WO1996035456A2

WO1996035456A2 - Use of polymeric contrast agents of medium molecular weight for differentiating between benign and malignant tumours using modern imaging techniques

Info

Publication number: WO1996035456A2
Application number: PCT/EP1996/001740
Authority: WO
Inventors: Heribert Schmitt-Willich; Johannes Platzek; Andreas Mühler; Gerhard Adam; Rolf Günther
Original assignee: Schering Aktiengesellschaft
Priority date: 1995-05-11
Filing date: 1996-04-26
Publication date: 1996-11-14
Also published as: DE19518222A1; ZA963749B; AU5692596A; WO1996035456A3; IL118214A0

Abstract

It has been found surprisingly that by using metal complex-conjugates with a polymeric basic structure and a mean molecular weight of between 10,000 and 40,000 dalton, it is possible to otbtain excellent differentiation between malignant and benign tumours in areas of the body without a blood-brain barrier. Contrast agents of this type not only permit reliable differentiation between malignant and benign tumours, they are also characterised by the fact that they can be completely eliminated from the body and thus have good tolerability. Suitable metal complex-conjugates with a polymeric basic structure and a means molecular weight in the range indicated above are disclosed in the German patent applications DE 39 38 992 and DE 195 25 924, as well as in the US patents US 5,338,532; 4,507,466; 4,631,337; 4,587,329; 4,568,737 and 4,507,466. Such polymeric basic structures are referred to as cascades or dendrimers, and contrast agents containing conjugates of these polymers with metal complexes can be referred to as cascade or dendrimeric contrast agents. Preference is given in particular to metal complex-conjugates carrying between 24 and 36 paramagnetic metal complexes, in particular metal complexes of gadolinium, at the end-position groups of the polymeric basic structure.

Description

Use of medium molecular weight polymeric contrast agents to differentiate benign and malignant tumors using modern imaging techniques

The invention relates to the subject characterized in the claims, that is, the use of paramagnetic metal complex conjugates with a polymeric backbone and a molecular weight of 10,000 to 40,000 daltons in modern imaging methods for differentiating between malignant and benign tumors.

In developed industrial countries, malignant tumors occupy second place in the type of death statistics (Statistical Yearbook 1991 for United Germany, ed .: Statistisches Bundesamt, Verlag Metzler-Poeschel (1992). In 1989, there were more than 200,000 people with malignant tumors in the Federal Republic of Germany They represent one of the most important medical problems. Because of the further development of the methods for the treatment of malignant tumors (eg percutaneous, interventional, and non-invasive therapies), it is becoming increasingly important for diagnostic methods to identify such malignant (malignant) Benign (benign) tumors in good time, reliably and above all without being able to differentiate the (otherwise necessary) use of biopsies and sample excisions.The modern imaging methods (ultrasound, X-ray CT, magnetic resonance imaging, scintigraphy) fail to differentiate between malignant and benign Tumors. That is why Contrast agents are used in all such imaging processes in order to obtain information on the dignity (well-being) of the tumor from the contrast agent kinetics in the tumor tissue.

The attempt to improve the meaningfulness of the images obtained by using different contrast media was - as discussed below - only partially successful.

The contrast agents currently clinically used in the magnetic resonance imaging such as Gd-DTPA (Magnevist ^®), spread out in general in the entire extracellular space of the body (intravascular space and interstitium). This distribution space covers about 20% of the body volume (Vss = 0.20 1 / kg).

A special situation with regard to the regional distribution area arises, however, in the cerebral and spinal area, since extracellular contrast agents in healthy brain tissue cannot leave the intravascular space due to the blood-brain barrier. In pathological processes with a disturbance of the blood-brain barrier (e.g. in malignant tumors), regions with increased blood vessel permeability (permeability) then arise within the brain (spinal cord) for these extracellular contrast agents (Schmiedl et al., MRI of blood-brain barrier permeability in astrocytic gliomas: application of small and large molecular weight contrast media, Magn. Reson. Med. 22: 288, 1991), whereby diseased tissue can be recognized. For these special areas (brain / spinal cord) there is thus the possibility of differentiating areas with highly permeable vessel walls from such areas without increasing permeability.

However, there is no such permeability barrier for small, extracellular contrast agents outside the brain and spinal cord (Canty et al., First-pass entry of nonionic contrast agent into the myocardial extravascular space. Effects on radiographic estimate of transit time and blood volume. Circulation ,

1991, 84: 2071-2078). In these areas, the enrichment of an extracellular contrast agent depends primarily on the vascularization and the size of the extracellular space in the corresponding tissue. As is known, both the vascularization and the size of the extracellular space do not differ in malignant and benign tumors; consequently, extracellular contrast media do not allow differentiation from malignant and benign tumors in clinical use (SH Heywang-Köbrunner, Contrast-enhanced magnetic resonance imaging of the breast, Invest. Radiol. 1994, 29: 94-104; SH Heywang, MR imaging of the breast with Gd-DTPA: use and limitations, Radiology 1989, 171: 95-103). Such extracellular contrast media are therefore particularly suitable for the detection (detection) of tumors.

However, a permeability barrier outside the brain exists for polymeric substances that cannot diffuse there from the intravascular space into the interstitium (Shames et al., Measurement of capillary permeability to macromolecules by dynamic magnetic resonance imaging: a quantitative noninvasive technique, Magn. Reson. Med. 29: 616, 1993). Such so-called blood pool contrast agents should therefore be suitable in the whole body to selectively display diseased tissue with increased vascular permeability - similar to the extracellular contrast agents in the brain (Brasch et al., Pulmonary oxygen toxicity: demonstration of abnormal capillary permeability using contrast-enhanced MRI, Pediatric Radiology 23: 495, 1993). Gd-DTPA-albumin with a molecular weight of 85,000 Daltons (U. Schmiedl et al., Albumin labeled with Gd-DTPA as an intravascular, blood pool-enhancing agent for MR imaging: biodistribution and imaging studies. Radiology, 1987, 162: 205-210) and Gd-DTPA-polylysine with 50,000 Daltons (Schuhmann-Giampieri et al., In vivo and in vitro evaluation of Gd-DTPA-polylysine as a macromolecular contrast agent for magnetic resonance imaging, Invest. Radiol. 1991, 26: 969-974).

It was generally agreed that the blood pool contrast media must have a molecular weight of at least 50,000 Daltons in order to achieve the desired blood pool properties (VS Vexier et al., Effect of varying the molecular weight of the MR contrast agent Gd-DTPA-polylysine on blood pharmacokinetics and enhancement pattern, JMRI 1994, 4: 381-388).

An accepted criterion for a blood pool contrast agent was an almost constant blood level over a certain period of time (30 min - 2 h) (RC Brasch, Rationale and applications for macromolecular Gd-based contrast agents. Magn. Reson. Med., 1991, 22: 282-287).

The use of such high molecular weight blood pool contrast media for the visualization of permeability disorders in malignant tumors has in principle already been shown in experimental studies (M.G.

Wikstrom et al., Contrast-enhanced MRI of tumors: Comparison of Gd-DTPA and a macromolecular agent, Invest. Radiol. 1989, 24: 609-15; R.C. Brasch et al. Quantification of capillary permeability to macromolecular magnetic resonance imaging contrast media in experimental mammary adenocarcinomas, Invest. Radiol. 1994, 29: 8-11).

Thereby one takes advantage of the tumor physiology. The blood vessels of malignant tumors differ significantly from those in benign tumors. In malignant tumors, the blood vessels grow from the surrounding tissue during the early growth of the tumor (so-called vasoneogenesis). No basement membrane is formed in these tumor vessels, which is why the vessel lining has larger gaps due to the lack of a supporting structure (GE Palade et al., Structural aspects of the permeability of the microvascular endothelium, Acta Physiol. Scand. Suppl. 1979, 463: 11). Through these gaps (endothelial pores), macromolecular substances (e.g. blood pool contrast agents) can penetrate into the interstitial space of the tumor and accumulate there (LW Seymour, Passive tumor targeting of soluble macromolecules and drug conjugates. Crit-Rev-Ther-Drug-Carrier -Syst 1992, 9: 135-187). This property is called hyperpermeability or leakiness of vessels in malignant tumors. In contrast, blood vessels of benign tumors do not arise from vasoneogenesis, but are normal vessels of the corresponding host tissue. Hyperpermeability does not occur. Macromolecular substances (such as blood pool contrast agents) can therefore not accumulate in such tumors.

A differentiation of malignant and benign tumors can in principle be achieved by the different enhancement patterns (enhancement kinetics) of polymeric contrast agents. So far, however, it has always been assumed that a reliable distinction of the enhancement kinetics only with a constant blood level, i.e. only possible with blood pool contrast media with a molecular weight greater than 50,000 Daltons (R.C. Brasch, Rationale and applications for macromolecular Gd-based contrast agents, Magn. Reson. Med, 1991, 22: 282-287). A gradual increase over a longer period of time is pathognomonic for malignant tumors. In contrast, constant contrast agent concentrations are observed in benign tumors. Blood pool contrast media with a molecular weight above 50,000 Daltons are therefore suitable for use in areas with increased vascular permeability, e.g. accumulate in malignant tumors. The almost constant blood level of the contrast medium required for this is primarily due to the size of the blood pool contrast medium above the kidney filtration threshold (approx. 40,000-50,000).

However, these high molecular weight blood pool contrast media did not find their way into clinical testing because the problem of missing or insufficient excretion could not be solved. The long retention of the contrast medium in the body can lead to side effects and tissue damage (T. Fritz et al., Detailed toxicity studies of liposomal gadolinium-DTPA, Invest. Radiol., 1991: 26, 960-968).

It was therefore an object of the present invention to find contrast media which are suitable for differentiating malignant and benign tumors in areas without a blood-brain barrier and which moreover have the disadvantages of the prior art Overcome technology, ie those that are excreted via the kidney and are therefore well tolerated.

This object is achieved by the present invention.

It has been found that surprisingly, when using metal complex conjugates with a polymeric backbone and an average molecular weight of 10,000 to 40,000 daltons, an excellent differentiation of benign and malignant tumors in body regions is possible without a blood-brain barrier.

Contrast agents of this type not only allow the reliable differentiation of malignant and benign tumors, they are also distinguished among other things. also by the fact that they are completely eliminated from the body and are therefore well tolerated.

Suitable metal complex conjugates with a polymeric backbone and an average molecular weight in the range mentioned are u.a. in German patent applications DE 39 38 992 and DE 195 25 924 and in US Patents 5,338,532; 4,507,466; 4,631,337; 4,587,329; 4,568,737 and 4,507,466.

Such polymeric backbones are also referred to as cascades or dendrimers, contrast media containing conjugates of these polymers with metal complexes as cascade or dendrimeric contrast media.

Particularly preferred are metal complex conjugates which carry between 24 and 36 paramagnetic metal complexes, in particular metal complexes of gadolinium, on the terminal groups of the polymeric backbone.

The polymeric blood pool contrast agents which can be used according to the invention have a molecular weight below the kidney filtration threshold in the range from 10,000 to 40,000 daltons, preferably in a range from 12,000 to 30,000 daltons, in particular in a range from 15,000 to 25,000 daltons.

With regard to their excretion behavior and their blood kinetics, such polymeric contrast media differ only insignificantly from low-molecular extracellular contrast media. For these contrast agents, too, the main route of elimination from the body is glomerular filtration, as with a polymer contrast agent (24 Gd DTPA cascade polymer) with one Molecular weight of 17,650 daltons could be shown in rats. When examining the blood level in the rat and evaluating it using a two-compartment model, it was found that the total blood clearance of the polymer (11.1 ml / min kg) changed only slightly compared to that of Gd-DTPA (13.4 ml / min kg) was. The drop in blood concentration of both substances was accordingly parallel, ie a very rapid decrease in the blood concentration was also seen for the polymer.

On the other hand, if one looks at the diffusion behavior from the blood space into the interstitial space, one finds in experiments on the rat - using the identical polymeric contrast agent - surprisingly that 10 minutes after injection still 82% and 15 minutes after injection still 73% injected dose remain in the blood space. In comparison, under otherwise identical conditions for the extracellular contrast medium Magnevist ^{® mentioned,} only 30% contrast medium can be detected in the vascular space after 3 minutes. The remaining percentages indicate the proportion that has already diffused into the interstitial space, since the diffusion behavior in animals with impaired kidney function (ie with renal ligament) was examined, ie the contrast medium could not be excreted via the kidney.

Although, as explained above, due to the rapid excretion of the polymeric contrast agent via the kidney, a constant blood level is not achieved, the agents mentioned - as shown in the study described below - are surprisingly excellent for differentiating malignant and benign tumors. So diffuses, despite the short residence time of the contrast medium in the

Blood circulation, surprisingly a sufficient amount of the imaging agent through the enlarged endothelial pores of the vessels of a malignant tumor in the interstitial space, with the consequence that the decrease in the contrast agent concentration in the tumor is much slower than that in the blood.

On the other hand, no mterstitial diffusion takes place in the case of male tumors, so the contrast agent concentration in the blood is associated with the contrast agent concentration in the male tumor (rapid increase after injection, rapid decrease due to renal elimination).

The following study serves to explain the subject of the invention in more detail, without wishing to restrict it to them. study

A study was conducted in 18 dogs (3.8-38 kg) suffering from a spontaneous, naturally occurring tumor of the mammary gland. It was unknown whether the tumor was benign or malignant.

The anesthetized dogs were examined by means of magnetic resonance tomography at 1.5 Tesla. A rapid Tl-weighted recording sequence (FLASH, TR 175 ms, TE 15 ms, α = 50 °) was carried out before and up to 20 minutes after contrast agent injection.

In the tests two different contrast agent was injected for comparison purposes in succession in the same animal: First Magnevist ^® (Schering) was a commercially available extracellular contrast agent at a dose of 50 mol / kg injected. After complete elimination of this contrast agent, a 24 Gd DTPA cascade polymer, a contrast agent with a molecular weight of 17,650 daltons, was injected at a dose of 25 μmol / kg body weight. The cascade polymer was prepared as in Wiener EC et al., Dendrimer-based metal chelates: a new class of magnetic resonance imaging contrast agents, MRM 1994; 31: 1-8.

This study design enabled an intra-individual comparison of the contrast agent kinetics.

After the MRI examination, the dog was operated on, the tumor was removed and the type of tumor was determined. It was found that 5 malignant (carcinomas) and 13 benign tumors (adenomas and mixed tumors) had been examined.

Figure 1 shows a sagittal MR slice before (top), 5 min p.i (middle) and 20 min p.i. (bottom) of the extracellular contrast agent. A blurred, oval tumor with central necrosis appears in the gland. The histology revealed a lobular breast cancer (malignant tumor). A clear and rapid enhancement of the tumor is observed five minutes after the injection of the extracellular contrast agent Gd-DTPA / dimeglumine. After 20 minutes, however, the signal intensity of the tumor has visibly decreased and is again approaching the pre-contrast values.

FIG. 2 shows the signal curves (intensity-time curves) of all tumors examined after injection of the extracellular contrast medium. In all tumors [benign Tumors (mixed tumors, adenomas) and malignant tumors (carcinomas)] a similar curve pattern was observed, ie a rapid signal increase and a subsequent rapid signal decrease were found first. It is not possible to differentiate between the different types of tumors because of the similar course of the curve.

Figure 3 shows a sagittal MR slice before (top), 5 min p.i (middle) and 20 min p.i. (bottom) of the polymeric contrast agent. In the gland, there is an oval, fuzzy tumor with central necrosis. The histology revealed a lobular breast cancer (malignant tumor). A marked enhancement of the tumor is observed five minutes after injection of the polymeric contrast medium. After 20 minutes the signal intensity of the tumor has still not decreased and the signal values 20 min p.i. seem to them 5 min p.i. to be similar.

Figure 4 shows the signal curves (intensity-time curves) of all tumors examined after injection of the polymeric contrast agent. After injection, a very similar signal pattern was observed in the benign tumors as in the extracellular contrast medium. In contrast, the malignant tumors showed a plateau phase of the signal for the first 20 minutes after injection. A reliable one

A distinction between benign and malignant tumors is possible.

Figure 5 gives the clearance half-lives for each contrast agent and tumor type. The clearance half-lives (HWZ) were determined by means of a non-linear adaptation to the signal intensity-time curves of the individual tumors and represent a measure of the speed of the signal drop (following the peak enhancement). As can be easily recognized, they differ the clearance-HWZ of male and malignant tumors significantly. The clearance-HWZ of the malignant tumors were significantly longer compared to those of benign tumors. This size is therefore best suited to differentiate between malignant and benign tumors after injection of the polymeric contrast medium, since there was no overlap between the values of the two groups, particularly with this size.

Claims

claims

1. Use of paramagnetic metal complex conjugates with a polymeric backbone and an average molecular weight of 10,000 to 40,000 daltons for the differentiation of benign and malignant tumors in

Regions of the body without a blood-brain barrier.

2. Use of paramagnetic metal complex conjugates with a polymeric skeleton according to claim 1, characterized in that the molecular weight of the conjugate is in the range from 12,000 to 30,000 daltons.

3. Use of paramagnetic metal complex conjugates with a polymeric backbone according to claim 1, characterized in that the molecular weight of the conjugate is in the range from 15,000 to 25,000 daltons.

4. Use of paramagnetic metal complex conjugates with a polymeric backbone according to claim 1 in NMR diagnostics for the differentiation of benign and malignant tumors in body regions without

Blood-brain barrier.

5. Use of paramagnetic metal complex conjugates with a polymeric backbone according to claim 1 containing as a polymeric backbone a cascade or a dendrimeric polymer.

6. Use of paramagnetic metal complex conjugates with a polymeric backbone according to claim 1 containing 24 to 36 paramagnetic metal ions.

7. Use of paramagnetic metal complex conjugates with a polymeric skeleton according to claim 6 containing as paramagnetic metal ions gadolinium ions.

8. Paramagnetic metal complex conjugates with a polymeric

Basic structure and an average molecular weight in the range of 10,000 to 40,000 daltons for the preparation of an agent for NMR diagnostics Differentiation of benign and malignant tumors in body regions without a blood-brain barrier.

9. Contrast media for NMR diagnostics for the differentiation of human and malignant tumors in body regions without a blood-brain barrier containing paramagnetic metal complex conjugates with a polymeric framework and an average molecular weight in the range from 10,000 to 40,000 daltons.

10. A method for differentiating between malignant and benign tumors, characterized in that the blood clearance half-lives are determined with the aid of paramagnetic metal complex conjugates with a polymeric backbone and a molecular weight of 10,000 to 40,000 daltons.