Research article
Somatostatin-receptor-targeted α-emitting 213Bi is therapeutically more effective than β-emitting 177Lu in human pancreatic adenocarcinoma cells

https://doi.org/10.1016/j.nucmedbio.2006.11.006Get rights and content

Abstract

Introduction

Advance clinical cancer therapy studies of patients treated with somatostatin receptor (sstr)-targeted [DOTA0-Tyr3]octreotide (DOTATOC) labeled with low-linear-energy-transfer (LET) β-emitters have shown overall response rates in the range of 15–33%. In order to improve outcomes, we sought to compare the therapeutic effectiveness of sstr-targeted high-LET α-emitting 213Bi to that of low-LET β-emitting 177Lu by determining relative biological effectiveness (RBE) using the external γ-beam of 137Cs as reference radiation.

Methods

Sstr-expressing human pancreatic adenocarcinoma Capan-2 cells and A549 control cells were used for this study. The effects of different radiation doses of 213Bi and 177Lu labeled to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid and sstr-targeted DOTATOC were investigated with a clonogenic cell survival assay. Apoptosis was measured using the Cell Death Detection ELISAPLUS 10× kit.

Results

Using equimolar DOTATOC treatment with concurrent irradiation with a 137Cs source as reference radiation, the calculated RBE of [213Bi]DOTATOC was 3.4, as compared to 1.0 for [177Lu]DOTATOC. As measured in terms of absorbance units, [213Bi]DOTATOC caused a 2.3-fold-greater release of apoptosis-specific mononucleosomes and oligonucleosomes than [177Lu]DOTATOC at the final treatment time of 96 h (P<.001) in sstr-expressing Capan-2 cells.

Conclusions

In conclusion, at the same absorbed dose, [213Bi]DOTATOC is therapeutically more effective in decreasing survival than is [177Lu]DOTATOC in human pancreatic adenocarcinoma cells due to its comparatively higher RBE.

Introduction

Pancreatic adenocarcinoma is the fourth leading cause of cancer deaths in the United States and has the lowest survival rate among nonhematological cancers [1]. The observed low survival rate in pancreatic adenocarcinoma is due to the advance and metastatic stages of the disease at the time of diagnosis and resistance to all forms of treatment [2], [3]. In an attempt to improve the treatment options for pancreatic adenocarcinoma, researchers have explored a number of novel therapies, such as endocrine therapy and immunotherapy. Unfortunately, most of these efforts have led, at best, to modest improvements in median survival, without any increase in cure rates [3].

The discovery that certain types of cancers and tumor types overexpress receptors for peptide hormones led to the field of peptide receptor scintigraphy and peptide receptor radionuclide therapy (PRRT) [4], [5]. One such target for peptide receptor cancer therapy is somatostatin receptor (sstr) [6]. Five sstrs (sstr1–sstr5) have been identified and characterized in humans, all of which are members of G-protein-coupled receptors [7]. Sstr subtypes 2 and 5 are highly overexpressed in exocrine pancreatic adenocarcinomas of neuroendocrine origin. Somatostatinomas account for <2% of all pancreatic cancers [8], [9]. Various somatostatin analogues have been developed to specifically target sstr2 and sstr5, including [DOTA0-Tyr3]octreotide (DOTATOC), for potential use as diagnostic and therapeutic radiopharmaceuticals in nuclear medicine and cancer therapy [4], [10], [11]. Studies of radiolabeled DOTATOC for the targeted therapy of sstr-expressing pancreatic tumors have demonstrated efficacy and possible toxicity with the β-emitting 90Y [11], [12]. [90Y]DOTATOC therapies have led to overall pancreatic tumor responses in 15–33% of patients evaluated, but not without renal or hematological toxicities [4], [12], [13]. Kidney failure has even been reported in patients treated with [90Y]DOTATOC in clinical studies [14], [15]. In an attempt to overcome this problem, [177Lu]DOTATOC and [177Lu]DOTATATE were studied in rats bearing small tumors [16]. For [177Lu]DOTATOC, the overall response rate ranged from 50% to 60%, and minimal nephrotoxicity was observed as a result of a lower radiation dose absorbed into the kidney due to lower energy and shorter mean path length [16].

A limiting factor associated with the moderate success of previous studies may be the linear energy transfer (LET) of the β-emitting radioisotopes used. Over the past decade, there has been a growing interest in the use of high-LET α-emitting radionuclides for cancer therapy [17], [18], [19].

It is believed that the most important factor in radiation damage is DNA double-strand breaks [20]. When these double-strand breaks are joined with wrong DNA ends during a subsequent nonhomologous end-joining event, the results may be deletions, chromosome aberrations, and reproductive cell deaths [21], [22]. It has been demonstrated that misjoining frequencies are higher with high-LET radiation than with low-LET radiation [23] and, as a consequence, the complex DNA damage produced by high-LET radiation is not easily repaired even at low radiation doses of 2 Gy [24]. Therefore, when compared to low-LET forms of radiation, high-LET forms of radiation pose a higher relative biological effectiveness (RBE) for cell killing and possess good localization characteristics resulting in energy deposition within 0.04–0.1 mm of a decay site [18], [19]. It has also been shown that high-LET radiation can generate reactive oxygen species [25] and can induce apoptosis and cell cycle arrest [26], [27], [28], [29]. Therefore, it is hypothesized that high-LET forms of radiation could be used effectively in treating quiescent and hypoxic tumors, which are usually resistant to conventional chemotherapy and radiotherapy.

In order to further improve the outcomes of sstr-targeted radionuclide therapy, we sought to compare the therapeutic effectiveness of sstr-targeted high-LET α-emitting 213Bi compared to that of the low-LET β-emitting 177Lu for treating human pancreatic adenocarcinoma cells by determining the effects on toxicity and the RBE using the external γ-beam of 137Cs as reference radiation. In the past few years, the study of radiation-induced apoptosis has gained immense interest; therefore, an apoptotic assay was performed to observe the differences in the induction of apoptosis by radionuclides of varying LET. High-LET radioisotopes have been shown to be more successful in treating small-sized carcinomas than in treating larger-sized carcinomas [18], [30], [31], [32]; therefore, for this study, we chose 177Lu instead of 90Y, as 177Lu was shown to be more successful than 90Y for the treatment of smaller-sized pancreatic carcinomas [33].

In the present study, we report a comparison of high-LET radiation versus low-LET radiation for use in PRRT.

Section snippets

Cell culture

Human pancreatic adenocarcinoma Capan-2 cells express high levels of sstr2 and sstr5 [34]. Therefore, for this study, Capan-2 was selected as the sstr-expressing test cell line. For control experiments, A549 lung carcinoma cells were chosen as they do not express sstr2 and sstr5. Capan-2 cells were cultured in McCoy's 5a medium and supplemented with 1.5 mM l-glutamine, 10% (vol/vol) fetal bovine serum, 100 U/ml penicillin and 100 μg/ml streptomycin. A549 cells were cultured in Ham's F12K medium

Radionuclide and radiochemical purity

The radionuclidic purity of 213Bi obtained from the 225Ac/213Bi generator was evaluated using γ-spectroscopy on a wide window, as well as on the 213Bi window (404 keV±20%), over a period of 7 days. The 213Bi eluted from the 225Ac/213Bi generator was >99% pure, as no γ-emission characteristic of 225Ac was detected over 7 days. The radionuclidic purity of 177Lu was >99%, as described in the certificate of analysis provided by the supplier.

The radioisotopes 213Bi and 177Lu were successfully

Discussion

The primary objective of this study was to compare the therapeutic effectiveness of sstr-targeted high-LET α-emitting 213Bi to that of low-LET β-emitting 177Lu in human pancreatic adenocarcinoma cells by determining their effects on toxicity and RBE using an external γ-beam 137Cs as reference radiation. We found that the sstr-targeted high-LET α-emitting 213Bi had a greater RBE and was more toxic than the sstr-targeted low-LET β-emitting 177Lu.

The critical feature of radiation damage is the

Acknowledgments

We are thankful to Rahul Poria (University of New Mexico Health Sciences Center, Albuquerque, NM) for technical support and laboratory assistance. Critical manuscript-writing inputs from Dr. Robert Glew and Dr. Dorothy VanderJagt (University of New Mexico Health Sciences Center, Albuquerque, NM) are highly appreciated. Gratitude is extended to Dr. Martin W. Brechbiel and Dr. Kayhan Garmestani (Radioimmune and Inorganic Chemistry Section, Radiation Oncology Branch, NCI) for providing the 225Ac/

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