Porphyrin-High-Density Lipoprotein: A Novel Photosensitizing Nanoparticle for Lung Cancer Therapy.

BACKGROUND: We have developed ultrasmall porphyrin-high-density lipoprotein (HDL) nanoparticles (<20 nm), called "porphyrinHDL," that have a high density of porphyrin molecules and dissociate rapidly upon tumor cell accumulation to become fluorescent and photoactive. This is introduced as a novel activatable photosensitizer for image-guided photodynamic therapy (PDT). Here, we report the studies of these nanoparticles targeted to scavenger receptor class B type I (SR-BI) expressed on lung cancer cells as a first step toward development of a minimally invasive treatment for peripheral lung cancer and metastatic lymph nodes of advanced lung cancer. METHODS: The in vitro uptake of porphyrinHDL and the corresponding PDT efficacy were evaluated in both SR-BI-positive and SR-BI-negative lung cancer cell lines. A clinically relevant orthotopic lung cancer model in mice was used to examine fluorescence activation and quantification of uptake in tumor. In addition, we investigated the effect of porphyrinHDL-mediated PDT. RESULTS: PorphyrinHDL promoted proper intracellular uptake in the H460 human lung cancer cell line. When irradiated with a 671-nm PDT laser, porphyrinHDL produced significant therapeutic effectiveness in vitro. After systemic administration in mice with orthotopic lung cancer xenografts, porphyrinHDL demonstrated selective accumulation and photoactivation in tumor with significantly enhanced disease-to-normal tissue contrast. Moreover, porphyrinHDL-PDT significantly induced cell apoptosis in lung tumors (73.2%) without toxicity in normal tissues or damage to adjacent critical structures. CONCLUSIONS: SR-BI-targeted porphyrinHDL-mediated PDT of lung cancer is selective and effective in vitro and in vivo. These initial proof-of-principle studies suggest the potential of a "smart" PDT approach for highly selective tumor ablation.

Preclinical investigation of folate receptor-targeted nanoparticles for photodynamic therapy of malignant pleural mesothelioma.

Photodynamic therapy (PDT) following lung-sparing extended pleurectomy for malignant pleural mesothelioma (MPM) has been investigated as a potential means to kill residual microscopic cells. High expression levels of folate receptor 1 (FOLR1) have been reported in MPM; therefore, targeting FOLR1 has been considered a novel potential strategy. The present study developed FOLR1­targeting porphyrin-lipid nanoparticles (folate-porphysomes, FP) for the treatment of PDT. Furthermore, inhibition of activated epidermal growth factor (EGFR)-associated survival pathways enhance PDT efficacy. In the present study, these approaches were combined; FP-based PDT was used together with an EGFR-tyrosine kinase inhibitor (EGFR-TKI). The frequency of FOLR1 and EGFR expression in MPM was analyzed using tissue microarrays. Confocal microscopy and a cell viability assay were performed to confirm the specificity of FOLR1­targeting cellular uptake and photocytotoxicity in vitro. In vivo fluorescence activation and therapeutic efficacy were subsequently examined. The effects of EGFR-TKI were also assessed in vitro. The in vivo combined antitumor effect of EGFR-TKI and FP-PDT was then evaluated. The results revealed that FOLR1 and EGFR were expressed in 79 and 89% of MPM samples, respectively. In addition, intracellular uptake of FP corresponded well with FOLR1 expression. When MPM cells were incubated with FP and then irradiated at 671 nm, there was significant in vitro cell death, which was inhibited in the presence of free folic acid, thus suggesting the specificity of FPs. FOLR1 targeting resulted in disassembly of the porphysomes and subsequent fluorescence activation in intrathoracic disseminated MPM tumors, as demonstrated by ex vivo tissue imaging. FP-PDT resulted in significant cellular damage and apoptosis in vivo. Furthermore, the combination of pretreatment with EGFR-TKI and FP-PDT induced a marked improvement of treatment responses. In conclusion, FP-based PDT induced selective destruction of MPM cells based on FOLR1 targeting, and pretreatment with EGFR-TKI further enhanced the therapeutic response.

Personalized siRNA-Nanoparticle Systemic Therapy using Metastatic Lymph Node Specimens Obtained with EBUS-TBNA in Lung Cancer.

Inhibiting specific gene expression with siRNA provides a new therapeutic strategy to tackle many diseases at the molecular level. Recent strategies called high-density lipoprotein (HDL)-mimicking peptide-phospholipid nanoscaffold (HPPS) nanoparticles have been used to induce siRNAs-targeted delivery to scavenger receptor class B type I receptor (SCARB1)-expressing cancer cells with high efficiency. Here, eight ideal therapeutic target genes were identified for advanced lung cancer throughout the screenings using endobronchial ultrasonography-guided transbronchial needle aspiration (EBUS-TBNA) and the establishment of a personalized siRNA-nanoparticle therapy. The relevance of these genes was evaluated by means of siRNA experiments in cancer cell growth. To establish a therapeutic model, kinesin family member-11 (KIF11) was selected as a target gene. A total of 356 lung cancers were analyzed immunohistochemically for its clinicopathologic significance. The antitumor effect of HPPS-conjugated siRNA was evaluated in vivo using xenograft tumor models. Inhibition of gene expression for these targets effectively suppressed lung cancer cell growth. SCARB1 was highly expressed in a subset of tumors from the lung large-cell carcinoma (LCC) and small-cell lung cancer (SCLC) patients. High-level KIF11 expression was identified as an independent prognostic factor in LCC and squamous cell carcinoma (SqCC) patients. Finally, a conjugate of siRNA against KIF11 and HPPS nanoparticles induced downregulation of KIF11 expression and mediated dramatic inhibition of tumor growth in vivoImplications: This approach showed delivering personalized cancer-specific siRNAs via the appropriate nanocarrier may be a novel therapeutic option for patients with advanced lung cancer. Mol Cancer Res; 16(1); 47-57. ©2017 AACR.

Nanoparticle targeted folate receptor 1-enhanced photodynamic therapy for lung cancer.

OBJECTIVE: Despite modest improvements, the prognosis of lung cancer patients has still remained poor and new treatment are urgently needed. Photodynamic therapy (PDT), the use of light-activated compounds (photosensitizers) is a treatment option but its use has been restricted to central airway lesions. Here, we report the use of novel porphyrin-lipid nanoparticles (porphysomes) targeted to folate receptor 1 (FOLR1) to enhance the efficacy and specificity of PDT that may translate into a minimally-invasive intervention for peripheral lung cancer and metastatic lymph nodes of advanced lung cancer. MATERIALS AND METHODS: The frequency of FOLR1 expression in primary lung cancer and metastatic lymph nodes was first analyzed by human tissue samples from surgery and endobronchial ultrasonography-guided transbronchial needle aspiration (EBUS-TBNA). Confocal fluorescence microscopy was then used to confirm the cellular uptake and fluorescence activation in lung cancer cells, and the photocytotoxicity was evaluated using a cell viability assay. In vivo fluorescence activation and quantification of uptake were investigated in mouse lung orthotopic tumor models, followed by the evaluation of in vivo PDT efficacy. RESULTS: FOLR1 was highly expressed in metastatic lymph node samples from patients with advanced lung cancer and was mainly expressed in lung adenocarcinomas in primary lung cancer. Expression of FOLR1 in lung cancer cell lines corresponded with the intracellular uptake of folate-porphysomes in vitro. When irradiated with a 671nm laser at a dose of 10J/cm2, folate-porphysomes showed marked therapeutic efficacy compared with untargeted porphysomes (28% vs. 83% and 24% vs. 99% cell viability in A549 and SBC5 lung cancer cells, respectively). Systemically-administered folate-porphysomes accumulated in lung tumors with significantly enhanced disease-to-normal tissue contrast. Folate-porphysomes mediated PDT successfully inhibited tumor cell proliferation and activated tumor cell apoptosis. CONCLUSION: Folate-porphysome based PDT shows promise in selectively ablating lung cancer based on FOLR1 expression in these preclinical models.

Development of a novel ex vivo porcine laparoscopic Heller myotomy and Nissen fundoplication training model (Toronto lap-Nissen simulator).

BACKGROUND: Surgical trainees are required to develop competency in a variety of laparoscopic operations. Developing laparoscopic technical skills can be difficult as there has been a decrease in the number of procedures performed. This study aims to develop an inexpensive and anatomically relevant model for training in laparoscopic foregut procedures. METHODS: An ex vivo, anatomic model of the human upper abdomen was developed using intact porcine esophagus, stomach, diaphragm and spleen. The Toronto lap-Nissen simulator was contained in a laparoscopic box-trainer and included an arch system to simulate the normal radial shape and tension of the diaphragm. We integrated the use of this training model as a part of our laparoscopic skills laboratory-training curriculum. Afterwards, we surveyed trainees to evaluate the observed benefit of the learning session. RESULTS: Twenty-five trainees and five faculty members completed a survey regarding the use of this model. Among the trainees, only 4 (16%) had experience with laparoscopic Heller myotomy and Nissen fundoplication. They reported that practicing with the model was a valuable use of their limited time, repeating the exercise would be of additional benefit, and that the exercise improved their ability to perform or assist in an actual case in the operating room. Significant improvements were found in the following subjective measures comparing pre- vs. post-training: (I) knowledge level (5.6 vs. 8.0, P<0.001); (II) comfort level in assisting (6.3 vs. 7.6, P<0.001); and (III) comfort level in performing as the primary surgeon (4.9 vs. 7.1, P<0.001). The trainees and faculty members agreed that this model was of adequate fidelity and was a representative simulation of actual human anatomy. CONCLUSIONS: We developed an easily reproducible training model for laparoscopic procedures. This simulator reproduces human anatomy and increases the trainees' comfort level in performing and assisting with myotomy and fundoplication.

Overexpression of MAGEA2 has a prognostic significance and is a potential therapeutic target for patients with lung cancer.

Melanoma-associated antigens (MAGE) are expressed in different type of cancers including lung cancer and have been shown to be functionally related to p53 tumor suppressor gene. Little is known about the relationship between MAGE genes and p53 aberrant expression in lung cancer. The aims of this study were to observe the expression of MAGEA2, examine the role of MAGEA2 in lung cancer survival, investigate its correlation between MAGEA2 and p53, and explore its clinicopathologic significance as a prognostic marker. Quantitative reverse transcription-polymerase chain reaction was performed to detect the expression of MAGEA2 using 36 primary tumors and 31 metastatic lymph nodes from patients with lung cancer. The role of MAGEA2 in cancer cell growth and in the regulation of p53 downstream genes were examined using small interfering RNA. The expression of MAGEA2 and p53 were analyzed immunohistochemically using tissue microarray from 353 resected lung specimens. High-level expression of MAGEA2 (High-MAGEA2) was confirmed in lung tumors with high frequency. Inhibiting MAGEA2 expression effectively suppressed cancer cell growth and decreased the expression of p53 downstream target genes in vitro. In adenocarcinoma, High-MAGEA2 was strongly associated with aberrant p53 expression (P<0.001) and was associated with worse clinical outcomes (5-year OS, 87.1% in low vs. 74.1% in high, P=0.014). Aberrant p53 expression was also significant worse prognostic factor (P=0.029). Among the adenocarcinoma patients with wild-type p53, High-MAGEA2 had poorer prognosis than low-level MAGEA2 groups (5-year OS, 90.1% vs. 72.1%, P=0.037), whereas had no difference in p53 aberrant tumors. On multivariate analysis, MAGEA2 was independently associated with survival (hazard ratio; 2.12, P=0.030). In conclusion, suppression of MAGEA2 in lung cancer cells significantly reduced the growth/survival of cancer cells. High-MAGEA2 was identified as an independent prognostic factor in lung adenocarcinoma. Specific inhibition of MAGEA2 may be a promising therapeutic strategy for patients with lung cancer.

Initial characterization of behavior and ketamine response in a mouse knockout of the post-synaptic effector gene Anks1b.

The human ANKS1B gene encodes an activity-dependent effector of post-synaptic signaling. It was recently associated with neuropsychiatric phenotypes in genome-wide studies. While the biological function of ANKS1B has been partly elucidated, its role in behavior is poorly understood. Here, we breed and characterize a full knockout (KO) for murine Anks1b. We found that the homozygous KO genotype was partially lethal, showing significant deviation from expected segregation ratios at weaning. Behaviorally, KOs exhibited no difference in baseline acoustic startle response, but showed deficits in prepulse inhibition (PPI). KOs also exhibited locomotor hyperactivity and increased stereotypy at baseline. Administration of ketamine, a non-competitive NMDA-receptor antagonist, greatly exacerbated locomotor activity in the KOs at lower doses, but genotype groups were almost indistinguishable as dose increased. Stereotypy showed a complex response to ketamine in the KOs, with elevated stereotypy at lower doses and markedly less at high doses, compared to wild type. Our study is the first to probe the behavioral phenotypes associated with ablation of Anks1b. Deficits in PPI, locomotor hyperactivity, elevated stereotypy and altered response to NMDA receptor antagonism are murine behavioral outcomes with translational relevance for psychiatric disorders. These findings are also consistent with the role of Anks1b as an effector of glutamatergic signaling. As an intermediary between post-synaptic receptor stimulation and long-term changes to neuronal protein expression, further investigation of Anks1b is warranted.

First Evaluation of the New Thin Convex Probe Endobronchial Ultrasound Scope: A Human Ex Vivo Lung Study.

BACKGROUND: Endobronchial ultrasonography (EBUS)-guided transbronchial needle aspiration allows for sampling of mediastinal lymph nodes. The external diameter, rigidity, and angulation of the convex probe EBUS renders limited accessibility. This study compares the accessibility and transbronchial needle aspiration capability of the prototype thin convex probe EBUS against the convex probe EBUS in human ex vivo lungs rejected for transplant. METHODS: The prototype thin convex probe EBUS (BF-Y0055; Olympus, Tokyo, Japan) with a thinner tip (5.9 mm), greater upward angle (170 degrees), and decreased forward oblique direction of view (20 degrees) was compared with the current convex probe EBUS (6.9-mm tip, 120 degrees, and 35 degrees, respectively). Accessibility and transbronchial needle aspiration capability was assessed in ex vivo human lungs declined for lung transplant. The distance of maximum reach and sustainable endoscopic limit were measured. Transbronchial needle aspiration capability was assessed using the prototype 25G aspiration needle in segmental lymph nodes. RESULTS: In all evaluated lungs (n = 5), the thin convex probe EBUS demonstrated greater reach and a higher success rate, averaging 22.1 mm greater maximum reach and 10.3 mm further endoscopic visibility range than convex probe EBUS, and could assess selectively almost all segmental bronchi (98% right, 91% left), demonstrating nearly twice the accessibility as the convex probe EBUS (48% right, 47% left). The prototype successfully enabled cytologic assessment of subsegmental lymph nodes with adequate quality using the dedicated 25G aspiration needle. CONCLUSIONS: Thin convex probe EBUS has greater accessibility to peripheral airways in human lungs and is capable of sampling segmental lymph nodes using the aspiration needle. That will allow for more precise assessment of N1 nodes and, possibly, intrapulmonary lesions normally inaccessible to the conventional convex probe EBUS.

Evaluation of a New Ultrasound Thoracoscope for Localization of Lung Nodules in Ex Vivo Human Lungs.

BACKGROUND: Localization of small, nonvisible and nonpalpable nodules is challenging during video-assisted thoracoscopic surgery. We evaluated the feasibility of using a new ultrasound thoracoscope to localize nodules in resected ex vivo human lungs. METHODS: The tumor was localized and measured in its greatest dimension with a prototype ultrasound thoracoscope (XLTF-UC180; Olympus Corporation, Tokyo, Japan) at different frequencies (5.0 to 12.0 MHz) and different lung specimen states (deflated, semiinflated). Measured tumor size and depth from lung surface were compared and correlated to the true diameter and depth from lung surface acquired from pathologic morphology. RESULTS: Ex vivo evaluation was performed on 16 solid nodules and nine part solid ground-glass nodules. All tumors were successfully localized in the deflated lung specimens (average size, 13.7 ± 5.2 mm). The tumor boundaries were best evaluated with an ultrasound frequency of 10 MHz. Solid nodules were more easily visualized than ground-glass nodules. Part solid ground-glass nodules were not easily detected in the semiinflated specimen owing to peritumoral air surrounding the tumor. Tumor boundaries were also difficult to identify in deeply situated tumors and in lungs with underlying disease. A strong positive correlation existed between the ultrasound measurement and true measurement of tumor size (R2 = 0.89, p < 0.001). CONCLUSIONS: The ultrasound thoracoscope can be used to localize nodules in resected human lungs. The clarity of the tumor boundaries is influenced by the tumor type and depth and the underlying pulmonary disease. Complete lung deflation and the use of 10 MHz ultrasound frequency optimize the visualization of target tumors.

SORORIN and PLK1 as potential therapeutic targets in malignant pleural mesothelioma.

Malignant pleural mesothelioma (MPM) is an aggressive type of cancer of the thoracic cavity commonly associated with asbestos exposure and a high mortality rate. There is a need for new molecular targets for the development of more effective therapies for MPM. Using quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and an RNA interference-based screening, we examined the SORORIN gene as potential therapeutic targets for MPM in addition to the PLK1 gene, which is known for kinase of SORORIN. Following in vitro investigation of the effects of target silencing on MPM cells, cell cycle analyses were performed. SORORIN expression was analyzed immunohistochemically using a total of 53 MPM samples on tissue microarray. SORORIN was found to be overexpressed in the majority of clinical MPM samples and human MPM cell lines as determined by qRT-PCR. Gene suppression of each SORORIN and PLK1 led to growth inhibition in MPM cell lines. Knockdown of SORORIN showed an increased number of G2M-phase population and a larger nuclear size, suggesting mitotic arrest. High expression of SORORIN (SORORIN-H) was found in 50.9% of all the MPM cases, and there is a tendency towards poorer prognosis for the SORORIN-H group but the difference is not significant. Suppression of SORORIN with PLK1 inhibitor BI 6727 showed a combinational growth suppressive effect on MPM cell growth. Given high-dose PLK1 inhibitor induced drug-related adverse effects in several clinical trials, our results suggest inhibition SORORIN-PLK1 axis may hold promise for the treatment of MPMs.

Multi-Modal Imaging in a Mouse Model of Orthotopic Lung Cancer.

BACKGROUND: Investigation of CF800, a novel PEGylated nano-liposomal imaging agent containing indocyanine green (ICG) and iohexol, for real-time near infrared (NIR) fluorescence and computed tomography (CT) image-guided surgery in an orthotopic lung cancer model in nude mice. METHODS: CF800 was intravenously administered into 13 mice bearing the H460 orthotopic human lung cancer. At 48 h post-injection (peak imaging agent accumulation time point), ex vivo NIR and CT imaging was performed. A clinical NIR imaging system (SPY®, Novadaq) was used to measure fluorescence intensity of tumor and lung. Tumor-to-background-ratios (TBR) were calculated in inflated and deflated states. The mean Hounsfield unit (HU) of lung tumor was quantified using the CT data set and a semi-automated threshold-based method. Histological evaluation using H&E, the macrophage marker F4/80 and the endothelial cell marker CD31, was performed, and compared to the liposomal fluorescence signal obtained from adjacent tissue sections. RESULTS: The fluorescence TBR measured when the lung is in the inflated state (2.0 ± 0.58) was significantly greater than in the deflated state (1.42 ± 0.380 (n = 7, p<0.003). Mean fluorescent signal in tumor was highly variable across samples, (49.0 ± 18.8 AU). CT image analysis revealed greater contrast enhancement in lung tumors (a mean increase of 110 ± 57 HU) when CF800 is administered compared to the no contrast enhanced tumors (p = 0.0002). CONCLUSION: Preliminary data suggests that the high fluorescence TBR and CT tumor contrast enhancement provided by CF800 may have clinical utility in localization of lung cancer during CT and NIR image-guided surgery.

Formulation of hydrophobic therapeutics with self-assembling peptide and amino acid: A new platform for intravenous drug delivery.

Clinical application of hydrophobic therapeutics is restricted by lack of an efficient vehicle which permits their solubility in aqueous environments. We have previously developed a novel formulation strategy to deliver a hydrophobic Src inhibitor, PP2, involving combinations of one self-assembling peptide (SAP) and one of 4 selected amino acids (AAs). The present study aims to develop a generalized drug delivery platform for intravenous application of hydrophobic drugs by combining self-assembling peptide, amino acid and low concentration of co-solvent. A multi-step screening pipeline is established which includes assessment of drug solubility and physicochemical characteristics, as well as functional efficacy and safety in vitro and in vivo. Using PP2 as an exemplary hydrophobic compound, 480 different combinations of 6 SAPs, 20 naturally existing AAs at 2 concentrations, and 2 co-solvents were evaluated. Among the combinations, 60 formulae dissolved PP2; 10 of which significantly reduced thrombin-induced IL-8 production, a sign of inflammatory response, in normal human lung epithelial BEAS2B cells. These formulations did not show cytotoxicity alone, but 2 reduced cell viability with presence of thrombin. We then performed a double-blinded test in a rat model of pulmonary ischemia-reperfusion. PP2 formulated with EAK16-I peptide plus methionine and 2% ethanol were administrated intravenously, significantly reducing severity of lung injury. The SAP-AA formulation strategy was also successfully applied to other hydrophobic compounds, suggesting this strategy could be applicable to other hydrophobics for a variety of clinical applications.

Kinesin family members KIF11 and KIF23 as potential therapeutic targets in malignant pleural mesothelioma.

Malignant pleural mesothelioma (MPM) is a rare and aggressive form of cancer commonly associated with asbestos exposure that stems from the thoracic mesothelium with high mortality rate. Currently, treatment options for MPM are limited, and new molecular targets for treatments are urgently needed. Using quantitative reverse transcription-polymerase chain reaction (RT-PCR) and an RNA interference-based screening, we screened two kinesin family members as potential therapeutic targets for MPM. Following in vitro investigation of the target silencing effects on MPM cells, a total of 53 MPMs were analyzed immunohistochemically with tissue microarray. KIF11 and KIF23 transcripts were found to be overexpressed in the majority of clinical MPM samples as well as human MPM cell lines as determined by quantitative RT-PCR. Gene knockdown in MPM cell lines identified growth inhibition following knockdown of KIF11 and KIF23. High expression of KIF11 (KIF11-H) and KIF23 (KIF23-H) were found in 43.4 and 50.9% of all the MPM cases, respectively. Patients who received curative resection with tumors displaying KIF23-H showed shorter overall survival (P=0.0194). These results provide that inhibition of KIF11 and KIF23 may hold promise for treatment of MPMs, raising the possibility that kinesin-based drug targets may be developed in the future.

Overexpression of KIF23 predicts clinical outcome in primary lung cancer patients.

OBJECTIVE: High-level expression of kinesin family member 23 (KIF23), a member of microtubule-dependent molecular motors that transport organelles within cells and move chromosomes during cell division, has been observed in a variety of human malignancies. The aims of the present study were to observe the expression of KIF23 in lung cancer, examine the role of KIF23 in lung cancer cell growth and/or survival by small interfering RNA experiments, and explore its clinicopathologic significance and evaluate KIF23 expression as a prognostic marker. MATERIALS AND METHODS: Quantitative reverse transcription-polymerase chain reaction analysis was performed to detect the expression of KIF23 mRNA using metastatic lymph nodes from patients with advanced lung cancer obtained by endobronchial ultrasonography-guided transbronchial needle aspiration (EBUS-TBNA) and primary lung tumors through surgical sample. The role of KIF23 in cancer cell growth was examined by small interfering RNA experiments. A total of 339 lung cancers were analyzed immunohistochemically on tissue microarrays to examine the expression of KIF23 protein and its clinicopathologic significance. RESULTS: KIF23 transcript was found to be overexpressed in the great majority of metastatic lymph nodes from advanced lung cancers and primary lung tumors. Inhibiting KIF23 expression effectively suppressed lung cancer cell growth. High-level KIF23 expression was observed in 67.8% of the 339 cases. Lung adenocarcinoma patients with tumors displaying a high-level of KIF23 expression was also identified as an independent prognostic factor by multivariate analysis (P=0.0064). CONCLUSION: KIF23 not only provides additional prognostic information for surgical treatment of lung cancer, but may also be a novel therapeutic target for these patients.

Effective delivery of a rationally designed intracellular peptide drug with gold nanoparticle-peptide hybrids.

A novel gold nanoparticle-peptide hybrid strategy was developed to intracellularly deliver a potent PKCδ inhibitor peptide for the treatment of acute lung injury. The gold nanoparticle-peptide hybrids showed good stability with high uptake, and demonstrated in vitro and in vivo efficacy. Our formulation strategy shows great promise in intracellular delivery of peptides.

Loss of glucagon-like peptide-2-induced proliferation following intestinal epithelial insulin-like growth factor-1-receptor deletion.

BACKGROUND & AIMS: Glucagon-like peptide-2 (GLP-2) is an intestinal hormone that promotes growth of the gastrointestinal tract. Although insulin-like growth factor (IGF)-1 and the IGF-1 receptor (IGF-1R) are required for GLP-2-induced proliferation of crypt cells, little is known about localization of the IGF-1R which mediates the intestinotropic actions of GLP-2. METHODS: We examined intestinal growth and proliferative responses in mice with conditional deletion of IGF-1R from intestinal epithelial cells (IE-igf1rKO) after acute administration (30-90 min) of GLP-2, in response to 24-hour fasting and re-feeding (to induce GLP-2-dependent adaptation), and after chronic exposure (10 days) to GLP-2. RESULTS: IE-igf1rKO mice had normal small intestinal weight, morphometric parameters, proliferative indices, and distribution of differentiated epithelial cell lineages. Acute administration of GLP-2 increased nuclear translocation of ß-catenin in non-Paneth crypt cells and stimulated the crypt-cell proliferative marker c-Myc in control but not IE-igf1rKO mice. Small intestinal weight, crypt depth, villus height, and crypt-cell proliferation were decreased in control and IE-igf1rKO mice after 24-hour fasting. Although re-feeding control mice restored all of these parameters, re-fed IE-igf1rKO mice had reductions in adaptive regrowth of the villi and crypt-cell proliferation. Control mice that were given chronic GLP-2 had increases in small intestinal weight, mucosal cross-sectional area, crypt depth, villus height, and crypt-cell proliferation. However, the GLP-2-induced increase in crypt-cell proliferation was not observed in IE-igf1rKO mice, and growth of the crypt-villus axis was reduced. CONCLUSIONS: The proliferative responses of the intestinal epithelium to exogenous GLP-2 administration and conditions of GLP-2-dependent adaptive re-growth require the intestinal epithelial IGF-1R.