Pepsin and bile acids in saliva in patients with laryngopharyngeal reflux - a prospective comparative study.

OBJECTIVES: Laryngopharyngeal reflux (LPR) and biliary duodenogastric reflux can cause damage to the laryngeal mucosa and voice disorders. The aim of this study was to find out whether levels of pepsin and bile acids in the saliva can serve as diagnostic markers of LPR. SETTING: A prospective comparative study. PARTICIPANTS: Twenty-eight patients with LPR proven via high-resolution manometry and combined multichannel intraluminal impedance and 24-h pH monitoring and 48 healthy controls without symptoms of LPR were included in the study. MAIN OUTCOME MEASURES: In the patients with LPR symptoms, oesophagogastroscopy with oesophageal biopsy was performed. The levels of total pepsin, active pepsin, bile acids and the pH of the saliva were determined in all participants and compared between the groups. Reflux symptom index (RSI) and reflux finding score (RFS) were also obtained and compared. The groups differed significantly in RSI (P = 0.00), RFS (P = 0.00), the levels of bile acids (P = 0.005) and total pepsin in saliva (P = 0.023). The levels of total pepsin and bile acids were about three times higher in the patients with LPR than in the healthy controls. There was a significant correlation between the RSI and RFS score and the level of total pepsin and bile acids in the saliva. Histopathological examination of the oesophageal biopsy taken 5 cm above the lower oesophageal sphincter confirmed reflux in almost 93% of patients with symptoms. CONCLUSIONS: The study results show that the levels of total pepsin and bile acids in saliva are significantly higher in patients with LPR than in the controls, thus suggesting this as a useful tool in the diagnosis of LPR and particularly biliary LPR.

Transjugular intrahepatic portosystemic shunt versus endoscopic sclerotherapy in the elective treatment of recurrent variceal bleeding.

The present study was designed to compare elective transjugular intrahepatic portosystemic shunts (TIPS) and endoscopic sclerotherapy (EST) in terms of their efficacy in preventing recurrent bleeding from gastro-oesophageal varices in patients with advanced liver cirrhosis and portal hypertension. Of 96 patients with at least three gastro-oesophageal variceal rebleeds, 50 were treated with elective TIPS and 46 with EST. Recurrent variceal bleeding was significantly more frequent in patients receiving EST treatment compared with those receiving TIPS (45.7% versus 6.3%, respectively). Cumulative 1- and 4-year survival in the TIPS group was 83.0% and 73.5%, respectively, compared with 69.8% and 39.8% in the EST group, respectively. The rate of portosystemic encephalopathy was 33.3% in the TIPS group and 37.0% in the EST group. Elective TIPS was more effective than EST in the prevention of gastro-oesophageal variceal rebleeding in cirrhotic patients, it improved survival and it was associated with a similar rate of portosystemic encephalopathy.

Effects of native human leukocyte interferon-alpha and recombinant human interferon-alpha on P3-X63-Ag8.653 mouse myeloma cell growth.

Multiple myeloma (MM) remains largely incurable, although traditional chemotherapy and new compounds have been shown to produce a clinical response. Clinical studies were performed to determine the effectiveness of interferon-alpha (IFN-alpha) in MM, which has also recently been shown to function as a survival factor for MM cells. The effects of different doses of native human leukocyte interferon-alpha (nhIFN-alpha), recombinant human interferon-alpha2a (rhIFN-alpha2a) and recombinant human interferon-alpha2b (rhIFN-alpha2b) on in vitro P3-X63-Ag8.653 mouse myeloma cell growth were compared. A statistically significant dose-dependent reduction in cell viability following cell culture with nhIFN-alpha was observed. On the other hand, a statistically significant increase in cell viability was observed following cell culture with rhIFN-alpha2a and rhIFN-alpha2b, but only in relation to the control group and seemingly without dose dependency. These results highlight the importance of the type of human IFN-alpha used in the treatment and study of MM, and suggest that nhIFN-alpha may have a role in future personalized therapy approaches.

Expression and prognostic significance of Cathepsin L in early cutaneous malignant melanoma.

Cathepsins are papain-like lysosome cysteine proteases involved in tumor growth, invasiveness and spread, angiogenesis and alteration in immune and inflammatory responses. We investigated the differences in cathepsin L (CatL) concentrations in primary cutaneous malignant melanoma stage I and normal skin and correlated these values with well-established malignant melanoma prognostic factors. The study was performed on 36 patients (17 men and 19 women; mean age 54 years; range 21-84 years) with histological confirmed primary malignant melanomas less than 1.5 mm thick. The CatL concentrations were measured in 36 pairs of triton extracts of cytosols prepared from the tumor and adjacent normal tissue samples (matched pairs). The CatL concentrations were determined by commercially available enzyme-liked immunosorbent (ELISA) assay from KRKA (Novo Mesto, Slovenia). Significantly higher concentrations of CatL were detected in malignant melanomas than in normal surrounding skin (6.73 vs. 1.42 ng/mg total protein (mgp), p<0.001). Significant correlations between malignant melanoma and normal skin concentrations for CatL were found. The malignant melanoma CatL concentrations correlated significantly with normal skin (r=0.38; p=0.021). CatL concentrations were significantly lower (p<0.01) in the malignant melanomas of Breslow thickness 0.75 mm, Clark invasion of >or=II and

Pitfalls in diagnosing small bowel carcinoid tumors.

Small bowel carcinoid usually presents with clinical symptoms and signs deriving from this endocrinological function and it rarely bleeds profusely. Below we present a case of a patient with intestinal bleeding. The bleeding tumor was finally diagnosed with the aid of wireless capsule endoscopy (CE) and histological examination showed a small bowel carcinoid.

Double modulation of 5-fluorouracil by leucovorin and low-dose methotrexate in advanced colorectal cancer.

A phase II study was carried out to evaluate the efficacy and toxicity of a double biochemical modulation of 5-fluorouracil (5-FU) by methotrexate (MTX) and leucovorin (LV) in patients with advanced unresectable colorectal cancer. Forty-two patients with measurable metastases of colorectal cancer were treated with 5-FU in daily doses of 600 mg/m2 given in a 6-hour intravenous (i.v.) infusion on days 1-5, LV 50 mg/m2 i.v. on days 2,3 and 5, and MTX 40 mg/m2 i.v. on days 1 and 4, every 4 weeks. Twenty-eight patients had a single metastatic site, eleven double, whereas three had more than two metastatic sites. Objective response (one complete response) was observed in 12 of 40 patients (30%) (95% confidence interval 16-48), stable disease in 19 patients (47%) and progression in 9 patients (23%). Overall median survival was 12 months. Median time to progression was 6 months. Treatment was generally well tolerated. The most frequent adverse reactions were stomatitis (38%), nausea and vomiting (35%), diarrhea (31%), leukopenia (18%), and plantar-palmar erythroderma (15%). The combination of 5-FU, LV and MTX seems to be an active regimen in advanced colorectal cancer.

Cathepsin D expression in early cutaneous malignant melanoma.

BACKGROUND: The aspartic proteinase cathepsin D is believed to be associated with proteolytic processes leading to the invasion and seeding of tumor cells. An association between cathepsin D tissue concentration and aggressiveness of tumors has been detected in different cancer types, as well as in metastatic melanoma. METHODS: The concentration of cathepsin D was measured immunoradiometrically (ELSA-CATH-D kit, CIS Bio International) in the cytosols of 51 primary cutaneous melanomas (with Breslow index < 4 mm) to estimate the tissue concentrations of cathepsin D in early cutaneous melanoma. RESULTS: A significantly elevated concentration of cathepsin D was measured in the tumor cytosols as compared to adjacent normal tissue (44.2 vs. 14.7 pmol/mg of total protein, P < 0.001). CONCLUSIONS: Our results indicate that cathepsin D is expressed at high levels by melanoma cells. The extremely high expression of cathepsin D in two of our patients, with later progression of the disease over a 42-month follow-up period, suggests a possible correlation between the cathepsin D tissue concentration and the prognosis of primary cutaneous malignant melanoma.

In vitro functional tests for evaluation of stimulating capacity of cultured human dendritic cells.

Basic functional test for evaluation of in vitro cultured human dendritic cells (DC) is primary allogeneic one-way mixed lymphocyte reaction (MLR). In this way, one can evaluate stimulating capacity, which is a basic characteristic of DC. The proliferation of cells is measured through incorporation of 3H-thymidine. Normally proliferation is measured at days 5-7. We studied kinetics of proliferative responses initiated with different stimulating cell suspensions to evaluate differences and possibly reduce time needed to perform this test. Gradual increase in response from days 1 to 7 and a significant difference from controls (peripheral blood mononuclear cells) seen from day 4 was noted if macrophages were used as stimulators. A consistently higher proliferation, compared to controls, was always found already on day 2 when mature DC were used as stimulators. The reaction peaked 2 to 3 days earlier and was also more than two times more intense. This maximal and significantly higher response, consistently seen already after 48 hours, allows us to confirm the presence of mature DC in stimulating suspensions much earlier than previously.

Soluble tumor necrosis factor receptor I (sTNFRI) as a prognostic factor in melanoma patients in Slovene population.

Tumor necrosis factor alpha (TNF-alpha) and its receptors (TNFRI and TNFRII) which exist in soluble form as a product of cleavage of the extracellular domain of membrane integrated receptors, still rise debate about their importance. It was reported that TNF-alpha has numerous actions in diseases such as inflammation, autoimmunity, infectious diseases, septic shock and many types of cancer [1, 2]. Several authors have reported the significance of sTNFRI level in serum of cancer patients [3, 4]. This study was performed in collaboration with the Institute of Oncology of Slovenia. At least two different mouse monoclonal antibodies (MAbs) against human sTNFRI have been prepared to obtain a sensitive and reliable sandwich ELISA. It was compared with commercially available R&D and Endogen ELISAs for the determination of sTNFRI. Groups of patients with different stages of melanoma and epithelial ovarian carcinoma were tested and their clinical records were reexamined. Levels of sTNFRI were measured and compared with the normal serum levels of sTNFRI.

Electrochemotherapy with cisplatin: the systemic antitumour effectiveness of cisplatin can be potentiated locally by the application of electric pulses in the treatment of malignant melanoma skin metastases.

The application of electric pulses to skin tumour nodules enhances the antitumour effectiveness of cisplatin. This treatment approach, known as electrochemotherapy, was employed in the treatment of skin metastases and lymph node metastases in malignant melanoma patients. Electric pulses were applied to tumour nodules in order to potentiate locally the antitumour effectiveness of systemic cisplatin-based chemoimmunotherapy. The study included nine malignant melanoma patients with skin metastases and metastases in lymph nodes not amenable to surgery, undergoing cisplatin-based chemoimmunotherapy. The antitumour effectiveness of the chemoimmunotherapy was compared with the antitumour effectiveness of electrochemotherapy, i.e. application of electric pulses to tumour nodules together with cisplatin-based chemoimmunotherapy. Application of electric pulses to the 27 skin tumour nodules potentiated locally the antitumour effectiveness of cisplatin. Four weeks after the treatment, 48% of the tumour nodules had an objective response (OR), compared with 22% of the 18 tumour nodules treated with cisplatin-based chemoimmunotherapy alone. Furthermore, the median time to progression was longer in the electrochemotherapy-treated nodules (21 weeks) than in the chemoimmunotherapy-treated nodules (4 weeks). This study shows that application of electric pulses to malignant melanoma tumour nodules can potentiate the antitumour effectiveness of cisplatin in patients undergoing systemic cisplatin-based chemoimmunotherapy. Therefore, electrochemotherapy may be used as an adjunct to systemic ongoing cisplatin treatment, predominantly in patients in whom antitumour effectiveness needs to be potentiated locally.

Serum values of tumour necrosis factor-alpha and of soluble tumour necrosis factor-R55 in melanoma patients.

Tumour necrosis factor-alpha (TNFalpha) is a cytokine with a variety of biological activities, including an effect on tumour growth. The soluble TNF receptor TNF-R55 (sTNF-R55) inhibits TNFalpha functioning. Serum values of TNFalpha and TNF-R55 have been observed in patients with different cancers. To determine the serum values of TNFalpha and its soluble receptors and to investigate the prognostic value of sTNF-R55, we studied the sera of 68 patients with metastatic melanoma, 109 patients with no recurrent disease after surgical removal of melanoma, and 69 healthy controls. At least four different monoclonal antibodies against human TNFalpha and human TNF-R55, respectively, were prepared to obtain sensitive and reliable sandwich enzyme-linked immunosorbent assays. We found that in patients with metastatic melanoma the serum values of sTNF-R55 were significantly higher (2.41 ng/ml; range 0.02 23.0 ng/ml; P < 0.05) than in the melanoma patients without recurrence (0.54 ng/ml; range 0.02-6.25 ng/ml) and healthy controls (0.5 ng/ml; range 0.02 5.0 ng/ml). The sTNF-R55 concentrations increased as the disease progressed. Patients with metastatic melanoma also had significantly higher concentrations of TNFalpha (3.34 ng/ml; range 0.03-30.0 ng/ml; P<0.05) than patients without recurrence (1.24 ng/ml; range 0.02 23.0 ng/ml). Patients with metastatic melanoma, a high sTNF-R55 concentration, a low TNFalpha concentration and a low TNF:sTNF-R55 ratio had the worst prognosis. Low values of sTNF-R55 (<0.6 ng/ml) were associated with favourable response to chemotherapy (P = 0.007). According to our findings, patients with metastatic melanoma have higher values of sTNF-R55 than the controls and melanoma patients without recurrence. sTNF-R55 values higher than 0.6 ng/ml and a TNF:sTNF-R55 ratio lower than 1.5 are unfavourable prognostic factors for response to chemotherapy.

Electrochemotherapy with cisplatin: clinical experience in malignant melanoma patients.

Electrochemotherapy consists of chemotherapy followed by local application of electric pulses to the tumor to increase drug delivery into the cells. The aim of this Phase II clinical study was to evaluate the antitumor effectiveness of electrochemotherapy using intratumoral cisplatin administration on cutaneous tumor nodules in malignant melanoma patients. In 10 patients, 133 tumor nodules of different sizes were treated: (a) 82 tumor nodules were treated with electrochemotherapy; (b) 27 tumor nodules were treated with cisplatin; (c) 2 tumor nodules were treated with electric pulses; and (d) 22 tumor nodules were untreated. Four weeks after therapy, 78% objective responses were obtained in the electrochemotherapy group, and 38% objective responses were obtained in the cisplatin group. Exposure of tumor nodules to electric pulses without cisplatin treatment had no effect on tumor growth. Electrochemotherapy was well tolerated by all patients, and a good cosmetic effect was obtained, with only minimal scarring and a slight depigmentation of the skin. At 124 weeks of follow-up, a 77% control rate of the tumor nodules treated by electrochemotherapy was observed, compared to 19% for those that were treated with cisplatin only (P < 0.0001). Our results clearly demonstrate that electrochemotherapy with cisplatin is a highly effective approach for treatment of cutaneous malignant melanoma nodules. The advantages of this therapy include its simplicity, the short duration of treatment sessions, low cisplatin doses, and insignificant side effects, as well as the fact that it can be done on an outpatient basis.

Improved prediction of decreased creatinine clearance by serum cystatin C: use in cancer patients before and during chemotherapy.

BACKGROUND: Serum cystatin C, a cysteine protease inhibitor, has been suggested as a new marker of glomerular filtration rate (GFR). This study explored the possibility of replacing the creatinine clearance (CrCl) estimation of GFR with cystatin C in early detection of renal impairment in cancer patients on chemotherapy. METHODS: Serum creatinine and cystatin C concentrations as well as 24-h CrCl were determined simultaneously in 72 cancer patients. Among them, 60 were treated with combined chemotherapy with cisplatin (CDDP). Creatinine was determined enzymatically with a spectrophotometric method. Serum cystatin C was determined by a particle-enhanced turbidimetric immunoassay. RESULTS: Cystatin C and creatinine correlated significantly (P = 0.001) with CrCl. The correlation was significantly better for cystatin C than creatinine (r = 0.84 vs 0.74; P = 0.01). Stepwise regression analysis identified no differences for the correlations between cystatin C and CrCl in patients with or without metastases (r = 0.82 and 0.84, respectively) as well as before treatment and before the fourth cycle of chemotherapy (r = 0.70 and 0.75, respectively). A cystatin C cutoff concentration of 1.33 mg/L had 87% sensitivity and 100% specificity for detecting CrCl <78 mL/min. ROC analysis indicated that cystatin C was superior to serum creatinine for predicting CrCl <78 mL/min (P <0.04). CONCLUSIONS: Serum cystatin C is superior to serum creatinine for detection of decreased CrCl and potentially for the estimation of GFR in cancer patients independent of the presence of metastases or chemotherapy.

Electrochemotherapy with cisplatin: potentiation of local cisplatin antitumour effectiveness by application of electric pulses in cancer patients.

This study was aimed at assessing the response to electrochemotherapy with cisplatin of cutaneous tumour nodules in patients with malignant melanoma, squamous cell carcinoma and basal cell carcinoma. In 4 patients, 30 tumour nodules of different sizes were treated; five without treatment, one with electric pulses, five with cisplatin injected intratumorally and 19 with electrochemotherapy, i.e. intratumoral administration of cisplatin followed by delivery of electric pulses to the tumour nodule. After 4 weeks, a complete response (CR) in all 19 electrochemotherapy treated nodules was obtained. All electrochemotherapy treated nodules remained in CR (range 7-11 months), regardless of histological type, except for the metastasis of a squamous cell carcinoma that progressed after 9 months. CR was also obtained in two of five tumour nodules treated with cisplatin intratumorally, but the other three nodules progressed within 3-7 months. Exposure of the tumour nodule to electric pulses without cisplatin treatment had no effect on tumour growth. Electrochemotherapy was well tolerated by all patients and a good cosmetic effect was obtained, with only minimal scarring and a slight depigmentation of the skin. Electrochemotherapy with cisplatin has proved to be effective in patients with cutaneous tumour nodules. Furthermore, electrochemotherapy is easy to perform and can be carried out on an out-patient basis.

Treatment options in Western hepatocellular carcinoma: a prospective study of 224 patients.

BACKGROUND/AIMS: Though hepatocellular carcinoma (HCC) is one of the most frequent malignant tumors in the world, the optimal therapeutic strategy is still poorly defined. This is mainly due to geographic differences in HCC which may affect the validity of treatment regimens in differents areas of the world. The aim of the present study was to analyze the natural course of the disease as well as to assess the efficacy of different therapeutical schemes in HCC observed in Ljubljana (Slovenia) and Trieste (Italy), two cities in Western Europe situated close to each other. METHODS: During the period from January 1988 to December 1993, 224 consecutive patients (132 in Trieste and 92 in Ljubljana) with HCC were enrolled in the study. Patients were treated with the following 3 schemes: surgery 39 (17.4%), transcatheter chemoembolization (TACE) 116 (51.8%), and no treatment 69 (30.8%). The tumor was classified by Okuda staging and the liver disease by Child-Pugh score. Patients were followed up for 12-60 months, with an average of 40 months. The response rate to TACE and recurrence following surgery were evaluated. Comparative analysis of survival between different treatment groups was performed. RESULTS: The natural course of the disease, and other characteristics of the HCC, showed a typical Western type of tumor. Liver disease was scored as Child A in 58%, Child B in 30% and Child C in 12%, and the tumor was staged as Okuda I in 52%, Okuda II in 37% and Okuda III in 11%, respectively. Treatment with TACE was followed by an objective response in 27%, with a median survival of 31 months. Surgery was followed by a recurrence rate of 77% within 19.5 months and median survival of 49 months. The overall median survival of nontreated patients was 8 months. Survival in each group of patients differed significantly between all three consecutive stages of Okuda (p<0.001). In contrast, the differences in survival were significant only between Child A and B (p<0.02). The differences between Child B and C were not significant. CONCLUSIONS: This study emphasizes the importance of staging in the choice of treatment modality and diffusion of HCC in affecting an overall response to treatment and survival. Surgery is highly effective in monofocal HCC of Okuda I and II without cirrhosis. TACE is effective in Okuda I and II and Child A cirrhosis only. The treatment of HCC in Child B cirrhosis needs further studies. In Child C and/or Okuda stage III of HCC, any treatment except pure symptomatic relief is detrimental and should not be used.

Effective treatment of cutaneous and subcutaneous malignant tumours by electrochemotherapy.

Electrochemotherapy (ECT) enhances the effectiveness of chemotherapeutic agents by administering the drug in combination with short intense electric pulses. ECT is effective because electric pulses permeabilize tumour cell membranes and allow non-permeant drugs, such as bleomycin, to enter the cells. The aim of this study was to demonstrate the anti-tumour effectiveness of ECT with bleomycin on cutaneous and subcutaneous tumours. This article summarizes results obtained in independent clinical trials performed by five cancer centres. A total of 291 cutaneous or subcutaneous tumours of basal cell carcinoma (32), malignant melanoma (142), adenocarcinoma (30) and head and neck squamous cell carcinoma (87) were treated in 50 patients. Short and intense electric pulses were applied to tumours percutaneously after intravenous or intratumour administration of bleomycin. The tumours were measured and the response to the treatment evaluated 30 days after the treatment. Objective responses were obtained in 233 (85.3%) of the 273 evaluable tumours that were treated with ECT. Clinical complete responses were achieved in 154 (56.4%) tumours, and partial responses were observed in 79 (28.9%) tumours. The application of electric pulses to the patients was safe and well tolerated. An instantaneous contraction of the underlying muscles was noticed. Minimal adverse side-effects were observed. ECT was shown to be an effective local treatment. ECT was effective regardless of the histological type of the tumour. Therefore, ECT offers an approach to the treatment of cutaneous and subcutaneous tumours in patients with minimal adverse side-effects and with a high response rate.

Enzyme-linked immunosorbent assay for the detection of total cathepsin H in human tissue cytosols and sera.

An enzyme-linked immunosorbent assay (ELISA) was constructed for the determination of total human cathepsin H concentration in clinical samples. Utilising monoclonal and polyclonal antibodies, raised to human liver cathepsin H, the assay is able to detect a mature protein, a precursor molecule and enzyme-inhibitor complexes. The test system permits sensitive and reliable detection of analyte either in tissue cytosols or in sera. The detection limit is 2 ng/ml (n = 10, mean of zero standard +/- 3 SD). The average recovery of cathepsin H, added to the low content samples, was 95.3% +/- 1.8%. The within-run and between-run coefficient of variance (CV) varied from 2.3% to 8.9% and 12.7% to 16.4%, respectively, indicating satisfactory reproducibility of the method. The level of cathepsin H was defined in tissue cytosols of human heart, muscle and kidney and in sera from 30 healthy individuals. Additionally, cathepsin H was measured in sera from 55 patients with primary skin melanoma and from 42 patients with metastatic melanoma. The mean cathepsin H level was significantly higher for both groups of patients compared to normal sera level, being highest for metastatic melanoma patients.

Cathepsins B, H, and L and their inhibitors stefin A and cystatin C in sera of melanoma patients.

The levels of cathepsins (Cats) B, H, and L and their inhibitors stefin A and cystatin C were determined in the sera of 43 patients with metastatic melanoma, in 54 patients with treated cutaneous melanoma with no evidence of metastatic disease, and in 30 healthy blood donors, using quantitative ELISAs. The levels of Cats B and H and cystatin C were significantly higher within the group of metastatic melanoma patients compared with the healthy controls. The median Cat B was 4.8 versus 3.6 ng/ml (P < 0.013), the median Cat H was 13.7 versus 4.9 ng/ml (P < 0.0001), and the median cystatin C was 470 versus 320 ng/ml (P < 0.02). Cat H was also significantly increased within the group of melanoma patients with no metastasis, with a median of 9.6 ng/ml. Cat B was found to correlate with Cat L (r = 0.36; P < 0.02) and cystatin C (r = 0.41; P < 0.008). The serum level of Cat H was significantly increased in patients showing no response to the chemoimmunotherapy as compared to the level in responders. Metastatic melanoma patients with high contents of Cat B and Cat H experienced significantly shorter overall survival rates than the patients with low levels of each enzyme (Cat B: P < 0.003 and relative risk, 2.5; Cat H: P < 0.006 and relative risk, 2.4, using medians as cutoff values). The other potential factors for prognosis for this group of patients revealed moderate (histological type and age) or no (tumor thickness, sex, and lymph node metastasis) prognostic significance. Similarly, no difference in survival was found for stefin A, cystatin C, and Cat L. These results suggest that the serum levels of Cats B and H could serve as prognostic factors for patients with advanced melanoma.

Ukrain with chemotherapy in malignant melanoma (case report).

A female patient was treated in 1991 for a metastatic malignant melanoma. She was treated chemotherapeutically, with interferon, and surgically. After recurrence in 1993 the chemotherapy regime was changed and Ukrain therapy was administered between the chemotherapy series. Whereas in 1993 the patient suffered from multiple metastases in the lungs, the combined therapy with Ukrain induced a complete remission which has lasted more than one year to data.