Deficiency of Procollagen C-Proteinase Enhancer 1 in Mice has No Major Impact on Cardiac Collagen and Function Under Basal Conditions.

ABSTRACT: Maturation of fibrillar collagen is known to play a crucial role in the pathophysiology of myocardial fibrosis. Procollagen C-proteinase enhancer 1 (PCPE1) has a key role in procollagen maturation and collagen fibril formation. The phenotype of both male and female PCPE1 knock-out mice was investigated under basal conditions to explore the potential of PCPE1 as a therapeutic target in heart failure. Global constitutive PCPE1-/- mice were generated. Serum procollagen I C-terminal propeptide, organ histology, and cutaneous wound healing were assessed in both wild type (WT) and PCPE1-/- mice. In addition, the cardiac expression of genes involved in collagen metabolism was investigated and the total and insoluble cardiac collagen contents determined. Cardiac function was evaluated by echocardiography. No differences in survival, clinical chemistry, or organ histology were observed in PCPE1-/- mice compared with WT. Serum procollagen I C-terminal propeptide was lower in PCPE1-/- mice. Cardiac mRNA expression of Bmp1, Col1a1, Col3a1, and Loxl2 was similar, whereas Tgfb and Loxl1 mRNA levels were decreased in PCPE1-/- mice compared with sex-matched WT. No modification of total or insoluble cardiac collagen content was observed between the 2 strains. Ejection fraction was slightly decreased in PCPE1-/- male mice, but not in females. Finally, wound healing was not altered in PCPE1-/- mice. PCPE1 deficiency does not trigger any major liabilities and does not affect cardiac collagen content nor its function under basal conditions. Further studies are required to evaluate its role under stressed conditions and determine its suitability as a therapeutic target for heart failure.

Deconvolution of Tc-99m-Mercaptoacetyltriglycine Renograms with the Concomitant Use of a Sparse Legendre Polynomial Representation and the Moore-Penrose Pseudo-inverse.

OBJECTIVES: This study aimed to introduce an improved deconvolution technique for Tc-99m-mercaptoacetyltriglycine renograms based on the combination of a sparse Legendre polynomial representation and the Moore-Penrose inversion matrix (LG). This method reduces the effect of noise on the measurement of renal retention function transit time (TT). METHODS: The stability and accuracy of the proposed method were tested using a renal database containing Monte Carlo-simulated studies and real adult patient data. Two clinical parameters, namely, split function (SF) and mean TT (meanTT), obtained with LG were compared with values calculated with the established method that combines matrix deconvolution and a three-point linear smoothing (F121) as recommended by the 2008 International Scientific Committee of Radionuclides in Nephrourology consensus on renal TT measurements. RESULTS: For simulated data, the root mean square error (RMSE) between the theoretical non-noisy renal retention curve (RRC) and the results of the deconvolution methods applied to the noisy RRC were up to two times lower with LG (p<0.001). The RMSE of the reconvoluted renogram and the theoretical one was also lower for LG (p<0.001) and showed better preservation of the original signal. The SF was neither improved nor degraded by the proposed method. For patient data, no statistically significant difference was found between the SF for the LG method compared with the database values, and the meanTT better agreed with the physician's diagnosis than the matrix or clinical software (Hermes) outputs. A visual improvement of the RRC was also observed. CONCLUSION: By combining the sparse Legendre representation of the renogram curves and the Moore-Penrose matrix inverse techniques, we obtained improved noise reduction in the deconvoluted data, leading to better elimination of non-physiological signals -as negative values- and the avoidance of the smear effect of conventional smoothing on the vascular peak, which both influenced the meanTT measurement.

Procollagen C-Proteinase Enhancer-1 (PCPE-1) deficiency in mice reduces liver fibrosis but not NASH progression.

BACKGROUND AND AIMS: Nonalcoholic Steatohepatitis (NASH) is a major cause of end-stage liver diseases such as cirrhosis and hepatocellular carcinoma resulting ultimately in increased liver-related mortality. Fibrosis is the main driver of mortality in NASH. Procollagen C-Proteinase Enhancer-1 (PCPE-1) plays a key role in procollagen maturation and collagen fibril formation. To assess its role in liver fibrosis and NASH progression, knock-out mice were evaluated in a dietary NASH model. METHODS: Global constitutive Pcolce-/- and WT male mice were fed with a Choline Deficient Amino acid defined High Fat Diet (CDA HFD) for 8 weeks. Liver triglycerides, steatosis, inflammation and fibrosis were assessed at histological, biochemical and gene expression levels. In addition, human liver samples from control and NASH patients were used to evaluate the expression of PCPE-1 at both mRNA and protein levels. RESULTS: Pcolce gene deficiency prevented diet-induced liver enlargement but not liver dysfunction. Furthermore, liver triglycerides, steatosis and inflammation were not modified in Pcolce-/- male mice compared to WT under CDA HFD. However, a significant decrease in liver fibrosis was observed in Pcolce-/- mice compared to WT under NASH diet, associated with a decrease in total and insoluble collagen content without any significant modifications in the expression of genes involved in fibrosis and extracellular matrix remodeling. Finally, PCPE-1 protein expression was increased in cirrhotic liver samples from both NASH and Hepatitis C patients. CONCLUSIONS: Pcolce deficiency limits fibrosis but not NASH progression in CDA HFD fed mice.

S62798, a potent TAFIa inhibitor, accelerates endogenous fibrinolysis in a murine model of pulmonary thromboembolism.

Enhancement of fibrinolysis constitutes a promising approach to treat thrombotic diseases. Venous thrombosis and thromboembolism risks are associated with increased plasma levels of TAFI (Thrombin Activatable Fibrinolysis Inhibitor) as well as its active form TAFIa. A new TAFIa inhibitor, namely S62798 has been identified. Its ability to enhance fibrinolysis was investigated both in vitro and in vivo in a mouse model of pulmonary thromboembolism, as well as its effect on bleeding. S62798 is a highly selective human, mouse and rat TAFIa inhibitor (IC50 = 11; 270; 178 nmol/L, respectively). It accelerates lysis of a human clot in vitro, evaluated by thromboelastometry (EC50 = 27 nmol/L). In a rat tail bleeding model, no effect of S62798 treatment was observed up to 20 mg/kg. Enhancement of endogenous fibrinolysis by S62798 was investigated in a mouse model of Tissue Factor-induced pulmonary thromboembolism. Intravenous administration of S62798 decreased pulmonary fibrin clots with a minimal effective dose of 0.03 mg/kg. Finally, effect of S62798 in combination with heparin was evaluated. When treatment of heparin was done in a curative setting, no effect was observed whereas a significantly decreased pulmonary fibrin deposition was observed in response to S62798 alone or in combination with heparin. This study demonstrates that S62798 is a potent TAFIa inhibitor with minimal risk of bleeding. In vivo, curative S62798 intravenous treatment, alone or associated with heparin, accelerated clot lysis by potentiating endogenous fibrinolysis and thus decreased pulmonary fibrin clots. S62798 is expected to be a therapeutic option for pulmonary embolism patients on top of anticoagulants.

Turnip mosaic virus RNA replication complex vesicles are mobile, align with microfilaments, and are each derived from a single viral genome.

Nicotiana benthamiana plants were agroinoculated with an infectious cDNA clone of Turnip mosaic virus (TuMV) that was engineered to express a fluorescent protein (green fluorescent protein [GFP] or mCherry) fused to the viral 6K2 protein known to induce vesicle formation. Cytoplasmic fluorescent discrete protein structures were observed in infected cells, corresponding to the vesicles containing the viral RNA replication complex. The vesicles were motile and aligned with microfilaments. Intracellular movement of the vesicles was inhibited when cells were infiltrated with latrunculin B, an inhibitor of microfilament polymerization. It was also observed that viral accumulation in the presence of this drug was reduced. These data indicate that microfilaments are used for vesicle movement and are necessary for virus production. Biogenesis of the vesicles was further investigated by infecting cells with two recombinant TuMV strains: one expressed 6K2GFP and the other expressed 6K2mCherry. Green- and red-only vesicles were observed within the same cell, suggesting that each vesicle originated from a single viral genome. There were also vesicles that exhibited sectors of green, red, or yellow fluorescence, an indication that fusion among individual vesicles is possible. Protoplasts derived from TuMV-infected N. benthamiana leaves were isolated. Using immunofluorescence staining and confocal microscopy, viral RNA synthesis sites were visualized as punctate structures distributed throughout the cytoplasm. The viral proteins VPg-Pro, RNA-dependent RNA polymerase, and cytoplasmic inclusion protein (helicase) and host translation factors were found to be associated with these structures. A single-genome origin and presence of protein synthetic machinery components suggest that translation of viral RNA is taking place within the vesicle.

Individualized Prophylactic Neck Irradiation in Patients with cN0 Head and Neck Cancer Based on Sentinel Lymph Node(s) Identification: Definitive Results of a Prospective Phase 1-2 Study.

PURPOSE: This prospective, nonrandomized, interventional phase 1-2 study investigated the individualization of elective node irradiation in clinically N0 head and neck squamous cell carcinoma by sentinel lymph node (SLN) mapping with single-photon emission computed tomography/computed tomography (SPECT/CT) and its impact on tumor control and radiation-related toxicity. METHODS AND MATERIALS: Forty-four patients with clinically N0 head and neck squamous cell carcinoma treated with definitive (chemo-)radiation therapy were imaged with SPECT/CT after 99mTc nanocolloid injection around the tumor. The neck levels containing up to the 4 hottest SLNs were selected for prophylactic irradiation. A comparative virtual planning was performed with the selection of neck levels based on the current international guidelines. Regional control was monitored as a function of the selected volume. Dosimetric data for the organs at risk were compared between the plans. Normal tissue complication probability (NTCP) rates were derived for xerostomia, dysphagia, and hypothyroidism to predict the clinical benefit and correlated to quality-of-life (QoL) assessments at 6 months. RESULTS: Sixteen percent of patients presented unpredicted lymphatic drainage, and 48% drained unilaterally. The nodal clinical target volume based on lymphoscintigraphy was smaller than the nodal clinical target volume based on international guidelines by a factor of 2 (P < .0001). After a median follow-up of 46 months, only 1 patient experienced a regional relapse in a nonirradiated area. Significant median dose reductions to organs at risk were observed, particularly to contralateral salivary glands in patients with unilateral drainage (14.6-28.1 Gy) and to the thyroid gland in all patients (22.4-48.9 Gy). Median NTCP reductions were observed for xerostomia (0.3% to 13.7%), dysphagia (1.7% to 10.8%), and hypothyroidism (14.0% to 36.1%). QoL at 6 months was improved, particularly in patients irradiated unilaterally. CONCLUSIONS: Neck SLN mapping with SPECT/CT individualizes and reduces the elective nodal target volumes without compromising the regional control. The NTCP rates were reduced and favorable QoL were observed in all patients, particularly in the case of unilateral irradiation.

Inconclusive triple diagnosis in breast cancer imaging: is there a place for scintimammography?

UNLABELLED: Scintimammography (SM) can be used as a complementary test to mammography in patients with suspected breast cancers. This study was undertaken to evaluate the impact of SM on the management of patients with a doubtful or discordant triple diagnosis-that is, mammography, ultrasound, and fine-needle aspiration cytology. The clinical question was variable: initial diagnosis of cancer, suspicion of recurrence, doubtful tumor extension, or search for a primary tumor. METHODS: We performed a retrospective study of 118 procedures in 104 patients with a suggestion of breast cancer, either at initial presentation or after treatment (relapse), with an inconclusive triple diagnosis. Planar and tomographic imaging was performed after injection of (99m)Tc-labeled methoxyisobutylisonitrile ((99m)Tc-MIBI). Results were compared with histopathologic analysis (surgery or core biopsy) in 82 cases and with clinical and imaging follow-up in 36 cases. RESULTS: Breast cancer was proven in 69 cases. SM-SPECT had a sensitivity of 88.4% and a specificity of 67%. Eleven cancers were detected by SPECT, although planar images were negative. SM-SPECT was more sensitive in patients scanned at initial presentation (95%) than in those with suspected recurrence (81%). SM-SPECT correctly evaluated multicentricity or bilaterality in 8 of 11 patients and resulted in an increased tumor size in 8 patients. Overall, SM-SPECT modified the patient management in 58 of 118 cases (49%): SM made the diagnosis of cancer in 30 cases with doubtful or discordant triple diagnosis and ruled out malignancy in 28 cases. CONCLUSION: SM-SPECT is a useful complementary tool for the diagnosis and evaluation of disease extent in patients with an inconclusive triple diagnosis including fine-needle aspiration. The procedure altered the patient management in 49% of the population. These results must be confirmed in a prospective trial.

Akt regulates COX-2 mRNA and protein expression in mutated-PTEN human endometrial cancer cells.

In human endometrial cancer, the fourth most common cancer in women, tumor suppressor phosphatase tensin homologue (PTEN) is frequently mutated. In the presence of a mutated PTEN protein, Akt phosphorylation levels are increased leading to the activation of this survival pathway. Numerous studies indicated that COX-2 is inappropriately induced and up-regulated in a number of malignant cancer cells. COX-2 plays an important role in tumor cell biology, taking part actively in angiogenesis particularly via the production of prostaglandin E2 (PGE2). The present study was undertaken to determine the involvement of PI 3-K/Akt pathway in the regulation of COXs expression and PGE2 synthesis. Three different human endometrial cancer cell lines known to have wild-type PTEN (HEC 1-A) or a mutated inactive PTEN protein (RL 95-2 and Ishikawa) were used for these studies. Results showed that Akt phosphorylation was high in mutated PTEN cells. RT-PCR studies revealed that Akt1 and Akt2 were the regulated forms whereas Akt3 mRNA was nearly undetectable. COX-2 mRNA expression and protein levels were high in these cells compared to wild-type PTEN cells as demonstrated by RT-PCR and Western analysis respectively. PGE2 production was higher in mutated-PTEN expressing phospho-Akt and COX-2 compared to wild-type PTEN cells. Inhibition of PI 3-K with Wortmannin and LY294002 blocked Akt phosphorylation and inhibited expression of COX-2 in mutated-PTEN cells. Inhibition of Akt phosphorylation with specific PI 3-K inhibitors and down-regulation of COX-2 increased apoptosis in human endometrial cancer cells. Likewise, transfection of mutated-PTEN cells with a dominant negative Akt vector, resulted in COX-2 down-regulation and activation of apoptosis, as demonstrated by Hoechst nuclear staining. On the opposite, activation of Akt using a constitutively active expression vector, resulted in the up-regulation of COX-2 protein expression. Specific inhibition of COX-2 with NS-398 induced apoptosis in COX-2 expressing human endometrial cancer cells. It is concluded that the PI 3-K/Akt survival pathway is involved in the regulation of COX-2 and PGE2 synthesis in human endometrial cancer cells.

SPECT/CT lymphoscintigraphy of sentinel node(s) for superselective prophylactic irradiation of the neck in cN0 head and neck cancer patients: a prospective phase I feasibility study.

BACKGROUND: Clinically node negative HNSCC patients have a risk ranging between 18 and 45% of occult metastases, making prophylactic irradiation mandatory. Selective irradiation of nodal target volume based on international guidelines is practice. Anyway, about half the tumours lying in an anatomical subsite known to potentially drain bilaterally effectively do so, leading to unnecessary large volume irradiation. Moreover, 15% of the tumours show drainage outside of predicted basin, increasing the risk for potential geographical misses. Three-dimensional SPECT/CT lymphoscintigraphy (LS) of sentinel node(s) may help to individualize nodal target volume selection. This prospective phase I study explores its feasibility and the dosimetric impact. METHODS: Ten cN0 HNSCC patients eligible for definitive radiotherapy were imaged with SPECT/CT after 99mTc nanocolloid injection around the tumour. The neck levels containing up to four hottest nodes were identified and selected for prophylactic irradiation (CTVn-LS) by volumetric modulated arc therapy. A comparative virtual planning was performed with volumes selected according to international guidelines (CTVn-IG). RESULTS: Migration was observed in all patients (one with gamma probe only). 2.9 sentinel nodes were detected per patient on average. In some patients, accurate localization was difficult when not using thermoplastic mask for SPECT/CT. CTVn-LS was totally encompassed by CTVn-IG in all patients but one (unpredicted drainage in retropharyngeal level). On average, CTVn-LS and related planning target volumes were two times smaller than IG ones. This led to significant dose decrease in identified organs at risk as well as remaining volume at risk. CONCLUSIONS: SPECT/CT LS is a promising tool to individualize prophylactic node CTV in cN0 HNSCC patients eligible for definitive radiotherapy. Oncological safety must be confirmed by ongoing phase II study.

Eukaryotic elongation factor 1A interacts with Turnip mosaic virus RNA-dependent RNA polymerase and VPg-Pro in virus-induced vesicles.

Eukaryotic elongation factor 1-alpha (eEF1A) was identified as an interactor of Turnip mosaic virus (TuMV) RNA-dependent RNA polymerase (RdRp) and VPg-protease (VPg-Pro) using tandem affinity purification and/or in vitro assays. Subcellular fractionation experiments revealed that the level of eEF1A substantially increased in membrane fractions upon TuMV infection. Replication of TuMV occurs in cytoplasmic membrane vesicles, which are induced by 6K-VPg-Pro. Confocal microscopy indicated that eEF1A was included in these vesicles. To confirm that eEF1A was found in replication vesicles, we constructed an infectious recombinant TuMV that contains an additional copy of the 6K protein fused to the green fluorescent protein (GFP). In cells infected with this recombinant TuMV, fluorescence emitted by 6KGFP was associated with cytoplasmic membrane vesicles that contained VPg-Pro, the eukaryotic initiation factor (iso) 4E, the poly(A)-binding protein, the heat shock cognate 70-3 protein, and eEF1A. These results suggest that TuMV-induced membrane vesicles host at least three plant translation factors in addition to the viral replication proteins.

Transforming growth factor-beta3 increases the invasiveness of endometrial carcinoma cells through phosphatidylinositol 3-kinase-dependent up-regulation of X-linked inhibitor of apoptosis and protein kinase c-dependent induction of matrix metalloproteinase-9.

Tumor cells often acquire intrinsic resistance to the growth inhibitory and pro-apoptotic effects of transforming growth factor-beta (TGF-beta); moreover, TGF-beta can confer invasive properties to established tumor cells. In the present study, we show that TGF-beta isoforms (TGF-beta1, TGF-beta2, and TGF-beta3) trigger proper Smad signaling in human endometrial carcinoma cell lines and efficiently inhibit cellular proliferation. These cells, however, exhibit a high degree of resistance to TGF-beta pro-apoptotic effects; we found that this resistant phenotype would be acquired through up-regulation of X-linked inhibitor of apoptosis protein (XIAP) levels. In addition, using RNA interference and pharmacological inhibitors, we show that TGF-beta increases cellular invasiveness via two distinct signaling pathways in endometrial carcinoma cells: phosphatidylinositol 3-kinase/AKT-dependent up-regulation of XIAP and protein kinase C-dependent induction of matrix-metalloproteinase-9 (MMP-9) expression. Additionally, these findings were correlated with clinical observations showing abundant TGF-beta immunoreactivity in human endometrial carcinoma tumors in vivo, extending from the epithelial compartment to the stroma upon acquisition of an invasive phenotype (gradually from grades I to III). Collectively our results describe for the first time a role for TGF-beta3 in tumor invasiveness.

AKT involvement in cisplatin chemoresistance of human uterine cancer cells.

OBJECTIVE: The aim of this study was to investigate the possible involvement of Akt activity and specific isoforms (Akt1, Akt2, and Akt3) in the resistance of human uterine cancer cells to cisplatin. METHODS: Two different endometrial (HEC-1-A and KLE) and one cervical (HeLa) cancer cell lines all known as wild-type PTEN (tumor suppressor phosphatase tensin homologue, a lipid phosphatase involved in the negative regulation of Akt activity) were used for these studies. RESULTS: Basal levels of Akt1, Akt2, and Akt3 mRNAs were determined by real-time quantitative RT-PCR studies and Western blot analyses were carried out to determine protein abundance of each isoforms. Akt1 mRNA and protein were present in all cell lines studied. Akt2 and Akt3 mRNAs and proteins were strongly expressed in KLE cells. Surprisingly, Akt phosphorylation was found in KLE expressing high levels of wild-type PTEN protein. KLE cells remained resistant to PI 3-K inhibitor, indicating that Akt phosphorylation might be, in part, independent of PI 3-K in this cell line. Cisplatin induced apoptosis in HeLa and HEC-1-A cells, but KLE cells expressing Akt2 and Akt3 remained more resistant to cisplatin. Knockout of Akt isoforms using specific siRNA technology increased the sensitivity of KLE cells toward cisplatin and caused a significant induction of cell death. CONCLUSION: Taken together, these results suggest that specific Akt isoforms such as Akt2 and Akt3 might be involved in chemoresistance to cisplatin and that these isoforms could be putative targets for gene therapy in uterine cancers.

Synthesis of 17beta-estradiol platinum(II) complexes: biological evaluation on breast cancer cell lines.

The synthesis of a novel series of 17beta-estradiol-linked platinum(II) complexes is described. The new molecules are linked with an alkyl chain at position 16alpha of the steroid nucleus and bear a 16beta-hydroxymethyl side chain. They are made from estrone in five chemical steps with an overall yield exceeding 28%. The biological activity of these compounds was evaluated in vitro on estrogen dependent and independent (ER+ and ER-) human breast cancers. The derivatives incorporating a 2-(2'-aminoethyl)pyridine ligand displayed good activity against the cell lines particularly when the connecting arm is 10 carbon atoms long.

Quantitation in PET using isotopes emitting prompt single gammas: application to yttrium-86.

Several yttrium-90 labelled somatostatin analogues are now available for cancer radiotherapy. After injection, a large amount of the compound is excreted via the urinary tract, while a variable part is trapped in the tumour(s), allowing the curative effect. Unfortunately, the compound may also be trapped in critical tissues such as kidney or bone marrow. As a consequence, a method for assessment of individual biodistribution and pharmacokinetics is required to predict the maximum dose that can be safely injected into patients. However, (90)Y, a pure beta(-)particle emitter, cannot be used for quantitative imaging. Yttrium-86 is a positron emitter that allows imaging of tissue uptake using a PET camera. In addition to the positron, (86)Y also emits a multitude of prompt single gamma-rays, leading to significant overestimation of uptake when using classical reconstruction methods. We propose a patient-dependent correction method based on sinogram tail fitting using an (86)Y point spread function library. When applied to abdominal phantom acquisition data, the proposed correction method significantly improved the accuracy of the quantification: the initial overestimation of background activity by 117% was reduced to 9%, while the initial error in respect of kidney uptake by 84% was reduced to 5%. In patient studies, the mean discrepancy between PET total body activity and the activity expected from urinary collections was reduced from 92% to 7%, showing the benefit of the proposed correction method.

86Y-DOTA0)-D-Phe1-Tyr3-octreotide (SMT487)--a phase 1 clinical study: pharmacokinetics, biodistribution and renal protective effect of different regimens of amino acid co-infusion.

The pharmacokinetics and dosimetry of (86)Y-DOTA(0)- d-Phe(1)-Tyr(3)-octreotide ((86)Y-SMT487) were evaluated in a phase I positron emission tomography (PET) study of 24 patients with somatostatin receptor-positive neuroendocrine tumours. The effect of amino acid (AA) co-infusion on renal and tumour uptake was assessed in a cross-over randomised setting. Five regimens were tested: no infusion, 4-h infusion of 120 g mixed AA (26.4 g l-lysine + l-arginine), 4 h l-lysine (50 g), 10 h 240 g mixed AA (52.8 g l-lysine + l-arginine) and 4 h Lys-Arg (25 g each). Comparisons were performed on an intra-patient basis. Infusions of AA started 0.5 h prior to injection of (86)Y-SMT487 and PET scans were obtained at 4, 24 and 48 h p.i. Absorbed doses to tissues were computed using the MIRD3 method. (86)Y-SMT487 displayed rapid plasma clearance and exclusive renal excretion; uptake was noted in kidneys, tumours, spleen and, to a lesser extent, liver. The 4-h mixed AA co-infusion significantly ( P<0.05) reduced (86)Y-SMT487 renal uptake by a mean of 21%. This protective effect was significant on the dosimetry data (3.3+/-1.3 vs 4.4+/-1.0 mGy/MBq; P<0.05) and was further enhanced upon prolonging the infusion to 10 h (2.1+/-0.4 vs 1.7+/-0.2 mGy/MBq; P<0.05). Infusion of Lys-Arg but not of l-lysine was more effective in reducing renal uptake than mixed AA. Infusion of AA did not result in reduced tumour uptake. The amount of (90)Y-SMT487 (maximum allowed dose: MAD) that would result in a 23-Gy cut-off dose to kidneys was calculated for each study: MAD was higher with mixed AA co-infusion by a mean of 46% (10-114%, P<0.05 vs no infusion). In comparison with 4 h mixed AA, the MAD was higher by a mean of 23% (9-37%; P<0.05) with prolonged infusion and by a mean of 16% (2-28%; P<0.05) with Lys-Arg. We conclude that infusion of large amounts of AA reduces renal exposure during peptide-based radiotherapy and allows higher absorbed doses to tumours. The prolongation of the infusion from 4 to 10 h further enhances the protective effect on the kidneys.