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Biomedical subjects

G Hinkle

Publications and source records attributed to G Hinkle.

At least 37 records · Page 2Linked to original sources

Do prokaryotes contain microtubules?

In eukaryotic cells, microtubules are 24-nm-diameter tubular structures composed of a class of conserved proteins called tubulin. They are involved in numerous cell functions including ciliary motility, nerve cell elongation, pigment migration, centrosome formation, and chromosome movement. Although cytoplasmic tubules and fibers have been observed in bacteria, some with diameters similar to those of eukaryotes, no homologies to eukaryotic microtubules have been established. Certain groups of bacteria including azotobacters, cyanobacteria, enteric bacteria, and spirochetes have been frequently observed to possess microtubule-like structures, and others, including archaebacteria, have been shown to be sensitive to drugs that inhibit the polymerization of microtubules. Although little biochemical or molecular biological information is available, the differences observed among these prokaryotic structures suggest that their composition generally differs among themselves as well as from that of eukaryotes. We review the distribution of cytoplasmic tubules in prokaryotes, even though, in all cases, their functions remain unknown. At least some tend to occur in cells that are large, elongate, and motile, suggesting that they may be involved in cytoskeletal functions, intracellular motility, or transport activities comparable to those performed by eukaryotic microtubules. In Escherichia coli, the FtsZ protein is associated with the formation of a ring in the division zone between the newly forming offspring cells. Like tubulin, FtsZ is a GTPase and shares with tubulin a 7-amino-acid motif, making it a promising candidate in which to seek the origin of tubulins.

Animals↗

Monophyletic origins of the metazoa: an evolutionary link with fungi.

A phylogenetic framework inferred from comparisons of small subunit ribosomal RNA sequences describes the evolutionary origin and early branching patterns of the kingdom Animalia. Maximum likelihood analyses show the animal lineage is monophyletic and includes choanoflagellates. Within the metazoan assemblage, the divergence of sponges is followed by the Ctenophora, the Cnidaria plus the placozoan Trichoplax adhaerens, and finally by an unresolved polychotomy of bilateral animal phyla. From these data, it was inferred that animals and fungi share a unique evolutionary history and that their last common ancestor was a flagellated protist similar to extant choanoflagellates.

Animals↗

Evaluation of NR-LU-10 with the Neoprobe gamma detector in a mouse model.

The Neoprobe Model 1000 hand-held gamma detector, in combination with the murine monoclonal antibody (MAb) B72.3 can successfully intraoperatively target both primary and recurrent colorectal cancer. Because of the shortcomings of this system (length of time needed to clear unbound MAb and heterogeneous staining of cancer cells), new MAbs are under investigation. NR-LU-10 IgG and its FAB fragment were evaluated with the Neoprobe gamma detector in a nude mouse model. NR-LU-10 is a pancarcinoma IgG2b antibody that recognizes an oncofetal glycoprotein antigen that is expressed by most carcinomas. Animals were injected intraperitoneally with 125I-labeled IgG, 125I, or technetium 99m (99mTc)-labeled Fab fragment. The Neoprobe gamma detector and standard gamma well counts provided biodistribution and pharmacokinetic data. The Fab fragment achieved a 5:1 tumor to blood-pool background (BPB) ratio in only 66 hr, whereas the whole MAb required 14 days. Technetium did not appear to be an adequate isotope for this system because of its short half-life. Autoradiographs performed for both the 125I-labeled NR-LU-10 IgG and its 125I-labeled Fab fragment (but not 99mTc-labeled Fab) localized well in this model and gave adequate tumor to BPB and tissue ratios. Use of the 125I-labeled Fab significantly decreased the time required for clearance of the MAb radionuclide complex from the blood and tissue background, thus providing earlier tumor localization in the Radioimmunoguided Surgery (RIGS) system.

Animals↗

Therapeutic efficacy of a high-affinity anticarcinoembryonic antigen monoclonal antibody (COL-1).

COL-1 is a murine IgG2a monoclonal antibody (MAb) with a high affinity (1.4 x 10(9) M-1) for carcinoembryonic antigen (CEA) and no detectable reactivity for CEA-related antigens, such as nonspecific cross-reacting antigen (NCA) and normal fecal antigen (NFA). 125I-labeled COL-1 IgG was shown to efficiently and specifically target the LS-174T human colon carcinoma xenograft in athymic mice. Dose titration studies in this same model with 131I-labeled COL-1 demonstrated reduction of tumor growth rate when 300 microCi of the immunoconjugate was used (0.005 > p > 0.001). Administration of higher levels as a single dose led to increased toxicity. Dose fractionation experiments with 131I-COL-1 demonstrated the ability to administer much higher levels of the immunoconjugate with little or no toxicity, which resulted in a greater therapeutic efficacy. For example, three fractions of 200 microCi of 131I-COL-1 given at weekly intervals (for a total of 600 microCi) resulted in the substantial reduction (p < 0.0005) of the growth of established tumors in 100% (7/7) of mice, and in no evidence of tumor growth in 71% (5/7) of mice, at the end of the 63-day observation period. These results thus demonstrate the potential therapeutic efficacy for radiolabeled COL-1 in clinical trials and demonstrate the principle of the advantage of dose fractionation protocols for this immunoconjugate.

Adenocarcinoma, Mucinous↗

Therapeutic advantage of high-affinity anticarcinoma radioimmunoconjugates.

The effect of the relative affinity (Ka) on the antitumor efficacy of monoclonal antibodies (MAbs) has been questioned. It has previously been shown in experimental models that the use of MAbs with higher relative Kas manifests itself in a higher percentage of injected dose of MAb bound to tumor. On the other hand, mathematical models have proposed that the use of higher affinity MAbs may be disadvantageous for antitumor effects, since higher Ka MAbs would bind more antigen and prevent penetration of MAb through tumor. To test this hypothesis, three MAbs reacting to the human pancarcinoma antigen TAG-72 were used as radioimmunoconjugates for therapeutic efficacy versus the LS-174T human colon carcinoma xenograft. MAbs B72.3, CC49, and CC83 have all been shown by depletion studies to react to the same molecule and to all react with overlapping epitopes. While the relative Ka of B72.3 is 2.5 x 10(9) M-1, the relative Kas of CC49 and CC83 are 16.2 and 27.7 x 10(9) M-1, respectively. Each MAb was radiolabeled with 131I, and each radioimmunoconjugate was assayed at five dose levels for therapeutic efficacy using the human xenograft model. The results of these studies demonstrate substantial therapeutic advantage of the higher affinity MAbs CC49 and CC83 versus B72.3 at every dose level. While 500 microCi of B72.3 were required to reduce tumor growth in only a minority of tumor-bearing animals, the use of the same amount or less of the radioimmunoconjugates of CC49 or CC83 resulted in strong antitumor effects in 80 to 100% of tumor-bearing animals. Thus, stronger antitumor effects were seen using as little as 2.5- to 3-fold less of the higher Ka immunoconjugates CC49 and CC83 as compared with B72.3. While we acknowledge the potential disadvantages of higher Ka MAbs in some situations, at least the experimental studies and model system described here show that a distinct therapeutic advantage exists with the use of higher affinity immunoconjugates.

Adenocarcinoma, Mucinous↗

Intraoperative detection of colorectal cancer with radioimmunoguided surgery and CC49, a second-generation monoclonal antibody.

Radioimmunoguided surgery (RIGS) has been employed intraoperatively in cases of colorectal cancer to assess the extent of local tumor spread and metastatic disease. This technique uses radiolabeled monoclonal antibodies (MAbs) directed against tumor-associated antigens, and a hand-held gamma-detection probe to detect the radiolabel fixed to tumor tissue. Recently introduced is an MAb directed against tumor-associated glycoprotein (anti-TAG), CC49. Sixty patients were entered into the initial study. Eighteen of 21 (86%) primary tumors were localized by the CC49 MAb and the gamma-detecting probe. Twenty-nine of 30 (97%) recurrent tumors were localized. Antibody dose did not affect localization. Specimens were divided into tissue types I through IV, based on antibody localization and hematoxylin and eosin (H&E) staining: type I, RIGS (-) and histologically (-); type II, RIGS (-) and histologically (+); type III, RIGS (+) and histologically (-); type IV, RIGS (+) and histologically (+). Type IV tissue were further classified by whether they were grossly apparent, IVa, or grossly inapparent, IVb (occult). Occult tumor found by RIGS and confirmed by H&E staining (type IV) had localization ratios similar to RIGS-positive, histology-negative tissue (type III). Traditionally found cancer (type IV) had significantly higher ratios. In 12 of 24 patients (50%) with primary tumors and 14 of 30 patients (47%) with recurrent tumors, RIGS with CC49 altered the planned operative procedure. Radioimmunoguided surgery with CC49 provides useful, immediate intraoperative information not available by other techniques.

Adult↗

Radioimmunoguided surgery challenges traditional decision making in patients with primary colorectal cancer.

BACKGROUND: Initial experience with the radioimmunoguided surgery system (RIGS) has been found to impact on decision making in patients with recurrent colorectal cancers. Reported here is experience with RIGS-influenced therapeutic decisions in patients with primary colorectal cancer. METHODS: Thirty-six evaluable patients with primary cancers were injected with the second-generation anti-tumor-associated glycoprotein antibody CC49 labeled with 1 to 2 mCi iodine 125. Pharmacokinetic determination and precordial counts were obtained after injection and weekly until levels were less than 20 counts/2 sec. At surgery abdominal and pelvic explorations were performed, first traditionally by inspection and palpation and then with the hand-held, gamma-detecting probe. RIGS-positive tissue was considered cancerous and removed if possible. RESULTS: Thirty patients (83%) had positive antibody localization at surgery. Of those patients with localization, in 24 (80%) additional information was obtained at the time of surgery. In 11 patients (34%) staging changes were made as a result of RIGS exploration. New findings resulted in operative changes in nine patients (25%). Eleven (30%) of the original 36 patients became eligible for adjuvant chemotherapy based on current recommendations because of RIGS findings. CONCLUSIONS: In conclusion, the RIGS system provides immediate staging information that impacts on therapeutic interventions, challenging the adequacy of traditional procedures alone for primary colorectal cancer exploration.

Colonic Neoplasms↗

Radioimmunoguided surgery using iodine 125 B72.3 in patients with colorectal cancer.

Preliminary data using B72.3 murine monoclonal antibody labeled with iodine 125 suggested that both clinically apparent as well as occult sites of colorectal cancer could be identified intraoperatively using a hand-held gamma detecting probe. We report the preliminary data of a multicenter trial of this approach in patients with primary or recurrent colorectal cancer. One hundred four patients with primary, suspected, or known recurrent colorectal cancer received an intravenous infusion of 1 mg of B72.3 monoclonal antibody radiolabeled with 7.4 x 10 Bq of iodine 125. Twenty-six patients with primary colorectal cancer and 72 patients with recurrent colorectal cancer were examined. Using the gamma detecting probe, 78% of the patients had localization of the antibody in their tumor; this included 75% of primary tumor sites and 63% of all recurrent tumor sites; 9.2% of all tumor sites identified represented occult sites detected only with the gamma detecting probe. The overall sensitivity was 77% and a predictive value of a positive detection was 78%. A total of 30 occult sites in 26 patients were identified. In patients with recurrent cancer, the antibody study provided unique data that precluded resection in 10 patients, and in another eight patients it extended the potentially curative procedure.

Antibodies, Monoclonal↗

99mTc-medronate uptake in the temporomandibular joints of young rats treated with a mandibular hyperpropulsor.

To correct maxillomandibular sagittal discrepancies in growing children, most functional appliances position the mandible more anteriorly than its habitual relation with the maxilla. The aim of this study was to evaluate the effects of such a procedure on the mineralization of the temporomandibular joints in young growing rats. Temporomandibular joint uptake of a radioactive bone marker, technetium 99m methylene diphosphonate (99mTc-MDP), was measured over time as a hyperpropulsor appliance was being worn intermittently 12 hours/day. As the uptake of 99mTc-MDP decreased in control animals, it increased significantly in the joints of treated rats before returning to the baseline level after 5 weeks of treatment. These results suggest that intermittent anterior positioning of the mandible induces a high rate of bone and cartilage remodeling in the joints of growing rats and that bone-seeking radiopharmaceutical uptake, with 99mTc-MDP, may be a useful technique to monitor joint adaptation to an experimental functional change.

Activator Appliances↗

Radioimmunoguided surgery (RIGS) in recurrent colorectal cancer.

Since 1986, 191 patients with recurrent colorectal cancer have undergone surgical exploration 2 to 43 days after injection of 1.0 to 0.25 mg of monoclonal antibody (MAb) (B72.3 or 17-1A) radiolabeled with 5.0 to 1.0 mCi of 125I. The intraoperative use of a hand-held gamma detector (Neoprobe 1000) demonstrated that MAb identified tumor in 73% of cases. Clearer intraoperative definition of tumor margins and identification of occult tumor assisted the surgeon in the resection of liver metastases as well as nodal and pelvic disease. Unsuspected nodal disease was identified. The external use of the Neoprobe to scan the sacral region and intrarectal and intravaginal use led to the avoidance of operative procedures by defining inoperable disease. In approximately 25% of cases, the surgical procedure was modified based on Neoprobe findings. RIGS system provides a method of immediate intraoperative staging which may prevent additional recurrences, lead to earlier institution of adjuvant therapy, and result in improved survival.

Antibodies, Monoclonal↗

Advantage of dose fractionation in monoclonal antibody-targeted radioimmunotherapy.

Monoclonal antibody (MAb) B72.3 IgG was radiolabeled with 131I and administered to female athymic NCr-nu mice bearing the LS-174T human colon adenocarcinoma xenograft to determine if fractionation of MAb dose had any advantage in tumor therapy. In the LS-174T xenograft, only approximately 30%-60% of tumor cells express the B72.3-reactive TAG-72 antigen. The LS-174T xenograft was used to reflect the heterogeneity of the TAG-72 antigen often seen in biopsy specimens from patients. In contrast to a single 600-muCi dose of 131I-B72.3 IgG where 60% of the animals died from toxic effects, two 300-muCi doses of 131I-B72.3 IgG (total of 600 muCi) reduced or eliminated tumor growth in 90% of mice, with only 10% of the animals dying from toxic effects. Dose fractionation even permitted escalation of the dose to three doses (each 1 wk apart) of 300 muCi of 131I-B72.3 IgG (for a total of 900 muCi), resulting in even more extensive tumor reduction or elimination and minimal toxic effects. The use of an isotype-matched control MAb revealed a nonspecific component to tumor growth retardation, but the use of the specific B72.3 IgG demonstrated a much greater therapeutic effect. Tumors that had escaped MAb therapy were analyzed for expression of the B72.3-reactive TAG-72 antigen with the use of the immunoperoxidase method; they were shown to have the same antigenic phenotype as the untreated tumors. We verified tumor elimination by killing the test animals after a 7-week observation period and performing histologic examination of tumor sites. We also monitored toxic effects by histologic examination of numerous organs, including bone marrow. These studies thus demonstrate the advantage of dose fractionation of a radiolabeled MAb for tumor therapy. We anticipate that the concept of dose fractionation can be practically applied in radioimmunotherapeutic clinical trials with the development and use of recombinant-chimeric MAbs and modified constructs.

Animals↗

Gamma-detecting probe and autoradiographic studies of radiolabeled antibody B72.3 in CX-1 colon xenografts.

Nude mice bearing CX-1 colon tumors were injected with 50 microCi 125I-labeled monoclonal antibody (MAb) B72.3. Radioactivity in tumors was studied with the gamma detecting probe (GDP) on days 1, 3, 7, and 10 after MAb injection. On each day, two mice were sacrificed and sections were examined with autoradiography (ARG), immunoperoxidase methods (IMP), and routine stains. Mean probe counts showed increasing tumor to background ratios and ARG demonstrated a progressive increase in radionuclide in the tumors. The distribution of 125I was primarily around the vascular spaces on day 1, but by day 3 and progressively it appeared in tumor gland lumina and necrotic areas. A regional correlation was shown between radionuclide in vascular spaces and its sequestration in tumor elements.

Animals↗

Mobilifilum chasei: morphology and ecology of a spirochete from an intertidal stratified microbial mat community.

Spirochetes were found in the lower anoxiphototrophic layer of a stratified microbial mat (North Pond, Laguna Figueroa, Baja California, Mexico). Ultra-structural analysis of thin sections of field samples revealed spirochetes approximately 0.25 micrometer in diameter with 10 or more periplasmic flagella, leading to the interpretation that these spirochetes bear 10 flagellar insertions on each end. Morphometric study showed these free-living spirochetes greatly resemble certain symbiotic ones, i.e., Borrelia and certain termite spirochetes, the transverse sections of which are presented here. The ultrastructure of this spirochete also resembles Hollandina and Diplocalyx (spirochetes symbiotic in arthropods) more than it does Spirochaeta, the well known genus of mud-dwelling spirochetes. The new spirochete was detected in mat material collected both in 1985 and in 1987. Unique morphology (i.e., conspicuous outer coat of inner membrane, large number of periplasmic flagella) and ecology prompt us to name a new free-living spirochete.

Animals↗

Intraoperative radioimmunodetection of ovarian cancer using monoclonal antibody B72.3 and a portable gamma-detecting probe.

To assess the value of radioimmunoguided surgery in the intraoperative detection of ovarian cancer, we used monoclonal antibody B72.3, radiolabeled with 125I, and a hand-held gamma-detecting probe in 13 women with ovarian cancer undergoing exploratory laparotomy. B72.3, which recognizes a tumor-associated glycoprotein, TAG 72, was injected 12-29 days preoperatively (intraperitoneally in four cases, intravenously in nine, and by both routes in one). Intraoperatively, the abdomen was surveyed with the probe and probe counts were correlated with biopsies and excised specimens studied by routine histologic stains. Probe counts were positive in four of seven evaluable patients with histologically confirmed disease. In three of these four patients, the probe detected cancer in specimens interpreted as normal on frozen histologic sections. The probe also identified microscopic cancer in the one patient who had no gross disease. The specificity of the probe was 70%. Preoperative computed tomography was normal in all patients, including those with tumors as large as 3 cm. This pilot study shows the ability of radioimmunoguided surgery to detect occult ovarian cancer.

Antibodies, Monoclonal↗

Radioimmunoguided surgery in primary colon cancer.

Radioimmunoguided surgery (RIGS), the intraoperative use of a hand-held gamma detecting probe (GDP) to identify tissue containing radiolabeled monoclonal antibody (MAb), was performed upon 30 patients with primary colon carcinoma. Each patient received an intravenous injection of MAb B72.3 (1.0 to 0.25 mg) radiolabeled with 125I (5.0 to 1.0 mCi) 8 to 34 days before exploration. The GDP was used to measure radioactivity in colon tissue, tumor bed, nodal drainage areas, and areas of suspected metastases. Antibody localized to histologically documented tumor in 23 of 30 patients (77%). Tumor margins were more clearly defined in 20 of 30 patients (67%). GDP counts led to major alterations in surgical resection in five patients (17%) and changes in adjuvant therapy in four (14%). GDP counts identified occult liver metastases in two patients (7%) and correctly indicated the benign nature of liver masses in three (10%). In four patients (13%), occult nodal metastases were identified. RIGS can precisely delineate tumor margins, define the extent of nodal involvement, and localize occult tumor, providing a method of immediate intraoperative staging that may lessen recurrences and produce higher survival rates.

Antibodies, Monoclonal↗

Intra-abdominal recurrence of colorectal cancer detected by radioimmunoguided surgery (RIGS system).

Since 1986, 32 patients with metastatic colorectal cancer have undergone second-look radioimmunoguided surgery (RIGS system). The primary tumor was located in the right and transverse colon in 11 patients, left and sigmoid colon in 16, and rectum in five. The carcinoembryonic antigen level was elevated in 30 patients (94%); all patients underwent a computed tomographic scan of the abdomen and pelvis. The overall sensitivity of the computed tomographic scan was 41% (abdomen other than liver, 27%; liver, 58%; and pelvis, 22%). The RIGS system identified recurrent tumor in 81% of the patients. The most common site of metastasis was the liver (41%), independent of the primary location. Local/regional recurrences alone accounted for 40% of all recurrences. In six patients (18%), recurrent tumor was found only with the RIGS system. The RIGS system is more dependable in localizing clinically obscure metastases than other methods, and carcinoembryonic antigen testing remains the most accurate preoperative method to indicate suspected recurrences.

Abdominal Neoplasms↗