Radiotherapy in mice with yttrium-90-labeled anti-Ly1 monoclonal antibody: therapy of established graft-versus-host disease induced across the major histocompatibility barrier

A monoclonal antibody recognizing Ly1, the murine homologue of CD5, was labeled with 90Y. In vivo biodistribution studies showed that 90Y-anti-Ly1 selectively localized in lymphoid tissue. Groups of B10,BR mice (H-2k) were lethally irradiated and given major histocompatibility complex-disparate C57BL/6 (H-2b) bone marrow and spleen cells to induce graft-versus-host disease (GVHD). Eight days later, mice with active GVHD were administered a single i.p. injection of 50 microCi90Y-anti-Ly1. Fifty % of these mice were alive 2 months after treatment. Long term (greater than 4-month) survival was significantly higher than in phosphate-buffered saline-treated mice. Survival was slightly improved in groups of mice receiving control irrelevant antibody labeled with 90Y or mice receiving free 90Y. However, survival in these groups was not significantly different from the phosphate-buffered saline-treated control group. The improved survival was supported by data showing improved mean animal weight. An anti-GVHD effect was confirmed by histopathological analysis. Unlabeled anti-Ly1 monoclonal antibody at comparable doses to 90Y-anti-Ly1 was not effective. Animals that died following 50-microCi treatment did not die of radiation toxicity, since all mice receiving 50 microCi 90Y-anti-Ly1 plus syngeneic bone marrow survived. The window of therapy was narrow in our studies, since 100 microCi 90Y-anti-Ly1 did not confer any survival advantage. Animals that did survive long term were studied for evidence of alloengraftment and found to have high levels of circulating donor mononuclear cells. 90Y-Anti-Ly1 localized in the spleen, thymus, liver, kidney and bone marrow but not in the bowel, lung, muscle, or skin. Animals given similar doses of free 90Y, 90Y-anti-Ly1, or labeled irrelevant antibody eliminated free 90Y fastest, followed by 90Y-anti-Ly1 and then labeled irrelevant antibody. Hematological analysis of peripheral blood from 90Y-anti-Ly1-treated mice showed reduction in total WBC counts, absolute lymphocyte numbers, and absolute neutrophil numbers on day 24 after treatment. Myelosuppression recovered by day 38. These findings indicate that Ly1-positive cells are involved in the effector phase of GVHD and that radiolabeled antibodies may be useful as cell-specific probes for studying the GVHD network. 90Y-Anti-Ly1 protected recipients long term from lethal GVHD, and the fact that it had a rather remarkable inhibitory and selective effect on the lymphoid system of mice suggests that these agents may have broader application in the field of transplantation.     

Radioimmunotherapy of human colonic cancer xenografts with 90Y labeled monoclonal antibodies to carcinoembryonic antigen

Monoclonal antibodies (MAbs) to carcinoembryonic antigen (CEA) or alpha-fetoprotein (AFP) were conjugated with diethylenetriaminepentaacetic acid and radiolabeled with 90Y at a specific activity of 4.0-6.0 mCi/mg. Approximately 50% of the radiolabeled anti-CEA antibody (90Y-labeled NP-2) bound to an immunoadsorbent containing CEA while analysis by high performance liquid chromatography revealed that 95-98% of the 90Y was associated with immunoglobulin. Less than 5% of the 90Y dissociated from either MAb after incubation in plasma for 48 h at 37 degrees C. After injection into nude mice, 98% of the circulating radioactivity remained associated with antibody and no loss of immunoreactivity was observed at 3 days. To evaluate 90Y-labeled NP-2 as a therapeutic agent, varied doses (10-100 microCi) were administered as a single i.v. injection into groups of nude mice bearing s.c. implants (0.3-0.4 g) of a CEA-producing human colonic cancer xenograft, GW-39. At the 10-microCi dose, no inhibition of tumor growth was observed. After 28 days, tumor growth was inhibited by as much as 77% in mice treated with 50 microCi of 90Y-labeled NP-2 as compared to tumor growth in control animals given 90Y-labeled anti-AFP. Doses higher than 50 microCi (75 and 100 microCi) were toxic to most of the animals, killing them within 2-3 weeks after administration. Marked suppression of circulating leukocytes was observed with 20 and 50 microCi by 1-2 weeks postinjection, but they returned to normal levels 3-4 weeks later. These studies show that treatment with 90Y-labeled MAbs against CEA can produce significant antitumor effects. However, toxicity to the bone marrow may limit the therapeutic efficacy of systemically administered 90Y-labeled MAbs.     

Therapy with unlabeled and 131I-labeled pan-B-cell monoclonal antibodies in nude mice bearing Raji Burkitt's lymphoma xenografts.

Clinical trials of radioimmunotherapy (RIT) of lymphoma have produced frequent tumor regressions and remissions, but it has been difficult to determine to what extent these tumor responses have been due to antibody-specific targeted radiation, nontargeted radiation, and/or cytotoxicity mediated by the carrier monoclonal antibody (MoAb). In this report, RIT was studied in athymic nude mice bearing s.c. Raji human Burkitt's lymphoma xenografts using two different pan-B-cell MoAbs, MB-1 (anti-CD37) and anti-B1 (anti-CD20), which differ in isotype (and thus the potential for interaction with host effector mechanisms) and isotype-matched control antibodies either in the unlabeled state or labeled with 131I. When a single i.p. injection of 300 microCi 131I-labeled MB-1 (IgG1) was compared to treatment with unlabeled MB-1 or 300 microCi 131I-labeled MYS control IgG1 MoAb, an antibody-specific targeted radiation effect of RIT was seen. 131I-labeled MB-1 produced a 44 +/- 19% (SEM) reduction in tumor size at 3 weeks posttreatment, while unlabeled MB-1 or 300 microCi 131I-labeled MYS control IgG1 antibody treatment resulted in continued tumor growth over this period of time. In vitro studies demonstrated that MB-1 was incapable of mediating antibody-dependent cellular cytotoxicity using Raji tumor cell targets and human peripheral blood mononuclear cells. Similar to the MB-1 studies, treatment with 300 microCi 131I-labeled anti-B1 produced a 64% reduction in mean tumor size, while 300 microCi of control antibody resulted in a 58% increase in tumor size over the same 3-week period. In contrast to MB-1, however, unlabeled anti-B1 (an IgG2a MoAb which in vitro studies showed to be capable of antibody-dependent cellular cytotoxicity) also had a substantial antitumor effect. Indeed, 300 microCi 131I-labeled anti-B1 and unlabeled anti-B1 treatment (using an equivalent amount of total protein in the treatment dose) produced a similar specific reduction in tumor size. Increasing the radionuclide dose of anti-B1 to 450 microCi in another experiment did not produce a significant difference in tumor regression compared to a 300-microCi dose. These results suggest that the antitumor effects of 131I-labeled anti-B1 treatment were dominated by antibody-mediated cytotoxicity mechanisms, such that an antibody-specific targeted radiation effect could not be distinguished. In contrast, antibody-specific targeting of radiation was the dominant mechanism of tumor killing with 131I-labeled MB-1.(ABSTRACT TRUNCATED AT 400 WORDS)     

Preclinical studies targeting normal and leukemic hematopoietic cells with Yttrium-90-labeled anti-CD45 antibody in vitro and in vivo in nude mice

A study was undertaken to investigate the suitability of using a high affinity (Kd = 1.1 nM) anti-CD45 monoclonal antibody for delivering the high energy beta-particle emitting isotope (90)Y to lymphohematopoietic target cells in vivo. The antibody, AHN-12, recognized the tyrosine phosphatase CD45 expressed on the surface of normal and malignant hematopoietic cells and studies showed that it reacted with both CD45-expressing normal peripheral blood cells and leukemia cells from patients. The antibody was readily labeled with (90)Y using the highly stable chelate 1B4M-DTPA and the radioimmunoconjugate was designated (90)Y-anti-CD45. The agent selectively bound to CD45(+) B cell line Daudi, but not CD45(-) control cells and significantly (p = 0.007) more bound to Daudi tumors growing in athymic nude mice than did a control non-reactive antibody. Moreover, biodistribution data correlated well to an anti-Daudi effect observed against established tumors in nude mice. The effect was dose dependent and irreversible with the best results in mice receiving a single dose of 137 microCi (90)Y-anti-CD45. These mice displayed a significantly (p < 0.0095) better anti-tumor effect than a control (90)Y-labeled antibody and survived over 135 days with no evidence of tumor. Histology studies showed no significant injury to kidney, liver, or small intestine even at 254 microCi, the highest dose tested. Because radiolabeled anti-CD45 antibody can be used to deliver radiation selectively to lymphohematopoietic tissue, these data indicate that this agent may be used to improve treatment of hematopoietic malignancies, particularly leukemia and lymphoma, when combined with hematopoietic stem cell transplantation in a future clinical trial.     

Therapeutic advantage of pretargeted radioimmunotherapy using a recombinant bispecific antibody in a human colon cancer xenograft.

PURPOSE: To assess if pretargeting, using a combination of a recombinant bispecific antibody (bsMAb) that binds divalently to carcinoembryonic antigen (CEA) and monovalently to the hapten histamine-succinyl-glycine and a (90)Y-peptide, improves therapeutic efficacy in a human colon cancer-nude mouse xenograft compared with control animals given (90)Y-humanized anti-CEA immunoglobulin G (IgG). EXPERIMENTAL DESIGN: Clearance and biodistribution were monitored by whole-body readings and necropsy. Animals were monitored for 34 weeks with a determination of residual disease and renal pathology in survivors. Hematologic toxicity was assessed separately in non-tumor-bearing NIH Swiss mice. RESULTS: Hematologic toxicity was severe at doses of 100 to 200 microCi of (90)Y-IgG, yet mild in the pretargeted animals given 500 or 700 microCi of the (90)Y-peptide. Evidence of end-stage renal disease was found at 900 microCi of the pretargeted (90)Y-peptide whereas animals given 700 microCi showed only mild renal pathology, similar to that seen in control animals given (90)Y-IgG. Biodistribution data indicated that the average amount of tumor radioactivity by a 700-microCi dose of the pretargeted peptide over a 96-hour period was increased 2.5-fold (48 microCi/g) compared with 150 microCi of (90)Y-IgG (18.9 microCi/g). At these doses, survival (i.e., time to progression to 2.5 cm(3)) was significantly improved (P < 0.04) compared with (90)Y-IgG, with ablation of about one third of the tumors, whereas viable tumor was present in all of the (90)Y-IgG-treated animals. CONCLUSION: Pretargeting increases the amount of radioactivity delivered to colorectal tumors sufficiently to improve the therapeutic index and responses as compared with conventional radioimmunotherapy.     

Radioimmunotherapy of human colorectal carcinoma xenografts using 90Y-labeled monoclonal antibody CO17-1A prepared by two bifunctional chelate techniques

Monoclonal antibody CO17-1A, which has specificity for colorectal and pancreatic carcinomas, was radiolabeled with the pure beta emitter, 90Y, by either the cyclic diethylenetriaminepentaacetic acid (DTPA) anhydride technique or by a site-specific bifunctional chelate technique using 1-(p-aminobenzyl)DTPA (p-NH2-Bz-DTPA). Female nude mice bearing SW 948 human colorectal carcinoma xenografts were given injections i.v. of 90Y-labeled monoclonal antibody CO17-1A at dosages of 100, 150, and 200 muCi/25 g body weight. Unlabeled CO17-1A (100 micrograms/25 g body weight) was coadministered. In animals receiving 90Y-CO17-1A prepared by the cyclic DTPA anhydride technique, tumor volume was unchanged from base line at a dose of 200 microCi/25 g. As the dosage of 90Y-CO17-1A increased, the rate of tumor growth decreased, but all experimental animals in this group died between 14 and 21 days. In contrast, CO17-1A radiolabeled with 90Y by the site-specific p-NH2-Bz-DTPA bifunctional chelate technique produced a maximum tumor volume reduction of 87% in the 200 microCi/25 g group by day 15, and no deaths were noted in any of the 90Y-CO17-1A-treated groups for 71 days. Dose-response curves again showed increased tumoricidal effects with increased dosages of 90Y-CO17-1A. S-2-(3-Aminopropylamino)ethylphosphorothioic acid, commonly known as WR-2721, is a radioprotective drug which has been shown to protect against bone marrow depression in irradiated humans. No protection was observed when WR-2721 was used as an adjunct to treatment with 90Y-CO17-1A prepared by either the cyclic DTPA anhydride technique or the site-specific p-NH2-Bz-DTPA technique. When the site-specific p-NH2-Bz-DTPA technique was used, the reduction in WBC and hemoglobin levels correlated with increasing bone marrow toxicity at higher doses. We conclude that CO17-1A labeled with 90Y via the site-specific p-NH2-Bz-DTPA technique has potential for radioimmunotherapy of human colorectal carcinoma.     

Gallium-67 radiotoxicity in human U937 lymphoma cells.

Promising clinical results have been obtained with radiolabeled antibodies in lymphoma patients. The higher uptake by lymphomas of 67Gallium (67Ga) compared with monoclonal antibodies makes selective radiotherapy by the widely available 67Ga appealing. However, the gamma radiation of 67Ga used in scintigraphy is considered to be almost non-toxic to lymphoma cells. However, in addition to photon radiation 67Ga emits low energy Auger electrons and 80-90 keV conversion electrons which could be cytotoxic. The objective of the present study was the assessment of radiotoxicity of 67Ga on a lymphoid cell line: U937. Proliferation (MTT-assay) and clonogenic capacity (CFU-assay) were measured after 3 and 6 days incubation with 10, 20 and 40 microCi ml-1 67Ga. Growth inhibition was 36% after 3 days incubation and 63% after 6 days incubation with 40 microCi 67Ga ml-1. Clonogenic capacity was reduced by 51% after 3 days and 72% after 6 days incubation with 40 microCi ml-1 67Ga. A survival curve showed an initial shoulder and became steeper beyond 200-250 pCi cell-1 (low linear energy transfer type). Iso-effect doses of 67Ga and 90Yttrium (90Y) were determined. The iso-effect dose of 40 microCi 67Ga ml-1 (cumulative dose of conversion electrons 306 cGy) was 2.5 microCi 90Y ml-1 (cumulative dose 494 cGy) and the iso-effect dose of 80 microCi 67Ga ml-1 was 5.0 microCi 90Y/ml. The main cytotoxic effect of 67Ga seems to be induced by the 80 keV conversion electrons. We conclude that the conversion electrons of 67Ga have a cytotoxic effect on U937 cells and that in our experiments a 16-fold higher microCi-dose of 67Ga than of 90Y was needed for the same cytotoxic effect. We believe that 67Ga holds promise for therapeutic use.     

Therapy of disseminated B-cell lymphoma xenografts in severe combined immunodeficient mice with an anti-CD74 antibody conjugated with (111)indium, (67)gallium, or (90)yttrium.

A radiolabeled antibody (Ab) to CD74 (the MHC class II invariant chain, Ii) was shown previously to effectively kill human B-lymphoma cells in vitro. Conjugates with both Auger electron and beta-particle emitters were able to kill cells, but the former displayed less nonspecific toxicity in the in vitro assay used. In this report, we have extended the studies to an in vivo model of tumor growth. The human B-cell lymphoma Raji was injected i.v. into severe combined immunodeficient mice, and radiolabeled Abs were injected at various times after tumor inoculation. The maximum tolerated dose (MTD), as well as lower doses, was tested. Tumor growth was monitored by hind-leg paralysis. With a 3-5-day interval before Ab injection, anti-CD74 conjugated to either (111)In or (67)Ga, at a dose of 240-350 microCi/mouse, produced a strong therapeutic effect, with greatly delayed tumor growth, and many of the treated mice were tumor free for >6 months. Control mice became paralyzed in 16-24 days, uniformly. Treatment at later time points (9-day interval) had little therapeutic effect. The MTD was required for optimal therapy. With the beta-particle emitter (90)Y, the MTD was much less, 25 microCi/mouse, and at this dose there was only a weak therapeutic effect. In conclusion, the data suggest that low-energy electrons are more effective than beta-particles in this model system. These results may be applicable to humans, particularly in the case of micrometastatic disease. This approach may also be effective with other Abs that accrete in large amounts.     

Radioimmunotherapy of intracerebral human glioma xenografts with 131I-labeled F(ab')2 fragments of monoclonal antibody Mel-14

The administration of radiolabeled monoclonal antibodies to improve the treatment of malignant gliomas is dependent upon achieving effective tumor radiation dose while sparing normal tissues. We have evaluated the efficacy of 131I-labeled F(ab')2 fragment of monoclonal antibody Mel-14, an IgG2a reactive with the chondroitin sulfate proteoglycan antigen of gliomas, melanomas, and other neoplasms, in prolonging survival of athymic mice transplanted intracerebrally with D-54 MG human glioma xenografts. Studies indicated that in vitro immunoreactivity, affinity, and tumor localization in vivo of radiolabeled Mel-14 F(ab')2 were maintained at specific activities of 10-13 microCi/micrograms. Intravenous injection of 1500 microCi/115 micrograms or 2000 microCi/154 micrograms 131I-labeled Mel-14 F(ab')2 into mice 6-7 days after xenograft implantation resulted in significant survival prolongation over control animals (P = 0.009 using Wilcoxon rank sum analysis). In another experiment, 1500 microCi/126 micrograms 131I-labeled Mel-14 F(ab')2 improved survival significantly over controls (P = 0.006), while 1500 microCi/220 micrograms 131I-labeled nonspecific antibody did not (P = 0.2). Increasing the injected radiation dose to 3000 microCi 131I-labeled Mel-14 F(ab')2 did not significantly increase survival in tumor-bearing mice, because of supervening radiation toxicity. However, giving 3000 microCi 131I-labeled Mel-14 F(ab')2 in two doses of 1500 microCi, 48 h apart, did significantly prolong animal survival over controls (P = 0.001). Estimated radiation dose to tumor was 915 rad after injection of 3000 microCi 131I-labeled Mel-14 F(ab')2 in two doses, a dose higher than that delivered to normal tissues. The results of this study suggest that radiolabeled Mel-14 F(ab')2 be evaluated as an agent for radioimmunotherapy trials.     

High-linear energy transfer (LET) alpha versus low-LET beta emitters in radioimmunotherapy of solid tumors: therapeutic efficacy and dose-limiting toxicity of 213Bi- versus 90Y-labeled CO17-1A Fab' fragments in a human colonic cancer model.

Recent studies suggest that radioimmunotherapy (RIT) with high-linear energy transfer (LET) radiation may have therapeutic advantages over conventional low-LET (e.g., beta-) emissions. Furthermore, fragments may be more effective in controlling tumor growth than complete IgG. However, to the best of our knowledge, no investigators have attempted a direct comparison of the therapeutic efficacy and toxicity of a systemic targeted therapeutic strategy, using high-LET alpha versus low-LET beta emitters in vivo. The aim of this study was, therefore, to assess the toxicity and antitumor efficacy of RIT with the alpha emitter 213Bi/213Po, as compared to the beta emitter 90Y, linked to a monovalent Fab' fragment in a human colonic cancer xenograft model in nude mice. Biodistribution studies of 213Bi- or 88Y-labeled benzyl-diethylene-triamine-pentaacetate-conjugated Fab' fragments of the murine monoclonal antibody CO17-1A were performed in nude mice bearing s.c. human colon cancer xenografts. 213Bi was readily obtained from an "in-house" 225Ac/213Bi generator. It decays by beta- and 440-keV gamma emission, with a t(1/2) of 45.6 min, as compared to the ultra-short-lived alpha emitter, 213Po (t(1/2) = 4.2 micros). For therapy, the mice were injected either with 213Bi- or 90Y-labeled CO17-1A Fab', whereas control groups were left untreated or were given a radiolabeled irrelevant control antibody. The maximum tolerated dose (MTD) of each agent was determined. The mice were treated with or without inhibition of the renal accretion of antibody fragments by D-lysine (T. M. Behr et al., Cancer Res., 55: 3825-3834, 1995), bone marrow transplantation, or combinations thereof. Myelotoxicity and potential second-organ toxicities, as well as tumor growth, were monitored at weekly intervals. Additionally, the therapeutic efficacy of both 213Bi- and 90Y-labeled CO17-1A Fab' was compared in a GW-39 model metastatic to the liver of nude mice. In accordance with kidney uptake values of as high as > or = 80% of the injected dose per gram, the kidney was the first dose-limiting organ using both 90Y- and 213Bi-labeled Fab' fragments. Application of D-lysine decreased the renal dose by >3-fold. Accordingly, myelotoxicity became dose limiting with both conjugates. By using lysine protection, the MTD of 90Y-Fab' was 250 microCi and the MTD of 213Bi-Fab' was 700 microCi, corresponding to blood doses of 5-8 Gy. Additional bone marrow transplantation allowed for an increase of the MTD of 90Y-Fab' to 400 microCi and for 213Bi-Fab' to 1100 microCi, respectively. At these very dose levels, no biochemical or histological evidence of renal damage was observed (kidney doses of <35 Gy). At equitoxic dosing, 213Bi-labeled Fab' fragments were significantly more effective than the respective 90Y-labeled conjugates. In the metastatic model, all untreated controls died from rapidly progressing hepatic metastases at 6-8 weeks after tumor inoculation, whereas a histologically confirmed cure was observed in 95% of those animals treated with 700 microCi of 213Bi-Fab' 10 days after model induction, which is in contrast to an only 20% cure rate in mice treated with 250 microCi of 90Y-Fab'. These data show that RIT with alpha emitters may be therapeutically more effective than conventional beta emitters. Surprisingly, maximum tolerated blood doses were, at 5-8 Gy, very similar between high-LET alpha and low-LET beta emitters. Due to its short physical half-life, 213Bi appears to be especially suitable for use in conjunction with fast-clearing fragments.     

Improved therapy of non-Hodgkin's lymphoma xenografts using radionuclides pretargeted with a new anti-CD20 bispecific antibody.

A comparison of the therapeutic efficacy of a new bispecific monoclonal antibody (bsMAb)-pretargeting system vs the conventional direct targeting modality was undertaken. A bsMAb was made by coupling the Fab' of a humanized anti-CD20 antibody to the Fab' of a murine antibody directed against the peptide histamine-succinyl-glycine (HSG). The tumor targeting of the bsMAb was separated from the subsequent delivery of the radionuclide-bearing HSG peptide conjugated with (111)In or (90)Y. Nude mice bearing s.c. Ramos human B-cell lymphomas were injected with the bsMAb and then, 48 h later, (111)In/(90)Y-HSG peptide was given. At 3 h postinjection, tumor/blood ratios for pretargeted (111)In-HSG-peptide were similar to that observed with the directly conjugated (111)In-anti-CD20 IgG at its highest level on day 7, but by day 1, tumor/blood ratios were about 10-fold higher than the IgG. Tumors progressed rapidly in animals given 800 microCi of (90)Y-HSG peptide alone, whereas 5/10 animals in the group pretargeted by the anti-CD20 bsMAb were tumor-free 18 weeks later. The antitumor response in animals administered the pretargeted (90)Y-HSG peptide was also significantly superior to treatment with the directly radiolabeled (90)Y-anti-CD20 IgG, whether given as a single injection (P<0.007) or as a divided dose (P=0.016). This bsMAb-pretargeting procedure significantly improves the therapeutic response of targeted radionuclides in non-Hodgkin's lymphoma, warranting further development of this method of radioimmunotherapy.     

Identification of a novel prostate tumor target, mindin/RG-1, for antibody-based radiotherapy of prostate cancer

Gene expression analysis showed that a human mindin homologue, mindin/RG-1, is expressed selectively in prostate tissues and that its expression level is elevated in some prostate tumors. Mindin/RG-1 protein expression is maintained in >80% of prostate cancers metastatic to bone or lymph nodes as well as in locally recurrent tumors in androgen-unresponsive patients. In contrast, mindin/RG-1 expression in other normal tissues is significantly lower than that seen in the prostate. A fully human antibody, 19G9, was generated against mindin/RG-1 protein and was shown to accumulate at high abundance in LNCaP tumor xenografts. Conjugates of this antibody with the chelator CHX-A'-DTPA were generated and radiolabeled with either 111In, 90Y, or 86Y. Small animal positron emission tomography imaging with the 86Y-radiolabeled conjugate showed very specific accumulation of the antibody in LNCaP tumor xenografts with clear tumor delineation apparent at 4 hours. The therapeutic efficacy of [90Y]-CHX-A'-DTPA-19G9 was evaluated in mice bearing LNCaP xenografts. A dose-finding study identified a nontoxic therapeutic dose to be approximately 75 microCi. Significant antitumor effects were seen with a single administration of radiolabeled antibody to animals bearing 200 to 400 mm3 tumors. Inhibition of tumor growth was observed in all treated animals over a 49-day period. At 49 days posttreatment, slow tumor growth recurred but this could be prevented for an additional 40-day period by a second administration of a 75 microCi dose at day 49. We conclude that [90Y]-CHX-A'-DTPA-19G9 is a novel antibody conjugate that has considerable promise for therapy of metastatic prostate cancer in androgen-unresponsive patients.     

186Re radioimmunotherapy of small cell lung carcinoma xenografts in nude mice.

A 186Re-labeled monoclonal antibody (MAb), NR-LU-10, was used for the radioimmunotherapy of a subcutaneous human small cell lung carcinoma xenograft, SHT-1, in nude mice. Biodistribution with specific and irrelevant labeled MAb demonstrated peak tumor uptake of 8% and 3% of the injected dose/g at 2 days, respectively. Dosimetry analysis predicted tumor:whole-body radiation-absorbed dose ratios of 2.43:1 for NR-LU-10 and 0.62:1 for irrelevant MAb. Single-dose toxicity screening estimated a 50% lethal dose within 30 days of 600 microCi (880 cGy of whole-body radiation). As anticipated, a multiple-dose regimen of 490 microCi in four doses over 10 days (720 cGy of whole-body radiation, eight of eight surviving greater than 30 days) was less toxic than a single bolus dose of 430 microCi (644 cGy of whole-body radiation), six of eight surviving greater than 30 days). A multidose radioimmunotherapy regimen was initiated in nude mice bearing 66-mm3 tumors (total dose, 500 to 600 microCi). Complete remissions (greater than 140 days) were achieved in three of 16 mice, and the remainder showed a mean tumor growth delay of 53 days. Matched doses with irrelevant MAb produced one remission, one treatment-related death, and a mean growth delay of only 20 days in six of eight mice. Thus, in this nonoptimal radioimmunotherapy model, significant antitumor responses were observed using a mildly toxic multiple dosing regimen.     

Combined modality radioimmunotherapy with Taxol and 90Y-Lym-1 for Raji lymphoma xenografts

Despite effective therapies for non-Hodgkin's lymphoma (NHL), the majority of patients are not cured. Radioimmunotherapy (RIT) has shown good results in preclinical and clinical trials even in patients that are non-responsive to standard chemotherapy. To make RIT more effective, agents such as paclitaxel (Taxol), that can enhance radiation effects, are being tested. Nude mice bearing human Burkitt's lymphoma (Raji) xenografts were treated with: 1) 150 or 200 microCi (5.5 or 7.3 MBq) of 90Y-2IT-BAD-Lym-1 alone, 2) 600 micrograms of Taxol alone, 3) 150 or 200 microCi of 90Y-2IT-BAD-Lym-1 plus 600 micrograms of Taxol given 24 hours after RIT, or 4) no treatment. Tumor size, survival, mouse weight and blood counts were monitored to assess efficacy and toxicity. Survival for mice treated in this 84 day trial was: 71% for 90Y-2IT-BAD-Lym-1 (200 microCi) plus Taxol, 29% for Taxol alone, 6% for 90Y-2IT-BAD-Lym-1 (200 microCi) alone and 14% in the untreated group. Average tumor volume in the 90Y-2IT-BAD-Lym-1 (200 microCi) plus Taxol group was reduced by 89 and 99% compared to the RIT alone and Taxol alone groups, respectively. Mice treated with 150 microCi had less toxicity than those treated with 200 microCi of 90Y-2IT-BAD-Lym-1, however, the higher radiation dose, and Taxol, were required for improved survival. Mouse weights and myelotoxicity in the combined modality (RIT plus Taxol) groups were similar to those receiving the same dose of RIT alone. In the Raji tumored nude mouse model, addition of Taxol to 90Y-2IT-BAD-Lym-1, in doses clinically achievable in humans, provided therapeutic synergy without increased or excessive toxicity.     

Therapeutic efficacy of anti-Lewis(y) humanized 3S193 radioimmunotherapy in a breast cancer model: enhanced activity when combined with taxol chemotherapy.

Monoclonal antibody therapy may provide new treatment options in the management of metastatic breast cancer by selectively targeting tumors and producing a therapeutic effect, by delivering radiation or other toxins directly to tumor cells, or by producing an intrinsic immune inflammatory response. The effect of 131I-labeled humanized anti-Lewis(y) monoclonal antibody 3S193 (hu3S193) was compared with that of placebo and radiolabeled huA33 control antibody in a series of radioimmunotherapy experiments in a MCF-7 xenografted BALB/c nude mouse breast cancer model. The maximum tolerated dose of 131I-labeled antibody occurred at 200 microCi/mouse, at which dose level three of six mice that received 131I-hu3S193 showed significant tumor growth inhibition in contrast to no responses in the comparable 131I-huA33 control treatment arm. Breast cancer is an ideal model to test the efficacy of combined modalities given its known sensitivity to both radiotherapy and chemotherapy. The synergy between radioimmunotherapy and chemotherapy was therefore also explored using a combination of 131I-labeled hu3S193 antibody and Taxol using subtherapeutic doses of each agent. The combination of Taxol and 100 microCi of 131I-hu3S193 produced significant tumor inhibition in 80% of mice, whereas no responses were seen with either treatment modality alone or the combination of Taxol and 131I-huA33. These results support a potential therapeutic role of radiolabeled hu3S193 in the treatment of breast cancer, including combination therapy with Taxol, and warrants further investigation of this promising new agent.     

131I-anticarcinoembryonic antigen therapy of LS174T human colon adenocarcinoma spheroids

LS174T human colon adenocarcinoma multicell spheroids were used to study the radiobiological aspects of radioimmunotherapy. The spheroids were incubated in 131I-anticarcinoembryonic antigen (B7) at an antibody concentration of 0.5 microgram/ml and at 131I concentrations of 2.5 and 7.5 microCi/ml. After incubation times of 90 h, clonogenic cells per spheroid were reduced by 1400-fold and 23-fold at the high and low 131I concentrations, respectively. 131I Nonspecific antibody (PX63) resulted in 2- and 1.2-fold reductions. Spheroid diameter was not significantly affected by therapy but histological examination revealed that there had been a significant reduction in the cell density, particularly near the spheroid surface. Using a theoretical model to estimate radiation dose, a radiation survival curve was constructed. The resulting curve was somewhat concave suggesting the presence of a resistant population of cells. It is likely that this observation is primarily due to the fact that the inner cells received a lower dose than the outer cells. A population of radiobiologically hypoxic cells in the inner portion of the spheroids may also have contributed to the decreasing slope of the curve as well as ongoing cell division leading to new cells which receive a lower radiation dose per cell cycle. Because of the ability to estimate radiation dose for a given biological effect, these types of experiments may allow predictions of the efficacy of radiolabeled antibody therapy for micrometastatic disease.     

Radioimmunotherapy of CD22-expressing Daudi tumors in nude mice with a 90Y-labeled anti-CD22 monoclonal antibody.

A study was undertaken to investigate the efficacy of a high affinity, rapidly internalizing anti-CD22 monoclonal antibody for selectively delivering high-energy (90)Y radioactivity to B lymphoma cells in vivo. The antibody, RFB4, was readily labeled with (90)Y using the highly stable chelate, 1B4M-diethylenetriaminepentaacetic acid. Labeled RFB4 selectively bound to the CD22(+) Burkitt's lymphoma cell line Daudi, but not to CD22(-) control cells in vitro as compared with a control antibody, and was more significantly bound (P = 0.03) to Daudi solid tumors growing in athymic nude mice. Biodistribution data correlated well with the antitumor effect. The therapeutic effect of (90)Y-labeled anti-CD22 (Y22) was dose-dependent, irreversible, and the best results were achieved in mice receiving a single i.p. dose of 196 microCi. These mice displayed a significantly better (P < 0.01) antitumor response than control mice and survived >200 days with no evidence of tumor. Histology studies showed no significant injury to kidney, liver, or small intestine. Importantly, tumor-bearing mice treated with Y22 had no radiologic bone marrow damage compared with tumor-bearing mice treated with the control-labeled antibody arguing that the presence of CD22(+) tumor protected mice from bone marrow damage. When anti-CD22 radioimmunotherapy was compared to radioimmunotherapy with anti-CD19 and anti-CD45 antibodies, all three antibodies distributed significantly high levels of radioisotope to flank tumors in vivo compared with controls (P < 0.05), induced complete remission, and produced long-term, tumor-free survivors. These findings indicate that anti-CD22 radioimmunotherapy with Y22 is highly effective in vivo against CD22-expressing malignancies and may be a useful therapy for drug-refractory B cell leukemia patients.     

Targeting of human glioma xenografts in vivo utilizing radiolabeled antibodies

Radiolabeled antibodies provide a potential basis for selective radiotherapy of human gliomas. We have measured tumor targeting by radiolabeled monoclonal and polyclonal antibodies directed against neuroectodermal and tumor-associated antigens in nude mice bearing human glioma xenografts. Monoclonal P96.5, a mouse IgG2a immunoglobulin, defines an epitope of a human melanoma cell surface protein, and specifically binds the U-251 human glioma as measured by immunoperoxidase histochemistry. 111In-radiolabeled P96.5 specifically targets the U-251 human glioma xenograft and yields 87.0 microCuries (microCi) of tumor activity per gram per 100 microCi injected activity compared to 4.5 microCi following administration of radiolabeled irrelevant monoclonal antibody. Calculations of targeting ratios demonstrate deposited dose to be 11.6 times greater with radiolabeled P96.5 administration compared to irrelevant monoclonal antibody. The proportion of tumor dose found in normal organs is less than 10%, further supporting specific targeting of the human glioma xenograft by this antibody. Monoclonal antibody ZME018, which defines a second melanoma-associated antigen, and polyclonal rabbit antiferritin, which defines a tumor-associated antigen, demonstrate positive immunoperoxidase staining of the tumor, but comparatively decreased targeting. When compared to the 111In-radiolabeled antibody, 90Y-radiolabeled P96.5 demonstrates comparable tumor targeting and percentages of tumor dose found in normal organs. To test the therapeutic potential of 90Y-radiolabeled P96.5, tumors and normal sites were implanted with miniature thermoluminescent dosimeters (TLD). Seven days following administration of 100 microCi 90Y-radiolabeled P96.5, average absorbed doses of 3770, 980, 353, and 274 cGy were observed in tumor, liver, contralateral control site, and total body, respectively. Shared cell surface antigens among neuroectodermally derived neoplasms provide a basis for exploration of human glioma radioimmunotherapy.     

Biodistribution and dosimetry following infusion of antibodies labeled with large amounts of 131I.

Dosimetry and treatment planning for therapeutic infusions of radiolabeled antibodies are usually performed by extrapolation from the biodistribution of trace-labeled antibody. This extrapolation assumes that the biodistribution of high specific activity antibody will be similar to that seen with trace-labeled antibody. However, high doses of radiation result in rapid depletion of lymphoid and hematopoietic cells in lymph nodes, spleen, and marrow with replacement by blood and plasma. If radiolabeled antibody is cleared slowly from blood, this replacement may result in increased radionuclide concentrations in these tissues following infusions of antibody labeled with large amounts of radionuclide. To examine the influence of deposited radiation on the biodistribution of radiolabeled antibody, we treated mice with a constant amount of antibody that was labeled with varying amounts of 131I. Survival was determined in normal specific pathogen-free AKR/Cum mice (Thy1.2+) after infusion of anti-Thy1.1 antibody labeled with 10 to 6500 muCi of 131I, to determine an appropriate range of 131I doses for further study. The dose producing 50% lethality within 30 days following infusion of 131I-labeled antibody was 530 muCi 131I. Biodistribution, bone marrow histology, and dosimetry were subsequently determined after infusion of 500 micrograms of antibody labeled with 10, 250, 500, or 3500 muCi 131I. The amount of 131I did not influence uptake or retention of antibody in blood, liver, lung, or kidney. In contrast, infusion of antibody labeled with 250 to 3500 muCi of 131I led to a dose-related increase in the concentration of 131I in marrow, spleen, lymph node, and thymus. For example, at 96 h after infusion of antibody labeled with 500 or 3500 muCi 131I, concentrations in marrow were 3- to 4-fold higher than after infusion of trace-labeled antibody. The increase in marrow 131I concentrations was associated with depletion of cells and hemorrhage within the marrow space. As a result, estimated mean absorbed doses to marrow, lymph node, spleen, and thymus were 1.2 to 3.1 times higher than would have been predicted from the biodistribution of trace-labeled antibody. These results suggest that the biodistribution of trace-labeled antibody should be an accurate predictor of the behavior of high specific activity antibody in blood and solid organs such as liver and kidney. In contrast, radiation from antibody labeled with large amounts of radionuclide can result in an alteration of the concentration of radiolabeled antibody in rapidly responding tissues such as marrow.(ABSTRACT TRUNCATED AT 400 WORDS)     

A comparison of 131I-labeled monoclonal antibody 17-1A treatment to external beam irradiation on the growth of LS174T human colon carcinoma xenografts

Inhibition of growth of LS174T human colon cancer xenografts in athymic nude mice due to 131I-labeled MoAb 17-1A treatment was compared to inhibition due to different single doses of 60Co external radiation. From those data, conditions which produced equivalent radiobiological end points could be identified and compared to dose estimates calculated using a technique analogous to the Medical Internal Radiation Dose (MIRD) Committee formalism. The tumor growth rate in mice injected with a single intraperitoneal administration of 300 microCi of 131I-labeled MoAb was reduced relative to tumor growth in untreated control animals and in mice administered unlabeled MoAb and was found to be similar to the growth rate of tumors given a single 6 Gy dose of 60Co radiation. Furthermore, the growth rate of tumors in mice that received three injections of 300 microCi of 131I-labeled MoAb on days 9, 16 and 28 after tumor cell injection was similar to the growth rate of tumors given a single 60Co dose of 8 or 10 Gy. The biodistribution data for 125I-labeled 17-1A MoAb were used to calculate total doses for the tumor and various normal tissues in animals given a single administration of 131I-labeled 17-1A MoAb. The absorbed radiation dose in tumor was approximately five times higher than in normal tissues. The results of the present study indicate that the tumor growth inhibition produced by the administration of radiolabeled antibody can equal that produced by up to 10 Gy of external beam radiation. In addition, the MIRD calculations allow comparison of this form of low dose radiation to external photon irradiation.