Chest
Volume 145, Issue 1, January 2014, Pages 108-112
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Original Research
Diffuse Lung Disease
Sirolimus Decreases Circulating Lymphangioleiomyomatosis Cells in Patients With Lymphangioleiomyomatosis

https://doi.org/10.1378/chest.13-1071Get rights and content

Background

Lymphangioleiomyomatosis (LAM), sporadic or in women with tuberous sclerosis complex (TSC), is characterized by cystic lung destruction, lymphatic involvement (eg, chylous pleural effusions, lymphangioleiomyomas), and renal angiomyolipomas (AMLs). The multisystem manifestations of LAM appear to result from metastatic dissemination of LAM cells bearing inactivating mutations or having loss of heterozygosity (LOH) of the tumor suppressor genes TSC1 or TSC2, which leads to hyperactivation of the mammalian target of rapamycin. Sirolimus slows the decline of lung function, reduces chylous effusions, and shrinks the size of AMLs. The purpose of this study was to determine the effect of sirolimus on circulating LAM cells.

Methods

Cells from blood were isolated by a density-gradient fractionation system and from urine and chylous effusions by centrifugation. Blood cells were incubated with anti-CD45-fluorescein isothiocyanate (FITC) and anti-CD235a-R-phycoerythrin (PE) antibodies, and urine and chylous effusion cells were incubated with anti-CD44v6-FITC and anti-CD9-R-PE antibodies. Cells were sorted and analyzed for TSC2 LOH.

Results

LAM cells with TSC2 LOH were identified in 100% of blood specimens and 75% of urine samples from patients before therapy. Over a mean duration of 2.2 ± 0.4 years of sirolimus therapy, detection rates of LAM cells were significantly decreased to 25% in blood (P < .001) and 8% in urine (P = .003). Following therapy, a greater loss of circulating LAM cells was seen in postmenopausal patients (P = .025).

Conclusions

Patients receiving sirolimus had a progressive loss of circulating LAM cells that depended on time of treatment and menopausal status.

Section snippets

Study Design

Twenty-three patients with LAM were enrolled between 2007 and 2012 at the National Institutes of Health (NIH) Clinical Center in clinical protocols (95-H-0186; 96-H-0100) approved by the National Heart, Lung, and Blood Institute Institutional Review Board. The diagnosis of LAM was based on clinical, radiographic, and histopathologic findings. Patients with only high-resolution CT scan-compatible cystic disease were not judged to have LAM, unless a biopsy specimen was obtained or the patients

Results

Twenty-three patients with LAM who fit the study inclusion criteria were enrolled between 2007 and 2012 at the NIH Clinical Center. Baseline demographic and clinical characteristics are shown in Table 1. Samples of blood, urine, and chylous effusions were collected at NIH before and during sirolimus therapy. Sirolimus was prescribed by local physicians and the dose adjusted to maintain serum levels between 5 and 15 ng/mL. Cells from blood, chylous effusions, and urine were sorted on the basis

Discussion

We studied the effect of sirolimus on phenotypically distinct circulating LAM cells. Patients treated with sirolimus had a decrease in circulating LAM cells in blood, chylous effusions, and urine that depended on time of treatment and the hormonal status of the patient. The LAM cells were identified by LOH in the tumor suppressor gene TSC2. This assay was used because nucleotide polymorphisms flanking the tumor suppressor gene, which were the basis for the LOH determinations, were shown

Acknowledgments

Author contributions: Drs Cai, Pacheco-Rodriguez, Stylianou, and Moss had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis.

Dr Cai: contributed to carrying out the research, analyzing the data, and writing the manuscript.

Dr Pacheco-Rodriguez: contributed to the research design, sample preparation, and writing of the manuscript.

Ms Haughey: contributed to the patient recruitment, data analysis, and review of the

References (18)

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Dr Cai is currently at the School of Pharmaceutical Sciences, Nanjing University of Chinese Medicine (Nanjing, Jiangsu, China).

Funding/Support: This study was funded in part by the Intramural Research Program of the National Institutes of Health, National Heart, Lung, and Blood Institute [to Dr Moss], and R01-AR062080 [to Dr Darling].

Reproduction of this article is prohibited without written permission from the American College of Chest Physicians. See online for more details.

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