Pediatric Radiology

, Volume 38, Issue 5, pp 529–537

MR imaging of ovarian tumors using folate-receptor-targeted contrast agents

Authors

    • Department of RadiologyUniversity of California San Francisco
  • Sophie Boddington
    • Department of RadiologyUniversity of California San Francisco
  • Michael Wendland
    • Department of RadiologyUniversity of California San Francisco
  • Reinhard Meier
    • Department of RadiologyUniversity of California San Francisco
  • Claire Corot
    • Research DivisionGuerbet
  • Heike Daldrup-Link
    • Department of RadiologyUniversity of California San Francisco
Original Article

DOI: 10.1007/s00247-008-0764-6

Cite this article as:
Wang, Z.J., Boddington, S., Wendland, M. et al. Pediatr Radiol (2008) 38: 529. doi:10.1007/s00247-008-0764-6

Abstract

Background

Because of its over-expression in many human tumors, the folate receptor (FR) is a promising target for tumor-specific imaging.

Objective

To evaluate the uptake of FR-targeted gadolinium (P866) and iron-oxide (P1048) agents in an ovarian tumor model.

Materials and methods

FR-positive ovarian cancer cells (IGROV-1) were incubated with FR-targeted agents (P866 or P1048) in the absence or presence of competing free folate. Intracellular gadolinium or iron-oxide concentrations were measured. MR imaging of implanted ovarian tumors in rats was performed following injection of FR-targeted (P866 and P1048) and nontargeted (P1001 and P904) agents. Changes in longitudinal and transverse relaxation rates (ΔR1 and ΔR2), which were proportional to the contrast agent concentration in the tumors, were compared between tumors injected with FR-targeted and nontargeted agents.

Results

IGROV-1 cells showed uptake of P866 and P1048, which decreased with competing free folate. The ΔR1 values were higher at 1 h following injection of P866 than following injection of P1001 (P < 0.05), indicating a higher amount of contrast agent retained in the tumor following P866 injection. There was a trend for higher ΔR2 values at 48 h following injection of P1048 than following injection of P904, but it was not statistically significant (P = 0.09).

Conclusion

Specific accumulation of the FR-targeted gadolinium agent P866 was suggested in an FR-positive ovarian tumor model, demonstrating the possibility of combining the specificity of receptor targeting with the improved anatomic resolution of MR imaging. This could improve diagnosis and treatment of FR-positive tumors.

Keywords

MRIContrast materialOvarian tumor

Introduction

The development of tumor-specific imaging agents is highly desirable, because they can provide earlier and more accurate diagnosis and can improve the assessment of the biological aggressiveness of the evaluated tumors and the monitoring of treatment response. The folate receptor (FR) has particular characteristics that make it a promising target for tumor-specific imaging and therapy.

The vitamin folate is required by all cells for metabolism and survival. Eukaryotic cells are unable to produce folate and therefore must acquire it from the environment. There are two routes by which folate can enter a cell (Fig. 1):
  1. 1.

    Reduced folate carrier (RFC) is a transmembrane transporter that is ubiquitously expressed throughout development and in normal adult tissue. It is the major route of entry for the reduced forms of folate [1, 2]. Upon oral ingestion of folate, the vitamin undergoes intestinal absorption and is rapidly taken up by the liver, where it is either stored in hepatocytes or converted to dihydrofolate, tetrahydrofolate, or methylene tetrahydrofolate [3]. The reduced form of folate is released by the liver and enters cells of normal organs via the RFC.

     
  2. 2.

    FR is a glycopolypeptide that binds folate with high affinity [4]. FR is over-expressed in various types of human carcinomas including ovarian, breast, colorectal and nasopharyngeal carcinomas in adults [57], as well as pediatric tumors such as choroid plexus tumors, ependymomas, osteosarcomas and leukemia [810]. There are relatively low levels of FR in normal tissues [11]. It is important to understand that the FR has a high affinity for folate but not its reduced forms (Fig. 1). There are several FR isoforms (α, β, and γ). The α isoform is over-expressed in cancer cells and has a high affinity for the folate ligand (Kd about 10−10 M) [12]. This makes folate a valuable vehicle for conjugation with specific tracers, thereby allowing the delivery of the tracers to FR-positive cancer cells and providing cancer-specific imaging.

     
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Fig. 1

Mechanisms of folate entry into cells

Successful tumor-selective FR targeting has been reported both in vitro and in vivo with a variety of radionuclides [1320] (Table 1). The FR-targeted radionuclides have several advantages, including relatively easy and cost-effective synthesis, potential broad applications for a large variety of tumors, and favorable tumor-to-background ratios. However, current radiotracer-based imaging techniques are limited in anatomic resolution, which necessitates additional imaging modalities to further localize the detected tumor. In addition, the radiotracer techniques cause a considerable radiation exposure. FR-targeted optical imaging probes have also been developed to provide a similar sensitivity and specificity for cancer diagnosis [2123] (Table 2). The optical imaging techniques, however, also have limited anatomical resolution and are not readily available for clinical applications.
Table 1

Selected references relating to successful in vivo tumor targeting using radionuclides conjugated with the FR

Reference

FR-targeted radionuclide

Targeted pathology

Targeted species

16

111In-DTPA folate

FR-positive KB cells (human nasopharyngeal carcinoma)

Mouse

15

99mTc-HYNIC folate

FR-positive 24JK-FBP cells (mouse sarcoma cell)

Mouse

18

99mTc-EC20

FR-positive M109 cells (murine lung carcinoma)

Mouse

20

99mTc-MAG3 folate

MCF-7 (human breast cancer)

Mouse

13

18F-FBA folate

FR-positive K31 cells (epidermal carcinoma)

Mouse

Table 2

Selected references relating to successful in vivo tumor targeting using fluorescent probes conjugated with the FR

Reference

FR-targeted fluorescent probe

Targeted pathology

Targeted species

21

Folate targeting NIR fluorochrome

FR-positive KB cells (human nasopharyngeal carcinoma)

Mouse

22

Folate–fluorescein conjugate

FR-positive M109 cells (murine lung carcinoma) and L1210 cells (leukemia cells)

Mouse

23

Fluorescent folate probe (FFP)

Activated macrophages on dysplastic intestinal adenomas

Mouse

More recently, efforts have been directed toward the development of imaging probes that are detectable with MR imaging [2427]. MRI provides a noninvasive means for tumor detection with excellent soft-tissue contrast and anatomic resolution, without radiation exposure. The aim of our study was to investigate the diagnostic performance of new formulations of FR-targeted MR contrast agents in an ovarian tumor model. We focused our studies on ovarian cancer as a representative tumor with high levels of FR over-expression [6] in order to evaluate the feasibility of FR-targeted MR imaging. The concept of receptor-targeted imaging, however, would be applicable to other FR-positive tumors.

Materials and methods

Contrast agents

All contrast agents were provided by the Research Division of Guerbet, Paris, France.

P866

P866 is a high-relaxivity dimeric gadolinium chelate that is surrounded by hydrophilic branches and conjugated to a folate moiety (Fig. 2). It has a molecular weight of 9.4 kDa. The r1 relaxivity is 42 s−1 mM−1 (i.e. 21 s−1 mM−1 Gd), and the r2 relaxivity is 60 s−1 mM−1 (i.e. 30 s−1 mM−1 Gd) in water at 60 MHz and 37°C. The P866 molecule is stable in plasma and is excreted whole in the urine (no metabolism) (unpublished data, Guerbet Research).
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Fig. 2

FR-targeted MR contrast agents: a gadolinium P866, b iron-oxide P1048. P866 is a high-relaxivity dimeric gadolinium chelate conjugated to folate. P1048 consists of an iron-oxide core conjugated to folate

P1001

P1001 is a non-FR-targeted analog of P866. It is composed of an identical gadolinium chelate without the folate moiety. It has the same molecular weight and R1 and R2 relaxivities as P866.

P1048

P1048 consists of an iron-oxide core conjugated to folate (Fig. 2). The iron-oxide core is coated with hydrophilic branches of amino-alcohol and grafted with eight to ten folate moieties per nanoparticle. The size of P1048 is 25 nm. The R1 relaxivity is 14 s−1 mM−1 Fe, and the R2 relaxivity is 92 s−1 mM−1 Fe in water at 60 MHz and 37°C. There is no loss of activity after incubation of P1048 in standard cell culture media, which is indirect proof of stability of the contrast material in biological media (unpublished data, Guerbet Research).

P904

P904 is a non-FR-targeted analog of P1048. Its size is 21 nm. It has the same R1 and R2 relaxivities as P1048.

Cell culture

FR-positive human ovarian cancer cells (IGROV-1; from Dr. Joe Gray, UCSF) were grown in folate-free RPMI-1640 medium (Gibco BRL, Grand Island, NY) supplemented with 10% fetal calf serum at 37°C. Ten million cells were incubated for 24 h with 50 μM of FR-targeted P866 in the absence or presence of 750 μM competing free folate, or with the non-FR-targeted analog P1001. Similarly, ten million cells were incubated for 24 h with 20 μM of FR-targeted P1048 in the absence or presence of 750 μM competing free folate, or with the non-FR-targeted analog P904. Intracellular gadolinium or iron-oxide concentrations were measured by inductively coupled plasma mass spectrometry (ICP-MS) (Perkin Elmer, Paris, France).

Animal model

The study was approved by the Committee for Animal Research at our institution. For tumor implantation and MR imaging, animals were anesthetized with 1.5–2% isoflurane (Narkomed, North American Drager) in oxygen administered via a face mask.

IGROV-1 ovarian tumor cells were implanted in nine 3- to 4-week-old female athymic Harlan rats. Approximately 6×106 tumor cells were injected subcutaneously into the right lower flank of each rat. All rats received a folate-deficient diet (Dyets, Bethlehem, PA) for 3–4 weeks prior to MR imaging in order to reduce the serum folate concentration to levels that are normally found in humans. The tumors were imaged when they reached a size of approximately 1 cm.

Animal experimental procedure

For the evaluation of the FR-targeted gadolinium agent P866, MR imaging was performed in a group of three rats before injection and up to 1 h after injection of P866 (0.03 mmol Gd/kg) on day 1. On day 2, the same group of rats was imaged before and up to 1 h after injection of non-FR-targeted analog P1001 (0.03 mmol Gd/kg). We evaluated the gadolinium-based contrast agents over 1 h, as a previous pharmacokinetic study done by our group showed the elimination half-life of P866 to be 36 min based on plasma measurements [28]. The same group of three rats was used for both the P866 and P1001 studies, because pilot studies showed no significant residual P866 present in the tumors after a 24-h delay on day 2.

For the evaluation of the FR-targeted iron-oxide agent P1048, MR imaging was performed in one group of three rats before, immediately after, and at 24 h and 48 h after injection of the FR-targeted P1048 (0.3 mmol Fe/kg). For comparison, MR imaging was performed in a separate group of three rats using the same protocol but with the non-FR-targeted P904 (0.3 mmol Fe/kg). We evaluated the iron-oxide contrast agents over 48 h and in two separate but comparable groups of rats because of the much longer elimination half-life (elimination half time about 15 h; unpublished data).

MR imaging

MR imaging was performed on a 2-T Omega CSI-II superconducting MR scanner (Bruker Instruments, Fremont, CA). The animals were placed in a bird-cage radiofrequency coil on a warming mattress filled with 37°C warm deuterium oxide, in order to keep the body temperature of the animals relatively constant.

For the evaluation of the gadolinium contrast agent P866, the following pulse sequences were used. Images before contrast agent administration were acquired using a single-slice inversion recovery (IR) snapshot FLASH sequence with the following parameters: TR 3 ms, TE 1.5 ms, field of view 55×55 mm, matrix 64×64, slice thickness 3 mm, flip angle 5°, TI (inversion times) 100–2,500 ms. Dynamic contrast-enhanced MR imaging was performed every 2 min for 1 h using a T1-weighted (T1-W) 3-D spoiled gradient recalled (SPGR) sequence with the following parameters: TR 30 ms, TE 4.83 ms, field of view 55×55×48 mm, matrix 128×128×16, effective slice thickness 3.6 mm, flip angle 90°. In addition, a 1-h IR snapshot FLASH sequence after contrast agent administration was acquired with the same parameters as used for the precontrast scan.

For the evaluation of the iron-oxide contrast agent P1048, multiecho (four echoes) T2*-weighted (T2*-W) 3-D SPGR sequences and multiecho (four echoes) T2-W 2-D spin-echo (SE) sequences were obtained. The T2*-W sequence had the following parameters: TR 200 ms, TE 5, 10, 15, 20 ms, field of view 50×50 mm, matrix 256×128, slice thickness 2 mm. The T2-W SE sequence had the following parameters: TR 2,000 ms, TE 20, 40, 60, 80 ms, field of view 50×50 mm, matrix 256×128, slice thickness 2 mm. Dynamic imaging was not performed because iron-oxide contrast agents are expected to have a much longer blood half-life.

Imaging analysis

MR images were analyzed using MR-Vision software (MR-Vision, Menlo Park, CA). The relative signal enhancement ΔSI (%) was quantified as \(\Delta {\text{SI}}{\left( {\text{\% }} \right)} = {\left\{ {{{\left( {{\text{SI}}_{{{\text{post}}}} - {\text{SI}}_{{{\text{pre}}}} } \right)}} \mathord{\left/ {\vphantom {{{\left( {{\text{SI}}_{{{\text{post}}}} - {\text{SI}}_{{{\text{pre}}}} } \right)}} {{\text{SI}}_{{{\text{pre}}}} }}} \right. \kern-\nulldelimiterspace} {{\text{SI}}_{{{\text{pre}}}} }} \right\}} \times 100\% \). For the evaluation of the gadolinium contrast agents, the relative signal enhancement of the tumors before and 1 h after injection of P866 was compared to that before and after injection of P1001. In addition, longitudinal relaxation rate R1 (1/T1) estimates for tumors were obtained by curve fitting based on one set of IR images. ΔR1, the difference between precontrast R1 and 1-h postcontrast R1, is assumed to be directly proportional to the concentration of the contrast medium in the tumor. ΔR1 values were compared between tumors injected with P866 and those injected with P1001. For the evaluation of the iron-oxide contrast agents, the relative signal losses of the tumors before, immediately after, and at 24 h and 48 h after injection of P1048 were compared with the signal losses of tumors injected with P904. In addition, transverse relaxation rate R2 (1/T2) estimates for tumors were obtained by curve fitting based on one set of multiecho T2 SE images. ΔR2, the difference between precontrast R2 and 24-h or 48-h postcontrast R2, which were proportional to the concentration of iron oxide in the tumors, were compared between tumors injected with P1048 and those injected with P904.

Histology

Immediately after the last MR imaging procedure, the animals were killed and the tumors were dissected and frozen at −80°C. Frozen sections were stained with FR-α antibody specific to humans (Santa Cruz no. sc-16386, 1:100 dilution) and counterstained with hematoxylin. As a negative control, implanted HT1080 human fibrosarcoma tumors from a different study were also stained with the same antibody. The IGROV-1 ovarian cancer cells have been reported to have a high level of FR-α expression [29], while the HT1080 human fibrosarcoma cells were reported to be FR-negative [30].

Statistics

Statistical analysis was performed using the Stata software package version 7.0 (Stata Corporation, College Station, TX). Differences in ΔSI (%) of tumor before and after injection of P866 versus P1001 or P1048 versus P904, differences in ΔR1 values between tumors injected with P866 and P1001, and differences in ΔR2 values between tumors injected with P1048 and P904 were tested with a two-tailed Student’s t-test. Results were considered statistically significant with P values <0.05.

Results

Cell culture

After incubation of IGROV-1 cells for 24 h with P866, the concentration of gadolinium in the cell lysate as measured by ICP-MS was 0.668 nmol/107 cells, which decreased by 25% to 0.499 nmol/107 cells in the presence of competing free folate (Table 3). Similarly, after incubation of IGROV-1 cells for 24 h with P1048, the concentration of iron in the cell lysate was 18.3 nmol/107 cells, which decreased by 32% to 12.3 nmol/107 cells in the presence of competing free folate (Table 3).
Table 3

Intracellular gadolinium or iron concentrations as measured by ICP-MS in the IGROV-1 cell lysate. The IGROV-1 cells were incubated with FR-targeted contrast agents in the presence or absence of free folate, or with the non-FR-targeted analogs

Intracellular gadolinium (nmol/107 cells)

Intracellular iron (nmol/107cells)

Incubation with P866

Incubation with P1001

Incubation with P1048

Incubation with P904

Without folate

With folate

Without folate

With folate

0.668

0.499

0.514

18.3

12.3

13.7

Animal studies

Gadolinium agents (P866 and P1001)

The implanted tumors showed mild peak enhancement at approximately 4 min following injection of either P866 or P1001 on the T1-W dynamic SPGR images (Fig. 3). This initial peak enhancement of the tumors was apparently a perfusion effect and was followed by a rapid decline, with minimal residual tumor enhancement at 1 h following injection of either contrast agent (Fig. 3). There was no significant difference in relative signal enhancement between tumors injected with P866 and those injected with P1001 (P = 0.70). However, the mean ΔR1 relaxation rates were significantly higher at 1 h following injection of P866 (mean ΔR1 0.214 s−1) than following injection of P1001 (mean ΔR1 0.112 s−1; P = 0.03), indicating a higher amount of contrast agent retention in the tumor following injection of P866 than following injection of P1001 (Fig. 4).
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Fig. 3

Representative axial T1-W 3D-SPGR MR images through the implanted ovarian tumor before injection and at 4 min and 1 h after injection of P866 (top row) or P1001 (bottom row). There is mild enhancement of the tumor at 4 min after injection of either P866 or P1001, with minimal residual enhancement at 1 h with either contrast agent

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Fig. 4

Mean ΔR1 relaxation rates at 1 h following injection of P866 (mean ΔR1 0.214 s−1) or P1001 (mean ΔR1 0.112 s−1; P < 0.05). The significant difference between the ΔR1 relaxation rates suggests at least a component of FR-specific uptake of P866 by the tumors

Iron oxide agents (P1048 and P904)

The iron-oxide sensitive T2*-W MR images showed moderate signal loss in the tumors immediately following injection of either P1048 or P904, likely representing a perfusion effect (Fig. 5). There was washout of both contrast agents over 48 h. No significant difference in relative signal change of the tumors was observed between tumors injected with P1048 and P904 at either 24 or 48 h (P = 0.85). At 24 h after injection, the mean ΔR2 relaxation rates, which were proportional to the iron-oxide concentration in the tumors, were also similar between tumors injected with P1048 and those injected with P904 (P = 0.76). At 48 h, however, there was a trend for higher mean ΔR2 values, suggesting a higher amount of retained contrast agent in the tumor following injection of P1048 (mean ΔR2 0.0027 s−1) than following injection of P904 (mean ΔR2 0.0011 s−1), but the difference was not statistically significant (P = 0.09; Fig. 6).
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Fig. 5

Representative axial T2-W SPGR MR images through the implanted ovarian tumor before injection, immediately after injection and at 24 h and 48 h after injection of P1048 (top row) or P904 (bottom row). There was moderate signal loss in the tumors immediately following injection of either P1048 or P904, with washout of both contrast agents over 48 h

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Fig. 6

Mean ΔR2 relaxation rates at 48 h following injection of P1048 (mean ΔR2 0.0027 s−1) or P904 (mean ΔR2 0.0011 s−1; P = 0.09)

Histopathology

FR-α strongly stained multiple tumor cells from the implanted IGROV-1 ovarian tumors, consistent with reported high levels of FR-α expression (Fig. 7). In contrast, no significant staining was noted in the HT1080 human fibrosarcoma cells, which have been reported to be FR-negative (Fig. 7).
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Fig. 7

FR-α staining of FR-positive IGROV-1 ovarian tumor (a) and FR-negative HT1080 human fibrosarcoma (b). The IGROV-1 ovarian tumor cells demonstrate dark, positive cytoplasmic and cytoplasmic membrane staining for the FR (arrows). The HT1080 human fibrosarcoma cells demonstrate negative staining for the FR

Discussion

The FR-targeted gadolinium chelate P866, but not its non-FR-targeted analog P1001, caused a significant increase in R1 relaxation rates of FR-positive ovarian tumors on delayed MR scans (1 h after injection). Because R1 relaxation rates reflect the amount of contrast agent in the tumors, our results suggest a component of specific accumulation of FR-targeted P866 in FR-positive tumors. This is also supported by an in vitro study that showed decreased gadolinium concentrations in cells incubated with P866 in the presence of competing free folate.

The FR-targeted iron-oxide agent P1048, when compared to its non-targeted analog P904, showed a trend for an increase in R2 relaxation rates, corresponding to a higher amount of retained contrast agent, in FR-positive ovarian tumors on delayed MR scans (48 h after injection), but the increase was not statistically significant (P = 0.09). The in vitro study showed decreased iron concentrations in cells incubated with P1048 in the presence of competing free folate, suggesting specificity of P1048 for the FR on the ovarian tumor cells in vitro. When the ovarian tumors were imaged in vivo, there might have been more nonspecific uptake of the iron-oxide contrast agents because of nonspecific leak of the iron-oxide particles into the tumor interstitium and the presence of other cell populations such as macrophages that are known to take up iron [31]. This may partially explain why we were unable to find a statistically significant difference in contrast uptake between P1048 and P904 in vivo. The detection of nonspecific uptake might be reduced by allowing unbound iron-oxide particles to wash out over time, which was the rationale behind imaging the tumors at 48 h in our study. This process is dependent on the concentration gradient across the vasculature and the pharmacokinetics of the FR-targeted iron oxide particles. Thus, further modification of the contrast agents that allow higher interstitial accumulation and longer interstitial residence, along with optimization of the imaging efficiency, are needed to improve the FR-specific uptake into tumor cells and to improve sensitivity and specificity of the FR-targeted iron-oxide contrast agents.

Few other investigators have studied FR-targeted MR contrast agents for tumor imaging in vivo. Konda et al. [24, 25] and Wiener et al. [32] have described uptake of a gadolinium polyamidoamine (PAMAM) folate-dendrimer in FR-α-positive ovarian cancers (OVCA 432). More recently, Choi et al. [27] have reported the use of iron oxide nanoparticles conjugated to folate as MR contrast agent for imaging of nasopharyngeal carcinomas (KB cells). Our results are in agreement with those of previous studies showing the feasibility of imaging FR-positive tumors in vivo using targeted MR contrast agents. In addition, our results suggest active FR-targeting by P866 because we were able to compare the uptake of targeted (P866) and nontargeted (P1001) contrast agents by tumors. The previous studies did not compare targeted and nontargeted contrast agents. Corot et al. [33] have recently reported specific uptake of P866 in a nasopharyngeal carcinoma tumor model (KB cells) in mice. The degree of specific uptake of P866 in the KB cells was much higher than that observed in our ovarian tumor model in rats. The differences in P866 uptake might be attributed to a much higher level of FR-α expression in KB cells than in IGROV-1 cells [34].

As shown in Fig. 1, the specific uptake of FR-targeted contrast agents in vivo was better evaluated using a non-FR-targeted analog as a control rather than performing folate competition experiments. For in vivo folate competition studies, an excessive amount of free folate would be required to saturate the metabolic pathways in the liver, resulting in a nonphysiological amount of folate in the blood. Although our study suggested a specific retention of the FR-targeted contrast agent P866 in ovarian tumors, we did not investigate whether the P866 was bound to FR on the cell surface or whether some of the contrast agent was internalized into the cells. The mechanism is important with respect to the elimination pathway of this diagnostic agent as well as potential future designs of FR-targeted therapy.

The depiction of the FR-specific uptake in our study was close to the detection limit of MR imaging. The differences between the FR-targeted and nontargeted agents could only be detected by direct measurements of the R1 relaxation rates for the gadolinium agent rather than signal intensity measurements. It is known that changes in relaxation rates are proportional to the concentrations of contrast agents in the tumors, and that relaxation rate measurements are more sensitive than actual signal intensity measurements. Compared to the FR-targeted radionuclide and optical imaging techniques, MR imaging of cell surface receptors still faces many challenges, mostly because of the relatively low signal yield of MR contrast agents. The gadolinium agent P866 used in our study is composed of folate coupled to a high relaxivity dimeric gadolinium chelate in order to reach a higher MR sensitivity. Similarly, P1048 is composed of a folate moiety coupled to an iron oxide core in an attempt to improve the sensitivity of signal detection [35]. Clearly our study represents work-in-progress, and further modification of the contrast agents, improvement in pulse sequences, and use of higher field MR scanners are needed to improve the sensitivity of FR-specific uptake in tumors. Nonetheless, our current study serves as proof-of-concept and demonstrates the possibility of combining the specificity of receptor targeting with the improved anatomic resolution of MR imaging.

The impetus behind the FR-targeted MR contrast research is the potential clinical applications, which include characterization and treatment monitoring of FR-positive tumors. FR-targeted contrast agents might allow more sensitive and specific diagnosis of FR-positive tumors by detecting additional sites of tumor that are missed on conventional imaging, and by better distinguishing between tumor and treatment-related fibrosis or scarring. Assessment of tumor FR levels with targeted contrast agents might also lead to improved characterization of tumor aggressiveness, provide a rational means for selecting patients who would most benefit from antifolate therapy, and allow better treatment monitoring. We focused our studies on ovarian tumors because FR is over-expressed in 90% of ovarian tumors [6], so they serve as a good model to evaluate the feasibility of FR-targeted MR imaging. Analyses of human ovarian tumors to date have demonstrated considerable variability in the expression levels of FR among patients as well as heterogeneity within the same tumor [3638]. FR-specific MR contrast agents might estimate the level of over-expression of the receptors, which has been correlated directly with biological aggressiveness such as the histological grade and S-phase fraction [6, 36]. The FR status has also been reported to reflect a tumor’s response to chemotherapy [39]. Although ovarian tumors are relatively rare in the pediatric population, our study is designed to test the feasibility and potential of FR-targeted MR imaging. The concept of FR-targeted MR contrast agents might be applied to various FR-positive tumors, including pediatric tumors such as choroid plexus tumors and ependymomas [8], osteosarcomas [9], and acute myelogenous leukemia [3, 10].

In conclusion, specific accumulation of the FR-targeted gadolinium agent P866 was suggested in a FR-positive ovarian tumor model. Further development in FR-targeted contrast agents and improvement in imaging efficiency are needed to improve the sensitivity and specificity of MR imaging of FR-positive tumors.

Acknowledgement

Zhen J. Wang is supported by NIBIB T32 Training Grant 1 T32 EB001631-01A1.

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© Springer-Verlag 2008