Insulin-like growth factor-I inhibition with pasireotide decreases cell proliferation and increases apoptosis in pre-malignant lesions of the breast: a phase 1 proof of principle trial

Baljit Singh, Julia A Smith, Deborah M Axelrod, Pietro Ameri, Heather Levitt, Ann Danoff, Martin Lesser, Cristina de Angelis, Irineu Illa-Bochaca, Sara Lubitz, Daniel Huberman, Farbod Darvishian, David L Kleinberg, Baljit Singh, Julia A Smith, Deborah M Axelrod, Pietro Ameri, Heather Levitt, Ann Danoff, Martin Lesser, Cristina de Angelis, Irineu Illa-Bochaca, Sara Lubitz, Daniel Huberman, Farbod Darvishian, David L Kleinberg

Abstract

Introduction: Estrogen inhibition is effective in preventing breast cancer in only up to 50% of women with precancerous lesions and many experience side effects that are poorly tolerated. As insulin-like growth factor I (IGF-I) underlies both estrogen and progesterone actions and has other direct effects on mammary development and carcinogenesis, we hypothesized that IGF-I inhibition might provide a novel approach for breast cancer chemoprevention.

Methods: In total, 13 women with core breast biopsies diagnostic of atypical hyperplasia (AH) were treated for 10 days with pasireotide, a somatostatin analog which uniquely inhibits IGF-I action in the mammary gland. They then had excision biopsies. 12 patients also had proliferative lesions and one a ductal carcinoma in situ (DCIS). Primary outcomes were changes in cell proliferation and apoptosis after treatment. Expression of estrogen receptor (ER), progesterone receptor (PR), and phosphorylated Insulin-like growth factor I receptor (IGF-1R), protein kinase B (AKT) and extracellular signal-regulated kinases 1/2 (ERK1/2) were also assessed. Core and excision biopsies from 14 untreated patients served as non-blinded controls. Hyperglycemia and other side effects were carefully monitored.

Results: Pasireotide decreased proliferation and increased apoptosis in all AH (from 3.6 ± 2.6% to 1.3 ± 1.2% and from 0.3 ± 0.2% to 1.5 ± 1.6%, respectively) and proliferative lesions (from 3.8 ± 2.5% to 1.8 ± 1.8% and from 0.3 ± 0.2% to 1.3 ± 0.6%, respectively). The DCIS responded similarly. ER and PR were not affected by pasireotide, while IGF-1R, ERK1/2 and AKT phosphorylation decreased significantly. In contrast, tissue from untreated controls showed no change in cell proliferation or phosphorylation of IGF-1R, AKT or ERK 1/2. Mild to moderate hyperglycemia associated with reduced insulin levels was found. Glucose fell into the normal range after discontinuing treatment. Pasireotide was well tolerated and did not cause symptoms of estrogen deprivation.

Conclusions: IGF-I inhibition by pasireotide, acting through the IGF-1R, was associated with decreased proliferation and increased apoptosis in pre-malignant breast lesions and one DCIS. Assuming hyperglycemia can be controlled, these data suggest that inhibiting the IGF-I pathway may prove an effective alternative for breast cancer chemoprevention.

Trial registration: NCT01372644 Trial date: July 1, 2007.

Figures

Figure 1
Figure 1
Trial profile. IGF, insulin-like growth factor; s.c., subcutaneous.
Figure 2
Figure 2
Effect of treatment with pasireotide on cell proliferation and apoptosis in pre-malignant lesions of the breast. Effects of pasireotide on proliferation rate as assessed by immunohistochemistry for Ki67 in 12 proliferative (A) and 6 atypical hyperplasia (B) Apoptosis, as assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), in 11 proliferative (C) and 6 atypical hyperplasia (D) lesions of the breast before and after 10 days of treatment with pasireotide.
Figure 3
Figure 3
Representative images of proliferation (Ki67) and apoptosis (terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)) from patients treated with pasireotide. Representative examples showing Ki67 immunostaining of atypical hyperplasia (A) and one case of ductal carcinoma in situ (DCIS) (B) from pasireotide-treated patients (top panel). (C) TUNEL staining of an atypical lobular hyperplasia (ALH) lesion before and after treatment with pasireotide. Positive cells are brown. (D) A small increase in the number of apoptotic cells (arrows) was also observed after treatment in one case of DCIS.
Figure 4
Figure 4
Effect of pasireotide on the estrogen and progesterone receptor. No significant differences were observed in the levels of estrogen receptor (ER) (A) or progesterone receptor (PR) (B) before and after treating with pasireotide.
Figure 5
Figure 5
Effect of treatment with pasireotide on phospho insulin-like growth factor -1 receptor (IGF-1R). Effect of pasireotide on mean intensity of phospho IGF-1R by immunofluorescence in four patients with atypical hyperplasia before and after treatment with pasireotide (A) and five controls (B). Representative sections from one of the patients before (C) and after treatment (D). DAPI: 4',6-diamidino-2-phenylindole; n.s., not significant.
Figure 6
Figure 6
Effect of treatment with pasireotide on phospho AKT and ERK1/2. (A) The mean intensity index was decreased by pasireotide from 42.5 ± 14 to 9.37 ± 3 after treatment as measured in eight patients. (B) Similar effects were observed with phospho ERK1/2 with a mean intensity index of 218 ± 29 before to 100 ± 18 after treatment, measured in six patients. Control tissues were not affected for p-AKT (C) or p-ERK 1/2 (D).
Figure 7
Figure 7
Effects of treatment with pasireotide on glucose, insulin, and insulin-like growth factor -1 (IGF-I). (A) Mean concentrations of plasma glucose and serum insulin during the 10-day treatment with pasireotide and the subsequent follow up. (B) Mean serum concentration of IGF-I before, at the end of treatment, and 90 days after treatment with pasireotide. Error bars are SDs in both graphs.

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