American Journal of Respiratory and Critical Care Medicine

Searching for early predictive markers of the therapeutic effects of high-dose corticosteroids (“pulse therapy”) on patients with rapidly progressing idiopathic pulmonary fibrosis (IPF), we evaluated 14 such patients, who had received weekly pulse therapy for at least 3 wk. Eight patients responded to the treatment and survived. However, six patients failed to respond, and all of them died within 3 mo after treatment. Serum levels of KL-6 (MUC1 mucin), neutrophil elastase (NE), and lactate dehydrogenase (LDH) were measured before, and at 1 wk and 3 wk after treatment. Levels of KL-6 decreased significantly in patients who lived, whereas KL-6 levels tended to increase in patients who died. The values of NE did not change significantly. LDH levels decreased significantly at 1 wk, and tended to decrease at 3 wk in patients who lived. However, in patients who died, they did not significantly change. At the first cycle of treatment when clinical effects may not be evident, the decrease in KL-6 but not LDH levels was significantly related to a favorable outcome, whereas their increase was related to a poor outcome. Results suggest that monitoring with KL-6 may contribute to early clinical decisions for alternative therapy in the management of rapidly progressing IPF.

Despite a worldwide effort, idiopathic pulmonary fibrosis (IPF) still remains a disease of unknown etiology with a poor prognosis. To evaluate the activity and monitor the course of the disease, chest roentgenogram, pulmonary function testing, gallium-67 lung scan, and bronchoalveolar lavage (BAL) are clinically used (1). However, there are problems with the sensitivity, effort–dependability, and ease of repetition of these examinations. As a more convenient and reliable indicator, soluble markers have been studied, not only to evaluate and monitor the disease activity, but also for differential diagnosis and prognosis (2-10). Establishment of such a circulating marker would improve the management of IPF.

To this end, Kohno and coworkers (11) found a circulating high-molecular-weight glycoprotein, termed KL-6. The epitope recognized by the anti-KL-6 monoclonal antibody (mAb) has been demonstrated to be on the MUC1, as determined by immunohistochemical analysis in an international workshop (IASLC) (12). The serum level of KL-6 is elevated in a majority of patients with a number of interstitial lung diseases, including IPF, active hypersensitivity pneumonia, interstitial pneumonitis (IP) associated with collagen vascular disease and sarcoidosis, radiation pneumonitis, and far advanced pulmonary tuberculosis. It is not elevated in patients with bacterial pneumonia or in normal control subjects (13-18). Recently, these observations were confirmed by other laboratories in a national multicenter study (19, 20). In the lungs of the patients with IP, the majority of cells stained by the KL-6 mAb were regenerating type II pneumocytes (11, 13). Lactate dehydrogenase (LDH) has also been studied as a marker for pulmonary inflammation (2) and could reflect the cellular destruction of lung cells. However, LDH is a nonspecific marker. Neutrophil elastase (NE) is derived from neutrophils and reflects the activity of the neutrophil population, which is an underlying feature of the pathology of IPF (1).

The clinical course of IPF is usually slowly progressive, but it varies widely. It can progress rapidly, and such exacerbation often proves fatal. Although the mainstay treatment for IPF is oral corticosteroids, intravenous high-dose corticosteroid therapy (pulse therapy) is often used for the rapidly progressive disease, usually repeating several cycles for weeks or even months. Such high-dose therapy can control neutrophilic alveolitis, which would not be regulated by conventional corticosteroid therapy (21). The clinical effect of the treatment may not be evident until it has been repeated for several cycles. If a marker reflecting the effectiveness of corticosteroids were available in such a situation, an early decision for alternative therapy could be made. Therefore, the present study was planned to evaluate the relationship between the effectiveness of “pulse therapy” and changes in the biochemical markers KL-6, NE, and LDH.

Between April 1995 and January 1998, 14 patients were evaluated for the present study. The patients were selected on the basis of two criteria: (1) Rapidly progressive IPF. “Rapid” progression was defined as an increased dyspnea (an increase of more than 1 grade in the Hugh-Johns classification system) or a decrease in PaO2 (⩾ 10 mm Hg in the same condition) during the prior 2 mo, and the presence of ground-glass attenuation in high-resolution computed tomography (HRCT). Because clinical deterioration may result from infection, heart failure, cancer, or thromboembolism, rather than disease progression (22), we excluded such patients by clinical evaluation, including CT, sputum examinations, and echo cardiography. (2) The likelihood of treatment with weekly high-dose corticosteroids at least 3 cycles. A cycle of high-dose corticosteroid pulse therapy consisted of intravenous methylprednisolone 1,000 mg/d for 3 consecutive days, followed by oral prednisolone 60 mg/d for 4 d. After 3 cycles of the treatment, several cycles of pulse therapy and/or immunosuppressive drugs were administered to most of the nonresponders, and a gradual tapering of the oral corticosteroid was started for the responders.

Of the 14 patients, nine were men and five were women. The age range was 47 to 73 yr (mean ± SD: 63.4 ± 7.2) (Table 1). Diagnosis for IPF was based on the histologic features in specimens obtained by postmortem examination (Patients 1 and 2), open lung biopsy (Patient 5), biopsy under thoracic endoscopy (Patients 4 and 6), or clinical and radiological features including honeycombing in HRCT with histology-proven interstitial pneumonia, which did not contradict the diagnosis of IPF, in specimens obtained by transbronchial lung biopsy (Patients 3 and 7–14). The patients who fulfilled the diagnostic criteria of collagen vascular diseases were excluded. Fine crackles of both lungs were noted in all patients. None of the patients had been treated with oral corticosteroids or cytotoxic agents in the past or at the time of deterioration. The duration of IPF was an estimation based on the time of the onset of the symptoms. The patients were classified as current smokers (S) if they had smoked within 1 yr; ex-smokers (Ex) if they had not smoked for 1 yr but had smoked previously; and never-smokers (N). The degree of dyspnea was described according to the Hugh-Johns classification. Arterial blood gas analysis was done at less than 24 h before treatment and calculated as the alveolar-arterial difference of oxygen (AaDo 2). It should be noted that the percent vital capacity shown in Table 1 was obtained up to 2 wk before treatment, not specifically just prior to treatment.


Patient No.Age/SexDuration (mo)Smoking* Dyspnea (Hugh-Johns)%VCAaDo 2 Circulating levels of:Outcome
KL-6 (U/ml)NE (μg/L)LDH (IU/L)
 173/M  1ExVNA 6171,150  7061,020Deceased
 264/M114SIV79 78  760  610  698Deceased
 357/M120ExV646182,600   96  604Deceased
 461/M 32SIV39 702,600  225  604Deceased
 573/F  3NV441631,8202,050  529Deceased
 663/F 62NIV68 603,800  387  586Deceased
 773/M 60SV981132,400  201  767Surviving
 866/M 82SV812022,100  800  440Surviving
 963/F 15NIII80 465,1002,260  531Surviving
1047/F 36NIII58 553,5001,010  537Surviving
1156/M 18SVNA2962,0001,460  682Surviving
1267/M 91SV911641,190  189  376Surviving
1364/M 36ExIV77 592,600  178  513Surviving
1461/F 24NIII83 402,400ND  563Surviving

*Smoking status as defined in Methods.

Not available.

Because we selected rapidly deteriorating patients as described previously, the therapeutic effect of pulse therapy was directly related to the prognosis in the present study. Eight patients (Patients 7–14, Table 1) responded to the treatment as evidenced by an increased PaO2 with the same condition (> 10 mm Hg) and decreased dyspnea (a decrease of more than 1 grade in the Hugh-Johns classification). The therapy seemed to stabilize the disease process, and all of the patients survived the deterioration (i.e., they survived at least the next 6 mo after treatment). Six patients (Patients 1–6, Table 1) failed to respond to the treatment and all six died from progressive respiratory failure within the next 3 mo.

Venous blood was obtained before, and at 1 wk and 3 wk after treatment. Serum and ethylenediaminetetraacetic acid (EDTA)– plasma were obtained and stored at −80° C until use. The concentrations of KL-6 in serum (normal value [n.v.]: < 520 U/ml) were determined by a specific ELISA, as described elsewhere (11, 14). Serum LDH level was determined according to the method of Wroblewski La Due (n.v. < 420 IU/L). Plasma NE concentration (n.v. < 154 μg/L) was determined by enzyme immunoassay using a commercially available kit (Merck KGaA, Darmstadt, Germany).

Data are shown as mean ± SD. For statistical evaluation of changes in the concentrations before, and at 1 wk and 3 wk after treatment, the Friedman test was applied. To test differences in the levels before and at 1 wk or 3 wk, the Wilcoxon test was used. The relationship between the outcome and changes in the concentrations of the circulating markers was estimated by the Fisher exact probability test. Statistical significance was defined as p < 0.05.

The backgrounds of the patients evaluated in the present study are shown in Table 1. There were no significant differences in age, sex, duration of the disease, dyspnea score, or AaDo 2 between nonresponders (deceased) and responders (surviving). The percent vital capacity (%VC) in patients who died was significantly lower than that in patients who lived (58.8 ± 16.8% versus 81.1 ± 12.5%; p < 0.05). None of the pretreatment KL-6, NE, or LDH concentrations differed between the patients who lived and the patients who died.

Changes in the circulating levels of KL-6, NE, and LDH before and after corticosteroid treatment are shown in Figure 1. Levels of KL-6 in patients who lived significantly decreased from 2,661 ± 1,178 U/ml to 2,160 ± 910 U/ml (−18.9 ± 14.4%; p < 0.05), and 1,801 ± 899 U/ml (−32.7 ± 20.9%; p < 0.05), at 1 wk and 3 wk after treatment, respectively. In patients who died, KL-6 tended to increase from 2,122 ± 1,110 U/ml to 2,352 ± 866 U/ml (+22.9 ± 34.5%) at 1 wk, and to 3,755 ± 2,288 U/ml (+93.7 ± 103%) at 3 wk. The concentrations of NE did not significantly change in patients who lived (871 ± 785 μg/L, 895 ± 756 μg/L [+ 9.9 ± 60.4%] and 736 ± 689 μg/L [−17.9 ± 38.5%]) or patients who died (679 ± 709 μg/L, 559 ± 791 μg/L [−25.2 ± 28.4%] and 524 ± 623 μg/L [−16.8 ± 30.5%]). LDH levels in patients who lived fluctuated, but significantly decreased from 551 ± 125 IU/L to 425 ± 56 IU/L (−21.0 ± 11.3%) at 1 wk, and tended to decrease to 438 ± 93 IU/L (−17.4 ± 23.4%) at 3 wk. However, in patients who died, the overall concentrations of LDH did not change significantly (p = 0.31; Friedman test) (674 ± 178 IU/L, 667 ± 156 IU/L, 825 ± 115 IU/L, before and at 1 wk and 3 wk, respectively).

The relationship between outcome and changes in circulating markers was estimated and is shown in Table 2. At 3 wk, the decrease in KL-6 and LDH concentrations was significantly related to a favorable outcome, whereas their increase was related to a poor outcome. However, only KL-6 was significantly related to the outcome at the 1-wk point. Changes in NE between prior to and after the treatment were not related to the outcome.


1 wk (increased/decreased)3 wk (increased/decreased)
Surviving (n = 8)0/7 2/50/81/71/61/7
Deceased (n = 6)5/12/42/45/11/55/1
p* < 0.050.9990.165< 0.050.999< 0.05

*Fisher exact probability test.

KL-6 level of one alive patient was not changed in 1 wk.

The present study demonstrates that the circulating level of KL-6 could be a marker to predict the efficacy of high-dose corticosteroid pulse therapy at 1 wk after treatment when overall clinical effect may not yet be evident. The concentrations of LDH, however, could predict the outcome only at the 3-wk point after treatment. The levels of LDH tended to fluctuate, whereas those of KL-6 continued to decrease in patients who lived and increased in patients who died. NE had no value as a prognostic indicator.

KL-6, a circulating MUC1 mucin, is supposed to reflect the number of regenerating type II epithelial cells in patients with IP (11, 13). Type II cells are regenerated over the alveolar basement membrane after the death of type I cells during the first stage of lung injury (23). It should be noted that KL-6 is not a simple marker, but has a pathophysiologic role in IP. Purified KL-6 is a chemoattractant for fibroblast, and is more potent than platelet-derived growth factor (PDGF) or fibroblast growth factor (FGF) (24). KL-6 has been demonstrated to be a better marker for the differential diagnosis of IP than LDH or collagen-relevant products, such as type III procollagen N-terminal peptide (PIIIP) or type IV collagen 7S, with respect to sensitivity and specificity (14, 16). In the present study, we demonstrated that KL-6 may be a useful marker for monitoring therapeutic effect in rapidly deteriorating IPF patients.

The mechanism for the decrease of KL-6 observed as a therapeutic effect of high-dose corticosteroids is unclear at present. Because the promoter region of the human MUC1 gene does not contain a glucocorticoid receptor binding site, direct effect of steroids is unlikely to occur (25). It is known that proinflammatory cytokines such as tumor necrotizing factor (TNF) and interferon-γ can augment the expression of MUC1 in human carcinoma cell lines (26). Corticosteroids could inhibit proinflammatory cytokines and then, secondarily, reduce MUC1 production. Alternatively, steroids may decrease permeability, which is caused by inflammation, to decrease leakage of KL-6 from the lungs to circulating blood. It is demonstrated that circulating levels of KL-6 may be dependent on alveolar–capillary permeability in patients with berylliosis (18).

As a marker for IP, a variety of enzymes, cytokines, adhesion molecules, collagen-relevant products, and products of type II epithelial cells have been evaluated (2-10). Among them, the phospholipids and surfactant protein A (Sp-A) in bronchoalveolar lavage fluid (BALF) have been reported as soluble markers for disease activity and prognosis of IPF (3, 4, 7). Recently, measurement of circulating Sp-A and Sp-D levels have been reported to be useful for differential diagnosis (6, 8). However, it is not known whether these concentrations have prognostic value or not. It should be noted that the patients in the present study were rapidly deteriorating, rather than the usual slowly progressive IPF patients who had been studied in BALF Sp-A. This may explain why we observed a significant relationship between %VC and outcome in our study, though it has been noted that the measure of lung function may not be a reliable indicator of survival in the usual IPF population (1).

The limitations of the present study should be mentioned. First, because rapid progression of IPF is not a common phenomenon, the number of patients in the present study was not sufficient for a valid statistical analysis. Second, measurement of KL-6 levels in the pretreatment period could not be used to predict outcome. However, we believe that the pretreatment values of KL-6 in both patients who lived and those who died, were already elevated by deterioration. In our experience, higher serum KL-6 seemed to be related to a worse prognosis in the typical IPF patient. Another study is ongoing to establish this point. Third, it should be noted that KL-6 concentrations may be increased in patients with some malignancy, such as adenocarcinoma of the lung, breast, or pancreas (27).

In conclusion, the present study suggests that circulating concentrations of KL-6 may contribute to early identification of patients with a high probability of resistance to corticosteroid treatment, and thus could predict outcome. Although part of the steroid refractory state could be overcome by immunosuppressive therapy, there is no consensus regarding this situation (28, 29). If the KL-6 level is increased after one cycle of pulse therapy, an alternative therapy may be considered to avoid both unnecessarily long exposure to drugs without a positive impact on the outcome and potentially hazardous side effects.

The writers thank Ms. Aibara for her assistance in measuring KL-6 concentrations and Dr. Yoshikazu Inoue for providing information of the patients.

Supported in part by a Grant-in-Aid for Scientific Research (No. 08670665) from the Ministry of Education, Science, Sports and Culture, Japan.

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Correspondence and requests for reprints should be addressed to Akihito Yokoyama, Second Department of Internal Medicine, Ehime University School of Medicine, Onsen-gun, Ehime 791-0295, Japan.


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