. It may be.

Recent approaches to evaluating efficacy of chemotherapy depend on tumor growth rate, tracking results for several months, visiting hospitals multiple times, limited sensitivity and high cost, including multiple radiographic scans and frequent treatment cycles. Is the approach.

Technetium labeled mitochondrial imaging agents are limited due to several limitations. More specifically, labeling molecules with ^{99 m} Tc requires a conjugate portion to form a technetium ion complex such that the Tc-based imaging agent has a high molecular weight that reduces the permeability of the imaging agent in the target region. In addition, technetium imaging agents are imaged with SPECT with relatively low spatial resolution and sensitivity compared to corresponding PET images.

There is a technetium complex which is a derivative of annexin V [ ^{99 m} Tc] using apoptosis using SPECT. The novelty of the proposed [ ¹⁸ F] phosphonium cations (PhCs) lies in the detection of apoptosis processes through changes in ΔΨm, whereas annexin V derivatives have these functions due to overexpression of certain membrane proteins. Do this.

[ ^{99 m} Tc] Annesine V detects apoptosis due to externalization of phosphatidylserine on the outer cell membrane. This phenomenon occurs at the end of the apoptosis process when fragmented cells are converted into molecular masses (apoptotic bodies). Immediately after the externalization of phosphatidylserine (called "eat me" phospholipids), apoptotic bodies are phagocytized by adjacent cells. Thus, detecting overexpression of phosphatidylserine is limited to a narrow time window that lasts only a few days. In addition, the appearance and duration of this time window may vary with different chemotherapeutic agents and subjects.

The breakdown of ΔΨm is not a turning point in the process of apoptosis. Thus, the disruption of ΔΨm provides the earliest time point for detecting apoptosis but not the last event in the case of Annexin V, and this disruption persists independently of time.

Current approaches to the assessment of cardiac muscle perfusion and viability include shielding of myocardial activity by accumulation in organs adjacent to the heart and short half-lives of radioactive isotopes ([ ¹³ N] -ammonina and ⁸² rubidium). Because of its limitations, it is limited to PET centers with on-site cyclotrons.

It is desirable to have a family of lipophilic salts having affinity for mitochondria, especially for mitochondria with abnormal activity.

In view of the high incidence of cancer (~ 1.3 million per year in the United States) and the high frequency of chemotherapy application and low frequency of chemotherapy success, noninvasive imaging probes of rapid and sensitive assessment of response to tumor therapy are urgently needed. Required.

The need for diagnostic means in oncology is best exemplified by the rapid conversion of [ ¹⁸ F] FDG PET from investigations of tumors in years to preferred diagnostic means.

There is a need to diagnose and image cardiovascular diseases and disorders, most of which are associated with mitochondrial dysfunction. Thus, there is an urgent need for non-invasive imaging seekers to quickly and sensitively measure the cardiac accumulation of imaging agents with affinity for dysfunctional mitochondria for imaging cardiovascular diseases such as cardiovascular perfusion.

The present invention relates to novel lipophilic salts, in particular lipophilic salts comprising pharmaceutically acceptable anions and at least one phosphonium or ammonium cation according to formula (I) and cations of formula (I) and at least one pharmaceutically acceptable carrier or additive It provides a pharmaceutical composition comprising a.

Preferred lipophilic salts of the present invention exhibit high affinity for mitochondria, especially dysfunctional mitochondria with increased or inhibited activity.

The present invention provides salts comprising at least one pharmaceutically acceptable anion and at least one cation corresponding to formula (I).

Formula I

Where

E is phosphorus or nitrogen;

X ¹ , X ² , X ³ and X ⁴ are independently selected from the group consisting of Ar and R, wherein at least ^one of X ¹ , X ² , X ³ and X ⁴ is an Ar group;

Ar is optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl;

R is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more radioisotope Contains an element.

Salts of the invention comprising a cation of formula (I) are suitable for imaging or evaluation of mitochondrial dysfunction in a patient, in particular for use in PET or SPECT imaging.

Preferred of the invention preferably salts comprising ¹¹ C, ¹⁸ F, ⁷⁶ Br or ¹²³ I, more preferably ¹⁸ F ⁷⁶ Br or ¹²³ I are labeled with one or more radioisotopes. The present invention provides a ¹¹ C- group, such as ^[11 C] a phosphonium cation tracer labeled with triphenylmethyl phosphonium (TPMP).

Although suitable for use in medical centers located close to cyclotrons, [ ¹¹ C] TPMP is limited in distant medical centers since ¹¹ C has a short half-life, such as about 20 minutes. Preferred salts of the present invention include ¹⁸ F, ⁷⁶ Br, ¹²³ I or combinations thereof, suitable for use in peripheral medical facilities and PET medical institutions.

The present invention preferably comprises a cation comprising a cation of formula (I) or a subformula thereof suitable for use in imaging or radiotherapy applications, accumulated by dysfunctional mitochondria, such as mitochondria with inhibited or increased activity. To provide an oily salt. The present invention comprises radiolabeled lipophilic cations, in particular lipophilic phosphonium or ammonium salts of the invention, having one or more radioisotopes capable of binding dysfunctional mitochondria, such as mitochondria with inhibited or increased activity. To provide an imaging agent. More specifically, the radiolabeled lipophilic phosphonium or ammonium salts of the present invention are various in vivo, in which images are formed by radiation emitted by radioisotopes of lipophilic phosphonium or ammonium salts. Used to measure mitochondrial membrane potential (ΔΨm) under conditions. In a preferred embodiment, the radiolabeled lipophilic phosphonium or ammonium salts of the invention comprise one or more radioactive isotopes capable of emitting positron emission and are suitable for use in positron emission tomography (PET) Do.

According to another aspect, the present invention provides a pharmaceutical composition comprising a radiolabeled salt of formula (I) or a pharmaceutically acceptable salt thereof or a solvate thereof, wherein the composition changes in mitochondrial surface potential (ΔΨm). It is useful for imaging cells or tissues with dysfunctional mitochondria and diseases or disorders associated with dysfunctional mitochondria. The invention also provides a method of imaging a patient suffering from any of the diseases or disorders mentioned above with an effective amount of a salt or composition of the invention.

In addition, the present invention provides a therapeutic agent for treating a disease or disorder associated with dysfunctional mitochondria with high affinity of the lipophilic phosphonium or ammonium salts of the present invention, such as disorders or disorders associated with dysfunctional mitochondrial activity. It relates to the use of salts (particularly labeled salts of the invention which emit high energy radiation). Typical diseases or disorders are cancer, cardiovascular and liver diseases, degenerative disorders, autoimmune diseases and disorders, aging, DNA mutations, oxidative stress disorders, various myopathy, HIV and AIDS, and the like.

Preferred lipophilic cations of the invention, including phosphonium or ammonium salts, are preferably located locally in mitochondria, such as dysfunctional mitochondrial activity, with increased or inhibited activity levels.

(Short description of the drawing)

1 to 19 show the results of Examples 9 to 19 described below.

1 shows the radiochromatogram of ¹⁸ F-FBnTP (quality control) and plasma 30 min after injection of the tracer into mice.

2 shows bone uptake of F-18-FBnTP and F-18 in mice.

3 shows mitochondrial membrane potential-dependent uptake of ¹⁸ F-FBnTP in vitro.

4 shows the accumulation of ¹⁸ F-FBnTP in malignant (tumor) and healthy (control) glandular and healthy (muscle) and inflammatory muscles in rats.

5 shows cell uptake of F-18-FBnTP in lung cancer A549 tumors.

6 shows the accumulation of F-18-FBnTP in the prostate lobe at day 4 post castration. Castration-induced apoptosis induced a significant lobe-specific decrease in the ventral side compared to the control.

9 shows the accumulation of F-18-FbnTP and F-18-FDG in prostate tumors in mice.

10 shows C-11 TPMP myocardial uptake.

11 shows the C-11-TPMP extraction fraction.

12 shows C-11-TPMP heart muscle uptake as a function of coronary blood flow.

13 shows the rapid removal of F-18-FBnTP from blood.

FIG. 14 shows the accumulation kinetics of F-18-FBnTP in the left and right ventricles in three consecutive axial slides 8 to 10. As demonstrated above, each trace is the mean value of ROIs. This figure is a good example of the rapid fact kinetics of FBnTP and its uniform distribution in the heart muscle.

15 shows F-18-FBnTP uptake kinetics in the left ventricle and lungs. ROIs for the generation of left ventricular time-activity curves are represented on each corresponding slice. These figures show an excellent example of obtaining high-quality images of the heart muscle as shown in the figure below due to the preferential accumulation of FBnTP in the heart muscle compared to adjacent lungs.

16 relates to F-18-FBnTP heart muscle accumulation in heart failure.

FIG. 17 relates to F-18-FBnTP PET monoaxial imaging of heart muscle before and after pacing. FIG.

FIG. 18 shows a decrease in F-18-FBnTP uptake in inferior walls following pacing-induced heart failure. FIG.

FIG. 19 relates to the time-active course of [ ¹⁸ F] FBnTP cardiac muscle uptake before and after four weeks of pacing (red). Each data point represents the average activity of four slices (slices 46-49). The localization of the region of interest (ROI) is shown in the lower left cardiac image. The same ROI template is used to collect data from all heart sections.

In addition to the salts of formula I mentioned above, the present invention relates to the lipophilic salts of formula I (shown above) wherein the compound provided by the invention is a lipophilic salt of formula I. From here,

Ar is an optionally substituted aryl having 6 to 18 carbon atoms and 1 to 3 rings, 3 to about 18 carbon atoms, 1 to about 3 rings and 1 to about 4 rings selected from N, O and S Optionally substituted heteroaryl having heteroatoms, and optionally substituted aralkyl having 7 to about 12 carbon atoms;

R is optionally substituted C _1-6 alkyl, optionally substituted C _2-6 alkenyl, optionally substituted C _2-6 alkynyl, optionally substituted C _1- with one or more F, Cl, Br, or I atoms ₆ haloalkyl, optionally substituted cycloalkyl having 3 to about 8 ring carbon atoms, optionally substituted aralkyl having 7 to about 12 carbon atoms,

In one or more instances of R above one or more radioisotopes.

Preferred salts of the present invention include salts having at least one phosphonium cation of formula (I) wherein E is phosphorus. Other preferred salts of the present invention include salts with one or more ammonium cations of formula I, wherein E is nitrogen. Another preferred salt comprises a mixture of cations according to formula (I), wherein each cation can be a phosphonium or ammonium cation.

Preferred salts of the invention include one or more R substituents comprising radioisotopes capable of releasing positrons. Typically preferred positron emitting radioisotopes suitable for use in the R substituents include ¹¹ C, ¹⁸ F, ¹²³ I or any combination thereof.

Other preferred salts provided by the present invention include salts comprising a cation of formula II.

Formula II

Wherein E is phosphorus or nitrogen;

Ar ¹ , Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl;

R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one It includes the above radioisotope.

More preferably the cation according to formula (II) provided by the present invention,

Ar ¹ , Ar ² , and Ar ³ are optionally substituted aryl having 6 to 18 carbon atoms and 1 to 3 rings, 3 to about 18 carbon atoms, 1 to about 3 rings, and N, O and S Independently selected from the group consisting of optionally substituted heteroaryl having 1 to about 4 ring heteroatoms selected from and optionally substituted aralkyl having 7 to about 12 carbon atoms;

R ¹ is optionally substituted C _1-6 alkyl, optionally substituted C _2-6 alkenyl, optionally substituted C _2-6 alkynyl, optionally substituted C _1- with one or more F, Cl, Br or I atoms ₆ haloalkyl, optionally substituted cycloalkyl having 3 to about 8 ring carbon atoms, optionally substituted aralkyl having 7 to about 12 carbon atoms,

Wherein at least one instance of R includes at least one radioisotope.

Particularly preferred cations of the invention according to formula (II) are selected from the group consisting of ¹¹ C-methyl, optionally substituted C _1-6 alkyl, optionally substituted C _7-12 aralkyl, optionally substituted C 6-12 aryl, and Each is substituted with one or more ¹¹ C-methyl, ¹¹ C-methoxy, ¹⁸ F, ⁷⁶ Br, ¹²³ I, ¹²⁵ I, ¹³¹ I or a combination thereof. More preferably, R ¹ is benzyl substituted by ¹¹ C- methyl, substituted by one or more F ¹⁸ C _2-6 alkyl or one or more of the ¹⁸ F, ⁷⁶ Br or ¹²³ I.

The invention also provides salts comprising at least one cation according to formula (I) or formula (II) wherein R or R ¹ comprises at least one radioisotope suitable for use in radiation therapy.

The invention further provides salts comprising a cation of formula I represented by formula III.

Formula III

Wherein E is phosphorus or nitrogen;

R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more radioactive Isotopes;

R ² , R ³ and R ⁴ are, in each case of R ² , R ³ and R ⁴ , hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally Independently selected from the group consisting of substituted alkoxy, optionally substituted (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides do.

Preferred R ¹ groups of formula III include halo-C _2-6 alkyl groups or halobenzyl groups, more preferably, R ¹ of formula III is ω-fluoro-C _2-6 alkyl, ω-iodine-C _{2- 6} alkyl group, ortho, meta or para-fluoro benzyl group or ortho, meta or para-iobenzyl group.

Other preferred salts of the invention having a cation according to formula (I) include salts comprising a cation according to formula (IV).

Formula IV

Wherein E is phosphorus or nitrogen;

Arl, Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl;

R ⁵ and R ⁶ in each case of R ⁵ and R ⁶ are selected from the group consisting of hydrogen, halogen, hydroxy, amino, optionally substituted alkyl, optionally substituted haloalkyl and optionally substituted alkoxy;

X is ¹¹ C-methyl or a radioisotope of fluorine or iodine;

m is a number from about 2 to about 6.

Another preferred salt of the invention having a cation according to formula (I) comprises a salt comprising a cation according to formula (V).

Formula V

Wherein E is phosphorus or nitrogen;

Ar ¹ , Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl;

X is ¹¹ C-methyl or a radioisotope of fluorine or iodine;

M is a number from about 1 to about 5.

Another preferred salt of the invention having a cation according to formula I includes salts comprising a cation according to formula VI.

Formula VI

Wherein E is phosphorus or nitrogen;

Z is chloro, fluoro, hydroxy or methoxy;

n is a number from 1 to about 12;

p and q are independently selected from 0 to about 5;

R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more isotopic An element;

R ² and R ³ in each instance of R ² and R ³ are hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alkoxy, optionally substituted Independently selected from the group consisting of (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides.

Other preferred salts of the invention having a cation according to formula (I) include salts comprising a cation according to formula (VII).

Formula VII

Wherein E is phosphorus or nitrogen;

p, q and r are independently selected from a number from 0 to about 4;

R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more Radioactive isotopes;

R ² , R ³ and R ⁴ are, in each case of R ² , R ³ and R ⁴ , hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally Independently selected from the group consisting of substituted alkoxy, optionally substituted (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides do.

Particularly preferred salts comprising at least one pharmaceutically acceptable salt and at least one cation according to formula (I),

¹¹ C-methyl-triphenylphosphonium ion,

¹¹ C-methyl-tri-ortho-tolylphosphonium ion,

¹¹ C-methyl-tri-meth-tolylphosphonium ion,

¹¹ C-methyl-tri-para-tolylphosphonium ion,

¹⁸ F-2-fluoroethyl-triphenylphosphonium ion,

¹⁸ F-2-fluoroethyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-2-fluoroethyl-tri-meth-tolylphosphonium ion,

¹⁸ F-2-fluoroethyl-tri-para-tolylphosphonium ion,

¹⁸ F-3-fluoropropyl-triphenylphosphonium ion,

¹⁸ F-3-fluoropropyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-3-fluoropropyl-tri-meta-tolylphosphonium ion,

¹⁸ F-3-fluoropropyl-tri-para-tolylphosphonium ion,

¹⁸ F-4-fluorobutyl-triphenylphosphonium ion,

¹⁸ F-4-fluorobutyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-4-fluorobutyl-tri-meth-tolylphosphonium ion,

¹⁸ F-4-fluorobutyl-tri-para-tolylphosphonium ion,

¹⁸ F-2-fluorobenzyl-triphenylphosphonium ion,

¹⁸ F-2-fluorobenzyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-2-fluorobenzyl-tri-meta-tolylphosphonium ion,

¹⁸ F-2-fluorobenzyl-tri-para-tolylphosphonium ion,

¹⁸ F-3-fluorobenzyl-triphenylphosphonium ion,

¹⁸ F-3-fluorobenzyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-3-fluorobenzyl-tri-meta-tolylphosphonium ion,

¹⁸ F-3-fluorobenzyl-tri-para-tolylphosphonium ion,

¹⁸ F-4-fluorobenzyl-triphenylphosphonium ion,

¹⁸ F-4-fluorobenzyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-4-fluorobenzyl-tri-meta-tolylphosphonium ion,

¹⁸ F-4-fluorobenzyl-tri-para-tolylphosphonium ion,

¹⁸ F-3-fluoro-4-formyl-benzyl-triphenylphosphonium ion,

¹⁸ F-3-fluoro-4-formyl-benzyl-tri-ortho-tolylphosphonium ion,

¹⁸ F-3-fluoro-4-formyl-benzyl-tri-meta-tolylphosphonium ion,

¹⁸ F-3-fluoro-4-formyl-benzyl-tri-para-tolylphosphonium ion,

( ¹⁸ F-4-fluorobenzyl)-(2-chloroethyl) -diphenylphosphonium ion,

( ¹⁸ F-4-fluorobenzyl)-(3-chloropropyl) -diphenylphosphonium ion,

( ¹⁸ F-4-fluorobenzyl)-(4-chlorobutyl) -diphenylphosphonium ion,

( ¹⁸ F-4-fluorobenzyl)-(6-chloropentyl) -diphenylphosphonium ion,

( ¹⁸ F-4-fluorobenzyl)-(5-chlorohexyl) -diphenylphosphonium ion,

¹⁸ F-2-fluoroethyl-tri (4-pyridyl) phosphonium ion,

¹⁸ F-3-fluoropropyl-tri (4-pyridyl) phosphonium ion,

¹⁸ F-4-fluorobutyl-tri (4-pyridyl) phosphonium ion,

¹⁸ F-2-fluorobenzyl-tri (4-pyridyl) phosphonium ion,

¹⁸ F-3-fluorobenzyl-tri (4-pyridyl) phosphonium ion,

¹⁸ F-4-fluorobenzyl-tri (4-pyridyl) phosphonium ion and

^And salts comprising at least one cation selected from the group consisting of ¹⁸ F-3-fluoro-4-formyl-benzyl-tri (4-pyridyl) phosphonium ions.

Preferred radiolabeled salts of the present invention comprising a cation according to any one of formulas I, II, III, IV, V, VI and VII, are mitochondria for background radiation, such as radiolabeled salts that are not absorbed into the mitochondria. And optionally located locally in the mitochondria such that the ratio of radiation emitted from the radiolabeled salt present at is at least about 5: 1. More preferably, the salts of the present invention are selectively absorbed by dysfunctional mitochondria such that the ratio to normal mitochondria is at least about 5: 1.

Salts of the present invention, in particular lipophilic salts of the present invention, have a distribution profile in the body that is a function of mitochondrial integrity and are suitable for use as diagnostic means in the identification and imaging of various diseases and disorders associated with mitochondrial dysfunction.

The salts of the present invention are also useful diagnostic tools for evaluating the efficacy of existing therapeutic drugs as well as the development of new drugs. For example, the effectiveness of a drug to induce apoptosis (e.g., an anticancer drug) or to inhibit apoptosis (e.g., a drug that inhibits the degenerative process in HIV) is such that administration of the drug uses the salts and imaging methods of the present invention. By measuring ΔΨm to determine whether it is monitored by observing a change (valid) or no change (not valid) for ΔΨm.

Preferred compounds of the invention, in particular compounds suitable for use in the imaging methods provided by the present invention, will emit one or more radiation forms suitable for detection with any standard radiation device such as PET, SPECT, gamma cameras, MRI, and the like. One or more radioactive isotopes.

Preferred radioisotopes include tritium and carbon isotopes, fluorine, technathium, iodine and other radioactive isotopes capable of emitting positrons. Particularly preferred radioisotopes are ¹¹ C, ¹⁸ F, ⁷⁶ Br and ¹²³ I.

The present invention provides a method comprising the steps of providing a radiolabeled salt comprising at least one pharmaceutically acceptable anion and at least one cation according to any one of Formulas I, II, III, IV, V, VI and VII;

Contacting the cell or tissue with a radiolabeled salt; And

An imaging method comprising the step of making a radiographic image is provided.

The imaging method provided by the present invention is suitable for evaluating mitochondrial membrane potential. More preferably, the imaging method of the present invention is suitable for measuring changes in mitochondrial membrane potential over time to assess the effectiveness of a treatment protocol or pharmaceutical treatment. Cells showing an inhibited or increased rate of apoptosis also exhibit reduced or increased mitochondrial activity. Salts provided by the present invention are typically located in cells in concentration proportional to the level of mitochondrial activity. Thus, when cells have decreased levels of apoptosis (e.g. cancer cells), most of the salts of the invention administered to the patient are located locally in these cells, in contrast cells with increased levels of apoptosis (e.g. autoimmune disorders, chemistry Tumor cells responsive to the therapeutic agent will accumulate less salt of the present invention than normal cells. Thus, the imaging methods of the present invention are suitable for use in the imaging of cells, tissues or other physiological targets that suffer from suppressed or increased apoptosis.

The imaging method of the present invention is generally suitable for imaging any disease, disorder or pathology associated with mitochondria. Preferred diseases and disorders suitable for imaging include cancer (including neoplasms), cardiovascular diseases (including crack fractures and blood vessels), liver disease, degenerative diseases or disorders, autoimmune disorders, aging, HIV infection, oxidative stress Or diseases and disorders associated with myopathy or mitochondrial dysfunction caused by DNA mutations.

In addition, the imaging methods of the present invention are suitable for use in measuring the efficiency of therapeutic agents that can cause or inhibit apoptosis. The imaging method of the present invention can also be used to assess the efficacy of a chemotherapy or radiation therapy protocol used to delay or eliminate the growth of cancer or other malignancies.

Imaging methods of the invention suitable for assessing the efficacy of a therapeutic drug may also be suitable for developing new therapeutic agents that can disrupt mitochondrial function in target tissues.

The radiolabeled lipophilic salts of the present invention and imaging methods using the same provide a non-invasive approach for rapidly and sensitively assessing the effectiveness of treatment within days of initiation of a treatment protocol as compared to current assessment methods requiring months.

The most important anticancer drugs (e.g., Taxol, Cisplatin, Vinblastine and Etoposide) have apoptosis effects through a series of cascaded responses, in which the breakdown of ΔΨm constitutes an early, compulsory and irreversible step in the process of apoptosis. Induce. The radiolabeled lipophilic salts of the present invention mainly accumulate in the mitochondria and are directly associated with ΔΨm. Cells affected by treatment accumulate less radiolabeled lipophilic salts of the present invention than unaffected cells. Thus, a significant change between pre- and post-treatment scans is indicative of tumors responding to treatment and, if there is no difference, to indicate non-responsive tumors. Disruption of ΔΨm occurs within a few hours after treatment with most therapeutic agents.

The ability to monitor the first event of the irreversible stage of the apoptosis process can provide a non-invasive method for the rapid and sensitive detection of tumors in response to treatment. The imaging method provided by the present invention in medical settings provides a powerful means for the foundation of chemotherapeutic strategies that most benefit patients with low mortality.

In addition, the radiolabeled lipophilic salts of the present invention are suitable for use in developing this new era of chemotherapeutic agents.

The radiolabeled salts of the present invention and imaging methods using the same are suitable for use in non-invasive techniques for rapidly and sensitively assessing therapeutic efficiency at the molecular level in clinical studies. In addition, the imaging methods of the present invention are suitable for use in selecting suitable malignant targets in test subjects based on the integrity of the mitochondria, from which new drugs can be tested.

The invention is also suitable for use in imaging tumors with one or more salts having a cation according to formula (I) or subformulae thereof. In a preferred tumor imaging method of the invention, radiolabeled salts administered to a patient preferentially accumulate in the mitochondria of malignant cells, thereby radiolabeling of formula I in the mitochondria of malignant cells rather than the concentration of cations in adjacent normal cells. The concentration of cation is higher.

The extent (stage) of cancer disease is a major prognostic factor, and the noninvasive stage using imaging techniques plays an important role in planning the treatment strategy (e.g., surgery versus radio-chemotherapy versus adjuvant chemotherapy).

The lipophilic salts of the invention, including salts with a cation according to formula (I), accumulate to a substantially greater extent in malignant cells than in normal cells. Administering the radiolabeled salts of the present invention is suitable for identifying and imaging malignant cells and tumors, and is also more suitable for measuring tumor developmental stages.

The tumor imaging method of the present invention is suitable in certain embodiments for imaging cancer, particularly preferably for neoplasia including various lung, chest and prostate cancers. In addition, the tumor imaging method of the present invention that can be used can measure the degree of cancer disease (cancer stage).

The present invention provides a method for distinguishing between malignant tumors such as neoplasms and tissues undergoing various inflammatory processes. The lipophilic salts of the present invention, including salts with a cation of formula (I), accumulate to a greater extent in malignant cells than in normal cells, and less in the cellular components of the inflammatory process than in normal cells, thereby causing malignant cells, normal cells and Inflammatory cells can be distinguished. By detecting and detecting malignant cells, unnecessary surgical procedures performed every year can be avoided and the cost-effectiveness of protection management in oncology is improved.

The tumor imaging method of the present invention can distinguish between inflammatory processes and malignant lesions. While not wishing to be bound by theory, it is possible to distinguish and detect malignant cells because malignant cells accumulate salts of the invention to a greater extent than normal cells and tissues, or cellular components of the inflammatory process are typically This is because salts accumulate at lower concentrations.

The present invention further provides a method of imaging cardiovascular disease, in particular a method of imaging myopathy. The cardiovascular imaging method of the present invention comprises administering one or more compounds of formula (I) or subformulae thereof to a patient susceptible to or already suffering from cardiovascular disease.

[ ¹⁸ F] phosphonium cations of the present invention are suitable for imaging various cardiovascular diseases, especially myopathy. Myocytes contain the highest concentration of mitochondria, so the heart is the major target organ of the phosphonium cation. In addition, phosphonium cations maintain good perfusion flow characteristics that allow high-contrast imaging of cardiac failure, including necrosis following cardiac infarction and apoptosis. [ ¹⁸ F] phosphonium cations can accurately distinguish between cardiac muscle areas where apoptosis processes cannot be stopped by medication and angiogenesis and those that can be mediated by intervention.

A preferred imaging method provided by the present invention is a ratio of the radiation intensity of the target to the background of at least 2: 1, more preferably about 5: 1, about 10: 1 or about 15: 1. The use of lipophilic salts according to formulas (I), (II), (III), (IV), (V), (VI) or (VII), which may give rise to proportions. In certain preferred methods the radiation intensity of the target tissue is stronger than that of the background. In another embodiment, the present invention provides a method wherein the strength of the target tissue is weaker than the strength of the background. In general, any radiation intensity difference between the target tissue and the background sufficient to identify and visualize the target tissue is sufficient for use in the methods of the present invention.

In a preferred method of the present invention the compound of the present invention is rapidly released from the tissues of the body to prevent prolonged exposure to the radiation of the radiolabeled compound administered to the patient. Typically compounds according to formula (I) or subformulae thereof are removed from the body within about 24 hours. More preferably, the compounds of the present invention are removed from the body within about 16 hours, 12 hours, 8 hours, 6 hours, 4 hours, 2 hours, 90 minutes or 60 minutes. Typically, preferred compounds are removed between about 60 minutes and about 120 minutes.

Preferred compounds of the present invention are stable in vivo, wherein substantially all of the injected compounds, such as at least about 50%, 60%, 70%, 80%, or more preferably at least 90%, are excreted in the body before Not metabolized

The compounds and salts of the present invention and the imaging methods of the present invention are useful for imaging various pathological conditions including cancer, cardiovascular and liver disease, HIV, AIDS, autoimmune diseases, degenerative disorders, neoplasms and the like.

Typical subjects to which the compounds of the present invention can be administered are mammals, particularly primates, especially humans. A wide variety of subjects are suitable for animal applications. Livestock such as, for example, cattle, sheep, goats, cows, swine, etc .; Poultry such as chickens, ducks, geese, turkeys and the like, domesticated animals, especially pets such as dogs and cats. For diagnostic or research applications, a wide range of mammals, including rodents (eg mice, rats, hamsters), rabbits, primates, and pigs such as inbred pigs, are suitable subjects. Additionally, for in vitro applications such as in vitro diagnostic and research applications, the subject's body fluids and cell samples may be used for blood, urine or tissue samples or animal applications of mammals, particularly primates such as humans. It is suitable for use such as blood, urine or tissue samples of the aforementioned animals.

The invention also provides a package comprising a pharmaceutically acceptable carrier and a salt comprising at least one pharmaceutically acceptable anion and a cation according to any one of formulas I, II, III, IV, V, VI and VII. It provides a pharmaceutical composition.

In certain embodiments, the packaged pharmaceutical composition comprises a reaction precursor necessary to generate a compound or salt according to Formula I or a subformulae thereof in combination with a radiolabeled precursor.

Other packaged pharmaceutical compositions provided by the present invention may further comprise

Instructions for use of the composition to image cells or tissues with increased or inhibited mitochondrial activity,

Instructions for use of the composition to evaluate the therapeutic effect of a drug protocol administered to a patient,

Instructions for use of the composition to selectively image malignant cells and tumors in the presence of inflammation and

Instructions including one or more of the instructions for use of the composition to measure mitochondrial membrane potential (ΔΨm).

In certain preferred embodiments, the present invention provides a kit comprising from about 1 to 30 mCi of the aforementioned radionuclide labeled imaging agent, in combination with a pharmaceutically acceptable carrier. The imaging agent and carrier may be provided in solution or lyophilized form. If the imaging agent and carrier of the kit are in lyophilized form, the kit may optionally include sterile, physiologically acceptable reconstitution media such as water, saline, buffered saline, and the like.

The invention also provides for the synthesis of ¹¹ C, ¹⁸ F, ⁷⁶ Br or ¹²³ I labeled salts of the invention comprising salts comprising a cation according to any one of formulas I, II, III, VI, V, VII and VIII. And devices and synthetic protocols for automating the preparation of pharmaceutical compositions comprising such salts. Similar to the rapidly developing distribution system adapted for [ ¹⁸ F] FDG, the half-life (120 minutes) of the F-18 allows the distribution of cation detectors from the central cyclotron to satellite PET scanners. By labeling the cationic detector with I-123, it can be distributed from the manufacturing center to the medical center with the SPECT.

Imaging agents of the invention can be used in accordance with the methods of the invention by those skilled in the art, such as those in the field of nuclear medicine, to image locations with dysfunctional mitochondria such as mitochondria that exhibit abnormal activity in a patient subject. Any location with dysfunctional mitochondria such as mitochondria showing abnormal activity can be imaged by the imaging methods and imaging agents of the present invention.

Imaging can be generated by differences in the spatial distribution of imaging agents that accumulate at locations with dysfunctional mitochondria such as mitochondria showing abnormal activity. The spatial distribution can be measured using any means suitable for a particular level, such as a gamma camera, a PET device, a SPECT device, and the like. The degree of accumulation of imaging agent can be quantified using known methods for quantifying radioactive release. Particularly useful imaging approaches use one or more imaging agents to perform concurrent studies. Alternatively, the imaging method can be used over a large amount of time, with increasing doses of salts according to formula (I) to conduct continuous studies using split-dose imaging subtraction.

Preferably, an effective amount of detectable degree of the imaging agent of the invention is administered to the subject. According to the present invention, an "detectable amount of effective amount" of an imaging agent of the present invention is defined as an amount sufficient to produce an acceptable image using a device usable for clinical use. A detectable effective amount of the imaging agent of the invention may be administered in one or more injections. The detectable effective amount of the imaging agent of the present invention may vary depending on factors such as the acceptability of the individual, age, sex, weight, specificity of the individual, dose measurement. In addition, an effective amount of detectable degree of the imaging agent of the present invention may vary depending on the device and film-related factors. Optimization of these factors is possible at the level of those skilled in the art. The amount of imaging agent used for diagnostic purposes and the duration of the imaging study are based on the radionuclide used to label the agent, the weight of the patient, the nature and severity of the pathological condition being treated, the treatment being performed by the patient, and the patient's specificity. Depends on the reaction. Ultimately, the attending physician determines the amount of imaging agent administered to each individual patient and the duration of the imaging study.

The compounds described herein may have one or more asymmetric centers or plates. Compounds of the invention comprising asymmetrically substituted atoms can be separated in optically active or racemic forms. Methods for preparing optically active forms are known by methods such as separation of racemic forms (racemates), asymmetric synthesis or synthesis of optically active starting materials. Separation of the racemate can be carried out by conventional methods such as, for example, crystallization in the presence of a dissolving agent or by chromatography, eg, using a chiral HPLC column. Numerous olefin geometric isomers, C═N double bonds and the like may be present in the compounds described herein, and all such stable isomers are contemplated herein. Cis and trans geometric isomers of the present invention are described, which can be separated in the form of mixtures of isomers or in the form of isolated isomers. Unless a specific structural chemistry or isomeric form is specifically designated herein, all geometrical isomeric forms as well as all chiral (enantiomers, diastereomers) and racemic forms are intended.

When any variable in a formula or formula for a compound appears more than once, its definition in each case is independent of its definition in all other cases. Thus, for example, if a group is represented as being substituted with 0-2 *, the group may be optionally substituted with up to two R * groups, in which case R * is selected independently of the definition of R * . In addition, combinations of substituents and / or variables are permissible only if such combinations form stable compounds.

Ar, Ar ¹ , Ar ² , Ar ³ , R, R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , X, X ¹ , X ² , X ³ , X ⁴ and Formulas I, II, Various substituents of the various formulas as indicated above, including the Z of III, IV, V, VI and VII and the substituents mentioned in sub-formulas such as formula I and subformulae (Formulas I, II, III, IV) , V, VI or VII) are optionally substituted. As used herein, the term "optionally substituted" means that one or more arbitrary hydrogens at a designated atom or group are designated groups of substituents, provided that they do not exceed the normal valence of the designated atom and form a stable compound. It means that it is selected from. If the substituent is oxo (keto, ie = O), two hydrogens of the atom are substituted. The present invention is intended to include all isotopes (including radioactive isotopes) of atoms represented in the present invention.

Ar, Ar ¹ , Ar ² , Ar ³ , R, R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , X, X ¹ , X ² , X ³ , X ⁴ and Formula I and their When such substituents as mentioned in the Z and sub-formulas of the subformulae are further substituted, they may also be substituted at one or more possible positions, typically at positions 1-3, or 4, by suitable one or more groups disclosed herein. It may be. Suitable groups that may be present in the "substituted" R ¹ , R ² , R or other gas phase include, for example, halogen; Cyano; Hydroxyl; Nitro; Azido; Alkanoyl (C _1-6 alkanoyl groups such as acyl, etc.); Carboxamido; Alkyl groups (including cycloalkyl groups having 1 to about 8 carbon atoms, preferably having 1, 2, 3, 4, 5 or 6 carbon atoms); Alkenyl and alkynyl groups (including groups having one or more unsaturated bonds and 2 to about 8, preferably 2, 3, 4, 5 or 6 carbon atoms); Alkoxy groups having at least one oxygen bond and from 1 to about 8, preferably 1, 2, 3, 4, 5 or 6 carbon atoms; Aryloxy, such as phenoxy; Alkylthio groups having at least one thioether linkage and 1 to about 8 carbon atoms, preferably 1, 2, 3, 4, 5 or 6 carbon atoms; Alkylsulfinyl groups having at least one sulfinyl bond and from 1 to about 8 carbon atoms, preferably 1, 2, 3, 4, 5 or 6 carbon atoms; Alkylsulfonyl groups comprising at least one sulfonyl bond and 1 to 8 carbon atoms, preferably 1, 2, 3, 4, 5 or 6 carbon atoms; Aminoalkyl groups comprising at least one N atom and 1 to about 8, preferably 1, 2, 3, 4, 5 or 6 carbon atoms; Carbocyclic aryl groups having at least 6 carbons and at least one ring (eg, phenyl, biphenyl, naphthyl and the like, each ring being substituted or unsubstituted aromatic); Arylalkyl having 1 to 3 separated or fused rings and 6 to about 18 ring carbon atoms, benzyl is preferably an arylalkyl group; Arylalkoxy having 0 to 3 separated or fused rings and 6 to about 18 ring carbon atoms, 0-benzyl is preferred arylalkoxy group; Or a saturated, unsaturated or aromatic heterocyclic group having 1 to 3 separated or fused rings having 3 to about 8 members per ring and at least one N, O or S atom such as coumarinyl, quinolinyl, Isoquinolinyl, quinazolinyl, pyridyl, pyrazinyl, pyrimidyl, furanyl, pyrrolyl, thienyl, thiazolyl, triazinyl, oxazolyl, isoxazolyl, imidazolyl, indolyl, benzofuranyl , Benzothiazolyl, tetrahydrofuranyl, tetrahydropyranyl, piperidinyl, morpholinyl, piperazinyl and pyrrolidinyl. Such heterocyclic groups may be further substituted, for example, with hydroxy, alkyl, alkoxy, halogen and amino.

"Alkyl" as used herein is intended to include branched and straight chain saturated aliphatic hydrocarbon groups having a certain number of carbon atoms. Examples of alkyl are methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl and s-pentyl. Preferred alkyl groups are C _1-6 alkyl groups. Particularly preferred alkyl groups are methyl, ethyl, propyl, butyl and 3-pentyl.

As used herein, the term C _1-4 alkyl includes alkyl groups consisting of 1 to 4 carbon atoms, which may include cyclopropyl groups. Suitable examples are methyl, ethyl and cyclopropylmethyl.

"Cycloalkyl" is intended to include saturated ventilation with a certain number of carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. Cycloalkyl groups typically have from 3 to about 8 reductions.

In the term “(C _3-8 cycloalkyl) C _1-4 alkyl” cycloalkyl and alkyl are as defined above and the point of attachment is at the alkyl group. Such terms include, but are not limited to, cyclopropylmethyl, cyclohexylmethyl, and cyclohexylmethyl.

"Alkenyl" is intended to include straight or branched hydrocarbon chains containing one or more unsaturated carbon-carbon bonds that may occur at any stable position along the chain, such as ethenyl and propenyl. Alkenyl groups typically have 2 to about 8 carbon atoms, more typically 2 to about 6 carbon atoms.

"Alkynyl" is intended to include straight chain or branched hydrocarbon chains comprising one or more carbon-carbon triple bonds that can occur at any stable position along the chain, such as ethynyl and propynyl. Alkynyl groups typically have 2 to about 8 carbon atoms, more typically 2 to about 6 carbon atoms.

"Haloalkyl" is intended to include branched and straight chain saturated aliphatic hydrocarbons having a specified number of carbon atoms, substituted with one or more halogen atoms. Examples of haloalkyl are mono-, di- or tri-fluoromethyl, mono-, di- or tri-chloromethyl, mono-, di-, tri-, tetra- or penta-fluoroethyl and mono-, di -, But not limited to, tri-, tetra- or penta-chloroethyl. Typical haloalkyl groups have 1 to about 8 carbon atoms, more typically 1 to about 6 carbon atoms.

"Alkoxy" refers to an alkyl group as defined above having the specified number of carbon atoms bonded through the bridge of oxygen. Examples of alkoxy are methoxy, ethoxy, n-propoxy, i-propoxy, n-butoxy, 2-butoxy, t-butoxy, n-pentoxy, 2-pentoxy, 3-pentoxy, Isopentoxy, neopentoxy, n-hexoxy, 2-hexoxy, 3-hexoxy and 3-methylpentoxy. Alkoxy groups typically have 1 to about 8 carbon atoms, more typically 1 to about 6 carbon atoms.

"Halooloxy" refers to a haloalkyl group as defined above having a certain number of carbon atoms bonded through an oxygen bridge.

As used herein, the term "alkylthio" includes groups having one or more thioether bonds and preferably 1 to about 8 carbon atoms, more typically 1 to about 6 carbon atoms.

As used herein, the term "alkylsulfinyl" includes one or more sulfoxide (SO) linking groups and groups typically having from 1 to about 8 carbon atoms, more typically from 1 to about 6 carbon atoms.

As used herein, the term “alkylsulfonyl” includes one or more sulfonyl (SO ₂ ) linking groups and groups typically having from 1 to about 8 carbon atoms, more typically from 1 to about 6 carbon atoms. .

As used herein, the term "alkylamino" refers to one or more primary, secondary and / or tertiary amine groups and typically from 1 to about 8 carbon atoms, more typically from 1 to about 6 Groups with carbon atoms.

As used herein, "halo" or "halogen" refers to fluoro, chloro, bromo or iodine, and "counter-ion" refers to small, negatively charged chemistry such as chlorides, bromide, hydroxides, acetates, sulfates and the like. Used to represent species.

As used herein, a "carbocyclic group" is any stable 3- to 7-membered monocyclic or bicyclic or 7- to 13-membered bicyclic or tricyclic group, any of which being It is intended to mean a group that is saturated, partially unsaturated or aromatic. In addition to those exemplified elsewhere in this document, examples of such carbocycles include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, cyclooctyl, [3.3.0] bicyclo Octyl, [4.3.0] bicyclononanyl, [4.4.0] bicyclodecanyl, [2.2.2] bicyclooctanyl, fluorenyl, phenyl, naphthyl, indanyl and tetrahydronaphthyl But it is not limited thereto.

As used herein, the term “heterocyclic group” refers to 1 to 3 (preferably fused) having at least one ring comprising an atom selected from N, O and S, which is 3 to about 8 members per ring. It is intended to include saturated, partially unsaturated or unsaturated (aromatic) groups with rings. Nitrogen and sulfur heteroatoms may be optionally oxidized. The term "heterocycloalkyl" is intended to denote a saturated heterocyclic group.

The heterocyclic ring is attached to its pendant group at any heteroatom or carbon atom to form a stable structure. The heterocyclic rings described in this document can be substituted on carbon or nitrogen atoms if the resulting compound is stable. Nitrogen in the heterocycle may be optionally quaternized. The term "aromatic heterocyclic system" as used herein refers to any stable 5- to 7-membered monocyclic including carbon atoms and 1 to 4 heteroatoms independently selected from the group consisting of N, O and S. Or 10- to 14-membered bicyclic heterocyclic aromatic ring systems. It is preferred that the total number of S and O atoms in the aromatic heterocyclic is not more than two, more preferably not more than one.

Examples of heterocycles include, but are not limited to, those exemplified elsewhere in this document, and in addition, acridinyl, azosinyl, benzimidazole, benzofuranyl, benzothiofuranyl, benzothiophenyl , Benzoxazolyl, benzthiazolyl, benztriazolyl, benztetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolinyl, carbazolyl, NH-carbazolyl, carbolinyl, chromanyl, cro Menyl, cynolinyl, decahydroquinolinyl, 2H, 6H-1,5,2-dithiazinyl, dihydrofuro [2,3-b] tetrahydrofuran, furanyl, furazanyl, imidazolidinyl , Imidazolinyl, imidazolyl, 1H-indazolyl, indolenyl, indolinyl, indolinyl, indolyl, 3H-indolyl, isobenzofuranyl, isochromenyl, isoindazolyl, isoindolinyl Isoindolyl, isoquinolinyl, isothiazolyl, isoxazolyl, morpholinyl, naphthyridinyl, jade Hydroisoquinolinyl, oxdiazolyl, 1, 2,3-oxadiazolyl, 1,2,4-oxadiazolyl; 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl , Oxazolidinyl, oxazolyl, oxazolidinyl, pyrimidinyl, phenantridinyl, phenanthrolinyl, phenazinyl, phenothiazinyl, phenoxatiinyl, phenoxazinyl, phthalazinyl, piperazinyl, piperidinyl Neil, Pterridinyl, Funinyl, Pyranyl, Pyrazinyl, Prazolidinyl, Pyrazolinyl, Pyrazolyl, Pyridazinyl, Pyridoxazole, Pyridimidazole, Pyridazole, Pyridinyl, Pyridyl , Pyrimidinyl, pyrrolidinyl, pyrrolinyl, 2H-pyrrolyl, pyrrolyl, quinazolinyl, quinolinyl, 4H-quinolininyl, quinoxalinyl, quinuclidinyl, tetrahydrofuranyl, tetrahydro Isoquinolinyl, tetrahydroquinolinyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thia Diazolyl, 1,3,4-thiadia Reel, thiantrenyl, thiazolyl, thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl, thiophenyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl , 1,2, 5-triazolyl, 1,3,4-triazolyl and xanthenyl.

Preferred heterocyclic groups include, but are not limited to, pyridinyl, pyrimidinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, pyrrolidinyl, morpholinyl, piperidinyl, piperazinyl and imidazolyl no. Such as fused rings and spiro compounds containing such heterocycles.

As used herein, the term “carbocyclic aryl” includes groups containing 1 to 3 isolated or fused rings and 6 to about 18 ring elements, which do not contain heteroatoms as reductions. Particularly preferred carbocyclic aryl groups include phenyl and naphthyl including 1-naphthyl and 2-naphthyl.

"Pharmaceutically acceptable carrier" refers to a biotissue compatible solution that has appropriate sterility, pH, isotonicity, stability, and the like, and includes any and all solvents, diluents (sterile saline, sodium chloride injection, Ringer's injection, dextrose) And sodium chloride injections, lactic ringer injections and other aqueous buffer solutions), dispersion media, coatings, antibiotics and antifungal agents, isotonic agents, and the like. In addition, pharmaceutically acceptable carriers may include stabilizers, preservatives, antioxidants or other additives known to those skilled in the art or known excipients.

As used herein, "pharmaceutically acceptable salts" refer to derivatives of the disclosed compounds wherein the parent compound is modified by the preparation of their non-toxic or basic salts. Examples of pharmaceutically acceptable salts include mineral or organic acid salts of basic residues such as amines; Alkali or organic salts of acidic residues such as carboxylic acids; And the like, but are not limited thereto. Pharmaceutically acceptable salts include, for example, conventional non-toxic salts or quaternary ammonium salts of the parent compound formed from non-toxic inorganic or organic acids. For example, conventional non-toxic salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitriic and the like; Acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic , Salicylic, mexilic, sulfanilic, 2-acetoxybenzoic, fumaric, toluenesulphonic, methanesulphonic, ethanedisulfonic, oxali, isethionic, HOOC- (CH ₂ salts prepared from organic acids such as) n-COOH and the like. Pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing basic or acidic groups. Generally, such salts react the free acid form of such compounds with stoichiometric amounts of suitable bases (Na, Ca, Mg or K hydroxides, carbonates, bicarbonates, etc.), or the free base forms of such compounds may be It can be prepared by reacting with stoichiometric amounts. Such reactions are typically carried out in water or organic solvents or mixtures thereof. In general, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile, which are feasible, are preferred.

Further suitable salts are described, for example, in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, PA, p. 1418 (1985).

Because of the correlation between increased or suppressed mitochondrial activity and various diseases and disorders, imaging agents capable of assessing changes in mitochondrial surface potential, and imaging methods using them, have various disease states associated with inducing or inhibiting apoptosis in cells. Is an effective diagnostic tool to test for the presence of Imaging agents suitable for use in imaging or evaluating changes in mitochondrial surface potentials are also used to study a variety of diseases including cancer, cardiovascular or epilepsy, HIV, AIDS, autoimmune diseases, degenerative disorders, neoplasms, and the like. Suitable for use

(Example)

The invention is illustrated in more detail by the following examples, which should not be construed in a limiting sense in any way. The contents of references (including cited documents, registered patents, and published patents) cited throughout this application are expressly incorporated herein by reference. The practice of the invention has been used in the art, unless otherwise indicated. Such techniques are explained in detail in the literature.

Example 1: [ ¹⁸ F] 3-fluoropropyltriphenyl-phosphonium ion ([ ¹⁸ F] FPTP)

Synthesis began with [ ¹⁸ F] fluoride from cyclotron targets transferred onto anion exchange columns (trap and discharge column (DW-TRC) D and W, Inc., Oakdale, TN, USA). The column was eluted with aqueous potassium carbonate (2.3 mg dissolved in 0.3 mL) into a 5 cc v-vial containing Kryptofix. Cryptofix, potassium carbonate, [ ¹⁸ F] fluoride mixture is dried at 120 ° C .; 7 mg of propyl ditosylate (Aldrich) was added to 0.5 mL acetonitrile. After 5 min heating at 80 ° C., 21 mg triphenylphosphine (Aldrich) in 0.5 mL toluene was added. Acetonitrile was evaporated and the toluene mixture was heated to boil for 3-5 minutes. After toluene was evaporated, the vial was cooled and 0.5 mL of high pressure liquid chromatography (HPLC) solvent [35: 65 acetonitrile: water (0.1 M ammonium formate)] was added to the vial. The mixture was filtered through a 0.45 μm Teflon HPLC filter and injected at 7 ml / min on an HPLC column (Waters Novapak C-18 6 URN, 7.8 × 300 mm) for purification. The product was collected on a modified rotary evaporator to allow the addition and removal of solvent, the HPLC solvent was evaporated, and the radiolabeled Fesponium salt was dissolved in sterile normal saline. The total decay correction radioactivity yield of [ ¹⁸ F] FPTP calculated from the starting [ ¹⁸ F] fluoride was 12 percent. After sterile filtration (PALL-Gelman 0.2 μm Tuffryn) into sterile vials, the solution is analyzed with known concentrations of non-radioactive standard solutions of characterized 3-fluoropropyltriphenyl-phosphonium bromide. Radiochemical, chemical purity and specific activity using HPLC [40: 60 acetonitrile: water (0.1 M ammonium formate), Waters Novapak C-18 60 μs 4 μm, 3.9 x 150 mm] at 3 ml / min ) Was investigated. [Physical data: mp 313-316 C; ¹ H NMR (CDCl 3, δ) 1.81-2.17 (m, 2H), 4.01-4.11 (m, 2H), 4.72-4.75 (m, 1H), 4.87-4.90 (m, 1H), 7.69-7.88 (m, 15H)]. Synthesis is summarized in Scheme 1.

Scheme 1: Synthesis of [ ¹⁸ F] alkyl phosphonium ions.

Example 2 [ ¹⁸ F] 2-Fluoroethyltriphenyl-phosphonium ion ([ ¹⁸ F] FETP)

Depending on the method used to produce the ^[18 F] FPTP described in Example 1 was prepared in the ^[18 F] FETP.

Example 3: [ ¹⁸ F] 2-fluorobutyltriphenyl-phosphonium ion ([ ¹⁸ F] FBTP)

Depending on the method used to produce the ^[18 F] FPTP described in Example 1 was prepared in the ^[18 F] FETP.

Example 4 [ ¹⁸ F] fluorobenzyltriphenylphosphonium ion

[ ¹⁸ F] fluoride in potassium carbonate / kryptofix (as described in Example 1) was collected, dried and trimethylammoniumbenzaldehyde triflate salt (7 mg) in 0.2 mL of dimethylsulfoxide (barium before use) Vacuum distillation from oxide) was added to the mixture.

After heating at 120 ° C. for 5 minutes, the aldehyde is diluted with 5 mL of HPLC water, recovered on a C-18 solid phase extraction cartridge (Waters C-18 PLUS Sep-Pak), then washed with 10 ml of HPLC water, and the cartridge The inert gas was passed through for 3 minutes to dry. The aldehyde was eluted from the cartridge using 2 mL of diethyl ether (Aldrich) and passed through 10% sodium borohydride (Aldrich-200-400 mg) on basic alumina to reduce it to alcohol. The alcohol was then converted to [ ¹⁸ F] fluorobenzyl bromide by mixing with triphenylphosphine dibromide (75-100 mg) in 1 mL of methylene chloride for 5 minutes. Passed through a silica solid phase extraction cartridge (Waters Silica Classic Sep-Pak), rinsed with 1 mL of methylene clyde, and then [ ¹⁸ F] fluorinated benzyl bromide 21 mg of tree dissolved in 0.5 mL toluene in a 5 ml v-vial. To phenylphosphine (ie analogues thereof). The methylene chloride / ether solvent was evaporated at low heat through an inert gas and the vial was capped and boiled for 3-5 minutes. After toluene was evaporated and the vial was cooled, 0.5 mL of high pressure liquid chromatography (HPLC) solvent [50: 50 acetonitrile: water (0.1 M ammonium formate)] was added. The mixture was filtered through a 0.45 μm Teflon HPLC filter (Altec 13 mm) and a preparative HPLC column (Waters Novapak C-18 6 μm, 7.8 × 300 mm) was performed at 7 ml / min for purification. The product was collected on a modified rotary evaporator to allow for the addition and removal of solvent, the HPLC solvent was evaporated and the radioactive phosphonium salt was dissolved in sterile normal saline. As in Example 1, after sterile filtration (PALL-Gelman 0.2 μm Tuffryn) into a sterile vial, the solution was analyzed by HPLC [40: 60 acetonitrile: water (0.1 M ammonium formate), Waters Novapak C- 18 60 × 4 μm, 3.9 × 150 mm] was used at 3 ml / min to investigate radiochemical, chemical purity and inactivity and compared to p-fluoropropyltriphenylphosphonium ion known standards. The total decay correction radioactivity yield of the [ ¹⁸ F] fluorbenzyltriphenyl phosphonium ion calculated from the starting [ ¹⁸ F] fluoride was 14 percent. [Physical data: mp 313-316 C; 1 H NMR (D6-DMSO, δ) 5.17-5. 21 (d, 2H), 6.99-7.08 (m, 4H), 7.67-7.94 (m, 15H)]. Synthesis is summarized in Scheme 2.

Scheme 2: Synthesis of [ ¹⁸ F] aryl phosphonium ions.

Example 5 Radiolabeled Diphenyl (Haloalkyl) ([ ¹⁸ F] fluorobenzyl) phosphonium ions

Radiolabeled haloalkylphosphonium ion derivatives were prepared by the synthesis of ¹⁸ F-fluorobenzyl halides as described in Example 4. Radiolabeled benzyl halide will be attached to diphenylphosphine to produce radiolabeled phosphine: The radiolabeled phosphine will then be converted to haloalkylphosphonium ions by reaction with a suitable haloalkyl iodine. . The synthesis of radiolabeled chloroalkyl phosphonium ions is summarized in Scheme 3. In addition, other haloalkyl species can be prepared, including chloro, bromo and iodine, attached to the alkyl chains of various lengths, branches and halide substituents.

Scheme 3: Synthesis of [ ¹⁸ F] -benzyl-chloroalkyl phosphonium ions.

Example 6 Radiolabeled [ ¹⁸ F] fluoroalkyl-tripyridylphosphonium ions and Radiolabeled [ ¹⁸ F] fluorobenzyl-tripyridylphosphonium ions.

Radiolabeled phosphonium ions comprising a pyridyl ring were prepared according to the generalized reaction scheme provided by Scheme 4. Radiosynthesis includes the reaction of ¹⁸ F-fluoroalkyl (Example 1) and benzyl (Example 4) groups as described in trippyridylphosphine.

Scheme 4: Synthesis of [ ¹⁸ F] -alkyl or aryl tripyridylphosphonium ions.

Example 7 Radiolabelled Ammonium Ion

The preparation process described in Examples 1-6 for the preparation of radiolabeled phosphonium ions is also applicable for the preparation of the quaternary ammonium ions described in Scheme 5. Quaternary ammonium ions have a biological distribution equivalent to phosphonium ions.

                R = phenyl and / or alkyl group

Scheme 5: Synthesis of Radiolabeled Ammonium Ion

Example 8 Whole Body PET / SPECT Imaging

One imaging protocol suitable for delivering salts of the present invention involves intravenously administering salts and acquiring a few minute stationary scan on the bed. The exact scan time may vary depending on the size of the patient, the salt dose and the nature of the tissue being imaged. However, imaging parameters for imaging primates, especially humans, can be modified as needed by those skilled in the radiation arts and those skilled in the art familiar with PET and / or SPECT imaging using other radiopharmaceutical agents.

Example 9 Stability in Plasma After Mouse Injection of Radiolabel and Agent of F-18-FBnTP.

Radiochemical purity of F-18-FBnTP and its stability in vivo were determined by chromatography.

Method: Preparation of F-18-fluorobenzyl triphenyl phosphonium (F-18-FBnTP):

The synthesis of labeling FBnTP with F-18 at the ortho position is shown schematically below. After F-18-fluoride was collected on the kryptofix (as in fluoroalkyl derivatives), nitrobenzaldehyde in acetonitrile was added to the mixture. After heating, the aldehyde was reduced to alcohol and then converted to radiolabeled benzylhalide. Fluorinated benzyl halides were reacted with triphenylphosphine or derivatives thereof in toluene. The mixture was purified as mentioned above and subjected to quality control.

Synthesis of o-F-18-fluorobenzyl triphenyl phosphonium ion

Mouse plasma samples were obtained from heparinized whole blood collection at 5, 15 and 30 minutes after tracer injection. To remove the binding of plasma proteins, solid urea was added to the plasma to a final urea concentration of 8M. Plasma-urea was loaded into a column switch HPLC system (Hilton 2000) in which plasma passed through a small capture column with lipophilic solutes (Oasis Sorbent, Waters Corp.), while the polar species did not bind and the positron detector Detected when passing through. After 4 minutes, the capture column is free of plasma proteins and polar species. The contents of the capture column were then sent onto an analytical column (Prodigy ODS-3, Phenomenex) by 40% acetonitrile, 60% triethylamine acetate buffer pH 4.1 at a rate of 1 mL / min, where the parent compound and Separation between lipophilic metabolites occurs. Eluate from the analysis column also passes through a positron flow detector. The proportion of each species is determined from the region below each chromatographic peak. Results: The radiochemical purity of ¹⁸ F-labeled FBnTP was over 95%. Chromatography of plasma showed no single peak and a single radio-peak containing 97% of total activity. 1 is for parent compound incubated for similar time (30 min) in plasma and buffered saline collected 30 min after injection. Plasma activity and parent compound peaked at the same time period (˜6 minutes).

This radiochromatogram showed that F-18-FBnTP can be labeled with high radiochemical purity and stable for at least 30 minutes after intravenous injection of the tracer in mouse in vivo.

Example 10 Stability of FBnTP Fluorination Process

Methods: Mice were injected with 25 μCi 18-PhC through the tail vein; Left or right femurs were removed 5-30 minutes post-injection and bone resorption with standard (injected dose 1: 10) was measured on a gamma counter. Parallel groups of mice were injected with free fluoride (F-18). Three mice in each group were studied at each time point. Radioactivity is expressed as a percentage of injected dose (% ID) and total bone uptake is calculated as activity x 20 in the femur.

Results: Figure 2 shows the total bone resorption at 5 and 30 minutes after injection. The lowest bone uptake of F-18-PhC compared to the bone uptake in mice injected with F-18 only indicates the stability of fluorination of the phosphonium compound. Minimum bone uptake of F-18-FBnTP indicates that no free fluoride is present, indicating the stability of fluorination.

Example 11 Mitochondrial Membrane Potential (MMP) -dependent Uptake

MMP-dependent cellular uptake of F-18-FBnTP was assessed using CCCP, a known protonophore that selectively disrupts MMP.

Methods: Human lung cancer cells H549 (10 / ML) were incubated with 0.1 μCi / ml F-18-FBnTP for 30 minutes. A suspension sample (1 mL) was transferred to an Eppendorf vial and placed at 37 ° C. CCCP was added to the suspension at various concentrations (30, 60, 90, 120 μΜ). After 30 minutes incubation with CCCP, the Eppendorf vial was centrifuged for 1 minute and the activity in the pellet and supernatant was measured immediately.

Results: Figure 3 shows the cellular uptake of F-18-FBnTP in the presence of various concentrations of CCCP. CCCP induces a dose-dependent decrease in F-18-FBnTP cell uptake. Most (86%) of F-18-FBnTP cell uptake is MMP-dependent.

Example 12: In vivo distribution of novel F-18-fluorophosphonium cations compared to various tracers.

Methods: In vivo distribution of novel phosphonium compound F-18-fluorobenzyl triphenyl phosphonium (FBnTP) and F-18-fluoropropyl triphenyl phosphonium cation (FPTP) was studied in adult mice and tracer C-11 -Triphenylmethyl phosphonium (TPMP), tetraphenyl phosphonium (TPP) and Tc-99m-setamibi (MIBI). F-18-FBnTP was prepared as described above. The preparation of FPTP is described next. TPMP was prepared as previously described (Madar I, J Nucl Med. 1999; 40: 1180-5). TPP and MIBI were purchased from NEN and Dupont, respectively. Three to five mice were used to study the in vivo distribution of each tracer. Animals that were not anesthetized were injected intravenously with tracer solution (FBnTP or FPTP, TPMP 25 μCi; MIBI 40 μCi; TPP 2 μCi, all in a volume of 0.2 mL saline), followed by neck thirty minutes after injection. Dislocated and died. The organs and tissues of interest were removed and measured using a radioactivity counter with a standard (1: 100) (LKB Wallac, 1282 Compugamma CS).

Preparation of F-18-fluoropropyl triphenyl phosphonium ion (FPTP):

A schematic illustration of F-18-FPTP synthesis is shown below. F-18-FPTP was prepared from 1-fluoro-3-bromopropane and triphenylphosphine as fluoropropyltriphenyl phosphonium bromide which is a non-radioactive compound. Radiolabelled compounds were characterized by proton and carbon-13 NMR as well as HPLC. This standard was used as a comparison during the purification, quality control and measurement of inactivation of radiolabeled F-18-FPTP. F-18-FPTP was synthesized as described in FIG. 3. Briefly, 1,3-ditosylpropane in acetonitrile was added to a dried vial containing F-18-fluoride, kryptofix and potassium carbonate. After heating at 80 ° C. for 4 minutes on a heat gun, triphenylphosphine in toluene was added to the vial. After 5 minutes, the volume of the solution decreased to the level of several microliters. After cooling to room temperature, HPLC solvent was added to the vial. The mixture was filtered through a 0.45 micron filter and injected on a semi-preparative HPLC column. The product F-18-FPTP was collected, the solvent was evaporated and the remaining dry F-18-FPTP dissolved in sterile normal saline. The solution was filtered through a 0.22 micron filter into a sterile empty vial. Aliquots were removed to determine chemical and radiochemical purity by analytical HPLC. Inactivation was also measured.

Synthesis of F-18-fluoropropyltriphenyl phosphonium ion

Results: The biodistribution of the tracer is shown in Table 1. A comparative study of biodistribution in mice showed that F-18-FBnTP works better than C-11-TPMP as a PET perfusion agent. F-18-FBnTP uptake in the heart was significantly higher than other tracers even C-11-TPMP, while removal from blood was as good as C-11-TPMP. Many researchers have reported that apoptosis plays an important role in the pathogenesis of acute heart muscle infarction and other forms of heart failure. These data suggest that it has potential utility in assessing cardiomyopathy, including apoptosis associated with MMP function as well as as a cardiovascular perfusion tracer.

TABLE 1 In vivo distribution of fluorophosphonium cations at 60 minutes post mouse intravenous injection compared to various tracers.

Example 13. Differentiation of Tumor from Inflammation Using F-18-Phosphorium Cation.

Carcinogen Nitrosomethylurea (NMU) causes cancer tumors in the mammary gland of female rats. Thus, orthotopic NMU breast cancer is an excellent model for assessing tumor tracer selectivity by contrasting radioactivity accumulating in the mammary gland where cancer cells are concentrated and in the healthy mammary gland. Freund's complete adjuvant (FCA) is an inflammatory agent that has been studied a lot in rats. By inducing FCA inflammation in NMU-bearing rats, the tracer ability to distinguish inflammation from cancer can be directly quantified.

Methods: 0.1 ml of nitrosomethyl urea (NUM) (150 g) was injected intraperitoneally into female rats. When tumors reached a size close to 1-1.5 cm, FCA (0.15 g) was injected into each of the posterior leg. Absorption analysis was performed after 3 days. F-18-FBnTP (0.25 mCi) was injected through the tail vein. After 60 minutes the tumor, inflammatory tissue and healthy muscle tissue (control) on the opposite leg were collected on ice, weighed and measured at the gamma counter along with the standard.

Results: Figure 3 shows F-18-FBnTP activity in malignant mammary gland (tumor), healthy mammary gland (control mammary gland), muscle normalized inflammation (inflammatory). F-18-FBnTP uptake in tumors was four times higher than in healthy mammary glands and three times higher in inflammation. The differential uptake of F-18-FBnTP in mammary tumors in healthy mammary and inflammatory muscles is evidence of the effectiveness of the present invention.

Novel F-18-phosphonium cations, F-18-FBnTP and F-18-fluoropropyl triphenyl phosphonium (F-18-) compared to F-18-FDG in FCA-induced inflammation and healthy tissue FPTP) is shown in Table 2.

Fluorophosphonium compounds accumulate less than FDG in inflammation (Table 2). These data provide strong evidence of the claims that the F-18-phosphonium compound is suitable for distinguishing between inflammation and tumors and solves the major disadvantage of F-18-FDG.

Table 2: Accumulation of Fluorophosphonium Compounds (FBnTP and FPTP) FDG in Inflammatory Tissues and Healthy Muscles Three Days After FCA Administration

Example 14 Detection of Lung Cancer Tumor Response to Chemotherapy TAXETER In Vivo

Methods: 2 x 10 6 human lung cancer A549 cells were inoculated into 12 nude mice. When the tumor size approached the size of 5-10 mm, 6 mice were injected intravenously with texsetters and 6 mice were used as controls. Absorption analysis was then performed 48 hours later. 25 μCi of F-18-FBnTP was injected intravenously and the tumor and muscle tissues dissected after 60 minutes.

Results: Figure 5 shows the tumor activity normalized to muscle. Texecter reduced F-18-FBnTP accumulation in tumors by nearly 50% compared to non-treated mice.

Example 15 Detection of Apoptosis Induced by Androgen Depletion

Methods: Male mice were castrated and uptake assayed 4 days later. Ventral (VP), anterior (AP), and dorsal lateral (DLP) lobes of the prostate gland were excised with posterior leg muscles. After measuring tracer activity in tissue samples, TUNEL staining was performed and apoptotic cell sections in the ventral and anterior lobe were measured.

Results: The data in FIG. 6 is the mean of 8 control rats normalized to 9 treatments and muscles. Castration induced a lobe-specific decrease in the ventral side but not in the anterior and dorsal lobes. This result is in line with the fraction of apoptotic cells as measured in the stained histological sections. In the VP lobe, 12.4% ± 3.8% of the cells showed DNA laddering compared to only 3.3% ± 1.7% of the AP lobe cells (see Figure 9) (see Figure 8).

Example 16: F-18-FBnTP Selectivity for Prostate Cancer

The data shown above show that F-18-PhC can detect apoptosis processes in all animals. In addition, in the prostate, changes in F-18-PhC accumulation correlate with the degree of apoptosis in target tissues. These data suggest the suitability of F-18-PhC as a PET tracer to measure the efficacy of chemotherapy in prostate cancer and other types of cancer. However, the tracer's ability to accurately report the degree of apoptosis may depend on the tracer's selectivity for the tumor. To address this issue, tumor selectivity of F-18-FBnTP was studied in a stereotactic model of prostate cancer.

Methods: 2 × 10 6 cells were injected under anesthesia into the prostate epithelial tissue of nude mice. When tumors approached the size of 5 mm, uptake analysis was performed as described above. Tumor selectivity of F-18-FBnTP and F-18-FDG was compared (3 mice in each group).

Results: Figure 9 shows the accumulation of tracers in normal and malignant prostate, normalized to muscle. The rate of absorption of malignant-to-normal prostate tissue was 2.5 for F-18-Phc and 1.25 for FDG. These data also further demonstrate that F-18-PhC is suitable for detecting prostate cancer and measuring response to treatment.

Example 17 C-11-TPMP Absorption Kinetics in Cardiac Muscle

We tested the ability of C-11-triphenyl phosphonium cations (TPMP) to measure partial cardiac muscle flow in dogs using PET (Kraus, 1994).

Methods: 4 mCi of C-11-TPMP was introduced intravenously and dynamic images of increased duration (15 seconds to 20 minutes) were obtained over a total of 85 minutes. Images were taken with a GE 4096+ PET scanner (15 slices, 6.5 mm slice thickness). Images were reconstructed using backprojection and corrected for attenuation. A detailed description of the method is disclosed in Kraus (1994).

Results: The axial cuts of the heart at 5, 30 and 60 minutes after injection are shown in FIG. 10. These images provide good visualization of the heart muscle with high contrast to surrounding lung tissue. The heart / lung ratio was> 14: 1 and the heart / blood ratio was> 100: 1 during the period of plateau time.

FIG. 11 shows the extracted fraction of C-11-TPMP in the dog heart as a function of heart muscle blood flow. Under basal conditions (blood flow = 69 ml / min / 100g), the extraction fraction was very high (91%). A 5-fold increase in blood flow (by adenosine) reduced 39% of the extract fraction.

To investigate the relationship between cardiac muscle blood flow and C-11-TPMP uptake in the heart, LAD was blocked and C-11-TPMP accumulation in tissue samples compared to microspheres determined partial cardiac muscle blood flow. A significant correlation (r = 0.93, p <0.01) was found 5 minutes after LAD closure (FIG. 12). The non-cracked / cracked heart muscle ratio was 12.1 ± 2.4.

These data point to the superior properties of phosphonium cations as perfusion agents in assessing heart muscle blood flow, compared to the SPECT perfusion agents currently used. Under basal flow, thallium 201 extraction was about 80% and decreased about 60% for a five-fold increase in flow rate. MIBI extraction for normal flow rate was about 60% and decreased to 40% for high flux. The benefits of C-11-TPMP PET technology for these SPECT agents include: (1) totally high cardiac muscle extraction of C-11-TPMP; (2) an extended retention period of C-11-TPMP in the myocardium; And (3) better temporal and spatial resolution of PET scanners for excellent graphical data of ischemic sites in the myocardium.

Example 18 F-18-FBnTP Absorption Kinetics in Cardiac Muscle

Methods: Mongrel dogs (BW = 35 kg) were injected with 3-18 mCi of F-18-FBnTP. Images were taken on a GE 4096 + scanner (15 slices, 6.5 mm slice thickness). PET scans of increased duration (15 seconds to 20 minutes) were obtained over a total time of 85 minutes post injection. Arterial blood samples (0.5 ml of volume) were collected every few seconds for the first 3 minutes and at increasing intervals (1-10 minutes) for the remaining time of the imaging study. The vascular and myocardial kinetics of F-18-FBnTP were analyzed using the ROI method.

Results: F-18-FBnTP demonstrated rapid removal from the blood pool (see FIG. 13). F-18-FBnTP accumulated rapidly in the myocardium, reached equilibrium in minutes and continued for the rest of the scanning time (see FIG. 14). Rapid absorption was seen in both the left and right ventricles (see FIG. 15). In contrast, F-18-FBnTP demonstrated rapid removal in the atria as well as in adjacent lungs (see FIG. 15). In the period of 60-85 minutes after injection, the ratio of myocardium to atrial and lung was> 15: 1. As a result, F-18-FBnTP provided high contrast cardiac images with good visible clarity (fig. 16).

Example 19 Myocardial Accumulation of F-18-FBnTP in Heart Failure.

The cardiac face of the mongrel at 4 weeks high speed (210 bpm) is an excellent model of heart failure. The advantage of this model is that cardiomyopathy only includes apoptosis without coronary artery stenosis or associated ischemia. Thus, in this model the effect of apoptosis on FBnTP uptake can be analyzed in detail.

Methods: Dog preparation and data acquisition were performed as mentioned above. The mongrel was installed with a face maker in the rib case and a FBnTP PET scan (shown above) was performed. Following the baseline scan, the dog heart was paced at 210 bpm for 4 weeks and a second scan was obtained.

Results: FIG. 17 shows mono-axial images of FBnTP before and after 4-week pacing. Facing causes a significant reduction in FBnTP uptake of 40-60% through most inferior walls (Figures 18-19, Table 3). Although the PET scan was not open, pacing-induction of cardiac muscle typical of heart failure, including thinning of the ventricular wall and dilatation of the left ventricular chamber, due to FBnTP's good blood flow capacity and thus good transparency of the left ventricular wall heart muscle. Remodeling was clearly observed.

In addition, a marked reduction in the accumulation of F-18-FBnTP throughout the entire inferior wall of pacing induction indicates an increased heart failure process that mediates apoptosis of cardiac cells in this region. 18 shows co-recorded images before and after pacing. There is a marked decrease in F- ¹⁸ FBnTP absorption in the inferior walls.

Quantification of FBnTP in cardiac muscle was performed using the portion of interest located on co-recorded cardiac muscle images, whose activity was normalized to the dose injected before and after pacing. An example of the ROI location is shown in FIG. 18.

Cardiac muscle pacing resulted in a significant reduction (p <0.01) of 40-60% in inferior walls (see Table 3).

Table 3: F-18-FBnTP cardiac muscle uptake before and after pacing.

See FIG. 18 for the location of the ROI. Data was derived as shown in FIG. 19.

* Average activity (% of dose injected) accumulated over the mean of the activity of the indicated coronal slices, for 36 to 85 minutes per slice.

The location of the ROI as described above is above the left image.

Templates of the same ROIs were used to retrieve all the data.

The disclosures of the documents and references mentioned in this application, including patents, are incorporated herein by reference.

The present invention, methods of making and using the same, and manufacturing processes are described in complete, clear, concise, and accurate terms so that those skilled in the art can make and use the same invention. The foregoing has described preferred embodiments of the invention, and it should be understood that modifications may be made without departing from the spirit or scope of the invention as set forth in the claims. To specifically point out and explicitly claim the subject matter of the present invention, the following claims are set forth as the conclusion of the above detailed description.

Claims (56)

A salt comprising at least one pharmaceutically acceptable anion and at least one cation according to formula (I). Formula I Where E is phosphorus or nitrogen; X ¹ , X ² , X ³ and X ⁴ are independently selected from the group consisting of Ar and R, wherein at least ^one of X ¹ , X ² , X ³ and X ⁴ is an Ar group; Ar is optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl; R is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more radioisotope Contains an element. The method of claim 1, Ar is an optionally substituted aryl having 6 to 18 carbon atoms and 1 to 3 rings, 3 to about 18 carbon atoms, 1 to about 3 rings and 1 to about 4 rings selected from N, O and S Optionally substituted heteroaryl having heteroatoms, and optionally substituted aralkyl having 7 to about 12 carbon atoms; R is optionally substituted C _1-6 alkyl, optionally substituted C _2-6 alkenyl, optionally substituted C _2-6 alkynyl, optionally substituted C _1- with one or more F, Cl, Br, or I atoms ₆ haloalkyl, optionally substituted cycloalkyl having 3 to about 8 ring carbon atoms, optionally substituted aralkyl having 7 to about 12 carbon atoms, wherein at least one instance of R represents one or more radioactive isotopes Salts containing. The salt of claim 1 wherein E is phosphorus. The salt of claim 1, wherein E is nitrogen. The salt of claim 1, wherein said R group is one or more positron radioactive isotopes. The salt of claim 1, wherein said R group comprises one or more isotopes selected from ¹¹ C, ¹⁸ F, ⁷⁶ Br, ¹²³ I or any combination thereof. A salt according to claim 1 wherein said cation is according to formula (II). Formula II Wherein E is phosphorus or nitrogen; Ar ¹ , Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl; R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one It includes the above radioisotope. The method of claim 7, wherein Ar ¹ , Ar ² , and Ar ³ are optionally substituted aryl having 6 to 18 carbon atoms and 1 to 3 rings, 3 to about 18 carbon atoms, 1 to about 3 rings, and N, O and Independently selected from the group consisting of optionally substituted heteroaryl having 1 to about 4 ring heteroatoms selected from S and optionally substituted aralkyl having 7 to about 12 carbon atoms; R ¹ is optionally substituted C _1-6 alkyl, optionally substituted C _2-6 alkenyl, optionally substituted C _2-6 alkynyl, optionally substituted C ₁ having one or more F, Cl, Br or I atoms _-6 haloalkyl, optionally substituted cycloalkyl having 3 to about 8 ring carbon atoms, optionally substituted aralkyl having 7 to about 12 carbon atoms, wherein at least one instance of R represents one or more radioactive isotopes Include. The salt of claim 8, wherein E is phosphorus. The salt of claim 8, wherein E is nitrogen. The compound of claim 8, wherein R ¹ is selected from the group consisting of ¹¹ C-methyl, optionally substituted C _1-6 alkyl, optionally substituted C _7-12 aralkyl, optionally substituted C _6-12 aryl, and Each salt substituted with one or more ¹¹ C-methyl, ¹¹ C-methoxy, ¹⁸ F, ⁷⁶ Br, ¹²³ I, ¹²⁵ I, ¹³¹ I or a combination thereof. The method of claim 8, wherein R ¹ is a benzyl, substituted by a ¹¹ C- salt methyl, at least one ¹⁸ F A C _2-6 alkyl or one or more substituted with ¹⁸ F. The method of claim 8, wherein R ¹ is a benzyl, substituted by a ¹¹ C- salt methyl, at least one ¹⁸ F A C _2-6 alkyl or one or more substituted with ¹⁸ F. 8. The salt of claim 7, wherein R ¹ comprises at least one radioisotope suitable for use in radiation therapy. The method of claim 1, Wherein said cation is a salt according to formula III. Formula III Wherein E is phosphorus or nitrogen; p, q and r are independently selected from a number from 0 to about 5; R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more radioactive Isotopes; R ² , R ³ and R ⁴ are, in each case of R ² , R ³ and R ⁴ , hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally Independently selected from the group consisting of substituted alkoxy, optionally substituted (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides do. The salt of claim 15, wherein R ¹ is a halo-C _2-6 alkyl group or halobenzyl group. The compound of claim 6, wherein R ¹ is an ω-fluoro-C _2-6 alkyl, ω-iodine-C _2-6 alkyl group, ortho, meta or para-fluoro benzyl group or ortho, meta or para-iobenzyl Attributed salts. A salt according to claim 1 wherein said cation is according to formula IV. Formula IV Wherein E is phosphorus or nitrogen; Ar ¹ , Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl; R ⁵ and R ⁶ in each case of R ⁵ and R ⁶ are selected from the group consisting of hydrogen, halogen, hydroxy, amino, optionally substituted alkyl, optionally substituted haloalkyl and optionally substituted alkoxy; X is ¹¹ C-methyl or a radioisotope of fluorine or iodine; m is a number from about 2 to about 6. The method of claim 1, Salts according to formula (V). Formula V Wherein E is phosphorus or nitrogen; Ar ¹ , Ar ² and Ar ³ are independently selected from the group consisting of optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl; X is ¹¹ C-methyl or a radioisotope of fluorine or iodine; M is a number from about 1 to about 5. The method of claim 1, ¹¹ C-methyl-triphenylphosphonium ion, ¹¹ C-methyl-tri-ortho-tolylphosphonium ion, ¹¹ C-methyl-tri-meth-tolylphosphonium ion, ¹¹ C-methyl-tri-para-tolylphosphonium ion, ¹⁸ F-2-fluoroethyl-triphenylphosphonium ion, ¹⁸ F-2-fluoroethyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-2-fluoroethyl-tri-meth-tolylphosphonium ion, ¹⁸ F-2-fluoroethyl-tri-para-tolylphosphonium ion, ¹⁸ F-3-fluoropropyl-triphenylphosphonium ion, ¹⁸ F-3-fluoropropyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-3-fluoropropyl-tri-meta-tolylphosphonium ion, ¹⁸ F-3-fluoropropyl-tri-para-tolylphosphonium ion, ¹⁸ F-4-fluorobutyl-triphenylphosphonium ion, ¹⁸ F-4-fluorobutyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-4-fluorobutyl-tri-meth-tolylphosphonium ion, ¹⁸ F-4-fluorobutyl-tri-para-tolylphosphonium ion, ¹⁸ F-2-fluorobenzyl-triphenylphosphonium ion, ¹⁸ F-2-fluorobenzyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-2-fluorobenzyl-tri-meta-tolylphosphonium ion, ¹⁸ F-2-fluorobenzyl-tri-para-tolylphosphonium ion, ¹⁸ F-3-fluorobenzyl-triphenylphosphonium ion, ¹⁸ F-3-fluorobenzyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-3-fluorobenzyl-tri-meta-tolylphosphonium ion, ¹⁸ F-3-fluorobenzyl-tri-para-tolylphosphonium ion, ¹⁸ F-4-fluorobenzyl-triphenylphosphonium ion, ¹⁸ F-4-fluorobenzyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-4-fluorobenzyl-tri-meta-tolylphosphonium ion, ¹⁸ F-4-fluorobenzyl-tri-para-tolylphosphonium ion, ¹⁸ F-3-fluoro-4-formyl-benzyl-triphenylphosphonium ion, ¹⁸ F-3-fluoro-4-formyl-benzyl-tri-ortho-tolylphosphonium ion, ¹⁸ F-3-fluoro-4-formyl-benzyl-tri-meta-tolylphosphonium ion, ¹⁸ F-3-fluoro-4-formyl-benzyl-tri-para-tolylphosphonium ion, ( ¹⁸ F-4-fluorobenzyl)-(2-chloroethyl) -diphenylphosphonium ion, ( ¹⁸ F-4-fluorobenzyl)-(3-chloropropyl) -diphenylphosphonium ion, ( ¹⁸ F-4-fluorobenzyl)-(4-chlorobutyl) -diphenylphosphonium ion, ( ¹⁸ F-4-fluorobenzyl)-(6-chloropentyl) -diphenylphosphonium ion, ( ¹⁸ F-4-fluorobenzyl)-(5-chlorohexyl) -diphenylphosphonium ion, ¹⁸ F-2-fluoroethyl-tri (4-pyridyl) phosphonium ion, ¹⁸ F-3-fluoropropyl-tri (4-pyridyl) phosphonium ion, ¹⁸ F-4-fluorobutyl-tri (4-pyridyl) phosphonium ion, ¹⁸ F-2-fluorobenzyl-tri (4-pyridyl) phosphonium ion, ¹⁸ F-3-fluorobenzyl-tri (4-pyridyl) phosphonium ion, ¹⁸ F-4-fluorobenzyl-tri (4-pyridyl) phosphonium ion and A salt comprising a pharmaceutically acceptable anion and cation selected from the group consisting of ¹⁸ F-3-fluoro-4-formyl-benzyl-tri (4-pyridyl) phosphonium ions. The salt of claim 1 wherein said salt exhibits a ratio of mitochondria to non-mitochondria of at least about 5: 1. The salt of claim 21, wherein said salt exhibits a ratio of dysfunctional mitochondria to normal mitochondria of at least about 5: 1. The salt of claim 1 suitable for use in positron emission tomography (PET) or single photon emission computed tomography (SPECT). The salt of claim 1 having bio-distribution related to mitochondrial integrity. The method of claim 1, Salts in which the cation is according to formula VI: Formula VI Wherein E is phosphorus or nitrogen; Z is chloro, fluoro, hydroxy or methoxy; n is a number from 1 to about 12; p and q are independently selected from 0 to about 5; R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more isotopic An element; R ² and R ³ in each instance of R ² and R ³ are hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alkoxy, optionally substituted Independently selected from the group consisting of (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides. The method of claim 1, Salts according to formula (VII). Formula VII Wherein E is phosphorus or nitrogen; p, q and r are independently selected from a number from 0 to about 4; R ¹ is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein at least one instance of R is one or more Radioactive isotopes; R ² , R ³ and R ⁴ are, in each case of R ² , R ³ and R ⁴ , hydrogen, halogen, cyano, nitro, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally Independently selected from the group consisting of substituted alkoxy, optionally substituted (cycloalkyl) alkyl, optionally substituted alkylthio, optionally substituted alkylsulfinyl or optionally substituted alkylsulfonyl and optionally substituted mono or dialkylcarboxamides do. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and the salt of claim 1. A pharmaceutical composition as defined in claim 27 is contained in a container, further comprising: Instructions for use of the composition to image cells or tissues with increased or inhibited mitochondrial activity, Instructions for use of the composition to evaluate the therapeutic effect of a drug protocol administered to a patient, Instructions for use of the composition to selectively image malignant cells and tumors in the presence of inflammation and A package comprising instructions including one or more of the instructions for use of the composition for measuring mitochondrial membrane potential (ΔΨm). Providing a radiolabeled salt comprising at least one pharmaceutically acceptable anion and at least one cation according to formula (I)     Formula I     Wherein E is phosphorus or nitrogen; X ¹ , X ² , X ³ and X ⁴ are independently selected from the group consisting of Ar and R, wherein at least one of X ¹ , X ² , X ³ and X ⁴ is an Ar group;     Ar is optionally substituted aryl, optionally substituted heteroaryl and optionally substituted aralkyl;     R is optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted haloalkyl, optionally substituted cycloalkyl, optionally substituted aralkyl, wherein one or more instances of R are one or more radioactive Includes isotopes; Contacting the cell or tissue with a radiolabeled salt; And An imaging method comprising making a radiographic image. The method of claim 29, suitable for measuring mitochondrial membrane potential (ΔΨm). The method of claim 30, wherein the imaging method is suitable for the measurement of inhibited or increased apoptosis. The method of claim 30, wherein the imaging method is suitable for imaging pathological conditions associated with increased or inhibited apoptosis. 33. The method of claim 32, suitable for imaging a disease associated with inhibited apoptosis. 33. The method of claim 32, suitable for imaging a disease associated with increased apoptosis. 33. The method of claim 32, wherein the imaging method is suitable for imaging a disease or disorder associated with myopathy or mitochondrial dysfunction due to cancer, degenerative disease or disorder, autoimmune disorder, aging, HIV transmission, oxidative stress or DNA mutation. The method of claim 29, suitable for imaging use of mitochondrial dysfunction. The method of claim 29, wherein the imaging method is suitable for assessing the efficacy of a therapeutic drug capable of inducing or inhibiting apoptosis. 38. The method of claim 37, wherein the imaging method is suitable for evaluating the efficacy of a therapeutic protocol. The method of claim 38, wherein said therapeutic protocol is selected from drugs, radiation, and combinations thereof. The method of claim 29, wherein the imaging method is suitable for imaging tumors. 41. The method of claim 40, wherein the radiolabeled salt preferentially accumulates in the mitochondria of malignant cells. The method of claim 29, wherein the imaging method is suitable for imaging cancer. The method of claim 42, wherein the degree of cancer disease (cancer development stage) can be measured. The method of claim 42, wherein said radiolabeled salt preferentially accumulates in the mitochondria of cancer cells. 43. The method of claim 42, wherein said cancer is a neoplasm. 43. The method of claim 42, wherein said cancer is lung cancer. The method of claim 42, wherein the tissue inflammation and tumor can be distinguished. The method of claim 29, wherein the imaging method is suitable for developing new therapeutic agents that can disrupt mitochondrial function in target tissue. 30. The method of claim 29, wherein the radiolabeled salt exhibits a target ratio to non-target of at least about 5: 1. The method of claim 29, wherein the radiolabeled salt is stable in vivo. The method of claim 29, wherein said radiolabeled salt is substantially localized at a location or locations with dysfunctional mitochondria within about 120 minutes after administration. 30. The method of claim 29, wherein said radiolabeled salt is substantially localized at a location or locations with dysfunctional mitochondria within about 60 minutes after administration. 30. The method of claim 29, wherein said radiolabeled salt is substantially localized at a location or locations with dysfunctional mitochondria within about 30 minutes after administration. 30. The method of claim 29, wherein said radiolabeled salt is detected by a gamma camera, positron emission tomography (PET) or single photon emission computed tomography (SPECT). The method of claim 29, wherein the subject is a human, rat, mouse, cat, dog, horse, sheep, cow, monkey, bird, or amphibian. An imaging agent comprising the radiolabeled salt of any one of claims 1-26 or the pharmaceutical composition of claim 27.