TRADITIONAL CHINESE MEDICINE COMPOSITION FOR TREATING EGFR-TKIS RELATED SKIN RASH AND USE THEREOF

Abstract

A traditional Chinese medicine composition for treating EGFR-TKIs related skin rash and a use thereof, characterized by comprising a therapeutically effective amount of the following components: honeysuckle, dandelion, radix sophorae flavescentis, Kochia scoparia, Dictamnus dasycarpus, tree peony root barks, Cortex Phellodendri, and peppermint. The traditional Chinese medicine composition for treating EGFR-TKIs related skin rash has a certain effect for the treatments of EGFR-TKIs related skin rash and paronychia. The raw materials coordinate with each other, thus can play medicinal effects to a maximum extent, thus significantly improving life quality of patients and prolonging survival period of patients. In addition, the traditional Chinese medicine composition also has advantages of non-toxic side effects and lower treatment cost.

Claims

1. A traditional Chinese medicine composition, comprising therapeutically effective amounts of following components: Lonicera japonica, Taraxacum mongolicum, Sophora flavescens, Kochiae fructus, dictamni cortex, moutan cortex, Phellodendri Chinensis Cortex, and Menthae Haplocalycis Herba; and the traditional Chinese medicine composition is used for treating EGFR-TKIs related skin rash.

2. The traditional Chinese medicine composition according to claim 1, comprising 5-40 parts of Lonicera japonica, 5-50 parts of Taraxacum mongolicum, 3-50 parts of Sophora flavescens, 5-50 parts of Kochiae fructus, 3-50 parts of dictamni cortex, 5-50 parts of moutan cortex, 5-50 parts of Phellodendri Chinensis Cortex, and 2-30 parts of Menthae Haplocalycis Herba.

3. The traditional Chinese medicine composition according to claim 1, comprising 15 parts of Lonicera japonica, 15 parts of Taraxacum mongolicum, 9 parts of Sophora flavescens, 15 parts of Kochiae fructus, 10 parts of dictamni cortex, 12 parts of moutan cortex, 12 parts of Phellodendri Chinensis Cortex, and 6 parts of Menthae Haplocalycis Herba.

4. A use of the traditional Chinese medicine composition according to claim 1 in preparation of a formulation for treating EGFR-TKIs related skin rash.

5. A use of the traditional Chinese medicine composition according to claim 1 in preparation of a formulation for treating EGFR-TKIs related paronychia.

6. The use of the traditional Chinese medicine composition according to claim 4, wherein the formulation is a liniment, a water aqua, a decoction, a granule, a paste, a mistura, an aerosol, a gel, an ointment, a facial mask, a cream or an unguent.

7. A method for treating EGFR-TKIs related skin rash or paronychia, comprising administering the traditional Chinese medicine composition according to claim 1 to a patient in need thereof, including a human.

8. A use of the traditional Chinese medicine composition according to claim 2 in preparation of a formulation for treating EGFR-TKIs related skin rash.

9. A use of the traditional Chinese medicine composition according to claim 3 in preparation of a formulation for treating EGFR-TKIs related skin rash.

10. A use of the traditional Chinese medicine composition according to claim 2 in preparation of a formulation for treating EGFR-TKIs related paronychia.

11. A use of the traditional Chinese medicine composition according to claim 3 in preparation of a formulation for treating EGFR-TKIs related paronychia.

12. The use of the traditional Chinese medicine composition according to claim 5, wherein the formulation is a liniment, a water aqua, a decoction, a granule, a paste, a mistura, an aerosol, a gel, an ointment, a facial mask, a cream or an unguent.

13. A method for treating EGFR-TKIs related skin rash or paronychia, comprising administering the traditional Chinese medicine composition according to claim 2 to a patient in need thereof, including a human.

14. A method for treating EGFR-TKIs related skin rash or paronychia, comprising administering the traditional Chinese medicine composition according to claim 3 to a patient in need thereof, including a human.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0037] FIG. 1 is a schematic diagram of a change trend of a part C score of a WoMo scale in treating EGFR-TKIs related skin rash with the traditional Chinese medicine composition of the present disclosure.

[0038] FIG. 2 is a schematic diagram of a change trend of a total score of the WoMo scale in treating EGFR-TKIs related skin rash with the traditional Chinese medicine composition of the present disclosure.

[0039] FIG. 3 is a comparison of EGFR-TKIS related paronychia before and after treatment in case 1 of Example 4 (a is before treatment; and b is after treatment).

[0040] FIG. 4 is a comparison of EGFR-TKIS related paronychia before and after treatment in case 2 of Example 4 (a is before treatment; and b is after treatment).

[0041] FIG. 5 is a comparison of EGFR-TKIS related skin rash before and after treatment in case 1 of Example 5 (a is before treatment; and b is after treatment).

[0042] FIG. 6 is a comparison of EGFR-TKIS related skin rash before and after treatment in case 2 of Example 5 (a is before treatment; and b is after treatment).

[0043] FIG. 7 is a comparison of EGFR-TKIS related skin rash before and after treatment in case 3 of Example 5 (a is before treatment; and b is after treatment).

DESCRIPTION OF THE EMBODIMENTS

[0044] In order to show the technical solution, objective and advantages of the present disclosure more concisely and clearly, the present disclosure will be further described in detail below with reference to specific examples and accompanying drawings of the present disclosure.

Example 1. This example provided a traditional chinese medicine composition for treating EGFR-TKIs related skin rash.

[0045] 15 g of Lonicera japonica, 15 g of Taraxacum mongolicum, 9 g of Sophora flavescens, 15 g of Kochiae fructus, 10 g of dictamni cortex, 12 g of the moutan cortex, 12 g of Phellodendri Chinensis Cortex, and 6 g of Menthae Haplocalycis Herba (added lately). The whole prescription was 94 g, which was decocted in water and concentrated to 50 ml, containing 1.88 g crude drug per ml.

Example 2. This example provided a traditional chinese medicine composition for treating EGFR-TKIs related skin rash.

[0046] 5 parts of Lonicera japonica, 5 parts of Taraxacum mongohcum, 3 parts of Sophora flavescens, 5 parts of Kochiae fructus, parts of dictamni cortex, 5 parts of moutan cortex, 5 parts of Phellodendri Chinensis Cortex, and 2 parts of Menthae Haplocalycis Herba.

Example 3. This example provided a traditional chinese ,edicine composition for treating EGFR-TKIs related skin rash

[0047] 15 parts of Lonicera japonica, 10 parts of Taraxacum mongolicum, 8 parts of Sophora flavescens, 10 parts of Kochiae fructus, 12 parts of dictamni cortex, 6 parts of moutan cortex, 8 parts of Phellodendri Chinensis Cortex, and 10 parts of Menthae Haplocalycis Herba.

Example 4. This example provided a traditional chinese medicine composition for treating EGFR-TKIs related skin rash.

[0048] 20 parts of Lonicera japonica, 18 parts of Taraxacum mongolicum, 20 parts of Sophora flavescens, 20 parts of Kochiae fructus, 25 parts of dictamni cortex, 10 parts of moutan cortex, 10 parts of Phellodendri Chinensis Cortex, and 12 parts of Menthae Haplocalycis Herba.

[0049] Example 5. This example provided a traditional chinese medicine composition for treating EGFR-TKIs related skin rash.

[0050] 40 parts of Lonicera japonica, 50 parts of Taraxacum mongolicum, 50 parts of Sophora flavescens, 50 parts of Kochiae fructus, 50 parts of dictamni cortex, 50 parts of moutan cortex, 50 parts of Phellodendri Chinensis Cortex, and 30 parts of Menthae Haplocalycis Herba.

[0051] Example 6. This Example provided a method for preparing a traditional Chinese medicine composition for treating EGFR-TKIs related skin rash, which included the following specific steps:

[0052] (1) the aforementioned composition was weighed, added with 10 times the volume of water for soaking, decocted for 40-60 minutes, and then filtered to obtain a first medical solution;

[0053] (2) the dregs of decoction obtained in the first step was added with additional 10 times the volume of water, decocted for 30-40 minutes, and then filtered to obtain a second medical solution; and

[0054] (3) the filtrates obtained in step (1) and step (2) were combined and centrifuged, and the filtrate was concentrated to obtain a water decoction.

[0055] In the aforementioned preparation steps, after the twice decocting for a specified time, in order to avoid some components from losing due to precipitation after the water decoction was cooled, the medical solutions obtained in the two steps were filtered while they were hot.

[0056] (4) The decoction in step (3) was to heat and boil with high fire first, and then slowly decoct with mild fire to keep the solution in a slightly boiling state, which could not only reduce the volatilization of water, prevent the solution from drying and scorching, but also promote the extracting of effective components in medicinal materials.

[0057] (5) As a preferred protocol, the filtrate was concentrated to a filtrate concentration of 1.88 g/ml (i.e., every 1 ml of the concentrated solution contained 1.88 g of a decoction product of drugs), which was beneficial to control the volume of subsequent administration and exert a better pharmaceutical effect.

Example 7

[0058] The traditional Chinese medicine composition prepared in Example 1 was used for treating EGFR-TKIs related skin rash, specifically as follows:

[0059] 1. Treatment protocol

[0060] (1) Characteristics of population to be treated: 40 cases of patients who took EGRF-TKI and developed related skin rash. The average age of the patients was 54.70±12.00 years old, with the youngest being 30 years old and the oldest being 82 years old, wherein there were 25 males and 15 females. 32 cases (80.00%) were enrolled into stage IV, and 8 cases (20.00%) were enrolled into stage III. Mutation of exon 19 occurred in 12 cases (30.00%), mutation of L858R occurred in 19 cases (47.50%), and the details of 9 cases (22.50%) were unknown. Targeted drugs included Alfatinib in 15 cases (37.50%), Erlotinib in 7 cases (17.50%), and Gefitinib in 7 cases (17.50%), Icotinib in 5 cases (12.50%), Osimertinib in 4 cases (10.00%) and Dacomitinib in 2 cases (5.00%). The time from medication to occurrence of skin rash was 13.03±15.18 days.

TABLE-US-00001 TABLE 1 Description of study population Number of persons (cases) 40 Age 54.70 ± 12.00 (30.00-82.00) (mean ± SD; min-max, years old) Gender (case, %) Male 25 (62.50%) Female 15 (37.50%) Stages (case, %) Stage IV 32 (80.00%) Stage III 8 (20.00%) Gene detection result (case, %) Exon 19 deletion mutation 16 (40.00%) L858R mutation 24 (60.00%) Targeted drug (case, %) Afatinib 15 (37.50%) Erlotinib 7 (17.50%) Gefitinib 7 (17.50%) Icotinib 5 (12.50%) Osimertinib 4 (10.00%) Dacomitinib 2 (5.00%) Time from medication to skin rash (days) 13.03 ± 15.18

[0061] (2) Using method: external use, applying to the skin of patients with skin rash. 50 ml was administrated each time for twice a day, with 30 minutes each time, by externally applying onto the affected part or soaking the affected part. Each subject was given an externally applied facial mask of a decoction of skin rash-eliminating and itching-stopping prescription on his/her face, once in the morning and once in evening of a day, for 2 weeks.

[0062] 2. Evaluation Indicators

[0063] (1) Main research outcomes: a WoMo scale was used for evaluating the effectiveness of the skin rash-eliminating and itching-stopping prescription in treating EGFR-TKI related skin rash. It was compared whether there was any difference in the decrease of the score of the WoMo scale before and after treatment. In addition, a standard error of 1.96 for the difference before and after treatment was set as the minimally important clinical difference (MICD).

[0064] (2) Adverse reactions: the adverse events occurred during the research were recorded.

[0065] 3. Statistical Analysis

[0066] A Chi-square test was used for all counting data, wherein pearson Chi-square was used when the expected count <5 was no more than 20% and the minimum expected count >1; continuity correction was used when the expected count <5 was more than 20% and the minimum expected count was >1; and a fisher or likelihood ratio method was used when the minimum expected count <1 or n<40. Measurement data were expressed by mean±standard error (mean ±SD), and all measurement data used a homogeneity test for variance. On the contrary, a Mann-Whitney U test or t′ test was used for inter-group comparison, and a Wilcoxon signed-rank test or Wilcoxon signed rank test was used for intra-group comparison.

[0067] The outcome indicators of the research included the score change of the WoMo scale, such as a baseline (TO), after the first week of intervention (T1) and after the second week of intervention (T2), which belonged to replicate measurement data. A Generalized Estimation equation (GEE) is used for calculation of the replicate measurements.

[0068] p<0.05 was considered to be statistically difference. All confidence intervals were bilateral 95% confidence intervals. The differences between the two groups were compared, and all statistical hypotheses were 2-side tests. Empowerstats (www.empowerstats.com; X&Y solutions Inc., Boston, Mass.) and R (http://www.R-project.org) would be used for analysis.

[0069] 4. Treatment Results

[0070] (1) The traditional Chinese medicine composition of the present disclosure could decrease the score of the WoMo scale

[0071] The GEE was used for comparing the score changes of the WoMo scale before and after two weeks of treatment on the patients. As could be seen from FIG. 1 and Table 2, in the analysis, a baseline time was taken as the baseline of dependent variables, and for comparison at different time points, the partial regression coefficients of the week 1 (Time 1) and the week 2 (Time 2) were −2.4809 and −3.5863, respectively, which were both statistically significant (both with p values<0.0001). It showed that over time, the part C score of the WoMo scale has a significant decreasing trend, suggesting that the treatment was effective.

TABLE-US-00002 TABLE 2 Estimation of fixed effect parameters of linear mixed effect model (part C of WoMo scale) Estimated Standard Degree of T P Value error Freedom value value 95% CI. low 95% CI. upp Intercept  9.5142 0.4878 48 19.5045 <0.0001  8.5581 10.4703 Week 1 −2.4809 0.5673 48 −4.3732 <0.0001 −3.5928 −1.369  Week 2 −3.5863 0.5365 48 −6.6844 <0.0001 −4.6379 −2.5347

[0072] As could be seen from FIG. 2 and Table 3, when the GEE was used for analyzing the total score of the WoMo scale, in the analysis, a baseline time was taken as the baseline of dependent variables, and for comparison at different time points, the partial regression coefficients of the week 1 (Time 1) and the week 2 (Time 2) were −10.4973 and −15.5509, respectively, which were both statistically significant (both with p values <0.0001). It showed that over time, the part C score of the WoMo scale has a significant decreasing trend, suggesting that the treatment was effective.

TABLE-US-00003 TABLE 3 Estimation of fixed effect parameters of linear mixed effect model (total score of WoMo scale) Estimated Standard Degree of 95% CI. Value error Freedom T value P value low 95% CI. upp Intercept  44.4161 2.5721 47 17.2682 <0.0001  39.3747  49.4575 Week 1 −10.4973 2.4437 47 −4.2957 <0.0001 −15.2869  −5.7077 Week 2 −15.5509 2.3103 47 −6.7312 <0.0001 −20.079  −11.0228

[0073] (2) Adverse reactions: no adverse drug reactions were found during the treatment.

Example 8

[0074] The traditional Chinese medicine composition prepared in Example 1 was used for treating EGFR-TKIs related paronychia, specifically as follows:

[0075] 1. Treatment Protocol

[0076] (1) Characteristics of population to be treated: 16 cases of patients who took EGRF-TKI and developed paronychia. The average age of the patients was 53.81 ±11.99 years old, wherein there were 8 males and 8 females. 12 cases (75.00%) were enrolled into stage IV, and 4 cases (25.00%) were enrolled into stage IIIB. Mutation of exon 19 occurred in 10 cases (62.50%), L858R mutation occurred in 5 cases (31.25%), and 861Q mutation occurred in 1 case (6.25%). Among them, 15 cases were treated with Afatinib, and 1 case was treated with

[0077] Gefitinib.

TABLE-US-00004 TABLE 4 Description of study population Number of persons (cases) 16 Mean ± SD  Age (years old) 53.81 ± 11.99 N (%) Gender (case, %) Female 8 (50.00%) Male 8 (50.00%) Gene mutation EGFR Exon 19 Deletion 10 (62.50%) L858 R. 5 (31.2.5%) 861Q 1 (6.2.5%) TNM staging IV 12 (75.00%) III B 4 (25.00%) Used drug Afatinib 15 (93.75%) Gefitinib 1 (6.25%)

[0078] (2) Using method: external use, wherein 50 ml was administrated each time for twice a day, with 30 minutes each time, by externally applying onto the affected part or soaking the affected part. The administration was conducted once in the morning and once in evening of a day, for 2 weeks.

[0079] 2. Evaluation Indicators

[0080] (1) Main research outcomes: the grading of paronychia referred to the CTCAE 5.0 standard grading issued by NCI in 2017. See Table 5 for details. It was set that it was effective if it was decreased by one grade after treatment, it had a remarkable effect if it was decreased by two grades, and it was recovered if the paronychia was disappeared.

TABLE-US-00005 TABLE 5 Grading standard of paronychia Grade 1 Grade 2 Grade 3 Grade 4 Grade 5 Nailfold edema or erythema; In need of local treatment; oral In need of surgical treatment; — — damaged stratum corneum administration of a drug (e.g., intravenous administration of antibiotics, antifungal, antiviral antibiotics for treatment; therapy) for treatment; nailfold affecting self-rational daily edema or painful erythema; nail life activities shedding or nail plate separation; affecting daily life and instrumental activities

[0081] (2) Adverse reactions: the adverse events occurred during the research were recorded.

[0082] 3. Statistical Analysis

[0083] (1) A Chi-square test was used for all counting data, wherein pearson Chi-square was used when the expected count <5 was no more than 20% and the minimum expected count >1;

[0084] continuity correction was used when the expected count <5 was more than 20% and the minimum expected count was >1; and a fisher or likelihood ratio method was used when the minimum expected count <1 or n<40. Measurement data were expressed by mean±standard error (mean±SD).

[0085] p<0.05 was considered to be statistically significant. All confidence intervals were bilateral 95% confidence intervals. the differences between the two groups were compared, and all statistical hypotheses were 2-side tests. Empowerstats (www.empowerstats.com; X&Y solutions Inc., Boston, Mass.) and R (http://www.R-project.org) would be used for analysis.

[0086] 4. Treatment Results

[0087] (1) The therapeutic effect on paronychia after 2 weeks of external application of the skin rash-eliminating and itching-stopping prescription

[0088] Results were as shown in Table 6, and 16 persons with paronychia belonged to CTCAE 5.0 grade 2 (100.00%). After treatment, 3 persons (18.750%) were recovered (grade 0), and 13 persons (81.250%) were reduced to grade 1 according to the CTCAE 5.0 grading. The composition was effective in 16 persons, with an effective rate of 100.00%; and had remarkable effect/recovery effect in 3 persons, with the rate of remarkable effect of 18.750%.

[0089] There was a statistical difference in CTCAE staging before and after treatment (p=0.012).

TABLE-US-00006 TABLE 6 Therapeutic effect of the traditional Chinese medicine composition of the present disclosure on paronychia (CTCAE 5.0 grading) Before After Therapeutic effect treatment treatment Remarkable Grading (Person, %) (Person, %) Effective effect/recovery No 0 (0%) 3 (18.75%) Grade 1 0 (0%) 13 (81.25%) 16 3 Grade 2 16 (100.00%) 0 (0%) (100.00) (18.75%) Note: the Chi-square test was used for comparing the differences before and after treatment, and the theoretical number of counting variables was < 10, as obtained by a Fisher accurate probability test, with p = 0.012.

[0090] (2) Adverse reactions: no adverse drug reactions such as allergy and infection were found during the treatment.

Example 4

Clinical Experiment of the Traditional Chinese Medicine Composition of the Present Disclosure in Treatment of EGFR-TKIs Related Paronychia

[0091] 1. Case 1

[0092] In case 1, the patient was Wu XX, a 49 years old female. In June, 2019, she was diagnosed as stage IV of lung adenocarcinoma. and had exon 19 mutation as detected by Gene detection. She took 40 mg QD of Afatinib, and paronychia occurred in her 2 months later, and gradually worsened with formation of granulation-like tissue (FIG. 3a). From Aug. 3, 2019, she was soaked and washed with a topical skin rash-eliminating prescription twice a day, and after 1 week, the granulation tissue subsided, leaving only nail fold edema (FIG. 3b). The CTCEA grade was reduced from grade 2 to grade 1.

[0093] 2. Case 2

[0094] In case 2, the patient was Guan XX, a 59 years old male. In August, 2019, he was diagnosed as stage IV of lung adenocarcinoma. and had exon 19 mutation as detected by Gene detection. He took 40 mg QD of Afatinib, and paronychia occurred in him 1 months later, and gradually worsened with nail fold edema accompanied with pain appeared on the lateral edges of both thumbs (FIG. 4a). From August 25th, 2019, he was soaked and washed with a topical skin rash-eliminating prescription twice a day, and after 1 week, the nail fold edema subsided without pain and discomfort, and he was basically recovered (FIG. 3b). The CTCAE score was reduced from grade 2 to recovery.

Example 9

Clinical Experiment of the Traditional Skin Rash Composition of the Present Disclosure in Treatment of EGFR-TKIs Related Skin Rash

[0095] 1. Case 1

[0096] The patient was Huang XX, a 57-year-old male, who was diagnosed as left upper lobe adenocarcinoma with systemic multiple bone metastases after operation. From December, 2017 to October, 2018, he was treated by oral administration of Iressa, after more than 10 months, the tumor was progressed, and a severe skin rash was occurred after more than 4 months of administration. In November, 2018, Xia gene detection: Exon-21 (L858R mutation) and TP53 pathogenic variation were detected. In November, 2018, he had the first visit to our outpatient department, and was changed to oral administration of Tagrisso. In March, 2019, when the patient visited, the skin rash of him was progressively worsened. At that time, the skin rash on the face was mainly located on the left and right cheeks, was mainly papules and erythema, and had a dark red color. The total score of WoMo was 50, and the C part score of WoMo was 9 (FIG. 5a). After two weeks of intervention by administration of this protocol, the skin rash on the face was improved, and the facial papules and erythema obviously subsided. The total score of WoMo was 30, and the C part score of WoMo was 5 (FIG. 5b).

[0097] 2. Case 2

[0098] The patient was Shen XX, a 60 years old male. He was diagnosed as adenocarcinoma of the lower lobe of the left lung complicated with multiple M (IVa stage) in both lungs, bilateral supraclavicular bone, mediastinum and right hilum. From Nov. 14, 2018, he was given 7 courses of a platinum-pemetrexed (PP) regimen of chemotherapy +Avastin and immunotherapy. From May 9, 2019, he was given Alfatinib (30 mg) +Anlotinib (8mg), and the skin rash was occurred on the 16th day. The skin rash on the face of the patient was mainly located in the nose, cheeks of both sides and lower jaw, and was especially obvious in the nose. Meanwhile, erythema, papules and abscesses were also existed (FIG. 6a). The total score of WoMo was 60, and the C part score of WoMo was 13. After two weeks of external application of the skin rash-eliminating and itching-stopping prescription, the total score of WoMo was 30 and the C part score of WoMo was 7. The abscess of the nose was basically disappeared, leaving only a few papules and erythema (FIG. 6b).

[0099] 3. Case 3

[0100] The patient was Chen XX, a 57 years old male. On Aug. 20, 2018, the CT examination of the patient showed a left upper lung occupying lesion. Pathological examination: Lung adenocarcinoma. EGFR detection: Exon 19 mutation. He was subsequently treated with the traditional Chinese medicine. On May 22, 2019, the CT showed that the upper left lung adenocarcinoma was larger than before, with liver metastasis. Then he took Alfatinib. On Sep. 10, 2019, the reexamination of CT showed that the upper left lung adenocarcinoma was reduced by 50%. Then he was subjected to left lung and liver metastases ablation, and the first small lesions of liver metastases appeared. Therefore, he was treated with “1 g of pemetrexed disodium+0.5 g of Bevacizumab” in the same period on Oct. 15, 2019 and

[0101] November 14th, 2019. During the treatment, the patient developed severe skin rash, accompanied by cysts, abscesses, papules and erythema at the same time. The total score of WoMo was 80, and the C part score of WoMo was 10 (FIG. 7a). From Oct. 27, 2019, he began to be treated with the skin rash-eliminating and itching-stopping prescription. On the fourth day, the condition began to show obvious improvement, abscess began to heal, papules and cysts subsided, and the area of erythema was decreased. The total score of WoMo was 50, and the C part score of WoMo was 5 (FIG. 7b), so that the treatment effect was remarkable.

[0102] The examples described above are merely illustrative of several embodiments of the present disclosure, the description of them is more specific and detailed, but cannot be construed as limiting the scope of the present disclosure accordingly. It should be noted that, several variations and modifications can be made by those of ordinary skills in the art, under the premise of not departing from the concept of the present disclosure, and these variations and modifications all fall within the claimed scope of the present disclosure. Therefore, the scope of protection of the present disclosure shall be determined by the appended claims.