Intraperitoneal pretarget radioimmunotherapy with CC49 fusion protein.

PURPOSE: This study examined a pretarget radioimmunotherapy strategy for treatment of an i.p. tumor model (LS174T). EXPERIMENTAL DESIGN: The strategy used regional administration (i.p.) of a novel targeting molecule composed of four CC49 anti-tumor-associated glycoprotein 72 (TAG-72) single-chain antibodies linked to streptavidin as a fusion protein (CC49 fusion protein); 24 hours later, a synthetic clearing agent was administered i.v. to produce hepatic clearance of unbound CC49 fusion protein/synthetic clearing agent complexes. Four hours later, a low molecular weight radiolabeled reagent composed of biotin conjugated to the chelating agent 7,10-tetra-azacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) complexed with (111)In-, (90)Y-, or (177)Lu-DOTA-biotin was injected. RESULTS: Radiolocalization to tumor sites was superior with i.p. administration of radiolabeled DOTA-biotin as compared with i.v. administration. Imaging and biodistribution studies showed excellent tumor localization of radioactivity with (111)In- or (177)Lu-DOTA-biotin. Tumor localization of (111)In-DOTA-biotin was 43% ID/g and 44% ID/g at 4 and 24 hours with the highest normal tissue localization in the kidney with 6% ID/g at 48 and 72 hours. Therapy studies with (90)Y-DOTA-biotin at doses of 400 to 600 microCi or (177)Lu-DOTA-biotin at doses of 600 to 800 microCi produced significant prolongation of survival compared with controls (P = 0.03 and P < 0.01). CONCLUSIONS: Pretarget radioimmunotherapy using regional administration of CC49 fusion protein and i.p. (90)Y- or (177)Lu-DOTA-biotin represents a successful therapeutic strategy in the LS174T i.p. tumor model and this strategy may be applicable to human trials in patients with i.p. ovarian cancer.

Mouse/human chimeric antibodies to a tumor-associated antigen: biologic activity of the four human IgG subclasses.

Variable region genes from mouse monoclonal antibody 17-1A (gamma 2a kappa) with specificity for human gastrointestinal malignancies have been paired with human immunoglobulin constant region genes (for heavy and light chains) to produce mouse/human chimeric immunoglobulin molecules (chIgG) for each of the four human IgG subclasses. Mouse 17-1A and the four chIgG bound similarly to two human colon cancer cell lines and had comparable binding affinities. The chIgG1 and chIgG3 molecules mediated lymphocyte and monocyte antibody-dependent cell-mediated cytotoxicity (ADCC) to colon cancer tumor cell lines comparable to that of the parent murine 17-1A. The chIgG2 and chIgG4 molecules were able to mediate ADCC to colon cancer cell lines but were clearly inferior to the chIgG1 and chIgG3 reagents. None of the chIgG antibodies or the murine 17-1A was able to mediate complement lysis of colon cancer cell lines. These studies demonstrate the ability to produce all four human IgG subclass chimeric molecules which retain biologic activity. We have confirmed the subclass preferences of human lymphocyte and monocyte Fc receptors for human IgG subclasses previously determined by studies with monomeric or aggregated IgG. These data may aid in the selection of chimeric antibodies for in vivo trials.

Phase I trial of multiple large doses of murine monoclonal antibody CO17-1A. I. Clinical aspects.

Twenty-five patients with metastatic gastrointestinal adenocarcinoma received one to four infusions (400 mg) of murine monoclonal antibody CO17-1A. Eleven patients had mild gastrointestinal symptoms, and one had a transient flushing episode. Two of five who received three weekly infusions had readily reversible anaphylactic reactions at the time of the third infusion (day 15). There were no other toxic effects. One patient had a complete remission and is surviving at greater than 104 weeks, and four had stable disease. The median survival for the whole group was 57 weeks. In general, the antibody infusions were well tolerated but had modest antitumor effects.

Phase I trial of multiple large doses of murine monoclonal antibody CO17-1A. II. Pharmacokinetics and immune response.

Twenty-five patients with metastatic gastrointestinal adenocarcinoma received one to four infusions of large doses (400 mg) of murine monoclonal antibody CO17-1A (17-1A). The pharmacokinetics of 17-1A at the time of first, second, third, or fourth infusion were not statistically different; plasma half-lives were 15.0 +/- 1.7 hours (n = 5), 15.1 +/- 1.8 (n = 10), 25.3 +/- 6.2 (n = 3), and 14.4 +/- 1.8 (n = 5), respectively. Most patients had an antibody response to 17-1A, with peak levels occurring 15-22 days after infusion. The presence of serum antibody to 17-1A at the time of the second or third infusion did not significantly alter the pharmacokinetics of this large dose of antibody. Four of 25 patients failed to develop an antibody response, but this did not correlate with the amount of 17-1A administered. The administration of four doses of 400 mg over 1 week provided continuously circulating 17-1A for 10 days.

Lack of T-cell immunity in humans with preexisting anti-mouse immunoglobulin reactivity.

We have identified five normal individuals without known exposure to mouse immunoglobulin with "preexisting" human anti-mouse antibody (HAMA) against a panel of four mouse monoclonal antibodies and polyclonal mouse IgG. Competitive inhibition by polyclonal mouse IgG of HAMA binding to monoclonal antibody coated beads demonstrates the mouse constant region specificity of these sera. Lack of such inhibition by polyclonal human IgG eliminates polyclonal rheumatoid factors as an explanation. Lymphoblastic transformation studies in these five persons failed to demonstrate a memory T-cell response to the four mouse monoclonals or polyclonal mouse IgG. Positive controls included T-cell responses to streptolysin O in these five individuals as well as responses to monoclonal antibody D612 in three patients following treatment with that antibody. This lack of T-cell immunity to mouse IgG suggests that "preexisting" HAMA is the product of inadvertent cross-reactivity with murine constant region by antibodies directed against other antigens. Therefore, "preexisting" HAMA should pose no risk of anamnestic or allergic response in patients considered for murine monoclonal therapy.

Use of chlorotoxin for targeting of primary brain tumors.

Gliomas are primary brain tumors that arise from differentiated glial cells through a poorly understood malignant transformation. Although glioma cells retain some genetic and antigenic features common to glial cells, they show a remarkable degree of antigenic heterogeneity and variable mutations in their genome. Glioma cells have recently been shown to express a glioma-specific chloride ion channel (GCC) that is sensitive to chlorotoxin (CTX), a small peptide purified from Leiurus quinquestriatus scorpion venom [N. Ullrich et al, Neuroreport, 7: 1020-1024, 1996; and N. Ullrich and H. Sontheimer, Am. J. Physiol. (Cell Physiol.), 270: C1511-C1521, 1996]. Using native and recombinant 125I-labeled CTX, we show that toxin binding to glioma cells is specific and involves high affinity [dissociation constant (Kd)=4.2 nM] and low affinity (Kd=660 nml) binding sites. In radioreceptor assays, 125I-labeled CTX binds to a protein with Mr=72,000, presumably GCC or a receptor that modulates GCC activity. In vivo targeting and biodistribution experiments were obtained using 125I- and (131)I-labeled CTX injected into severe combined immunodeficient mice bearing xenografted gliomas. CTX selectively accumulated in the brain of tumor-bearing mice with calculated brain: muscle ratios of 36.4% of injected dose/g (ID/g), as compared to 12.4% ID/g in control animals. In the tumor-bearing severe combined immunodeficient mice, the vast majority of the brain-associated radioactivity was localized within the tumor (tumor:muscle ratio, 39.13% ID/g; contralateral brain:muscle ratio, 6.68%ID/g). Moreover, (131)I-labeled CTX distribution, visualized through in vivo imaging by gamma ray camera scans, demonstrates specific and persistent intratumoral localization of the radioactive ligand. Immunohistochemical studies using biotinylated and fluorescently tagged CTX show highly selective staining of glioma cells in vitro, in situ, and in sections of patient biopsies. Comparison tissues including normal human brain, kidney, and colon were consistently negative for CTX immunostaining. These data suggest that CTX and CTX-conjugated molecules may serve as glioma-specific markers with diagnostic and therapeutic potential.

Fractionated radioimmunotherapy of human colon carcinoma xenografts with 131I-labeled monoclonal antibody CC49.

Radiolabeled monoclonal antibodies (MoAbs) have been used for radioimmunotherapy (RIT) of human colon cancer xenografts in an attempt to develop promising clinical approaches to improving therapy success. Several strategies have been investigated to accomplish this task while decreasing toxicity. The CC49 antibody was selected for the present studies because of its relatively high affinity constant (16.2 x 10(9) M-1) for the high molecular weight TAG-72 mucinous antigen secreted by human colon cancer cells. In previous studies, when CC49 was labeled with 131I, it demonstrated a substantial therapeutic advantage over the lower affinity antibody (B72.3) reactive with TAG-72. One of the chief problems in achieving cures with RIT is that hematological toxicity limits the quantity of radionuclide that can be administered. In other studies of dose fractionation using athymic nude mice and 131I-labeled intact MoAbs reactive with human colon cancer xenografts, multiple administrations at approximately 1-week intervals were found to produce more prolonged tumor growth inhibition and less toxicity than single administrations. The purpose of this study was to investigate the therapeutic efficacy and toxicity of 131I-labeled CC49 MoAb administered with short fractionation schedules against human colon cancer xenografts to determine the optimal treatment schedule, with the ultimate aim of applying this approach in clinical trials. The results reported here demonstrate that in an animal colon cancer xenograft model, RIT delivered in a fractionated schedule clearly presents a therapeutic advantage. For example, one administration of 600 microCi 131I-labeled CC49 to LS174T tumor-bearing mice was lethal to approximately 25% of mice but produced no tumor regressions. Fractionation of this dose to two administrations of 300 microCi 131I-labeled CC49 at a 3-day interval resulted in tumor regression in approximately 30% of the animals, accompanied by a similar 25% death rate. The administration of 300 microCi 131I-labeled CC49 at a 7-day interval resulted in 15% animal lethality, but no complete tumor regressions. When three administrations of 300 microCi 131I-labeled CC49 were given over a 1-week period on days 0, 3, and 7, tumor regressions occurred in approximately 40% of the animals, accompanied by a 30% death rate. Moreover, three administrations of 300 microCi 131I-labeled CC49 resulted in 20% tumor recurrence, whereas two administrations of 131I-labeled CC49 resulted in 60% tumor recurrence.(ABSTRACT TRUNCATED AT 400 WORDS)

Pharmacokinetics and immune response of 131I-chimeric mouse/human B72.3 (human gamma 4) monoclonal antibody in humans.

Chimeric B72.3, composed of the V-regions of murine B72.3 and the constant regions of human immunoglobulin G4 heavy and kappa light chain, was administered as a 131I-labeled conjugate to 12 patients with metastatic colon cancer. Seven of these patients had an antibody response after initial infusion, and the immune response was primarily directed to the murine V-region, although a small proportion of the antibody response was directed to topographical epitopes requiring the presence of both murine V-region and human CH-1 and kappa constant regions (neo-epitopes). The pharmacokinetics included a plasma disappearance curve best fit by a two-compartmental model with an alpha t 1/2 of 18 +/- 7 h and a beta t 1/2 of 224 +/- 66 h. A second infusion of the same dose of 131I-chimeric B72.3 was administered to four of these patients 8 wk after the first infusion. Two patients who had a high antibody response to initial infusion had an anamnestic antibody response, and the infused ch-B72.3 rapidly disappeared from the circulation with associated immune complexes and free 131I in the plasma. One patient with no initial antibody response had no antibody response and identical pharmacokinetics on second infusion. One patient with a modest transient antibody response to initial infusion had no antibody response on second infusion and a modest shortening of plasma circulation. Thus, the human immunoglobulin G4 isotype chimeric B72.3 monoclonal antibody has a plasma half-life 6 to 8 times as long as murine B72.3 and retains considerable immunogenicity in some patients which can adversely affect repetitive infusions.

A phase I clinical trial of murine monoclonal antibody D612 in patients with metastatic gastrointestinal cancer.

In a phase I study, 21 patients with metastatic adenocarcinoma of the gastrointestinal tract received the murine monoclonal antibody D612. This antibody is directed at a M(r) 48,000 antigen restrictively expressed on tumors of the gastrointestinal tract and to a limited degree on normal gastrointestinal mucosa. Patients received total doses of 10-180 mg/m2 administered as single or multiple doses of 1-100 mg/m2 over an 8-day period. Dose-limiting toxicity was secretory diarrhea. A single dose of 100 mg/m2 exceeded guidelines for maximal tolerated dose. Higher total doses were achieved in subsequent patients by using repeated administration of lower doses. Three of five patients receiving 60 mg/m2 for 3 doses (180 mg/m2 total dose) experienced grade 3 diarrhea and could not complete the prescribed course. The dose of 40 mg/m2 administered on days 1, 4, and 8 (total dose, 120 mg/m2) has been selected as the dose for phase II studies. The pharmacokinetics of D612 is best described by a one-compartment model with a mean t1/2 of 48 +/- 3 h (SEM). Eighteen of 21 patients developed human anti-mouse antibody (HAMA). Patients who developed high levels of HAMA demonstrated a more rapid clearance of the day 8 dose than those who developed low levels of HAMA. In all patients studied, a component of HAMA was directed at the D612 variable region. With one exception, serum from all patients with detectable antibody to the D612 variable region demonstrated anti-paratope reactivity. Thirty-four % of known metastatic sites demonstrated uptake of radiolabeled D612. There were no objective antitumor responses in this phase I trial. The antitumor effect of D612 in vitro has been shown to be potentiated by interleukin 2 and recombinant human macrophage colony-stimulating factor. A phase II study of D612 administered in combination with cytokines that enhance human effector function is presently ongoing.

Phase II trial of murine monoclonal antibody D612 combined with recombinant human monocyte colony-stimulating factor (rhM-CSF) in patients with metastatic gastrointestinal cancer.

In a Phase II study, 14 patients with metastatic gastrointestinal cancer received the mAb D612 (40 mg/m2, days 4, 7, and 11) in combination with recombinant human monocyte colony-stimulating factor [(rhM-CSF) 80 micrograms/kg/days 1-14]. The combined treatment was well tolerated and resulted in characteristic biological activity associated with each of the agents. Thus, 10 of 14 patients experienced D612-associated secretory diarrhea, which responded to the prostaglandin inhibitor Indomethacin in 5 of 7 patients. rhM-CSF therapy was associated with peripheral monocytosis (peak absolute monocyte count, 1444 +/- 394/mm3) and thrombocytopenia (nadir count, 78 +/- 10/mm3). Monocyte surface marker analysis revealed a high baseline expression of CD16+ cells in our patient population with an additional increase with rhM-CSF therapy. We observed a correlation between the degree of thrombocytopenia and the pretreatment CD16+ monocyte count. Of the plasma cytokines assayed, serum Neopterin demonstrated the most consistent increase during rhM-CSF therapy. There was a significant difference in the half-life of the first and last dose of D612 (35.8 +/- 2 versus 27 +/- 2.9 h; P < 0.05). Eleven of fourteen patients developed low-moderate levels of anti-D612 antibody. Despite the observed biological activity of both rhM-CSF and D612 and the previously described in vitro synergy, no clinical antitumor responses were observed in this Phase II study.

Phase I clinical and pharmacological study of suppression of human antimouse antibody response to monoclonal antibody L6 by deoxyspergualin.

Development of human antimouse antibody (HAMA) is a major limiting factor in the application of murine mAb for clinical use. A novel immunomodulatory drug, deoxyspergualin (DSG), has shown potential to suppress antimouse antibody response in preclinical model systems. We conducted a Phase I trial to determine the effect of DSG on HAMA response to murine mAb L6 administered to patients with advanced cancers (in previous trials, this antibody elicited HAMA in two-thirds of the treated patients). L6 mAb was administered at a fixed dose of 200 mg/m2 on days 1-5. DSG was administered at doses of 50 mg/m2 [dose level (dl) 1] or 150 mg/m2 (dls II and III) on days 1-7. Treatment courses were repeated every 6 weeks (dls I and II) or every 3 weeks (dl III). HAMAs were quantitated by a commercially available ELISA assay (ImmuSTRIP; anti-isotypic antibodies) and a radiometric assay (antiisotypic and anti-idiotypic antibodies). Pharmacokinetics of L6 and DSG was also studied in all consenting patients. Among 24 evaluable patients, 2 patients developed detectable HAMAs using the ELISA (one each at dls I and II) after a median follow-up of 122 days (P = 0.0001 as compared to historical controls). Even in the two patients who developed HAMA, the HAMA levels were quite low (160 and 181 ng/ml; historical experience, 70-38,744 ng/ml). The radiometric assay detected anti-L6 antibodies in 13 patients (4, 6, and 3 at dls I-III, respectively) after a median of 82 days. The median highest anti-L6 antibody level was 129 ng/ml (range, 21-2150). The highest anti-L6 antibody level at dl III was only 44 ng/ml. The results suggest suppression of anti-idiotypic response also. No clinical antitumor activity was observed, and no significant changes in T4/T8 subsets or immunoglobulins occurred (suggesting a lack of generalized immunosuppression). We conclude that DSG can suppress HAMA response to L6. A starting dose of 150 mg/m2/day is recommended for Phase II trials to confirm this observation.

Phase I trial of the murine monoclonal anti-GD2 antibody 14G2a in metastatic melanoma.

In a phase I trial, 12 patients with GD2 antigen-positive metastatic melanoma received the murine anti-GD2 monoclonal antibody 14G2a. The monoclonal antibody was administered in four doses over an 8-day period with total dose ranging from 10 to 120 mg. All patients receiving greater than 10 mg of 14G2a experienced transient abdominal/pelvic pain during the antibody infusion. Five patients had a delayed extremity pain syndrome following the third and fourth antibody infusion. Four of the five patients developed neurological toxicity, including two patients with significant although reversible motor neuropathy. Two of the patients developed hyponatremia secondary to a syndrome of inappropriate antidiuretic hormone. All 12 patients developed high levels of human anti-14G2a antibody. The plasma half-life of 14G2a was 42 +/- 6 (SD) h. One patient each had a partial response, mixed response, and stable disease, respectively. The very modest antitumor activity accompanied by dose-limiting neurological toxicity at total doses greater than 80 mg may restrict the clinical utility of murine 14G2a.

Phase I study of anti--epidermal growth factor receptor antibody cetuximab in combination with radiation therapy in patients with advanced head and neck cancer.

PURPOSE: To evaluate the safety, pharmacokinetics, and efficacy of a chimeric anti-epidermal growth factor receptor monoclonal antibody, cetuximab, in combination with radiation therapy (RT) in patients with advanced squamous cell carcinoma of the head and neck. PATIENTS AND METHODS: We treated 16 patients in five successive treatment schedules. A standard dose escalation procedure was used; three patients entered onto the study at each dose level of cetuximab received conventional RT (70 Gy, 2 Gy/d), and the final three patients received hyperfractionated RT (76.8 Gy, 1.2 Gy bid). Cetuximab was delivered as a loading dose of 100 to 500 mg/m(2), followed by weekly infusions of 100 to 250 mg/m(2) for 7 to 8 weeks. Circulating levels of cetuximab during therapy were determined using a biomolecular interaction analysis core instrument. Human antichimeric antibody response was evaluated with a double-antigen radiometric assay. The recommended phase II/III dose was defined as the optimal cetuximab dose level based on the pharmacologic parameters and adverse events. RESULTS: The most commonly reported adverse events were fever, asthenia, transaminase elevation, nausea, and skin toxicities (grade 1 to 2 in most patients). Skin toxicity outside of the RT field was not strictly dose-dependent; however, grade 2 or higher events were observed in patients treated with higher dose regimens. There was one grade 4 allergic reaction. Most acute adverse effects were associated with RT (xerostomia, mucositis, and local skin toxicity). No antibodies against cetuximab were detected. All patients achieved an objective response (13 complete and two partial remissions). CONCLUSION: Cetuximab can be safely administered with RT. The recommended dose for phase II/III studies is a loading dose of 400 to 500 mg/m(2) and a maintenance weekly dose of 250 mg/m(2).

Patient-specific dosimetry of pretargeted radioimmunotherapy using CC49 fusion protein in patients with gastrointestinal malignancies.

UNLABELLED: Pretargeted radioimmunotherapy (RIT) using CC49 fusion protein, comprised of CC49-(scFv)4 and streptavidin, in conjunction with 90Y/111In-DOTA-biotin (DOTA = dodecanetetraacetic acid) provides a new opportunity to improve efficacy by increasing the tumor-to-normal tissue dose ratio. To our knowledge, the patient-specific dosimetry of pretargeted 90Y/111In-DOTA-biotin after CC49 fusion protein in patients has not been reported previously. METHODS: Nine patients received 3-step pretargeted RIT: (a) 160 mg/m2 of CC49 fusion protein, (b) synthetic clearing agent (sCA) at 48 or 72 h later, and (c) 90Y/111In-DOTA-biotin 24 h after the sCA administration. Sequential whole-body 111In images were acquired immediately and at 2-144 h after injection of 90Y/111In-DOTA-biotin. Geometric-mean quantification with background and attenuation correction was used for liver and lung dosimetry. Effective point source quantification was used for spleen, kidneys, and tumors. Organ and tumor 90Y doses were calculated based on 111In imaging data and the MIRD formalism using patient-specific organ masses determined from CT images. Patient-specific marrow doses were determined based on radioactivity concentration in the blood. RESULTS: The 90Y/111In-DOTA-biotin had a rapid plasma clearance, which was biphasic with <10% residual at 8 h. Organ masses ranged from 1,263 to 3,855 g for liver, 95 to 1,009 g for spleen, and 309 to 578 g for kidneys. The patient-specific mean 90Y dose (cGy/37 MBq, or rad/mCi) was 0.53 (0.32-0.78) to whole body, 3.75 (0.63-6.89) to liver, 2.32 (0.58-4.46) to spleen, 7.02 (3.36-11.2) to kidneys, 0.30 (0.09-0.44) to lungs, 0.22 (0.12-0.34) to marrow, and 28.9 (4.18-121.6) to tumors. CONCLUSION: Radiation dose to normal organs from circulating radionuclide is substantially reduced using pretargeted RIT. Tumor-to-normal organ dose ratios were increased about 8- to 11-fold compared with reported patient-specific mean dose to liver, spleen, marrow, and tumors from 90Y-CC49.

Biodistribution study of 188Re-labeled trisuccin-HuCC49 and trisuccin-HuCC49deltaCh2 conjugates in athymic nude mice bearing intraperitoneal colon cancer xenografts.

The trihydroxamate bifunctional chelating agent (BCA), trisuccin, has been shown to be a potential ligand for radiolabeling of monoclonal antibodies (MAbs) with rhenium radioisotopes, through an indirect postconjugation approach. The use of this trihydroxamate BCA made it possible to prepare stable BCA-MAb conjugates in pure form that could be radiolabeled with carrier-free 188Re. The anti-TAG-72 murine MAb, CC49, and its humanized derivatives are promising agents in the treatment of a number of malignancies with the CH2 domain-deleted MAb (HuCC49deltaCH2), which is of particular interest due to its rapid blood clearance. The biodistribution of 188Re-labeled conjugates of trisuccin with both humanized CC49 (HuCC49) and HuCC49deltaCH2 in athymic nude mice implanted i.p. with LS174T human colon carcinoma was studied. Trisuccin-MAb conjugates were synthesized at different BCA:MAb ratios by the 6-oxoheptanoic acid method using trisuccin hydrazide. The conjugates were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy for the number of incorporated trisuccin molecules. The conjugates were radiolabeled with carrier-free, generator-produced 188Re and purified by gel filtration on Sephadex G-25. Labeling yields and homogeneity of the labeled conjugates were analyzed by high-pressure liquid chromatography and instant TLC. Athymic nude mice were injected i.p. with LS174T human colon carcinoma cells, 7 days prior to injection of the labeled antibodies. 188Re-labeled MAbs were injected i.p., and the mice were sacrificed 24 h postinjection. Matrix-assisted laser desorption/ionization time-of-flight analyses showed stable incorporation of trisuccin into each MAb, with the measured ligand:MAb values positively correlating with the theoretical ratios. Labeling of the conjugates with 188Re proceeded with high yields, producing homogeneous 188Re-MAbs with good stabilities as shown by instant TLC and biodistribution analyses. Biodistribution of the radiolabeled MAbs at 24 h after injection showed median tumor uptake values of 23.5%ID/g and 17.6%ID/g for the 188Re-HuCC49deltaCH2 and 188Re-HuCC49, respectively. The blood clearance of the domain-deleted MAb was faster than that of the intact antibody. The blood values at 24 h after injection were 0.7%ID/g for 188Re-HuCC49deltaCH2 and 3.2%ID/g for 188Re-HuCC49. The results indicate that trisuccin is a promising agent for postconjugation labeling of antibodies with 188Re. Additionally, these results illustrate the potential of 188Re-HuCC49deltaCH2 in radioimmunodiagnosis and radioimmunotherapy of cancer.

Comparison of multiple bolus and continuous injections of 131I-labeled CC49 for therapy in a colon cancer xenograft model.

One of the problems in achieving cures with radioimmunotherapy is that hematological toxicity limits the quantity of radiolabeled monoclonal antibody (MAb) that can be administered. The MAb CC49 binds with high affinity to the TAG-72 antigen expressed in many human adenocarcinomas. We investigated tumor growth inhibition, survival, and tumor and bone marrow dosimetry after multiple bolus injections or continuous infusion of 131I-labeled CC49 MAb in a human colon cancer xenograft model to determine which method of administration results in the highest therapeutic ratio. Groups of athymic nude mice bearing established s.c. LS174T human colon cancer xenografts received three i.p. bolus injections (3X) of 131I-labeled CC49 (3X, days 0, 3, and 7) or were implanted i.p. with mini-osmotic pumps delivering 131I-labeled CC49 over 7 days. The total radionuclide doses administered were broken down into low-dose (< or = 450 microCi), medium-dose (450-800 microCi), and high-dose (> 800 microCi) groups. At the medium-dose level, the bolus-therapy animals did not have a significantly longer survival time but did have a significantly longer time-to-tumor doubling than the pump-therapy animals. The median survival for medium-dose bolus and pump therapy was 157 and 105 days, respectively, and the median time-to-tumor doubling was at least 114 and 77 days, respectively. At the low-dose level, the bolus-therapy animals had a significantly longer survival time but not a significantly longer time-to-tumor doubling than the pump-therapy animals. The median survival for low-dose bolus and pump therapy was 95.5 and 59 days, respectively, and the median time-to-tumor doubling was 73 and 38 days, respectively. The high-bolus dose was toxic. A comparison of the overall survival rate of pump therapy versus bolus therapy, excluding high-dose, resulted in the bolus-therapy animals having a longer survival time and a longer time-to-tumor doubling than the pump-therapy animals. Serial section autoradiography was used to reconstruct tumor activity density distributions over time. Average dose values calculated from total uptake data for 900 microCi administered activity yielded 158 Gy (3X) and 141 Gy (pump). Average three-dimensional doses using the radial histograms to calculate the absorbed fractions were 139 Gy and 123 Gy, respectively. This calculation includes energy loss external to the tumor. With cell proliferation parameters set to single fraction 60Co recurrence results, the effective dose (D(eff)) for local control was 11 Gy and 9 Gy, respectively. Three bolus injections resulted in a more uniform dose rate over a longer period, resulting in a calculated 19% improvement in D(eff) compared with pump administration. Dose to bone marrow was calculated assuming an activity concentration in bone marrow of 0.24 times the concentration in blood and an absorbed fraction of 0.63. For the 900-microCi 131I-labeled CC49 injected activity, pump administration resulted in an 80% higher calculated D(eff) to bone marrow compared with 3X bolus injection. These results demonstrate that 3X bolus injections were clearly superior to pump administration in terms of survival, tumor growth inhibition, tumor absorbed dose, and bone marrow dose.

A phase I study of anti-kinase insert domain-containing receptor antibody, IMC-1C11, in patients with liver metastases from colorectal carcinoma.

PURPOSE: Angiogenesis plays an important role in colorectal cancer progression. Stimulation of vascular endothelial growth factor receptor (VEGFR), a transmembrane glycoprotein, results in endothelial mitogenesis. Within this family of receptors, VEGFR 2/kinase-insert-domain-containing receptor (KDR) appear to be principally up-regulated during tumorigenesis. A chimeric anti-KDR antibody, IMC-1C11, blocks VEGFR-KDR interaction and inhibits VEGFR-induced endothelial cell proliferation. This trial seeks to assess the safety, tolerability and feasibility of targeting an important pathway in tumorigenesis. EXPERIMENTAL DESIGN: In a dose-escalation, single-agent study of IMC-1C11, we enrolled 14 patients with colorectal carcinoma and hepatic metastases. Safety-, pharmacokinetic-, immunogenicity-, and magnetic resonance imaging-assessed alteration of vascular effects of IMC-1C11 were evaluated in this trial. IMC-1C11 was infused weekly at 0.2 mg/kg (n = 3), 0.6 mg/kg (n = 4), 2.0 mg/kg (n = 3), and 4.0 mg/kg (n = 4) for 4 weeks, which constituted a cycle. RESULTS: No grade-3 or -4 IMC-1C11-related toxicities were observed. Minor grade-1 bleeding events were observed in four patients [0.2 mg/kg (n = 1) and 0.6 mg/kg (n = 3)]. Each resolved quickly and required no intervention. The starting dose of IMC-1C11 was selected to achieve a C(max) of approximately 5 micro g/ml. This concentration prevented KDR phosphorylation in vitro. Pharmacokinetic analysis demonstrated that the plasma t(1/2) and C(max) were dose dependent with a plasma t(1/2) of 67 +/- 3 h at the 4-mg/kg dose level. Human antichimeric antibodies were detected in 7 of 14 patients. The antibodies to IMC-1C11 inhibited the circulation of the agent in two patients. One patient had prolonged stable disease for seven cycles (28 weeks). The mean changes in tumor-influx volume-transfer constant k(in) (min(-1)) and enhancement factor after 4 weeks of therapy were significantly decreased compared with pretreatment values in 11 patients. CONCLUSION: IMC-1C11 was both safe and well tolerated. Drug levels of IMC-1C11 were reliably predicted. Further clinical investigation of anti-VEGFR/KDR agents is warranted.

Adenoviral-mediated delivery of gastrin-releasing peptide receptor results in specific tumor localization of a bombesin analogue in vivo.

Radioimmunotherapy is hindered by a variety of factors linked to the utilization of monoclonal antibodies. These limitations include restricted tumor penetration as well as low levels of intratumoral antigen expression. To address the latter problem, we used a gene therapy approach to induce tumor cells to express enhanced levels of receptor with high binding affinity for a radiolabeled peptide. In this regard, a radiolabeled bombesin analogue was used in conjunction with a recombinant adenoviral vector encoding the murine gastrin-releasing peptide receptor (mGRPr). A panel of human carcinoma cell lines was infected in vitro with the recombinant adenoviral vector encoding the mGRPr vector to examine the induced binding of a 125I-labeled bombesin peptide. All cell lines examined displayed high levels of induced peptide binding, with approximately 60-80% of the radioactivity bound to the cells, in a live-cell binding assay. The human ovarian carcinoma cell line SKOV3.ip1 was chosen for in vivo analysis of radiolabeled bombesin analogue tumor localization in biodistribution and pharmacokinetic studies in athymic nude mice. Genetic induction of mGRPr in vivo resulted in selective tumor uptake of the radiolabeled peptide and high tumor:blood ratios. The biodistribution results compared favorably to those obtained with 131I-labeled e21 anti-erbB-2 monoclonal antibody in animals bearing i.p. SKOV3.ip1 tumors that endogenously express erbB-2. Thus, a novel method to combine gene transfer and radioimmunotherapy may result in augmented tumor cell targeting of radiopharmaceuticals.

Phase II study of interferon-enhanced 131I-labeled high affinity CC49 monoclonal antibody therapy in patients with metastatic prostate cancer.

Adjuvant Interferon (IFN) was given to increase tumor antigen expression and enhance localization with 131I-labeled CC49 radioimmunotherapy in a Phase II trial for hormone resistant metastatic prostate cancer. Patients received four doses of alpha-IFN (3 x 10(6) IU) s.c. on alternate days, from day -5 to day +1 of 75 mCi/m2 131I-CC49 treatment. Toxicity was well tolerated, with the majority of patients experiencing transient grade 3 or 4 neutropenia and/or thrombocytopenia (maximal at 4-6 weeks). The absorbed dose was >25 Gy in four of eight tumors visualized, which represents an increase of >20 fold over whole body radiation dose. Two patients had radiographic minor responses by 6 weeks post-therapy, whereas five of six patients experiencing pain had symptom relief without radiographic changes. The protocol provided modest antitumor effects (pain relief in five of six patients and two minor radiographic responses). This study suggests that the addition of IFN enhanced tumor uptake and antitumor effects as compared to a prior Phase II trial of 131I-CC49 alone.

Phase II study of dual 131I-labeled monoclonal antibody therapy with interferon in patients with metastatic colorectal cancer.

The combination of COL-1 (anti-CEA) and CC49 (anti-TAG-72) has shown an increase in binding and distribution in colon cancer by immunoperoxidase staining compared to either antibody alone. To overcome tumor heterogeneity and allow delivery of higher radiation dose, 131I-labeled COL-1 and CC49 at a total dose of 75 mCi/m2 (2775 MBq/m2) were simultaneously administered to 14 patients with metastatic colon cancer. alpha-IFN (3 x 10(6) IU) was given s.c. on days -5 to +3 to increase carcinoembryonic antigen and TAG-72 antigen expression. Most patients had mild symptoms during IFN therapy, including mild neutropenia, fever, and malaise, which rapidly subsided after IFN cessation. No acute allergic reactions occurred with radioimmunotherapy; two patients experienced transient, delayed grade 2 arthralgias. Transient neutropenia and/or thrombocytopenia, which was maximal at 4-6 weeks, were consistent side effects without adverse events. All patients had tumor localization, and 13 of 14 patients achieved 4+ (highest grade) localization readings to at least one known site of disease. No objective responses occurred; 4 patients were stable and 10 progressed. Tumor dose estimates varied from 393 to 1327 cGy, including liver and extrahepatic sites. Combining two complementary antibodies and IFN administration appeared to increase localization intensity and radiation doses at tumor sites as compared to historical controls. The amount of radiation delivered to tumor sites was still below that required to cause tumor regressions in metastatic colorectal cancer.