Hepatotoxicity in Cancer Patients Receiving erb-38, a
`Recombinant Immunotoxin That Targets the erbB2 Receptor
` 
`Lee H. Pai-Scherf, Jacy Villa, Deborah Pearson, et al.
`Clin Cancer Res  
`
`1999;5:2311-2315.
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`on October 30, 2014. © 1999 American Association for Canceron October 30, 2014. © 1999 American Association for Cancer
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`Research. Research.
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`IMMUNOGEN 2029, pg. 1
`Phigenix v. ImmunoGen
`IPR2014-00676
`
`

`

`Vol. 5, 2311–2315, September 1999
`
`Clinical Cancer Research
`
`2311
`
`Advances in Brief
`
`Hepatotoxicity in Cancer Patients Receiving erb-38, a Recombinant
`Immunotoxin That Targets the erbB2 Receptor
`
`Lee H. Pai-Scherf, Jacy Villa, Deborah Pearson,
`Thelma Watson, Edison Liu,
`Mark C. Willingham, and Ira Pastan1
`Laboratory of Molecular Biology, Division of Basic Sciences
`[L. H. P-S., I. P.], and Laboratory of Molecular Signaling and
`Oncogenesis [J. V., E. L.] and Medicine Branch [D. P., T. W.],
`Division of Clinical Sciences National Cancer Institute, NIH,
`Bethesda, Maryland 20892; and Department of Pathology, Wake
`Forest University, Winston-Salem, North Carolina 27157 [M. C. W.]
`
`Abstract
`To exploit overexpression of erbB2 in human cancers,
`we constructed a single-chain immunotoxin (erb-38) that
`contains the Fv portion of monoclonal antibody e23 fused to
`a truncated form of Pseudomonas exotoxin A. In a Phase I
`study, five breast cancer patients and one esophageal cancer
`patient received three doses of erb-38 at 1.0 and 2.0 mg/kg.
`Hepatotoxicity was observed in all patients. Immunohisto-
`chemistry showed the presence of erbB2 on hepatocytes
`explaining the liver toxicity of erb-38. We suggest that tar-
`geting of tumors with antibodies to erbB2 armed with ra-
`dioisotopes or other toxic agents may result in unexpected
`organ toxicities due to erbB2 on normal cells.
`
`Introduction
`erbB2/HER2/neu encodes an Mr 185,000 cell membrane
`glycoprotein with tyrosine kinase activity. Overexpression of
`the erbB2 protein and its inherent tyrosine kinase activity causes
`loss of growth control and has an important role in the devel-
`opment of breast cancer and several other human cancers (1–3).
`erbB2 has been reported previously to be minimally expressed
`in normal adult tissues (4). For this reason, erbB2 is an attractive
`target for antibody-directed therapies. To exploit the overex-
`pression of erbB2 in many human cancers (breast, stomach,
`lung, and ovary), we made a single-chain immunotoxin erb-38
`using MAb2 e23 (5), which reacts with erbB2.
`erb-38 is a Mr 63,000 recombinant protein composed of the
`Fv portion of MAb e23 fused to PE38, a truncated form of PE
`with a molecular weight of 38,000. MAb e23 was selected from
`
`Received 2/18/99; revised 5/14/99; accepted 5/26/99.
`The costs of publication of this article were defrayed in part by the
`payment of page charges. This article must therefore be hereby marked
`advertisement in accordance with 18 U.S.C. Section 1734 solely to
`indicate this fact.
`1 To whom requests for reprints should be addressed, at Laboratory of
`Molecular Biology, National Cancer Institute, NIH, 37 Convent Drive,
`MSC 4255, Bethesda, MD 20892-4255. Phone: (301) 496-4797; Fax:
`(301) 402-1344; E-mail: pasta@helix.nih.gov.
`2 The abbreviations used are: MAb, monoclonal antibody; PE, Pseudo-
`monas exotoxin A; AST, aspartate aminotransferase; ALT, alanine
`aminotransferase; NCI, National Cancer Institute; MTD, maximum tol-
`erated dose.
`
`a panel of antibodies to erbB2 because it produced very active
`conventional immunotoxins when it was chemically linked to
`PE (6). PE is a Mr 66,000 protein secreted by Pseudomonas
`aeruginosa. PE is composed of three major structural domains
`(7): an NH2-terminal cell binding domain (domain Ia), a central
`translocation domain (domain II), and a COOH-terminal cata-
`lytically active domain (domain III). Domain III catalyzes the
`ADP ribosylation and inactivation of elongation factor 2, which
`inhibits protein synthesis and leads to cell death. Deletion of
`domain Ia of PE (amino acids 1–252) and part of domain Ib
`produces a Mr 38,000 protein (PE38) that has low cellular and
`animal cytotoxicity because it cannot bind to PE-specific cellu-
`lar receptors. The gene encoding PE38 was fused to a gene
`encoding the Fv portion of MAb e23 to form erb-38
`[e23(dsFv)PE38]. In this recombinant immunotoxin,
`the Fv
`fragment contains a disulfide bond connecting the light and
`heavy chains of MAb e23 (8). The IC50 of erb-38 is 0.2– 4 ng/ml
`on the various tumor cells lines that express the erbB2 antigen.
`erb-38 is capable of causing complete remission in nude mice
`bearing epidermoid carcinoma (A431) and breast cancer
`(MCF-7; Ref. 9). Preclinical toxicity experiments indicated that
`the LD50 of erb-38 in mice was 450 mg/kg every other day times
`three doses. Death was caused by acute hepatic necrosis. He-
`patic and renal injury was observed in cynomolgus monkeys at
`three doses of $500 mg/kg.3
`On the basis of the biological activity and safety testing
`results, we concluded that erb-38 had excellent antitumor activ-
`ity and acceptable animal toxicities. We initiated a Phase I study
`of erb-38 in patients with advanced carcinoma who had failed
`standard therapy.
`
`Patients and Methods
`Eligibility. Adult patients with metastatic carcinoma
`known to express erbB2 were eligible for this study. Tumors were
`considered positive if they expressed erbB2 on $30% of the tumor
`cells, as determined by immunohistochemistry on formaldehyde-
`fixed, paraffin-embedded tumor blocks. Tumor histology was con-
`firmed by a NIH pathologist. Other eligibility criteria included:
`advanced unresectable disease, failed conventional therapy, Eastern
`Cooperative Oncology Group performance status of 0 or 1, AST
`and ALT levels of ,1.5 times the upper limits of normal, total
`bilirubin levels within normal limits, absolute granulocyte counts of
`.2,000/mm3, and platelet counts of .100,000/mm3. Patients with
`positive hepatitis B antigen, history of any other prior liver disease
`(e.g., alcohol liver disease), central nervous system metastasis, or
`seizure disorders were excluded. Patients with positive antibodies
`to erb-38 were excluded. The clinical protocol and the consent form
`were approved by the Institutional Review Board of the NCI. Prior
`to therapy, patients underwent complete physical examination,
`
`3 Unpublished data.
`
`
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`on October 30, 2014. © 1999 American Association for Cancer
`Research.
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`IMMUNOGEN 2029, pg. 2
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`IPR2014-00676
`
`

`

`2312 Phase I Study of Anti-erbB2 Immunotoxin
`
`Table 1 Patient characteristics
`
`Diagnosis
`Age/sex
`Breast cancer
`49/F
`Breast cancer
`36/F
`58/M Esophageal cancer
`35/F
`Breast cancer
`57/F
`Breast cancer
`
`erbB2
`expression
`(%)
`80
`80
`80
`80
`100
`
`Patient
`no.
`1
`2
`3
`4
`5
`
`6
`
`Sites of metastasis
`Prior therapy
`XRTa, Doxo, cyclophosphamide, paclitaxel
`Lung, pleura, bone, skin, LN
`Pleura, pericardium, skin, LN
`Surgery, XRT, CMF, Doxo, 5-FU, taxotere, navalbine
`Liver, lung, LN
`XRT, cisplatin, paclitaxel
`Surgery, CAF, Taxol, Ctx, etoposide/melphalan (PBSC) Lung, skin, LN
`Surgery, XRT, CAF, TAM, arimidex, magace,
`LN
`cyclophosphamide/novantrone/5-FU
`Cyclophosphamide/thiotepa/carboplatin (PBSC)
`Surgery, XRT, CAF, paclitaxel,
`Etoposide/melphalan (PBSC)
`a XRT, radiation therapy; Doxo, doxorubicin; CMF, cyclophosphamide, methotrexate and 5-FU; 5-FU, 5-fluorouracil; CAF, cyclophosphamide,
`Doxo, and 5-FU; Ctx, cyclophosphamide; PBSC, peripheral blood stem cell support; TAM, tamoxifen; LN, lymph node.
`
`40/F
`
`Breast cancer
`
`100
`
`Lung, bone, skin
`
`Table 2 Dosing and drug-related toxicity
`
`Patient
`no.
`1
`2
`3
`4
`5
`6
`
`Dose
`(mg/kg)
`2.0
`2.0
`2.0
`2.0
`1.0
`1.0
`
`AST/ALT
`2
`3
`2
`3
`3
`3
`
`Toxicity grade
`
`Nausea/vomiting
`
`Skin rash
`
`2
`
`1
`
`3
`
`complete blood count, determination of chemistry profile, urinaly-
`sis, chest X-ray, electrocardiogram, tumor staging work-up, and
`tumor measurement. Informed consent was obtained from all pa-
`tients.
`erb-38.
`erb-38 (NSC 683039) was produced and placed
`in vials by NCI-Frederick Cancer Research & Development
`Center. erb-38 was supplied through the Cancer Therapy Eval-
`uation Program, NCI, under Investigational New Drug No.
`7373. erb-38 in phosphate buffer solution was supplied as a
`sterile solution at 0.5 mg/ml in 2-ml vials containing 1.0 mg of
`erb-38. Test and treatment doses were diluted in 0.9% normal
`saline and 0.2% albumin.
`Trial Design.
`erb-38 was administered by i.v. infusion
`over 30 min on days 1, 3, and 5. All patients received a 10-mg test
`dose on day 1 given as a bolus over 2 min. The second and third
`doses were delayed or withheld if any measure of toxicity was not
`less than grade II on the scheduled day of administration. A cycle
`could be repeated every 28 days, provided that the patient had not
`developed neutralizing antibodies against erb-38 and had no dis-
`ease progression. Patients were to be entered in groups of three.
`Dose escalation was to based on the modified Fibonacci series.
`While on study, patients were followed with complete
`blood count, blood chemistry and urine. Tumor imaging was
`repeated 1 month after therapy. Toxicity and response to treat-
`ment were graded by the NCI Common Toxicity Criteria. Lim-
`iting toxicity was defined as any grade III or grade IV toxicity.
`Pharmacokinetics. Blood samples were collected at the
`following times for pharmacokinetics: pretreatment; 2, 15, 30,
`45, 60, and 90 min; and 2, 4, 8, 12, 24, and 48 h. After the
`second and third doses of erb-38, samples were collected at 2
`min, 30 min, and 4 h. The concentration of erb-38 was deter-
`
`mined by incubating various dilutions of serum with N87 cells,
`a gastric cancer cell line known to express erbB2, and measuring
`its ability to inhibit protein synthesis. A standard curve was used
`to determine the amount of erb-38 in each sample. Data were
`weighted inversely, and the fit program used was RSTRIP
`(MicroMath Scientific Inc., Salt Lake City, UT).
`Immunogenicity of erb-38. Antibodies against erb-38
`were assessed pretreatment, on day 21, and then bimonthly
`thereafter by a serum neutralization assay: erb-38 was added to
`serum samples at 0.0 (control), 0.1, 0.5, and 1.0 mg/ml and
`incubated at 37°C for 15 min. The activity of erb-38 was
`assayed by incubating the samples with MCF-7 cells and meas-
`uring its ability to inhibit protein synthesis. A standard curve
`with erb-38 was used to determine IC50. A serum sample was
`considered negative for antibodies against erb-38 when the
`cytotoxic activity of erb-38 was not neutralized when incubated
`with sera at concentrations of .0.1 mg/ml erb-38.
`Immunohistochemistry of Normal Liver. Fresh normal
`human liver samples were frozen by immersion in liquid nitro-
`gen. Cryostat sections (;6 mm) were thaw-mounted on slides
`and fixed in 3.7% formaldehyde in PBS for 10 min at 23°C,
`followed by washing in PBS. Sections were not treated with
`H2O2 or any organic solvents. After blocking in 1% BSA in PBS
`for 10 min, sections were incubated in mouse monoclonal anti-
`erbB2 (CB11; Novocastra Laboratories, Novocastra antibody,
`Vector Laboratories, Burlingame, CA) at 10 mg/ml in BSA-PBS
`for 30 min at 23°C. Alternatively, sections were incubated in
`affinity-purified rabbit anti-erbB2 (code A 0485; Dako Corpo-
`ration, Carpinteria, CA) at 7 mg/ml in BSA-PBS. After further
`washing in PBS, sections were incubated, as appropriate, with
`either goat antimouse IgG or goat antirabbit IgG conjugated
`with horseradish peroxidase (Jackson ImmunoResearch) at 25
`mg/ml in BSA-PBS for 30 min. Peroxidase was detected using
`diaminobenzidine (1 ng/ml) with 0.01% H2O2 in PBS for 10
`min. Controls included deletion of the first antibody step. Sec-
`tions were mounted using dehydration and Permount without
`further counterstains and viewed using bright-field microscopy.
`
`Results
`Six patients were enrolled in this trial. Patient characteris-
`tics are shown in Table 1. The initial dose level was 2 mg/kg
`times three, which was 1/250 of the MTD in monkeys and 1/225
`
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`on October 30, 2014. © 1999 American Association for Cancer
`Research.
`
`IMMUNOGEN 2029, pg. 3
`Phigenix v. ImmunoGen
`IPR2014-00676
`
`

`

`Clinical Cancer Research
`
`2313
`
`Fig. 1 Time course of AST
`(E) and ALT (F) of patient 2,
`who received 2.0 mg/kg (A) and
`patient 5, who received 1.0
`mg/kg erb-38 (B).
`
`of the LD50 in mice. This conservative starting dose was chosen
`due to the fact that the toxicity of immunotoxins in humans has
`thus far been found to be unpredictable. Furthermore, 2.0 mg/kg
`times three has been shown to be safe with two other recombi-
`nant immunotoxins made with PE (LMB-7 and LMB-2).3
`Toxicity. Transient elevation of AST and ALT was ob-
`served in all patients. At 2.0 mg/kg, two patients had grade II
`and two patients had grade III elevations of AST and ALT. The
`toxicity observed in the first cohort prompted us to amend the
`protocol to decrease the dose by 50%. Two patients were treated
`at the 1.0 mg/kg dose level. Both patients had a grade 3 trans-
`aminase elevation after one cycle of erb-38 (Table 2).
`Increases in AST and ALT usually occurred within 24 h
`after the first or second dose of erb-38, and peak levels were
`reached 24 – 48 h after the last dose. Two patients required a
`50% dose reduction for the third dose (patients 2 and 6), and in
`two patients, the third dose was held (patients 4 and 5). Hepatic
`transaminases returned to baseline levels 14 –21 days following
`therapy. Fig. 1 shows the time course of AST and ALT of
`patient 2, who received 2.0 mg/kg erb-38, and patient 5, who
`received 1.0 mg/kg erb-38. There was no evidence of liver
`enlargement or right upper quadrant tenderness on exam. Two
`patients had symptoms of nausea and vomiting during therapy,
`with unclear relation to the hepatic injury. No significant drug-
`related changes in bilirubin, alkaline phosphatase, serum albu-
`min, or prothrombin time were observed. Other erb-38-related
`toxicities include a skin rash in one patient 4 days after erb-38
`and nausea and vomiting in two patients.
`As expected in a group of patients with advanced carci-
`noma, several untoward events occurred while the patients were
`on study: pain (n 5 3), malignant pleural effusion (n 5 2),
`malignant pericardial effusion (n 5 1), catheter infection (n 5
`2), venous thrombosis (n 5 1), and proteinuria (n 5 1). One
`patient (patient 6) died of acute respiratory failure in her home-
`town 10 days after discharge, due to pulmonary emboli. An
`autopsy was not performed. These events were attributed to the
`patient’s underlying disease and not related to erb-38.
`Localization of erbB2 in Normal Liver. Although prior
`literature had suggested that erbB2 was not expressed in normal
`liver (4), we used a sensitive immunohistochemical method with
`antibodies to erbB2 that were not available until recently. Rap-
`
`idly frozen normal human liver samples were sectioned with a
`cryostat and postfixed using formaldehyde but were not exposed
`to H2O2 or any organic solvents. Mouse monoclonal CB11 and
`rabbit polyclonal A0485 were used for the detection of erbB2
`using affinity-purified antiglobulins labeled with horseradish
`peroxidase. When viewed without counterstains, the localization
`of erbB2 to the sinusoidal surface of normal hepatocytes was
`clearly visible using either primary antibody (CB11 and AD485;
`Fig. 2). The same result was observed from samples of normal
`liver derived from at least three different patients. Similar results
`were also obtained with antibody E23 (data not shown).
`Pharmacokinetics. Blood samples were collected for
`pharmacokinetics in all patients. erb-38 is cleared monoexpo-
`nentially from the circulation, with a T1/2 that varies from 2.4 to
`10.3 (mean, 3.6 h). A high degree of variation was observed
`from patient to patient. The peak concentration achieved at the
`end of infusion ranged from ,20 ng/ml (patient 1) to 105 ng/ml
`(patient 3). At 1.0 mg/kg, the peak plasma levels were 38.1 and
`39.4 ng/ml. The volumes of distribution of central compartment
`and area under the concentration versus time curve were found
`to be 28.0 6 8.6 ml/kg and 248.7 6 139.6 ng/mlzh, respectively.
`Although full pharmacokinetic analysis was not possible due to
`limited sampling after second and third doses, clearance rates
`appeared to be similar to that after the first dose.
`Immunogenicity. Patients had no neutralizing antibodies
`against erb-38 prior to therapy, as required by protocol. At the
`end of one cycle, only one patient (patient 3) developed neu-
`tralizing antibodies (.1.0 mg/ml) against erb-38 14 days after
`treatment. Retreatment was not possible in this patient, due to
`disease progression.
`Response. No objective responses were observed in the
`trial. One patient had stable disease for 3 months, four patients
`progressed after one cycle of therapy, and one patient was not
`evaluable for response. Patient 2 had improvement of chest wall
`pain after erb-38 that lasted for 3 weeks. Fentanyl transdermal
`patch requirement decreased from 225 to 25 mg/h in this patient.
`
`Discussion
`We have shown that, when a recombinant immunotoxin,
`erb-38, which targets erbB2, is given to cancer patients, it causes
`hepatic injury, manifested by transient elevation of transami-
`
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`on October 30, 2014. © 1999 American Association for Cancerclincancerres.aacrjournals.org
`
`
`Research.
`
`IMMUNOGEN 2029, pg. 4
`Phigenix v. ImmunoGen
`IPR2014-00676
`
`

`

`2314 Phase I Study of Anti-erbB2 Immunotoxin
`
`Fig. 2 Localization of erbB2
`to the surface of normal human
`hepatocytes. Frozen sections of
`normal human liver were incu-
`bated with either a mouse MAb
`to erbB2 (CB11; B) or a rabbit
`polyclonal affinity-purified an-
`ti-erbB2 (A0485; D). Corre-
`sponding controls with deletion
`of the first step but with the
`second step included are shown
`for antimouse IgG (A) or anti-
`rabbit IgG (C). The characteris-
`tic pattern, especially at inter-
`cellular
`junctions,
`of
`the
`sinusoidal surface of hepato-
`cytes is evident (arrows) with
`both antibodies
`(B and D).
`These sections were not coun-
`terstained, and the nuclear lo-
`calization seen in D represents a
`nonspecific pattern seen only
`with the rabbit anti-erbB2 anti-
`(Magnification, 3375;
`body.
`scale bar, 50 mm).
`
`nases. The dose-limiting toxicities of erb-38 occurred at 1.0 and
`2.0 mg/kg when it was given i.v. every other day times three.
`These doses are significantly below the doses necessary to show
`antitumor effects in animals. Although the MTD was not deter-
`mined, based on our preclinical experience with this agent,
`effective serum levels cannot be achieved at doses ,1.0 mg/kg
`(6). In addition, we found that the erb-38 was cleared monoex-
`ponentially from the circulation, with a T1/2 that varied from 2.4
`to 10.3 h. As compared with other biological agents of this
`nature, a great deal of variation was observed from patient to
`patient. At 2.0 mg/kg, the maximum concentration varied from
`,20 to 105 ng/ml. No clear correlation can be made between the
`pharmacokinetic findings and the tumor burden, toxicity, or
`expression of erb-38 on tumor cells (all patients had tumors that
`expressed erbB2 on $80% of the cells).
`The toxicity of erb-38 is most likely due to the presence of
`erbB2 on hepatocytes, not detected by immunohistochemical
`staining in earlier publications (4). We have now repeated these
`assays using newly available antibody to erbB2 (affinity-puri-
`fied rabbit anti-erbB2, code A 0485; Dako Corp.) and have
`shown here that erbB2 is clearly present on the surface of
`normal hepatocytes. The expression of erbB2 is known to be
`very high in ;30% of breast cancers, often due to gene ampli-
`
`fication. Despite the fact that there is a very large difference in
`the amount of erbB2 on the surface of cancer cells relative to the
`small amount present on liver cells, liver toxicity was the first
`biological effect seen in this study. Several factors contribute to
`this finding. One is that hepatocytes are more rapidly exposed to
`agents injected into the circulation than tumor cells. There is a
`barrier to proteins entering tumors because tumors do not have
`lymphatics and there is no connective flow. As a consequence,
`mixing within tumors is solely by diffusion and, therefore, very
`slow. In addition, tumors are often poorly vascularized (10).
`There are several ways in which antibodies are being used
`as antitumor agents. Antibodies have been used by themselves
`to produce antitumor activity, taking advantage of their ability to
`carry out complement-mediated cell killing or ADCC (antibody
`dependent cell- and complement-mediated cytotoxicity) or to
`induce apoptosis directly. One or more of these effects explains
`how Rituximab, which binds to CD20, causes regression of
`lymphomas (11). Recently, clinical data concerning the safety
`and efficacy of a humanized antibody to erbB2 termed Hercep-
`tin were presented (12, 13). The antibody alone has been found
`to produce objective responses in breast cancer and when com-
`bined with chemotherapy results in an increased response rate.
`It is likely that the antitumor activity of the antibody in this
`
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`IMMUNOGEN 2029, pg. 5
`Phigenix v. ImmunoGen
`IPR2014-00676
`
`

`

`Clinical Cancer Research
`
`2315
`
`setting is dependent on genetic perturbations that alter the con-
`figuration of downstream signaling events, and, thereby, the
`responsiveness of the cancer cells to Herceptin. Thus, the mech-
`anism of killing depends on a genetic abnormality present in the
`cancer cells. In contrast, if the antibody is used to deliver a
`cytotoxic agent, such as a bacterial toxin or radioisotope, the
`death of the target cell will be principally dependent on the
`amount of agent delivered to the cell. In the case of erb-38, it
`appears that liver cells are the initial site of drug action. It has
`also been found in the Herceptin trial that there is evidence of
`cardiotoxicity, perhaps due to the presence of small amounts of
`erbB2 on myocytes or endocardial cells. Unfortunately, the
`information concerning the cardiotoxicity of Herceptin was not
`yet reported at the time of this trial. We did not include a
`throughout cardiac evaluation with echocardiogram or MUGA
`(Multigated Radionuclide Angiography) scan in the initial
`work-up. However, none of the patients had clinical evidence of
`myocardial dysfunction or arrhythmia before or after therapy
`with erb-38 based on their history, physical exam, chest X-ray,
`and electrocardiogram. It is quite possible that, because the
`concentrations of erb-38 given were low, overt cardiotoxicity
`was detected in this small trial. This might have been observed
`if higher doses of erb-38 had been administered.
`The new findings of reactivity with anti-erbB2 antibodies
`on hepatocytes are, for the most part, the result of the availabil-
`ity of the affinity-purified rabbit anti-erbB2. With this antibody,
`which has been shown to be 2– 4-fold more sensitive in detect-
`ing erbB2 in paraffin sections of breast tumors, we were able to
`evaluate carefully the conditions used for histochemistry to
`optimize localization of erbB2 in breast cancer specimens. With
`these new conditions, which include the use of frozen sections
`of freshly frozen tissues, formaldehyde postsectioning fixation,
`the deletion of treatments with any peroxide or organic solvents,
`and the deletion of counterstains, the localization of erbB2 was
`optimal in tumor specimens as well as in normal human liver.
`These results allowed us to go back and test previously available
`monoclonal antibodies, such as CB11 and E23, both of which
`also showed specific surface labeling at a low level in hepato-
`cytes. As shown in Fig. 2, this labeling was homogeneous,
`surface related, and absent in parallel controls for all of the
`antibodies, implying that it was truly specific for erbB2.
`Two other recombinant immunotoxins developed by our
`group have recently been evaluated in Phase 1 trials. Both can
`be given at much higher doses than erb-38. One of these is
`LMB-7 [B3(Fv)PE38], which targets a carbohydrate antigen
`present in colon, breast, and other epithelial cancers (14). The
`MTD of LMB-7 was found to be 24 mg/kg when given every
`other day times three. The dose-limiting toxicities were nausea,
`vomiting and diarrhea, secondary to the targeting of the carbo-
`hydrate antigen present on the gastric mucosa.4 LMB-2 [anti-
`Tac(Fv)PE38] is a recombinant immunotoxin that targets CD25
`(15), the a-subunit of the interleukin 2 receptor. In patients with
`
`leukemia and lymphoma, the MTD was 40 mg/kg, at which
`several clinical responses have been observed.5
`We conclude that the toxicity observed with erb-38 is most
`likely due to the presence of erbB2 on hepatocytes. The target-
`ing of tumors with antibodies to erbB2 that are armed with
`radioisotopes and other toxic agents may result in unexpected
`organ toxicities due to erbB2 expression on normal tissues.
`
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`4 L. H. Pai-Scherf and I. Pastan, manuscript in preparation.
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`5 R. J. Kreitman, T. Waldmann, and I. Pastan, manuscript in preparation.
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`on October 30, 2014. © 1999 American Association for Cancer
`Research.
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`IMMUNOGEN 2029, pg. 6
`Phigenix v. ImmunoGen
`IPR2014-00676
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