CN102961747A - Treatment of metastatic breast cancer - Google Patents

Abstract

The present invention relates to the use of an anti-EpCAM antibody for the manufacture of a medicament for the treatment of metastatic breast cancer. The present invention further relates to a method of treating metastatic breast cancer comprising administering said anti-EpCAM antibody.

Description

The treatment of metastatic breast cancer

The present invention is to be on 02 09th, 2007 the applying date, and application number is that the 200780009489.7(international application no is PCT/EP2007/001127), name is called the dividing an application of application for a patent for invention of " treatment of metastatic breast cancer ".

Invention field

The present invention relates to anti-EpCAM antibody for the preparation of the purposes of the medicine for the treatment of metastatic breast cancer.The invention still further relates to the method for the treatment of metastatic breast cancer, comprise and use described anti-EpCAM antibody.

Background technology

Breast carcinoma is the modal cancer of women, and is the second largest reason of woman cancer death.Calendar year 2001 is 90-100/100 at the sickness rate of U.S.'s breast carcinoma, 000, and be 50-70/100 at European sickness rate, 000.The sickness rate of this disease increases in the world wide.The risk factor of breast carcinoma comprise ethnic group (race), age and tumor suppressor gene BRCA-1 and-2 and p53 in sudden change.Excessive drinking, rich fatty diet, do not get enough athletic exercise, hormone and ionizing radiation also increase the risk of growing breast carcinoma after the menopause of external source.Estrogen receptor and progesterone receptor negative breast cancer (being respectively " ER-" and " PR-"), larger tumor size, cytology's height pathological changes (high grade cytology) relevant with poor prognosis (bad prognosis) with the age that is lower than 35 years old (Goldhirsch etc. (2001) .J.Clin.Oncol.19:3817-27).2005, estimate and to diagnose out the new case of 212,000 routine wellability (invasive) breast carcinoma and the new case of 58,000 routine non-infiltration breast carcinoma that wherein expection will have 40,000 women to die from breast carcinoma.

Breast carcinoma can be divided into several Main Stage usually: (the locally advanced) in early stage, local late period, local recurrence with metastatic.Breast carcinoma of early stage comprises non-infiltration breast carcinoma, for example lobular carcinoma in situ (lobular carcinoma in situ) (" LCIS ") and ductal carcinoma in situ(DCIS) (ductile carcinoma in situ, " DCIS ").The most normally, carry down according to tumor lympha and to move (Tumor Node Metastasis, " TNM ") categorizing system is stages for breast cancer, this categorizing system is provided by american cancer joint committee (the American Joint Committee on Cancer) (AJCC cancer staging handbook, the 6th edition).The TNM categorizing system has defined 7 different stages of breast carcinoma: 0, I, IIA, IIB, IIIA, IIIB and IV.The subclass of stage 0, I and Phase is usually relevant with breast carcinoma of early stage.The subclass of Phase I and Phase is usually relevant with advanced breast cancer.Phase IV is usually relevant with metastatic breast cancer.More details of the TNM classification of relevant breast carcinoma are shown in Fig. 1.Can measure and monitor tumor size by replying evaluation criterion (" RECIST ") (Therasse etc. (2000) .J.Natl.Cancer Inst.92:205-16) in the solid tumor.

The 5-survival prognosis of breast carcinoma of early stage surpasses 60% usually, and for advanced breast cancer, this number falls between the 40-60%.For metastatic breast cancer 5-survival prognosis common about 15%.The common site that the breast carcinoma far-end shifts comprises lung, liver, skeleton, lymph node, skin and CNS (brain).In case metastatic breast cancer is made a definite diagnosis, can expect that the patient on average lived 18-24 month again.The unlikely healing of metastatic breast cancer, the mode for the treatment of this systemic disease is (palliative) of slowing down property in essence mostly.

More than emphasized the especially importance of the new development in the treatment of metastatic breast cancer of breast cancer treatment.

Current treatment breast carcinoma comprises that the treatment of metastatic breast cancer selects to comprise operation (excision for example, autologous bone marrow transplantation), X-ray therapy, chemotherapy (anthracycline such as doxorubicin (doxorubicin) for example, alkylating agent is cyclophosphamide and ametycin (mitomycin C) for example, taxane is paclitaxel (paclitaxel) and Docetaxel (docetaxel) for example, antimetabolite is capecitabine (capecitabine) for example, microtubule inhibitors is vinblastine nvelbine (navelbine) for example, endocrine therapy (antiestrogen such as tamoxifen (tamoxifen) for example, progesterone is medroxyprogesterone acetate (medroxyprogesterone acetate) and megestrol acetate (megastrol acetate) for example, aromatase inhibitor (aromatase inhibitor) is aminoglutethimide (aminoglutethamide) and letrozole (letrozole) for example) and biological preparation (cytokine for example, immunotherapeutic agent is such as monoclonal antibody).Metastatic breast cancer is treated by a kind of chemotherapeutics (the most effective medicine comprises cyclophosphamide, doxorubicin, nvelbine, capecitabine and ametycin) or the combination of chemotherapeutics and endocrine therapy in the most common ground.

The standard care method of breast carcinoma is the exenterate tumor and carried out radiotherapy according to tumor stage and risk factor before or after hormone therapy or chemotherapy.Can be in Phase I to the patient of Phase IA (see below, and Fig. 1) by the treatment of adjuvant chemotherapy or hormonotherapy.For the patient who suffers from inoperable wellability Phase IB disease or Phase IV metastatic breast cancer, chemotherapy only alleviates described symptom.

Found that recently taxane and anthracycline significantly improve patient with breast cancer's survival rate.Capecitabine (Xeloda

Capecitabine, Roche Ltd:Summary of Product Characteristics (product feature general introduction)) has been approved for the second line (second-line) or the more senior treatment among the failed patient of cytotoxicity chemotherapy, described cytotoxicity chemotherapeutant comprises anthracycline and/or taxane, especially because its hypotoxicity and oral formulations (O ' Shaughnessy (2002) .Oncology 16:17-22).Yet although the treatment feature (modalities) of these improvement, advanced breast cancer patient's survival rate is still very low, and chemotherapy is slowing down property only.

Monoclonal anti-Her-2/neu antibody trastuzumab (trastuzumab) (Herceptin Trastuzumab, Roche, Ltd: product feature general introduction, in March, 2002) be first target therapeutic agent biology that is approved for the treatment patient with breast cancer, wherein said patient with breast cancer's tumor is crossed expression Her-2/neu.The combination of it and paclitaxel, the First Line (first-line) that is called as the metastatic breast cancer patient as single agent in same patient colony are treated and are called as the second line or more senior treatment (Cardoso etc. (2002) .Clin.Breast Cancer 3:258-9; Tan-Chiu and Piccart (2002) .Oncology 63:57-63).Yet, only have fraction patient with breast cancer (about 20%) high level to cross expression Her2/neu, therefore be suitable for using this Antybody therapy.

Therefore, exploitation new anti-cancer drug (in particular for treating not (not indicated) patient with breast cancer's of display effect new anti-cancer drug of trastuzumab) is important medical science needs.

Summary of the invention

A kind of promising immunotherapeutic agent is people's antibody, comprises such as SEQ ID NO.3,4 and 5 aminoacid sequences of listing at its variable region of heavy chain, and/or comprises such as SEQ ID NO.6,7 and 8 aminoacid sequences of listing at its variable region of light chain.Hereinafter this antibody is called " Anti-EpCAM ", and further characterizes with its heavy chain of listing such as SEQ ID NO.1 and 2 and the aminoacid sequence in the light chain respectively.This antibodies epithelial cell adhesion molecule (epithelial cell adhesion molecule, " EpCAM " is also referred to as 17-1A antigen, KSA, EGP40, GA733-2, ks 1-4 and esa).EpCAM is the surface glycoprotein of high conservative, and it is crossed in many cancers (comprising breast carcinoma) of Different Origin and expresses.List in tissue Microarray and to utilize sensitivity immunohistochemical staining analysis of test methods to express from the EpCAM of the tumor sample of 3722 colons, stomach, lung, ovary or patients with prostate cancer.All surpassing 88% in the tumor sample and reported that moderate to strong EpCAM expression, is 94% in the ovarian cancer, is 94% in the colon cancer, is 92% in the gastric cancer, is 90% in the carcinoma of prostate, is 71% in the pulmonary carcinoma.These results confirm that EpCAM usually is present in the epithelial tumor cell, have given prominence to Anti-EpCAM as potential diagnosis and treatment target.

In two researchs of primary breast tumor, be respectively 36% to show that strong EpCAM expresses (Tandon etc. (1990) Cancer Res.50:3317-24) in 384 sections, and 59% show that strong EpCAM expresses (Edwards etc. (1986) Cancer Res.46:1306-17) in 128 samples.In another research (Spizzo etc. (2002) Int.J.Cancer 98:883-8), find to have in 205 primary breast tumor samples 73 (36%) to exist strong EpCAM to express, the author reports EpCAM in the breast carcinoma and crosses and express relevant with the overall survival rate without tumor survival rate (disease-free survival) with reduction.It is also relevant with the hormone receptor negativity with tumor size that EpCAM crosses expression; It is the highest in conduit breast carcinoma and histology III level hypotype.In another series, show that about 90% breast carcinoma sample is expressed EpCAM to a certain extent, and the strong EpCAM of 40% above demonstration expresses.

External, Anti-EpCAM causes the cell toxicant (" ADCC ") of antibody dependence and the cell toxicant (" CDC ") of Complement Dependent.Anti-EpCAM is by raising patient's natural killer cell to tumor locus in conjunction with the EpCAM-positive tumor cell, and this is most probable mechanism of action.Then via the activation of patient's immune effector cell, the EpCAM-positive tumor cell can be removed.

Some therapeutic scheme that utilizes Anti-EpCAM is (WO 2005/080428) known in the art.Especially, WO 2005/080428 has described and has related to the therapeutic scheme that Anti-EpCAM is applied to the cancer patient.This paper is expected in the treatment of breast carcinoma for example or the remaining disease (minimal residual disease) of Minimum Residual and uses Anti-EpCAM.In the latter's context, the remaining disease of Minimum Residual is appreciated that tumor by local and the non local recurrence that causes for by single tumor cell survival.

Target of the present invention is to improve existing breast carcinoma therapy.

Particularly, the present invention relates to following aspect:

1. contain SEQ ID NO.3,4,5,6,7 and/or 8 anti-EpCAM antibody for the preparation of the purposes of the medicine for the treatment of metastatic human breast carcinoma.

2. 1 purposes, wherein said anti-EpCAM antibody contains SEQ ID NO.1 and/or 2.

3. 1 or 2 purposes, wherein said treatment comprises the long term stabilization of metastatic breast cancer.

4. according to each purposes of item 1-3, wherein metastatic breast cancer is carried down according to tumor lympha and is moved (" TNM ") system and classify as the IV phase.

5. aforementioned each purposes wherein with the load doses of at least one 2mg/kg body weight, is used described medicine for the form of a plurality of maintenance dosies afterwards, and each maintenance dose is the 2mg/kg body weight.

6. each purposes of aforementioned 1-4 wherein with the load doses of at least one 6mg/kg body weight, is used described medicine for the form of a plurality of maintenance dosies afterwards, and each maintenance dose is the 6mg/kg body weight.

7. the purposes of item 5 or 6 is wherein used weekly described/each load doses, and per two weeks are used each described maintenance dose.

8. 7 purposes, wherein when beginning, the 1st, 2 and 3 all each for the treatment of use a load doses, be 11 maintenance dosies afterwards, when the 4th, 6,8,10,12,14,16,18,20,22 and 24 all each for the treatment of begins, use a maintenance dose.

9. aforementioned each purposes is wherein used described antibody in the solution that contains 0.9% sodium chloride solution.

10. aforementioned each purposes, wherein intravenous is used described antibody.

11. comprising the people used, the method for the treatment of metastatic human breast carcinoma, described method comprise SEQ ID NO.3,4,5,6,7 and/or 8 anti-EpCAM antibody.

12. the method for item 11, wherein said anti-EpCAM antibody contains SEQ ID NO.1 and/or 2.

13. the method for item 11 or 12, wherein said treatment comprises the long term stabilization of metastatic breast cancer.

14. according to each method of item 11-13, wherein metastatic breast cancer is carried down according to tumor lympha and is moved (" TNM ") system and classify as the IV phase.

15. each method of 11-14 wherein with the load doses of at least one 2mg/kg body weight, is used described anti-EpCAM antibody for the form of a plurality of maintenance dosies afterwards, each maintenance dose is the 2mg/kg body weight.

16. each method of 11-14 wherein with the load doses of at least one 6mg/kg body weight, is used described anti-EpCAM antibody for the form of a plurality of maintenance dosies afterwards, each maintenance dose is the 6mg/kg body weight.

17. the method for item 15 or 16 is wherein used weekly described/each load doses, per two weeks are used each described maintenance dose.

18. the method for item 17, wherein when beginning, the 1st, 2 and 3 all each for the treatment of use a load doses, be 11 maintenance dosies afterwards, wherein when the 4th, 6,8,10,12,14,16,18,20,22 and 24 all each for the treatment of begins, use a maintenance dose.

19. each method of aforementioned 11-18 is wherein used described antibody in the solution that contains 0.9% sodium chloride solution.

20. each purposes of 11-19, wherein intravenous is used described antibody.

Detailed Description Of The Invention

Therefore, one aspect of the present invention relates to the anti-EpCAM antibody (" Anti-EpCAM ") that comprises SEQ ID NOs.3,4,5,6,7 and/or 8 listed aminoacid sequences for the preparation of the purposes of the medicine for the treatment of metastatic human breast carcinoma.

Term used herein " metastatic breast cancer " will be interpreted as a kind of disease, at least one cell from the conversion of breast primary tumor is that cancerous cells is separated from described primary tumor in this kind disease, and continues to grow into tumor being different from the residing position of described primary tumor (hereinafter referred to as " diverse location ").For example described diverse location can be in the residing same breast of described primary tumor (homonymy breast) or in another breast (offside breast).As a further example, described different parts can be in one or more lymph nodes, no matter its for movably or fixing, be homonymy or offside, SC, the oxter or other with respect to described primary tumor.In the context of TNM staging system (shown in Figure 1), " metastatic breast cancer " used herein will comprise particularly (i.a.) it will comprise that by stages M=1 (is Phase IV breast carcinoma; Referring to Fig. 1) all tumors, namely all exist any degree to the remote location tumor of the transfer of lung, liver, skeleton, lymph node, skin, brain and/or homonymy and/or the intramammary diverse location of offside for example.

Term used herein " breast carcinoma " refers to exist in single or two breast disease of primary tumor or multiple independent primary tumor (multiple individual primary tumors).This often means that (going back) do not isolate cancerous cell in the primary tumor from breast, and do not spread to " diverse location ".In this context, should be noted that the existence interior or homonymy and the interior Multiple primary carcinomas tumor of offside breast of same breast itself can not be interpreted as the implication that falls into " metastatic breast cancer ".This be because one or simultaneously two intramammary a plurality of cells can produce the Multiple primary carcinomas tumor, be not metastatic or not yet become metastatic.On the other hand, in the single breast a plurality of primary tumors only one of them separates cancerous cell, this individual cells that is positioned at " diverse location " buds into independently that tumor will consist of " metastatic breast cancer " used herein subsequently, no matter have one or more primary tumors at least one breast.

Should be noted in the discussion above that term used herein " metastatic breast cancer " and do not mean that the specific primary tumor generation of any one in breast of described transfer that is present in " diverse location ".That is to say that the origin of the transfer of described " diverse location " is immaterial for the title of the disease that is called " metastatic breast cancer ", rise in breast tissue as long as produce the primary tumor that shifts.For this reason, term " breast tissue " is interpreted as lobule (lobule) and the conduit that comprises breast, namely produces the modal tissue of mammary neoplasms.

The applicant has found surprisingly that Anti-EpCAM not only is very suitable for the treatment that such breast carcinoma namely relates to the breast carcinoma of at least one primary tumor in the breast, also is suitable for the treatment of metastatic breast cancer.Anti-EpCAM can use by this way fully and can't expect, because the tumor load relevant with metastatic breast cancer (tumor load) (is the quantity of cancer in cancerous cell number, tumor size or the body; Be also referred to as " tumor load (tumor burden) ") usually greater than viewed in the non-metastatic breast carcinoma.This be because single primary breast tumor can be at an easy rate-usually really-produce the transfer of many diffusions at whole body.Therefore, in metastatic breast cancer, be present in the absolute number of the lip-deep EpCAM molecule of whole body malignant cell usually greater than the absolute number in the non-metastatic breast carcinoma.The data show that provides among the appended embodiment is used the significant prolongation of the disease TTP (to the time (time to progression) of development) that the pharmaceutical composition that comprises anti-EpCAM antibody causes treating.It is relevant with the lip-deep EpCAM expression of the malignant cell for the treatment of that actual effect seems.Can be according to (2000) Lancet 356 such as Gastl, 1981-2 assigns to different EpCAM expresser (expressors) groups with the patient.Briefly, Gastl etc. has analyzed the EpCAM that separates from different patients' tumor cell and has expressed in immunohistochemical staining.Total immunostaining fractional computation is the product of representative fraction (proportion score) and intensity mark (intensity score).According to Gastl etc., described representative fraction describe estimated positive staining tumor cell mark (0, nothing; 1,＜10%;＜10%; 2,10%-50%; 3,50%-80%; 4,〉80%).According to Gastl etc., and the estimated staining power of described intensity mark representative (0, dye-free; 1, a little less than; 2, moderate; 3, strong).The gained total points is between 0 to 12.High EpCAM is defined as total points greater than 4, because described patient's sample shows the bimodal distribution (low and high EpCAM expresser) that EpCAM expresses, distinguishes minimum point (discriminating nadir) and is in total score value of 3 to 4.For the prognosis of accepting to comprise the patient who uses the anti-EpCAM Antybody therapy, the patient who is accredited as high EpCAM expresser will be more more optimistic than the patient who is accredited as moderate or low EpCAM expresser.Consistent with this observed result, the patient who is accredited as high EpCAM expresser (compares with high EpCAM expresser's average, the patient who has shown higher EpCAM amount on the malignant cell surface, i.e. total points 〉=8) compare the further prolongation that shows TTP with the patient who is accredited as high EpCAM expresser and accepts identical treatment.In addition, it is directly relevant with prognosis to be applied to the amount of anti-EpCAM antibody in patient's the pharmaceutical composition.Particularly, use the anti-EpCAM antibody of low dosage with the patient who organizes for identical EpCAM expresser and compare, the anti-EpCAM antibody of administered with high dose causes the prolongation of the disease TTP that treats.

Anti-EpCAM antibody A nti-EpCAM is people's antibody in a preferred embodiment.

Anti-EpCAM comprises all SEQ ID NO.3,4,5,6,7 and 8 in more preferred embodiment.Anti-EpCAM comprises SEQ ID NO.1 and/or 2 in more preferred embodiment.Anti-EpCAM comprises SEQ ID NO.1 and 2 in particularly preferred embodiments.

According to preferred embodiment, the treatment of metastatic breast cancer comprises the long term stabilization of metastatic breast cancer." long term stabilization " is interpreted as that disease progression is stabilized in it in the course for the treatment of of anti-EpCAM antibody and begins level or be stabilized in to be lower than the situation that it begins level.This can be understood as to the prolongation of used time of disease progression." long term stabilization " also comprises the situation of wherein tumor atrophy (part is replied)." long term stabilization " but comprise that also but disease progression wherein is reduced to detection level or is lower than the situation of detection level, namely the patient has completely treatment and replys, disease is cured (replying fully).In this case, if need indefinitely continual cure of prevent disease recurrence, perhaps can be according to doctor's judgement stopped treatment.

According to preferred embodiment, described medicine is suitable for so-called " low dosage is used (low dose administration) ".Use for described low dosage, the dosage range of at every turn using is 1 to 3mg anti-EpCAM antibody/kg body weight.Preferred described low dosage is used and is comprised that scope is at least one load doses of 1 to 3mg/kg body weight, is a plurality of maintenance dosies afterwards, and each maintenance dose scope is 1 to 3mg/kg body weight.Also each dosage range of using of preferred low dose is 1.5 to 2.5mg/kg body weight, and more preferably scope is 1.75 to 2.25mg/kg body weight.Most preferably each dosage of using of low dosage is the 2mg/kg body weight.Use for low dosage, at least one load doses that preferred low dose is used is the 2mg/kg body weight, is a plurality of maintenance dosies afterwards, and each maintenance dose is the 2mg/kg body weight.Perhaps described medicine is suitable for so-called " high dose is used ".Use for high dose, the dosage range of at every turn using is 4.5 to 8mg anti-EpCAM antibody/kg body weight.Preferred described high dose is used and is comprised that scope is at least one load doses of 4.5 to 8mg/kg body weight, is a plurality of maintenance dosies afterwards, and each maintenance dose scope is 4.5 to 8mg/kg body weight.Also each dosage range of using of preferred high dose is 5 to 7mg/kg body weight, and more preferably scope is 5.5 to 6.5mg/kg body weight, and further preferable range is 5.75 to 6.25mg/kg body weight.Most preferably each dosage of using of high dose is the 6mg/kg body weight.Use for high dose, at least one load doses that preferred high dose is used is the 6mg/kg body weight, is a plurality of maintenance dosies afterwards, and each maintenance dose is the 6mg/kg body weight.Unexpectedly such load and maintenance dose bring the treatment benifit can for the treatment of metastatic breast cancer, and wherein a plurality of transfers of whole body will be uprooted.

According to another embodiment of the present invention, determine that the metastatic breast cancer patient is higher degree or more to express EpCAM may be useful on low degree ground.Such as according to (2000) Lancet 356 such as Gastl, 1981-2 is described, the patient can be divided into non-EpCAM expresser group, moderate EpCAM expresser group, low EpCAM expresser group and high EpCAM expresser group.In general, it is useful that amount and the described patient's that observes the EpCAM expression that will be applied to metastatic breast cancer patient's Anti-EpCAM associates, high EpCAM expresser accepts the more Anti-EpCAM of high dose, and low EpCAM expresser accepts the more Anti-EpCAM of low dosage.Use the higher person in two above-mentioned dosage of Anti-EpCAM to the patient who expresses high-level EpCAM, namely the 6mg/kg body weight may be useful especially.

According to another embodiment of the present invention, persistent period between each load doses and another continuous load dosage or first maintenance dose, (duration of time) will be no longer than a week, and the persistent period between each maintenance dose and the ensuing maintenance dose will be no longer than two weeks.Preferably use weekly described/each load doses, per two weeks are used each described maintenance dose.Use weekly the Anti-EpCAM load doses at " load phase (loading phase) " and guarantee to keep Anti-EpCAM floor level (draining and the lasting removing of the form of elimination is taken into account) in the sufficiently high serum that can cause required therapeutic effect always.This required Anti-EpCAM floor level of therapeutic effect is called as " serum line of rabbet joint level (serum trough level) ", hereinafter will so censure.In case reach this serum levels, further use the Anti-EpCAM of maintenance dose at " maintenance phase (maintenance phase) " subsequently in two-weekly interval, the serum levels of guaranteeing (removing that will continue is equally taken into account) Anti-EpCAM never is reduced to and is lower than the required level of continued treatment effect.The required pharmacokinetics calculating of serum line of rabbet joint level of measuring Anti-EpCAM is (referring to the WO 2005/080428) that this area had been described.

According to particularly preferred embodiment, when beginning, the 1st, 2 and 3 all each for the treatment of use a load doses, be 11 maintenance dosies afterwards, when the 4th, 6,8,10,12,14,16,18,20,22 and 24 all each for the treatment of begins, use a maintenance dose.Here found surprisingly 3 load doses in the above-mentioned interval, afterwards for 11 maintenance dosies in the above-mentioned interval to be combined in treatment metastatic breast cancer aspect effective especially.This means that the always treatment phase is 24 weeks (not considering to check after relevant with this treatment of any classification under normal circumstances any treatment) from start to finish.In other preferred embodiment, the always treatment phase is 30 weeks, 40 weeks, 50 week or 60 weeks from start to finish.Further preferably using EpCAM antibody a period of time according to such scheme then is the time of not using EpCAM antibody to use the treatment phase of EpCAM antibody with another.Such cycle order can repeat several times.

Perhaps, another embodiment expection of the present invention is used Anti-EpCAM with above-mentioned load doses, uses maintenance dose according to the required consumption of control tumor development afterwards.In this embodiment, as long as the size of one or more monitored metastatic tumo(u)r does not increase and just can be considered as tumor development and be brought under control.In the case of the best, the size of one or more tumors of monitoring in fact may atrophy (as in part is replied).The tumor of this place monitoring can extremely not exist in atrophy, and (as in replying fully) namely can disappear.The tumor of monitoring can keep formed objects, and the time to disease progression may increase (as in the disease of stabilisation) like this.Therefore, according to this embodiment, in above-mentioned interval, can give continuously indefinitely the Anti-EpCAM of maintenance dose, as long as exist part to reply or stable replying, be continued until in described case, to record fully and reply.Further develop in tumor in the situation of (size of the tumor of namely monitoring during the treatment or number increase), can stop the treatment with Anti-EpCAM, if suitable, replace the Anti-EpCAM treatment with the replacement therapy form.

According to another embodiment of the present invention, Anti-EpCAM uses as the solution that comprises 0.9% sodium chloride.

According to another embodiment of the present invention, the Anti-EpCAM intravenous is applied to the metastatic breast cancer patient.

Another aspect of the present invention is the purposes that Anti-EpCAM is used for the treatment of metastatic breast cancer.

Another aspect of the present invention relates to the method for the treatment of metastatic human breast carcinoma, described method comprise with comprise SEQ ID NOs.3,4,5,6,7 and/or 8 anti-EpCAM antibody is applied to the people.This antibody is further characterized by SEQ ID NOs.1 and 2 listed heavy chain and light-chain amino acid sequences respectively.

Described in the preferred embodiment of the method for this treatment metastatic breast cancer such as the context of above purposes of the present invention; These embodiments that are applied to the inventive method have been done necessary correction (mutatis mutandi).

Now will be by following accompanying drawing and non-limiting example explanation the present invention.

The accompanying drawing summary

The summary of Fig. 1 breast carcinoma TNM classification/Staging System

Fig. 2 is to time (" the TTP ") figure line of development, and it showed without the probability of disease progression with respect to the time that gives respectively the low and high application dosage Anti-EpCAM of 32 and 35 patients

Fig. 3 (" TTP ") figure line, it showed without the probability of disease progression with respect to the time that gives respectively the low and high application dosage Anti-EpCAM of 27 and 40 patients

Fig. 4 (" TTP ") figure line, it shows the time of using high and low dosage Anti-EpCAM without the probability of disease progression with respect to the patient of and low-level EpCAM high to expression

Fig. 5 (" TTP ") figure line, its demonstration is compared with all other patients, without the probability of disease progression with respect to the time to the patient's administered with high dose Anti-EpCAM that expresses high-level EpCAM.

Fig. 6 (" TTP ") figure line, it showed without the probability of the disease progression time with respect to the Anti-EpCAM that gives respectively the low and high application dosage of 54 and 55 patients

Fig. 7 (" TTP ") figure line, it shows the time of using high and low dosage Anti-EpCAM without the probability of disease progression with respect to the patient of and low-level EpCAM high to expression

Fig. 8 (" TTP ") figure line, it shows with the low dosage Anti-EpCAM that uses to the patient who expresses low-level EpCAM compares, without the probability of disease progression with respect to the time to the patient's administered with high dose Anti-EpCAM that expresses high-level EpCAM.

Embodiment

General

Following examples are used for illustrating different aspect of the present invention, never mean to limit the scope of the invention.Substantially, described embodiment has described for the clinical research program of the complete human IgG1's antibody design that is called " Anti-EpCAM " and the result who is obtained by this clinical research program.SEQ ID NO.3,4 and 5 listed respectively the Anti-EpCAM variable region of heavy chain first, the aminoacid sequence of second and the 3rd complementary determining region (CDR).SEQ ID NO.6,7 and 8 listed respectively the Anti-EpCAM variable region of light chain first, the aminoacid sequence of second and the 3rd CDR.SEQ ID NO.1 has listed the heavy chain amino acid sequence of Anti-EpCAM, and SEQ ID NO.2 has listed the light-chain amino acid sequence of Anti-EpCAM.Below use following term and abbreviation among all embodiment:

Embodiment 1: for the II phase clinical research program of Anti-EpCAM design

Embodiment 1.1: the clinical research general introduction

Clinical research for the Anti-EpCAM design is summarized in following table (table 1).

Table 1: clinical research general introduction

Embodiment 1.2: utilize the non-clinical study general introduction of Anti-EpCAM

Utilize nine breast cancer cell lines to study the effect that Anti-EpCAM and trastuzumab are induced ADCC.Anti-EpCAM can mediate the ADCC specificity cracking with the trastuzumab par.The concentration of the EpCAM molecule on the cell surface has when adopting Anti-EpCAM strong related between the sensitivity to ADCC.All obtain maximum specificity cracking in the Anti-EpCAM concentration that is lower than 10 μ g/mL under all situations, the described Anti-EpCAM concentration that is lower than 10 μ g/mL is the minimum target line of rabbet joint concentration (minimal trough concentration targeted in patients) in the patient body.

During (intended) intravenous route administration of implementing to plan, Anti-EpCAM has shown fabulous part tolerance in rabbit, do not observe the change of macroscopic view, only observes less microscopic change.

Embodiment 1.3: the ultimate principle that dosage is selected

Based on preclinical test, for the Anti-tumor activity of Anti-EpCAM, expect that the serum line of rabbet joint level of 10 μ g/mL is effective.Yet can not get rid of higher dosage may be more effective.Therefore will assess as calculated the second dosage with the serum line of rabbet joint level that obtains 30 μ g/mL in this research.

The dosage of plan is no more than the maximum dose level that the I phase is applied to the patient in studying in this research.Utilize pharmacokinetic model to calculate load phase and maintenance phase to obtain in a short time target line of rabbet joint serum-concentration, avoided maximal plasma concentration to surpass the situation that the I phase assesses in studying.

Embodiment 1.4: known benifit

Preclinical data hint metastatic breast cancer and other tumor patients may be benefited from by Anti-EpCAM and remove the tumor cell of expressing EpCAM and the development that causes delay (stable disease).

Embodiment 1.5: the clinical research target

Primary goal

● the clinical benifit of two different Anti-EpCAM dosage among the assessment EpCAM positive, the metastatic breast cancer patient

By-end

● reply parameter for other of two various dose of Anti-EpCAM assessment

● assess safety and the toleration of two different Anti-EpCAM dosage

● measure the pharmacokinetics of two different Anti-EpCAM dosage

● assess the pharmacodynamics of two different Anti-EpCAM dosage

● measure the pharmacokinetics of Anti-EpCAM among the metastatic breast cancer patient

● the pharmacodynamics of assessment Anti-EpCAM (NK cell)

Embodiment 1.6: project

The research terminal point; Main terminal pointClinical benifit rate (SD+PR+CR) during the 24th week

According to RECIST (referring to embodiment 1.18), described clinical benifit rate is defined as stable disease (SD)+part and replys (PR)+reply fully ratio of (CR) patient.

The research terminal point; Secondary endpoints

● the clinical benifit rate (SD+PR+CR) during the 12nd week

● best overall tumor response rate (overall tumor response rate, OTR)

● reply the persistent period/to the development time

● the incidence rate of adverse events and laboratory abnormalities

● the serum-concentration of Anti-EpCAM

● the number of periphery NK cell (NK) cell

The holistic approach design

This studies for open-label, multicenter, randomization, parallel group, II phase that research low/moderate or high EpCAM in 24 week treatments express the effect of two Anti-EpCAM therapeutic doses among the patient and safety.

There are altogether 112 patients to join in this research.After the screening phase, satisfy the patient of all ambiguities (illegibility) standard will be in each EpCAM group one in randomization to two treatment group: low dose group and high dose group.Anti-EpCAM will use weekly (the 1st day, the 8th day and the 15th day) as 60 minutes intravenous (i.v.) infusions during the load phase, and after this per two weeks use, and amount to 24 week or until disease progressions.

According to its EpCAM expression with patient's classification (stratified).Two groups are arranged here: low/moderate EpCAM expression group, and high EpCAM expression group.Table 2 shows the summary of this extra classification.

Table 2: the treatment group of classification and Anti-EpCAM dosage

Treatment group	EpCAM expresses	The Anti-EpCAM dosed administration
			The I group	Moderate EpCAM on the primary tumor expresses	2mg/kg Anti-EpCAM i.v., per two weeks
The II group	Moderate EpCAM on the primary tumor expresses	6mg/kg Anti-EpCAM i.v., per two weeks
			The III group	High EpCAM on the primary tumor expresses	2mg/kg Anti-EpCAM i.v., per two weeks
The IV group	High EpCAM on the primary tumor expresses	6mg/kg Anti-EpCAM i.v., per two weeks

By clinical assessment and the per 6 weeks monitoring of laboratory test patient, until the 24th week, after this per 8 week monitorings (during the follow-up study).Tumor response be will more assess to estimate if detect the skeleton pathological changes during screening, thorax CT scan or chest X-ray, abdominal CT scan or MRI and bone scintigraphy art comprised.

With every group effect of assessment, with reply the persistent period/compare with the expected value in this patient colony to the time of development.To assess tumor response and be used for statistical analysis, wherein by RECIST (referring to embodiment 1.18) standard test tumor response.

The main terminal point of this research is that each organizes the clinical benifit rate (CR+PR+SD) when the 24th week in four groups.With assessment clinical benifit rate (SD+PR+CR), the best overall tumor response rate (OTR) in the 12nd when week, reply the persistent period/to time of development as secondary endpoints.Safety and toleration result when comparing for the 24th week between the treatment group.

For effect or toleration is former thereby speech, if whenever one of them dosage becomes obviously is superior during the research, then will correspondingly revise this research rules.

Treating the patient who has recorded its SD, PR or CR after 24 weeks and wherein do not reported unacceptable toxicity (and not interrupting greater than the treatment in 4 weeks) will submittedly participate in the follow-up study that continuous Anti-EpCAM treats.With evaluate parameter for example long-term tolerance, clinical progress and overall survival.

Embodiment 1.7: clinical research colony

Comprise standard (Inclusion Criteria)

1. only have as all following standard where applicable patients and just will be suitable for participating in the research:

2. the metastatic breast cancer of confirming through the histology, it is measured by SABC when screening, has positive EpCAM and express in (archived) tissue sample that files

3. have at least a pathological changes (namely shifting), it can measure (according to RECIST (referring to embodiment 1.18)) at least in one dimension (one dimension)

4. life expectancy (Life Expectancy) 〉=12 months

5.ECOG physical situation (performance status) 0-1

6. age 〉=18 year old

7. understand and be ready to sign the ability of written agreement of informing

Exclusion standard (Exclusion Criteria)

If following arbitrary standard is applicable then the patient will be unsuitable for participating in this research:

1. according to the assessment of researcher, when being included, it recommends/preferred any other treatment

2.CNS transfer medical history

3. according to the assessment of researcher, be used for trastuzumab (Herceptin

) treatment indication (indication)

4. except following, treatment begins immunization therapy, irradiation, chemotherapy or any other anticancer therapies in front 4 weeks:

● the 1st time medical before the localization radiotherapy (target lesion can not be in irradiation area, and the expection radiotherapy should not cause suppressing such as the marrow of definition in the exclusion standard 6) of beginning

● hormone therapy, the patient who treats is progressive, and stops before research treatment beginning

5. treatment begins any research products in front 4 weeks

6. such as undefined unusual organ or bone marrow function:

● hemoglobin concentration≤90g/L ot 9.0g/dL

● leukocyte＜3x10 ⁹/ L (3000/mm ³)

● platelet count＜100x10 ⁹/ L (100,000/mm ³)

● AST (SGOT) or ALT (SGPT)〉upper limit (ULN) of 2x normal value

(if having hepatic metastasis then〉5x ULN)

● serum creatinine〉1.5x ULN

● serum lipase〉1.5x ULN

● serum amylase〉1.5x ULN

7. except the malignant tumor medical history except breast carcinoma in front 5 years of the treatment beginning, the basal cell carcinoma or carcinoma in situs of cervix of skin

8. according to the judgement of researcher, be considered to any other complication or the medical disorder that to disturb this research to carry out

9. treatment begins front 4 all interior desired immunosuppressant for example demand or regular use of systemic corticosteroid

10. known HIV (human immunodeficiency virus) (HIV) infects and/or hepatitis B virus (HbsAg is positive) or hepatitis C virus (HCV positive resistance) infection

11. gestation or nursing women or during participating in this research and the women that reproductive potential is arranged who was unwilling to use effective contraceptive method in after this at least 3 months

12. the known allergy for any other component of immunoglobulin or this drugs preparation

Embodiment 1.8: the clinical trial material

Goods

The product A nti-EpCAM that is used for this research of clinical trial purposes provides as the solution of GMP quality, the Anti-EpCAM such as in waiting phosphatizing acid buffer, comprising 10mg/mL, be stored in+2 and+8 ℃ between

Calculate for the preparation of the amount to the Anti-EpCAM of the final solution of patient's infusion according to patient's body weight and treatment group (referring to top table 2).

To in clean, gnotobasis (laminar flow hood), be diluted in Anti-EpCAM in 500mL 0.9% sodium chloride solution.In order to be mixed for the final Anti-EpCAM solution of infusion, should put upside down gently sack, avoid producing foam.The concentrated solution and the final solution that are used for infusion all only are used for special purpose.

Treatment designated program (Treatment Assignment Procedure)

Will be as far as possible near the initial randomization that carries out of Anti-EpCAM treatment.Must satisfy all qualifications (eligibility) standard during randomization.To provide concentrated randomization operation by ICRS (interactive computer answering system).Researcher must sign in on the safe network address with identiflication number and password separately, must provide basic patient's data (patient's numbering, garbled data, date of birth, EpCAM result of the test) at described network address s/he, to accept to relate to the immediate acknowledgment for the treatment of appointment.Randomization operation is not to express by the center but by EpCAM to treat prescribed fractionated, to guarantee high in two EpCAM expression groups and low dose therapy balanced distribution.In case obtained required patient's number in the treatment group, just can not have more patient to be randomized this individual groups.

Should in 60 minutes, the flow velocity with 500mL/h the solution intravenous be applied to the patient.

Embodiment 1.9: treatment

The treatment schedule

Each patient will accept altogether 14 Anti-EpCAM infusions in the treatment in 24 weeks, unless disease progression occurs or treat restricted toxicity.The Anti-EpCAM solution that is used for infusion will be applied to the patient in the angular vein at 60 minutes, and will be weekly during the load phase, during the maintenance phase whenever biweekly.The 2nd time medical (the 1st day) is front, will arrive following one of them treatment group to patient's randomization in each EpCAM expression group:

● I and III group (low dosage): load phase 2mg Anti-EpCAM/kg body weight, weekly (the 1st, 2 and 3 week) is the maintenance dose of 11 2mg Anti-EpCAM/kg body weight, whenever biweekly afterwards

● II and IV group (high dose): load phase 6mg Anti-EpCAM/kg body weight, weekly (the 1st, 2 and 3 week) is the maintenance dose of 11 6mg Anti-EpCAM/kg body weight, whenever biweekly afterwards

The EpCAM assay of carrying out during according to screening is concentrated randomization operation and classification.The interactive computer answering system will be for registration EpCAM result and patient's data.In case registered described data, will give patient treatment and specify.Will each center calculation patient separately dosage and for the preparation of the final solution of infusion.

Treating the patient who has recorded its SD, PR or CR after 24 weeks and wherein do not reported unacceptable toxicity (and not interrupting greater than the treatment in 4 weeks) will submittedly participate in the follow-up study that continuous Anti-EpCAM treats.

Because the treatment of adverse events is ended

According to the order of severity and the cause effect relation of adverse events, can interrupt or give up the study of medication, perhaps the patient for high dose group can reduce dosage.

If researcher has sufficient evidence to show that the not described research medication of described adverse events causes, should continual cure.Yet, if can not get rid of adverse events and the relation of studying medication, should change dosage.

Patient's abort criterion

Particularly in following any event, should end to utilize the treatment of described research product:

● disease progression, such as RECIST defined (referring to embodiment 1.18)

● what the patient agreed withdraws from

● patient or researcher are not observed described research rules

● the medical disorder development, the suggestion of researcher is to stop the further participation of patient in described research

● use unauthorized parallel medication

● researcher concludes that the change treatment meets patient's maximum benefit

If ●

Zero researcher thinks that described adverse events is significant clinically, With

Zero does not solve to≤2 grades before next time administration, And

Zero at least may be relevant with the research medication,

3 grades of adverse events have then occured.

● 4 grades of adverse events occur, and dosage interrupted surpassing 4 weeks

● according to researcher and/or patient's suggestion, the generation of any adverse events is so that need maybe essential the termination.

The patient also should carry out the safety follow-up investigations that all are ranked before ending, to allow evaluation studies terminal point (referring to embodiment 1.11).

Parallel medication

All parallel medications all should be recorded in the case report form (CRF).Following medication and treatment are unallowed during the whole research:

● any Anti-tumor except this research product is for example treated:

Zero hormone therapy

Treatment zero biology

Zero chemotherapy

Zero radiotherapy (exception: the 1st time medical before the localization radiotherapy (target lesion can not be in irradiation area, and the expection radiotherapy will can not cause suppressing such as the marrow of definition in the exclusion standard 6) of beginning)

If get along with at p-GLU-HIS-TRP-SER-TYR-D-TRP-LEU-ARG-PRO-GLY-NH2 before entering research with patient's (hormone receptor+) before the menopause of p-GLU-HIS-TRP-SER-TYR-D-TRP-LEU-ARG-PRO-GLY-NH2 treatment, they can continue this treatment during the research so.

● long-term systemic high dose corticosteroid treatment and other immunosuppressant therapy

● any other research agent

According to standard operation, if necessary, should use the supporting treatment (supportive therapy) that medically needs, and should be recorded among the CRF.Shift the patient for bone, allow when entering research with bisphosphonates (salt) class (bisphosphonates) treatment.

Embodiment 1.10: assessment

Qualification (Eligibility)/efficacy assessment

The agreement of informing: must before any particular studies program, obtain written agreement of informing there from each patient.

Comprise/get rid of: the qualification of the assess patient that should summarize such as above-mentioned embodiment 1.7-clinical research colony comprises the summary of all experimental determinations that carry out for screening.

Medical history/Medical illness: generally comprise past and present medical disorder history with medical history specified disease; The complete history of record patient cancer course of disease during screening comprises tumor stage, other prognostic marker, the front information of Anti-tumor treatment.

Parallel medication: will during whole research, record all parallel medications.

Hepatitis B and hepatitis C detect: will carry out hbs antigen (HBsAg) and hepatitis C virus (HCV) antibody test during screening, to get rid of the Active infection of B-mode or hepatitis C virus.

Tumor assessment: will be according to RECIST (referring to embodiment 1.18) standard tumor response

● bone scintigraphy art: will when screening and final going to a doctor, implement the bone scintigraphy art with existing that the assessment bone shifts.If when screening exists bone to shift or shows that clinically (pain and alkali phosphatase for example occurring raises) exists bone to shift then carry out more scannings.

● thorax computerized tomography (Computerized-Tomography, CT) scanning or chest X-ray: will carry out thorax CT or chest X-ray with existing that the record far-end shifts in when screening, and periodical evaluation is replied when in treatment phase process and for the last time medical.

● abdominal CT scan or MRI: will carry out abdominal CT scan or MRI with existing that the assessment far-end shifts in when screening, and periodical evaluation is replied when during treating and for the last time medical.

Safety evaluation

Adverse events: will record the process neutralization for the treatment of phase until the adverse events (AE) that 4 weeks occurred behind the last Anti-EpCAM infusion.During the screening phase, also should report the AE relevant with search procedure.

Serious adverse events: comprise during the whole research phase that screening and tracking phase will record all serious adverse events (SAE).

Physical examination: carry out comprehensive physical examination that all airframe systems comprise vital sign when going to a doctor at regular intervals and for the last time during screening, during the treatment phase.Suitable words will be carried out the physical examination for symptom in whole research process, and will record all clinical correlated results.On the infusion date, should before the Anti-EpCAM infusion, implement physical examination.

Vital sign (Vital Signs) monitoring: will be in whole research following mensuration body temperature (oral cavity or tympanum), heart rate and blood pressure (contraction/relaxation):

● before the infusion

● during the 2nd to 6 infusion when medical per 15 minutes

● after the 2nd to 6 time the infusion when medical finishes per hour until 4 hours

ECOG mark: the physical situation that will utilize east tumor cooperative groups (ECOG) mark (referring to the table 6 among the embodiment 1.19) assess patient.

Electrocardiogram (ECG): will be in when screening, treatment phase process and carry out standard 12-wire (12-lead) ECG when medical for the last time.All should obtain two printouts for each ECG, one is used at the working space record, and another cardiologist by the center assesses.

Safety experiment chamber assessment: when each research that will be in the morning is medical (the infusion date before infusion) obtain the blood sample for the assessment of safety experiment chamber, and will carry out following analysis:

● clinical chemistry: aspartate transaminase (AST), alanine aminotransferase (ALT), gamma glutamyltransferase (GGT), alkali phosphatase (AP), lactic acid dehydrogenase (LDH), total bilirubin, total protein, creatinine, carbamide, uric acid, glucose, calcium, sodium, potassium, chloride, phosphate ester (salt), amylase, lipase, albumin, proteins C reactive (CRP).

● hematology: red blood cell count(RBC) (RBC), hemoglobin, hematocrit (hematocrit), numeration of leukocyte (WBC), differential blood count (differential blood count) and platelet count.

● blood coagulation: prothrombin time (Prothrombin time, PT, international normalized ratio [INR]), part thromboplastin time (PTT) and fibrinogen (fibrinogen).

Urinalysis (Urinalysis): will when each going to a doctor, assess the existence of glucose, protein and blood in the urine by dipstick (dipstick).On the Anti-EpCAM infusion date, should before the Anti-EpCAM infusion, carry out urinalysis.

Pregnancy tests: will when screening and last going to a doctor, have the women of reproductive potential to carry out pregnancy tests (β-human chorionic gonadotropin [β-HCG]) to all.

Immunogenicity: will when when screening, the 6th and 24 weeks and last twice the 17th and 18 secondary trackings are gone to a doctor, obtain the immunogenic blood sample for assessment of Anti-EpCAM.

The pharmacodynamics assessment

Natural killer cell: will measure by the cell sorter (FACS) of fluorescence-activation the number of NK cell (NK) cell.

The pharmacokinetics assessment

The mensuration Anti-EpCAM serum line of rabbet joint and peak level when per 6 to 8 weeks and last three the 16th to 18 secondary trackings are medical when will the in the 2nd to 6 time medical, in the treatment phase process.

The EpCAM expression analysis

The tumor material evaluation EpCAM that will utilize the patient to file when screening expresses.To measure EpCAM by SABC in central laboratory expresses.

The patient who only has EpCAM result can proceed all screening sequences, and described EpCAM result is low/moderate or high.Patient with negative EpCAM testing result does not satisfy the qualification standard, will be considered to screen unsuccessfully.

If carry out slicer (for example in the situation of the new transfer that detects) during the research, also should collect tumor tissues and express for analyzing EpCAM.

Embodiment 1.11: medical schedule

The calculating that all researchs are gone to a doctor is take baseline (the 1st day) as the basis, and described baseline is defined as that day that gives first Anti-EpCAM infusion dosage.

The screening phase (the-28 days to the-1 day)

Only have those patients that satisfy all qualification standards to understand designated patient's numbering (referring to embodiment 1.8-treatment designated program).

All screening assessments all must be carried out in 28 days using for the first time described research product (the 1st day).All results comprise EpCAM detection of expression result, all should be obtainable announcing that the patient is suitable for participating in before the research.In case can obtain to satisfy all qualification standards from result and the patient of Laboratory Evaluation, the randomize routine that researcher should be proceeded to concentrate (centralized randomization procedure) (ICRS).

The treatment phase (the 2nd to 15 time medical)

Should carry out following program and assessment during the treatment phase:

Zero vital sign

Zero physical examination

The assessment of zero safety experiment chamber

Zero urinalysis

Zero pharmacokinetics

Zero NK cell

Zero immunogenicity

Zero parallel medication

Zero adverse events and serious adverse events

Zero chest X-ray/CT scan

Zero abdominal CT scan or MRI

Zero bone scintigraphy art (if detecting osseous lesion when screening)

Zero ECG when medical (only the 6th and 15 time)

Phase and final PK/PD assessment (the one 16-18 time medical) are followed the tracks of in safety

To treat rear two weeks of end and carry out the safety tracking all around and go to a doctor.The 17th time medical is that last safety tracking is medical.

Last research medical (the 18th time medical) is the final PK/PD assessment in 12 weeks after treatment finishes.

The medical end of research (the 17th time medical/last)

If should follow the tracks of the last time when medical or the patient ends then at any time to finish research medical from research in advance.

● comprehensive physical examination

● vital sign

● 12-wire ECG

●ECOG

● the assessment of safety experiment chamber

● urinalysis

● pregnancy tests

● chest X-ray/CT scan

● the bone scintigraphy art

● abdominal CT scan or MRI

● pharmacokinetics

● immunogenicity

● parallel medication

● adverse events and serious adverse events

Effect is followed the tracks of the phase

To finish to participate in rear until followed the tracks of patient's disease progression, other treatment of cancer and survival rate in 1 year per 3 months in the research process.The patient who have stable disease during the 24th week, partially or completely replys will have an opportunity to enter open-label research, will carry out the assessment of disease progression and overall survival in open-label research.

Embodiment 1.12: sample storage

The blood serum sample that is used for immunogenicity, NK cell counting and PK will be in-20 ℃ of freezer storage in the research place.

Embodiment 1.13: security consideration

Researcher is responsible for satisfying detection and the record of the event of adverse events (AE) or serious adverse events (SAE) definition.This comprise assessment its seriousness (seriousness), its order of severity (severity) and with the cause effect relation of research product and/or concurrent treatment.

Embodiment 1.14: adverse events, serious adverse events

Adverse events

Use during the described Research of Medical product or the following event that detects afterwards or diagnose out is adverse events:

● in the deterioration of the disease that pre-exists or permanent disorder,

● at frequency or the strength increase of the incident that pre-exists or illness,

● use afterwards S or S that detect or that diagnose out (Signs or symptoms) of described Research of Medical product, even they may occur or exist before participation research,

● through hematology and other laboratory abnormalities and any event that causes intervening of labelling, comprise that withdrawing from testing drug/research product treats, reduces dosage or significantly extra concurrent treatment.

Serious adverse events

Serious adverse events (experience) or reaction are any undesirable medical events or the effects that is in any dosage:

● cause death, (1)

● be life-threatening, (2)

● need hospitalization or prolong existing inpatient's hospital stays,

● cause lasting or significantly incapability or impotentia,

● be birth defect or birth defect.

Severity: according to treatment of cancer appraisal procedure (Cancer Therapy Evaluation Program), with the generic term standard (CTCAE) of adverse events, the order of severity (or intensity) of the classification scale assessment AE that version 3 .0 provides.

Embodiment 1.15: statistical analysis

The mensuration of statistical method and research variable

The target of this research is to measure effect and the safety of Anti-EpCAM.Therefore, carry out the randomization II phase of two groups (low and high dose Anti-EpCAM) and test, wherein each treatment group will be considered to the standard single phase II phase and study.To make separately the judgement whether the treatment evidence has shown enough activity for each treatment group.Sample size assessment and main statistical analysis are designed to the basis with standard single group II phase single phase.

Table 3 has been listed the mensuration that will carry out and/or data management operations to determine main and less important research terminal point.

Table 3: main and less important goal in research, derive variable and assay method.

Statistical hypothesis: make following supposition for the statistical hypothesis of described research:

Estimate the clinical benifit rate of background of best supportive care (supportive care)≤5% (p ₀).If true clinical benifit rate (π) 〉=25% (p ₁), will there be sizable benefit in the following patient colony (positive EpCAM expresses) that is used for describing of Anti-EpCAM.

For each treatment group (the low or high dose group in low/moderate and the high EpCAM group), use following hypothesis:

H _0ij(π≤p _0ij): p _Ij≤ 5% (the clinical benifit probability of background)

H _1ij(π 〉=p _1ij): p _Ij〉=25% (for the interested clinical benifit probability of Anti-EpCAM)

P wherein _iFor with dosage level i treatment low/the clinical benifit rate observed in the middle of moderate EpCAM expression group (j=1) and the high EpCAM expression group patient (j=2), i=1 represents low dosage level, i=2 represents the high dose level.

Significance level, multiple comparisons (Multiple Comparisons) and multiplicity (Multiplicity): think that the II type error (85% ability) of 5% I type error and 15% is enough to determine described clinical benifit rate.There is no need to adjust significance level, this is neither because multiple comparisons neither be because of multiplicity (the not comparison of confirmation property between the treatment group).

The mensuration of sample size: the sample size assessment is based on the standard single stage operation of Fleming, but utilize accurate binomial distribution (A ' Hern (2001) .Statistics in Medicine 20:859-66), but not to the normal approximation (Fleming (1982) .Biometrics 38:143-51) of binomial distribution.These means are preferred, because normal approximation is incorrect for little sample size, and have following advantages based on sample size and the cut-off point of accurate distribution, if namely obtained described cut-off point then the confidence interval of calculating does not comprise p ₀Therefore the sample size table that A ' Hern (A ' Hern (2001) .Statistics in Medicine 20:859-66) is provided is used for this research.

Calculate according to these, studying every group for this needs 24 patients that can estimate effect, if like this in 24 weeks true whole response rate be 25% (p ₁), so for response rate, there is 85% chance (being ability=85%) to show that 95% one-sided confidence interval (being I type error=5%) gets rid of 5% (p ₀).

Suppose that about 10% patient can not estimate effect (person of abandoning (drop-outs)), then should (low/moderate EpCAM group has 54 with at least 108 patients altogether, wherein use low dose therapy for 27, use high-dose therapy for 27, high EpCAM expresses level 54, wherein use low dose therapy for 27, use high-dose therapy for 27) randomization is in this research.

The analysis of plan

Main and the secondary variable of detection property ground assessment.To analyze in descriptive manner all related datas (from data, the laboratory data of CRF) about the patient according to treatment group and medical grouping.

Each patient's data (according to treatment group and patient's number class) are provided in tabulation.List collect among the CRF with the data base in all data of comprising.

Demography and other baseline characteristic: summarize demographic and other baseline characteristic with integral body with by treatment group by the tabulate statistics (patient's number, average, standard deviation, minima, intermediate value, maximum) of continuous variable and according to the absolute and relative frequency of classified variable.Baseline characteristic is defined as the result of all inspections of carrying out before using Anti-EpCAM first.

The analysis for main terminal point of planning: Main Analysis will be based on complete analysis bank (full analysis set), described complete analysis bank refer to will carry out as sensitive analysis based on the analysis according to (per-protocol) groups of rules.The main terminal point of this research is clinical benifit rate (patient who has stable disease+CR+PR according to RECIST (referring to embodiment 1.18))

In the first step, assess separately the clinical benifit rate in each treatment group.In each treatment group, calculate 95% one-sided confidence interval for clinical benifit rate.If the lower limit of 95% one-sided confidence interval of response rate is greater than p in the treatment group ₀=5%, will get rid of described null hypothesis for this treatment group so.Cut-off point is 4 in this research, in case the meaning is to have obtained 4 the patient of clinical benifit is arranged then can to get rid of described null hypothesis for this individualized treatment group in treatment group.The further analysis of main terminal point will comprise:

● if all treatment groups have all shown enough activity, compile the treatment group with same dose level, and allow the dosage level of different clinical benifit rates the patient who relatively expresses for low/moderate and high EpCAM.Relatively being undertaken by the Logic Regression Models that possesses two key elements (being that EpCAM expresses and dosage level) of two dosage levels therefrom calculated the suitable odds ratio (odds ratios) of described two levels.

● if two dosage levels only show enough activity in a patient colony (have low/moderate EpCAM expression or have high EpCAM and express), then compare in descriptive manner these two dosage levels.

The analysis for secondary endpoints of planning: assess the best overall tumor response rate in the 24th when week for each treatment group, and with carry out for the described same way as of main terminal point.

Summarize adverse events according to treatment group and dosage level with integral body, its Major Systems organ classification according to them, high level item, preference and order of severity grouping.

Not between treatment group and dosage level to the comparison that takes statistics of the whole incidence rate of adverse events.

According to treatment group and dosage level for all samples date comprehensive summing up data.For the sample date behind all baselines by treatment group and the dosage level comprehensive summing up absolute change from baseline value.The displacement table of showing no increases in output and being conigenous baseline according to treatment group and dosage water for urinalysis data integral body.

Whole for all Study dates, according to treatment group and dosage level, by initial data being provided and analyzing in descriptive manner pharmacodynamic parameter from the tabulate statistics (average, standard deviation, minima, intermediate value and maximum) of the variation of baseline.

The administration data are checked

In order to determine further strategy for follow-up Anti-EpCAM, carry out the administration analysis of best overall response rate,

● after 70 patients have accepted at least one times infusion and have passed through the 9th medical the/the 12 week or ended in advance described research, and

● the data that relate to the best overall response rate are considered to quite clean analyzing for this administration and will stop recruiting (recruitment).

Embodiment 1.16: quality control and quality assurance

Before adding, the registration of research place must can obtain specific authority file the patient, for example the approval of independent Ethics Committee (independent ethics committee, IEC) and researcher and research office worker's resume.

By the qualified of research sponsor appointment and the people's study on monitoring through suitably training.

Embodiment 1.17: law and Ethical Demand

ICH the Tripartite Guideline Guidelines that coordinates and laws and regulations that all are suitable for according to good clinical practice carry out these researchs, the Declaration of Helsinki that comprises in June, 1964, it was modified by the 48th AMM that holds at the Somerset of Republic of South Africa West in October, 1996.

Patient information and the agreement of informing

The process of obtaining the agreement that the patient informs meets all applicable rule requests.Before being included in any patient in the described clinical research, must obtain in writing the agreement of informing that he/her freely expresses.

Embodiment 1.18:RECIST standard

It below is the summary of the RECIST standard used during the whole aforementioned research.

Qualification

The patient who only suffers from measurable disease at the baseline place just should be included in the rules, and wherein to reply be main terminal point to target tumor.

At least a measurable pathological changes of measurable disease-exist.Become if described measurable disease is limited to the single-shot sexually transmitted disease (STD), should confirm its neoplasm character by cytology/histology so.

The pathological changes of measurable pathological changes-can in one dimension at least, accurately measure, longest diameter 〉=20mm or longest diameter 〉=10mm when measuring with Spiral CT scan when utilizing conventional art to measure.

Immeasurablel pathological changes-all other pathological changes, comprise that (longest diameter＜20mm or when measuring with Spiral CT scan when measuring with conventional art＜10mm), namely osseous lesion, leptomeningeal sarcoma (leptomeningeal) disease, ascites, pleura/pericardial effusion, struvite mastopathy, lymphangitis skin/lung (pulmonis), cystic lesion (cystic lesions) also have the abdominal part caking (masses) that confirms and follow the tracks of without imaging technique to small lesions.

Should utilize ruler or clamp to obtain and with all measured values of metric system symbol record.All baseline estimates carry out in the time of all should approaching as far as possible the treatment beginning, must not surpass for 4 weeks before the treatment beginning.

The baseline place with follow the tracks of during should use identical appraisal procedure and identical each evaluation of characterized by techniques and the pathological changes of report.

Only have when clinical disease becomes (for example, skin nodules and the palpable lymph node) of shallow, just think measurable.For the case of dermatosis, recommend by the colour phhotograpy record, comprise the size of estimating pathological changes with ruler.

Measuring method

CT and MRI are current for measuring for the available of the best of replying the target lesion that assessment selects and method repeatably.Should be that conventional CT and MRI are carried out in 10 millimeters or less cutting continuously with slice thickness.Should utilize the continuous algorithm for reconstructing of 5mm to carry out spiral CT.This is applicable to the tumor of breast, abdominal part and pelvis.Those tumors of head and neck tumor and limbs need specific rules usually.

When pathological changes on chest X-ray is clearly determined and surrounded by gas lung (aerated lung), can accept it and be measurable pathological changes.Yet CT is preferred.

When the main terminal point of research is the target response assessment, should not use ultrasonic (US) to measure neoplastic lesion.Yet the possible replacement scheme of its shallow table tangibly lymph node that is clinical measurement, subcutaneous pathological changes and thyroid nodule.US also can be used for confirming the complete obiteration of superficial foci, and it is assessed by clinical examination usually.

The application in the target tumor assessment of splanchnoscopy and laparoscopy is not also ratified (validate) fully and widely.Their application in this particular range need only some center obtainable sophisticated equipment of ability and highly qualified specialist.Therefore these technology are replied for target tumor and should be only limited to the purpose that specialization is ratified in the heart.Yet these technology can be used for confirming that pathology are replied completely when having obtained slicer.

Only tumor markers can not be for assessment of replying.If mark is higher than the upper limit of normal value at first, so when all pathological changes have all disappeared, for the patient that will consider they must be in normalization in the complete clinical response.

Cytology and histology can be used for distinguishing PR and CR (for example, for example distinguishing remaining benign lesion and remaining malignant change in the germ cell tumor at tumor type after the treatment) in rare case.

The baseline record of " target " and " non-target " pathological changes

Until maximum is all measurable pathological changes that five pathological changes of each organ amount to 10 pathological changes, it represents all organs that involve, and all should be accredited as target lesion and at baseline place record with measure.

The suitability of should be according to its size (longest diameter of pathological changes) and being used for (by imaging technique or clinically) accurate repeated measure comes the select target pathological changes.

Calculate all target lesions longest diameter (LD) summation and be reported as baseline summation LD.Described baseline summation LD will as benchmark, characterize target tumor by it.

All other pathological changes (or disease location) should be accredited as non-target lesion, and also should be at baseline place record.Do not need to measure these pathological changes, but during whole tracking, should record existence or the disappearance of each pathological changes.

Reply standard

(1) although the obvious development of " non-target " pathological changes is exceptional case, in these cases, treatment doctor's suggestion should be preponderated, and should confirm described state of development by checking group (or research chairman) after a while.

The assessment that best overall is replied

Described best overall is replied and is referred to reply the minimum measured value of record (will be from treatment begins as the benchmark of PD) from treating initial until the best that disease progression/recurrence is recorded.Usually the best of patient is replied the acquisition that appointment will be depended on measured value and validation criteria.

The integral body of health status worsens to need to end treatment and should classify as the patient who does not at that time have the objective evidence of disease progression and to have " severity of symptoms ".Should make every effort to the record object development, even if after treatment is ended.

May be difficult in some cases residual disease is distinguished with normal structure.When this mensuration was depended in the assessment of replying fully, the suggestion described residual disease of research (fine needle aspiration thing/slicer) was to confirm the situation of replying fully.

Confirm

The main purpose of confirming target response is to avoid the viewed response rate of too high estimation.Reply in infeasible situation confirming, these researchs of report as a result the time should clear and definite described replying without affirmation.

In order to specify the situation of PR or CR, must be by the repeatedly variation of assessment affirmation measurement of tumor value, described repeatedly assessment should be no less than after first fit is replied standard to be carried out in 4 weeks.It is suitable also to can be according to the determined longer interval of research rules.

With regard to SD, the tracking measurement value has satisfied the SD standard at least during the minimum interval after just entering research (usually being not less than 6-8 week), and described minimum interval is determined in described research rules.

The persistent period that integral body is replied

The persistent period that integral body is replied began to measure from the time (being as the criterion with the situation that is recorded at first) of the measurement standard that satisfies CR or PR, until recorded objectively the first date of recurrence or PD, will certainly treat the minimum measured value of home record as the benchmark of PD.

The persistent period of stable disease

SD initially begins to measure until satisfied the standard of disease progression from treating, will be from the minimum measured value for the treatment of home record as benchmark.

For different tumor types and grade, the Clinical Correlation of SD persistent period is different.Therefore, advise that strongly described rules appointment is for required minimum interval between two measured values measuring SD.This interval should may bring this situation the clinical benifit of expection of Research Group and take into account.

Reply and check

Be the test of main terminal point for response rate, strong suggestion is checked all replying by the expert who is independent of described research when research is finished.Archives and the radiation image of checking simultaneously the patient are optimal paths.

Result's report

Must assess all patients of being included in described research to the replying for the treatment of, be underproof even great rules treatment deviation or they are arranged.A kind of with in the designated following classification of each patient: 1) reply fully, 2) part is replied, 3) stable disease, 4) PD, 5) because malignant disease and Deaths, 6) because toxicity and Deaths, 7) because other former thereby Deaths, perhaps 9) unknown (can not assess, data are incomplete).

All patients that satisfy the qualification standard should be included in the Main Analysis of response rate.Being in the patient who replys classification 4-9 should be considered to treatment nonreply (disease progression).Therefore incorrect treatment plan or medicament administration do not cause described response rate analysis is excluded.The explication of classification 4-9 will be that rules are specific.

All conclusions should be take all qualified patients as the basis.

Then can carry out the Asia as the basis take patient's subgroup and analyze (subanalyses), described patient's subgroup has been got rid of those patients (for example, owing to the Deaths of other reason, the early abort for the treatment of, great rules are run counter to etc.) that identify great rules deviation.Yet these inferior analyses can not be as the basis of the conclusion that draws relevant therapeutic efficiency, and report reason that the patient is excluded from described analysis with should be understood that.

95% confidence interval should be provided.

The general introduction of embodiment 1.19:ECOG muscle power situation

ECOG muscle power situation grade is disclosed in Oken, M.M. etc. (1982) Am J Clin Oncol5:649-655.

The administration data of embodiment 2:Anti-EpCAM II phase clinical research are checked

Embodiment 2.1: research overview and introduction

Such as clinical research as described in carrying out as described in the above embodiment 1.The result of now this research follows among the embodiment 2.

Methodology:

Randomization, open-label, multicenter, parallel group, II phase are studied.This research design comprises positive EpCAM test (with positive EpCAM testing) for assessment of effect and the safety of Anti-EpCAM in the treatment of 24 weeks of two various dose.The EpCAM result of the test of carrying out during according to screening is with the randomization classification at center.In case be registered in one of them EpCAM group, the patient has just been randomly assigned low dose therapy group or high-dose therapy group.

Patient's number of randomization and treatment: 112 (28 patients are still underway)

Patient's number of analyzing: the patient of 73 treatments (37 Anti-EpCAM high doses, 36 Anti-EpCAM low dosages)

The data of analyzing for aforementioned analysis:

Require to have monitored all data that cause these results according to GCP, but do not finish data scrubbing.Finished all until the 12nd week medical and all patients' CRF (n=23) with all tumors assessments in the 6th and 12 required weeks are the objects that medical science is checked, the emphasis that medical science is checked is to find out great rules deviation and cleaning data of safety.

Analyze for this, checked these 23 patients' radioactivity data at the center.

Be used for the definition of the colony of analysis:

● safety analysis group (SAF): all have accepted the patient of the appointment drugs of at least one dosage.

● complete analysis bank (FAS): from the patient of safety analysis group, wherein these patients have the EpCAM positive (low/moderate or high expressed) tumor, and owing in the former thereby situation about withdrawing from ahead of time of other, have at least a tumor to assess except clinical disease development after the treatment beginning.

The analysis of data of safety is take SAF as the basis.

The analysis of base-line data and effect terminal point is take complete analysis bank as the basis.

The terminal point (with reference to above-described embodiment 1.1) of analyzing:

● best overall tumor response (OTR) rate during the 12nd week (according to RECIST, having the patient that complete remission [CR] or partial symptoms are alleviated [PR])

● the clinical benifit rate (CBR) in the 12nd week (according to RECIST, having the patient that stable disease or complete remission [CR] or partial symptoms are alleviated [PR])

● the clinical benifit rate (CBR) in the 24th week (according to RECIST, having the patient that stable disease or complete remission [CR] or partial symptoms are alleviated [PR])

● to time (TTP) of development for from the time the randomization and time from treatment begins alternatively.

The subgroup of analyzing:

According to (2000) .Lancet 356 such as Gastl, 1981-2 has identified height, moderate and low EPCAM expresser.

Analyzed following subgroup:

● use the EpCAM of low dosage Anti-EpCAM treatment low/moderate expresser's (" low dosage Anti-EpCAM/ hangs down EpCAM ")

● use the EpCAM of high dose Anti-EpCAM treatment low/moderate expresser's (" high dose Anti-EpCAM/ hangs down EpCAM ")

● with the EpCAM high expressed person (" the high EpCAM of low dosage Anti-EpCAM/ ") of low dosage Anti-EpCAM treatment

● with the EpCAM high expressed person (" the high EpCAM of high dose Anti-EpCAM/ ") of high dose Anti-EpCAM treatment

● the EpCAM with low or high dose Anti-EpCAM treatment is low/moderate expresser (" low EpCAM ")

● with EpCAM high expressed person (" high EpCAM ") low or high dose Anti-EpCAM treatment

● EpCAM is low/low dosage Anti-EpCAM (" low dosage Anti-EpCAM ") among moderate or the EpCAM high expressed person

● EpCAM is low/high dose Anti-EpCAM (" high dose Anti-EpCAM ") among moderate or the EpCAM high expressed person

The analysis of carrying out:

Following analyzed above definition reply (CBR or OTR) terminal point:

● each subgroup (referring to above-mentioned) and all patients' CBR/OTR lead.

● be used for the Fei Xier that the CBR/OTR of following comparison leads (Fisher ' s) Precision Test:

● each subgroup (referring to above-mentioned) is with respect to all other patients of combination

● " low dosage Anti-EpCAM/ hangs down EpCAM " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

● " low dosage Anti-EpCAM/ hangs down EpCAM " is with respect to " high dose Anti-EpCAM/ hangs down EpCAM "

● " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

● " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " high dose Anti-EpCAM/ hangs down EpCAM "

To the timing definition of clinical disease development for date of Anti-EpCAM infusion first (perhaps in sensitive analysis: the randomized date) with the clinical disease development be the persistent period of date between separately of the first generation of PD.If do not observe the clinical disease development, with the corresponding time span of research date of expiry measuring and calculating (censor).In the disappearance situation of research date of expiry, with the date replacement of going to a doctor for the last time.

The following analysis to time (Time-To-Progression, the TTP) terminal point of development:

● each subgroup and all patients' intermediate value TTP (if can estimate).

● be used for the Log-Rank check of the TTP of following comparison:

Zero each subgroup is with respect to all other patients of combination

Zero " low dosage Anti-EpCAM/ hangs down EpCAM " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

Zero " low dosage Anti-EpCAM/ hangs down EpCAM " is with respect to " high dose Anti-EpCAM/ hangs down EpCAM "

Zero " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

Zero " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " high dose Anti-EpCAM/ hangs down EpCAM "

Embodiment 2.2: result-research patient

The data set of analyzing

Table 4 is made by the colony that is used for analyzing.

Fully analysis bank (FAS) (n=67) refers to be used to replying and to the analysis colony of the time assessment of development.In two Anti-EpCAM dosage groups, the EpCAM situation is equal distribution, and 38% low EpCAM expresser and 57% high EpCAM expresser have an appointment in each treatment group.

Table 4: the colony that is used for analysis ¹

¹Be included in the patient number (%) of administration data in checking

²Complete analysis bank (FAS): from the patient of safety analysis group, wherein these patients have the EpCAM positive (low/moderate or high expressed) tumor, and owing in the former thereby situation about withdrawing from ahead of time of other, have at least a tumor to assess except clinical disease development after the treatment beginning.

Embodiment 2.3: the efficiency analysis of result-whole tumor response (OTR) and clinical benifit rate (CBR)

According to research rules, the clinical benifit rate when the main terminal point of research was the 24th week.

" best overall tumor response (OTR) "

Provide in the following table 5 and 6 " best overall tumor response (OTR) rate during the 12nd week " among the high and low expresser of EpCAM and the whole FAS, " clinical benifit rate (CBR) during the 12nd week " and the result of when all " the 24th clinical benifit rate (CBR) ".

In the assessment of the radiology at center, can not confirm that any patient among the FAS is according to the relevant PR of RECIST standard or replying of CR.

" clinical benifit rate (CBR) " during the 12nd week

Generally, there are 16 (24%) when the 12nd week, to show stable disease (SD) among 67 patients among the FAS.Not seeing that there were significant differences aspect clinical benifit rate between the treatment group during the 12nd week, is 21.9% in the low dose group, is 25.7% in the high dose group.CBR is higher than in low EpCAM group in high EpCAM group, yet this difference is not remarkable statistically.

Can't detect only to comprise according to RECIST and have stable disease as the patient's of remission clinical benifit rate (referring to above-mentioned).

Table 5: the clinical benifit rate (CBR) in the 12nd when week (according to RECIST, have stable disease or Remission [CR] or partial symptoms are alleviated the patient of [PR] fully)-complete analysis bank

¹Can assess the patient's number that has clinical benifit in the middle of the patient of its clinical benifit according to RECIST

²Bilateral Fei Xier Precision Test

³N in low dose group/high dose group

" clinical benifit rate (CBR) " during the 24th week

Compare with low dose group (6.3%), in high dose group (14.3%), seen higher clinical benifit rate during the 24th week.In high dose group, CBR low and high EpCAM subgroup equates (14.3%).In low dose group, high and low EpCAM expresser's CBR is similar to (5.3 pairs 7.7%).

Only take patient with stable disease as the basis, it does not detect PR or CR to CBR.

Table 6: the clinical benifit rate (CBR) in the 24th when week (according to RECIST, have stable disease or Remission [CR] or partial symptoms are alleviated the patient of [PR] fully)

¹Can assess the patient's number that has clinical benifit in the middle of the patient of its clinical benifit according to RECIST

²Bilateral Fei Xier Precision Test

³N in low dose group/high dose group

Efficiency analysis: to the time of development

The timing definition of near clinical disease development for according to RECIST first the date of Anti-EpCAM infusion (perhaps in sensitive analysis: be persistent period between date of first generation of PD with the clinical disease development the randomized date).If do not observe the clinical disease development, obtain to the time span of research date of expiry.Can not obtain to study in the situation of date of expiry, with the date replacement of going to a doctor for the last time.

For hanging down and high dose group and EpCAM subgroup, table 7 (beginning to the time of clinical disease development from treatment) and table 8 (time of development from the randomization to the clinical disease) provide the FAS to the Median Time that develops.

Generally, compare with the Anti-EpCAM low dose therapy, the Anti-EpCAM high dose shows to the Median Time of development (being calculated as the time from infusion first) and has obviously prolonged 43 to 78 days (as shown in Figure 2).Check " survival " curve (p=0.0348; Log rank check) this difference is statistically significant (as shown in Figure 3) time.Similarly, express the patient when comparing when a low EpCAM being expressed patient and high EpCAM, the difference of observing Median Time to development (being calculated as the time from infusion first) (was respectively 42 to 80 days; P=0.0431; Log rank check).Expressed the highest Median Time observed among the patient to development (being calculated as the time from infusion first) (90 days at the high EpCAM with high dose Anti-EpCAM treatment; P=0.0238; Log rank check-compares with all other patients) (as shown in Figure 5).

Table 7: to the Median Time of development (from infusion begin to clinical disease development time [my god])-divide fully Analyse group

¹In the Median Time of sky to development

²Log Rank check for the difference between the treatment group

³N in low dose group/high dose group

The data of Fig. 4 graphic table 7.

For the time to development that begins to calculate from the randomized date, found comparable result.Compare with the Anti-EpCAM low dose therapy, whole Anti-EpCAM high dose has shown that again extremely the Median Time (being calculated as the time from randomization) of development obviously prolongs 46 to 79 days.When check during " survival " curve this difference be significant (p=0.0441 statistically; Log rank check).Treatment difference is also more remarkable in high EpCAM group, and for low dosage and the high EpCAM group of high dose, extremely the Median Time of development was respectively 63 and 91 days, was respectively 43 and 53 days in the low EpCAM group of low dosage and high dose.

Table 8: to the Median Time of development (from randomization play clinical disease development time [my god])-divide fully Analyse group

The effect conclusion

Based on the research rules, in whole colony and low and high EpCAM subgroup, high dose Anti-EpCAM group organized with low dosage Anti-EpCAM and compare, analyzed

-best overall is replied (OTR)

-clinical benifit rate (CBR); With

-to the time (TTP) of developing.

Can be when each time point determines to comprise all patients that show stable disease in the 12nd and 24 week the clinical benifit rate of (being respectively W12 and W24).

-in the 12nd when week high dose Anti-EpCAM group, in a little higher than low dose group of CBR (25.7% pair 21.9%), for high EpCAM subgroup, also simultaneously in two dosage groups, shown higher ratio.

-CBR higher (14.3%, be 6.3% in the low dosage Anti-EpCAM group) in the high dose Anti-EpCAM group when the 24th week.Fail between the EpCAM subgroup, to detect significant difference.

In whole sample, the Anti-EpCAM high dose is compared with the Anti-EpCAM low dosage, Median Time to development has shown obvious prolongation (43 to 78 days), and this difference is statistically significant (p=0.0348, log rank check) when the check survival curve.Expressed the Median Time observed among the patient to development (being calculated as the time from infusion first) (90 days at the high EpCAM with high dose Anti-EpCAM treatment; P=0.0238; Log rank check-compares with all other patients).

Whole conclusion

Data available has shown long-term the 24th week of stable disease (〉 at least 7 patients), some patients are still underway.

Time to development is assessed the significant prolongation that has shown " time-to-live ", supports that (in favor of) high dose Anti-EpCAM colony reaches statistical significance, can be clear that this point in Fig. 2-5.Particularly as shown in Figure 5, the high EpCAM that accepts high dose Anti-EpCAM expresses the patient and has obviously prolonged without development survival (90 days, with respect to 41-49 days in other group).

The final research report of embodiment 3:Anti-EpCAM II phase clinical research

Embodiment 3.1: research overview and introduction

As described in top embodiment 1, carry out clinical research.The result of now this research follows among the embodiment 3.

Methodology:

Randomization, open-label, multicenter, parallel group, II phase are studied.This research design is for assessment of effect and the safety of Anti-EpCAM in 24 weeks of two various dose, and it comprises positive EpCAM test.The EpCAM result of the test of carrying out during according to screening is with the randomization classification at center.In case be positioned in the EpCAM group, the patient just has been assigned to low dose therapy group or high-dose therapy group at random.

Patient's number of randomization and treatment: 112

Patient's number of analyzing: the patient of 112 treatments (56 Anti-EpCAM high doses, 56 Anti-EpCAM low dosages), wherein 109 patients are EpCAM+ after testing.

The analytical data that is used for aforementioned analysis:

According to the requirement of GCP monitoring and cleared up all data that cause these results, and before carrying out final analysis lock database.

Be used for the definition of the colony of analysis:

● safety analysis group (SAF): all have accepted the patient of the appointment drugs of at least one dosage.

● complete analysis bank (FAS): from the patient of safety analysis group, wherein these patients have the EpCAM positive (low/moderate or high expressed) tumor, and owing in the former thereby situation about withdrawing from ahead of time of other, have at least a tumor to assess except clinical disease development after the treatment beginning.

The analysis of data of safety is take SAF as the basis.

The analysis of base-line data and effect terminal point is take complete analysis bank as the basis.

The terminal point (with reference to above-described embodiment 1.1) of analyzing:

● the clinical benifit rate (CBR) during the 24th week (according to RECIST, having the patient that stable disease [SD] or complete remission [CR] or partial symptoms are alleviated [PR])

● best overall tumor response (OTR) rate during the 12nd week (according to RECIST, having the patient that complete remission [CR] or partial symptoms are alleviated [PR])

● the clinical benifit rate (CBR) during the 12nd week (according to RECIST, having the patient that stable disease [SD] or complete remission [CR] or partial symptoms are alleviated [PR])

● to time (TTP) of development for from the time the randomization and time from treatment begins alternatively.

The subgroup of analyzing:

According to (2000) .Lancet 356 such as Gastl, 1981-2 has identified high and low/moderate EPCAM expresser.

Analyzed following subgroup:

● use the EpCAM of low dosage Anti-EpCAM treatment low/moderate expresser's (" low dosage Anti-EpCAM/ hangs down EpCAM ")

● use the EpCAM of high dose Anti-EpCAM treatment low/moderate expresser's (" high dose Anti-EpCAM/ hangs down EpCAM ")

● with the EpCAM high expressed person (" the high EpCAM of low dosage Anti-EpCAM/ ") of low dosage Anti-EpCAM treatment

● with the EpCAM high expressed person (" the high EpCAM of high dose Anti-EpCAM/ ") of high dose Anti-EpCAM treatment

● the EpCAM with low or high dose Anti-EpCAM treatment is low/moderate expresser (" low EpCAM ")

● with EpCAM high expressed person (" high EpCAM ") low or high dose Anti-EpCAM treatment

● EpCAM is low/low dosage Anti-EpCAM (" low dosage Anti-EpCAM ") among moderate or the EpCAM high expressed person

● EpCAM is low/high dose Anti-EpCAM (" high dose Anti-EpCAM ") among moderate or the EpCAM high expressed person

The analysis of carrying out:

Following analyzed above definition reply (CBR or OTR) terminal point:

● each subgroup (referring to above-mentioned) and all patients' CBR/OTR lead.

The time of near clinical disease development is defined as respectively date of Anti-EpCAM infusion first (perhaps in sensitive analysis: be persistent period between date of first generation of PD with the clinical disease development the randomized date).If do not observe the clinical disease development, calculate corresponding time span with studying the date of expiry.In the disappearance situation of research date of expiry, with the date replacement of going to a doctor for the last time.

The following analysis to time (Time-To-Progression, the TTP) terminal point of development:

● each subgroup and all patients' intermediate value TTP (if can estimate).

● be used for the Log-Rank check of TTP of the treatment group difference of following comparison:

Zero each subgroup is with respect to all other patients of combination

Zero " low dosage Anti-EpCAM/ low/moderate EpCAM " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

Zero " low dosage Anti-EpCAM/ low/moderate EpCAM " is with respect to " high dose Anti-EpCAM/ low/moderate EpCAM "

Zero " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " the high EpCAM of low dosage Anti-EpCAM/ "

Zero " the high EpCAM of high dose Anti-EpCAM/ " is with respect to " high dose Anti-EpCAM/ low/moderate EpCAM "

Zero " low dosage Anti-EpCAM/ low/moderate EpCAM " is with respect to " the high EpCAM of high dose Anti-EpCAM/ "

Zero " low dosage Anti-EpCAM " is with respect to " high dose Anti-EpCAM "

Zero " high EpCAM " is with respect to " low/moderate EpCAM "

Embodiment 3.2: result-research patient

The data set of analyzing

Table 9 is made by the colony that is used for analyzing.

Fully analysis bank (FAS) (n=109) refers to be used to replying and to the analysis colony of the time assessment of development.

Table 9: the colony 1 that is used for analysis

¹Patient's number, average ± standard deviation (minima-maximum, intermediate value)

²Patient's number, %

Embodiment 3.3: the efficiency analysis of result-whole tumor response (OTR) and clinical benifit rate (CBR)

According to research rules, the clinical benifit rate when the main terminal point of research was the 24th week.

" best overall tumor response (OTR) "

The result of " best overall tumor response (OTR) rate during the 12nd week " among the high and low expresser of EpCAM and the whole FAS, " the clinical benifit rate (CBR) during the 12nd week " and when all " the 24th clinical benifit rate (CBR) " is provided in the following table 10 and 11.

Diagnosed two kinds by this locality radiation assessment and replied (according to the RECIST standard, about PR or CR), but in any patient of FAS, can not assess to confirm with the radiology at center.

" clinical benifit rate (CBR) " during the 24th week

Compare with low dose group (4.5%), the 24th when week in high dose group (7.9%) though in seen not obvious trend towards higher clinical benifit rate (CBR).Similarly, compare with low/moderate EpCAM expresser, high EpCAM expresser's CBR has shown the trend that leads towards higher CBR (7.3% with respect to 3.7%).

Only take patient with stable disease as the basis, it does not detect PR or CR to CBR.

Table 11: the clinical benifit rate (CBR) in the 24th when week (according to RECIST, have stable disease or Remission [CR] or partial symptoms are alleviated the patient of [PR] fully)

¹Patient's number (%)

²According to ' final ' assessment

³Still be in the patient in the research, its separately reply the center of being evaluated at /be classified as ' can not assess ' in the IRAB assessment

" clinical benifit rate (CBR) " during the 12nd week

Generally, there are 17 (16%) when the 12nd week, to show stable disease (SD) among 109 of FAS patients.

Compare with low dose group (14.5%), the 12nd when week in high dose group (16.7%) though seen not obvious trend towards higher clinical benifit rate (CBR).Similarly, compare with low/moderate EpCAM expresser, high EpCAM expresser's CBR has shown the trend that leads (18.9% with respect to 8.6%) towards higher CBR.

Table 10: the clinical benifit rate (CBR) during the 12nd week (according to RECIST, has stable disease or complete Full remission [CR] or partial symptoms are alleviated the patient of [PR])-complete analysis bank

¹Patient's number (%)

²According to ' final ' assessment

³Still be in the patient in the research, its separately reply the center of being evaluated at /be classified as ' can not assess ' in the IRAB assessment

Efficiency analysis: to the time of development

The timing definition of near clinical disease development be according to RECIST, and the date of Anti-EpCAM infusion is (perhaps in sensitive analysis: be persistent period between date of first generation of PD with clinical disease development the randomized date) first.If do not observe the clinical disease development, obtain to the time span of research date of expiry.Can not obtain to study in the situation of date of expiry, with the date replacement of going to a doctor for the last time.

Kapp orchid-Meyer (KM-) tracing analysis to the time of developing is provided in (time of development from the begin treatment to the clinical disease) among Fig. 6-8.

Compare with the Anti-EpCAM low dose therapy, whole Anti-EpCAM high dose shown along with passage of time to the development time obvious prolongation (as shown in Figure 6; Dangerous ratio (HR)=0.666).When check during " survival " curve this difference be significant (p=0.0465 statistically; Log rank check).Similarly, express the patient when comparing when low/moderate EpCAM being expressed patient and high EpCAM, observed trend (Fig. 7 that difference appears in time to development (being calculated as the time from infusion first); HR=0.706; P=0.1157; Log rank check).The excessive risk of expressing time of observing among the patient to development (being calculated as the time from infusion first) at the high EpCAM with high dose Anti-EpCAM treatment reduces (HR=0.433; P=0.0057; Log rank check-express the patient with the low EpCAM that treats with low dosage Anti-EpCAM to compare) (as shown in Figure 8).

The effect conclusion

Based on the research rules, in whole colony and low and high EpCAM subgroup, high dose Anti-EpCAM group organized with low dosage Anti-EpCAM and compare, analyzed

-best overall is replied (OTR)

-clinical benifit rate (CBR); With

-to the time (TTP) of developing.

Can be when each time point determines to comprise all patients that show stable disease in the 12nd and 24 week the clinical benifit rate of (being respectively W12 and W24).

-at the 12nd when week CBR (16.7% with respect to 14.5%) in high dose Anti-EpCAM group in a little higher than low dose group, for high EpCAM subgroup, also in two dosage groups, shown higher ratio (18.9% with respect to 8.6% among low/moderate EpCAM expresser).

-CBR higher (7.9% with respect to 4.5% in the low dosage Anti-EpCAM group) in the high dose Anti-EpCAM group when the 24th week, for high EpCAM subgroup, also in two dosage groups, shown higher ratio (7.3% with respect to 3.7% among low/moderate EpCAM expresser).

The Anti-EpCAM high dose is compared with the Anti-EpCAM low dosage, the time series analysis to development in whole sample demonstrates prolongation (R=0.666), this difference is statistically significant (p=0.0465, log rank check) when the check survival curve.Express time of observing among the patient to development the maximum extension (HR=0.433 of (being calculated as the time from infusion first) at the high EpCAM with high dose Anti-EpCAM treatment; P=0.0057; Log rank check-express the patient with the low EpCAM that treats with low dosage Anti-EpCAM to compare).

Whole conclusion

Radioactivity according to the center is checked, and at least 6 patients, data available has shown long-term the 24th week of stable disease (〉).

Time to development is assessed the significant prolongation that has shown " time-to-live ", supports high dose Anti-EpCAM colony to reach statistical significance, can be clear that this point in Fig. 6-8.Particularly as shown in Figure 8, the high EpCAM that accepts high dose Anti-EpCAM expresses the patient and has obviously prolonged without the development survival.

Claims (11)

1. contain SEQ ID NO.3,4,5,6,7 and 8 anti-EpCAM antibody for the preparation of the purposes in the medicine for the treatment of metastatic breast cancer, wherein described medicament administration is expressed and shown for the immunostaining total points of EpCAM greater than 4 experimenter in having high EpCAM, wherein said medicine is used according to low dosage application program or high dose application program, described low dosage application program comprises the load doses of at least one 1 to 3mg/kg body weight, be a plurality of maintenance dosies afterwards, each maintenance dose is 1 to 3mg/kg body weight; Described high dose application program comprises the load doses of at least one 4.5 to 8mg/kg body weight, is a plurality of maintenance dosies afterwards, and each maintenance dose is 4.5 to 8mg/kg body weight.

2. the purposes of claim 1, wherein said anti-EpCAM antibody contains SEQ ID NO.1 and/or 2.

3. claim 1 or 2 purposes, wherein said treatment comprises the long term stabilization of metastatic breast cancer.

4. each purposes according to claim 1-3, wherein metastatic breast cancer is carried down according to tumor lympha and is moved (" TNM ") system and classify as the IV phase.

5. the purposes of aforementioned arbitrary claim, wherein said medicine is used as follows: the load doses of at least one 2mg/kg body weight is a plurality of maintenance dosies afterwards, and each maintenance dose is the 2mg/kg body weight.

6. each purposes of aforementioned claim 1-4, wherein said medicine is used as follows: the load doses of at least one 6mg/kg body weight is a plurality of maintenance dosies afterwards, and each maintenance dose is the 6mg/kg body weight.

7. claim 5 or 6 purposes are wherein used weekly described/each load doses, and per two weeks are used each described maintenance dose.

8. the purposes of claim 7, wherein when beginning, the 1st, 2 and 3 all each for the treatment of use a load doses, be 11 maintenance dosies afterwards, when the 4th, 6,8,10,12,14,16,18,20,22 and 24 all each for the treatment of begins, use a maintenance dose.

9. the purposes of aforementioned arbitrary claim is wherein used described antibody in the solution that contains 0.9% sodium chloride solution.

10. the purposes of aforementioned arbitrary claim, wherein intravenous is used described antibody.

11. contain the purposes in the medicine of SEQ ID NO.3,4,5,6,7 and 8 anti-EpCAM antibody metastatic human breast carcinoma in for the preparation of the treatment experimenter, wherein said experimenter presents high EpCAM and expresses the immunostaining total points that shows for EpCAM greater than 4, wherein said medicine is used as follows: the load doses of at least one 6mg/kg body weight, be a plurality of maintenance dosies afterwards, each maintenance dose is the 6mg/kg body weight; Wherein use weekly each load doses, and per two weeks are used each maintenance dose; Wherein using a load doses when the 1st, 2 and 3 all each for the treatment of begins, is 11 maintenance dosies afterwards, uses a maintenance dose when the 4th, 6,8,10,12,14,16,18,20,22 and 24 all each for the treatment of begins.

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