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Recently, Gilead Sciences (Foster City, CA, USA) presented a potential cytostatic drug GS-9219. It is a novel lipophilic prodrug of cyprPMEDAP, in vivo releasing the active compound PMEG in a two-step process. GS-9219 has shown a substantial therapeutic potential in treatment of spontaneous non-Hodgkin's lymphoma in dogs and its utilization in the human medicine is prospective. Hence, cyprPMEDAP represents a key intermediate in the intracellular activation of GS-9219. Both acyclic nucleoside phosphonates PMEG and cyprPMEDAP, serving as the basis for development of GS-9219, were discovered and their mechanism of action was investigated in detail at the Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic. The biological studies using the rat lymphoma were carried out at the First Faculty of Medicine, Charles University.

Contemporary views on the mechanisms of aging and aging variations--progeria and longevity are presented. Replicative aging, the roles of glycation and free radicals on the genetical determination of progeria and longevity are discussed.

More than a decade of experimental work in an inbred subline of Sprague-Dawley rats having high incidence of spontaneous T-cell lymphoma/leukaemia is reviewed. Longitudinal follow-up of biological characteristics (growth, survival, haematology) of both multiple cases of primary disease and s.c. passaged lymphomas as well as comparative immunophenotypic and karyotypic studies are concluded. In these T-cell lymphomas (mostly CD4 positive), arising on the same genetic background of the inbred SD strain, the aberrations involving chromosome 11 have been recognized as a typical non-random cytogenetic marker. This unique rat model of lymphoblastic lymphomas/leukaemias, relevant to human pathology, seems to be very suitable for testing different anticancer therapeutic strategies, as it is documented by results of a number of various protocols conducted in our laboratory.

O-Phosphonatomethylcholine, an isopolar phosphocholine analogue with a phosphonomethyl ether group replacing a phosphomonoester residue, was prepared by reaction of diisopropyl 2-chloroethoxymethylphosphonate with dimethylamine followed by quaternization of the thus-obtained diisopropyl 2-dimethylaminoethoxymethylphosphonate with iodomethane; the ester groups in the quaternary intermediate were cleaved with bromotrimethylsilane. Replacement of dimethylamine in the reaction sequence by morpholine and/or pyrrolidine gave the N-methylmorpholinium or N-methylpyrrolidinium analogues of O-phosphonatomethylcholine. Reaction of O-phosphonomethylcholine monotetrabutylammonium salt with 1-bromoalkanes in acetonitrile afforded a series of the corresponding monoalkyl (C10-C16) esters. None of these compounds except for the hexadecyl ester exhibited any appreciable cytostatic activity against DU-145, H460, HT-29, or MES-SA cell lines in vitro (evaluated by 3H-Thd incorporation assay). The hexadecyl ester exhibited modest in vitro cytotoxic activity comparable to that of the anticancer drug miltefosine (hexadecyl O-phosphocholine). In vivo evaluation of hexadecyl O-phosphonomethylcholine [transplanted SD lymphoma in inbred SD/cub rats, 10 mg kg(-1) day(-1) intratumoral injection for 10 days] resulted in a 40% decrease in lymphoma mass.

Antitumour efficiency of combined therapy with N-9-[2-(phosphonomethoxy)ethyl]-2,6-diaminopurine (PMEDAP) and docetaxel (DTX) was studied in an in vivo model of s.c. transplanted Sprague-Dawley (SD/Cub) rat T-cell lymphoma (phenotype SD10/96). The effect of the combined treatment of DTX with PMEDAP was significantly higher than that of DTX or PMEDAP alone. The s.c. administration of DXT into the vicinity of growing lymphoma together with i.p. administration of PMEDAP was found to be the most efficient combination. In this case, two out of four rats did not develop any lymphoma and remained alive. An irregular expression of Bcl2 protein was found in untreated and treated lymphomas, while the expression of protein p53 as well as MDM2 was not observed. All three types of the above-mentioned treatments (PMEDAP, DXT, DXT+PMEDAP) increased significantly the number of p21-positive cells, compared with untreated tumours.

The antitumour activity of four N6-substituted PMEDAP derivatives, Me2NEt-PMEDAP, allyl-PMEDAP, Me2-PMEDAP and cypr-PMEDAP, selected on the basis of their in vitro cytostatic activity, was studied in an in vivo model of haematological malignancy of inbred Sprague-Dawley rats. These compounds are believed to serve as the prodrugs of another (phosphonomethoxy)ethyl derivative, PMEG (9-[2-phosphonomethoxy) ethyl] guanine. We compared their toxicity and ability to inhibit tumour development in two different dosage regimes with those of their parent compound PMEDAP, as well with PMEG. The study confirmed the anticancer efficacy of the parental compound PMEDAP. Unlike PMEDAP, its N6-mono- and disubstituted congeners Me2NEt-PMEDAP, allyl-PMEDAP and Me2-PMEDAP were less potent or exhibited the same antineoplastic effect as PMEDAP. cypr-PMEDAP significantly decreased the survival of lymphoma-bearing rats due to high toxicity, which was approximately the same as that of PMEG. Therefore, these acyclic nucleoside phosphonates substituted at the 6-position of 2,6-diaminopurine ring do not seem to be promising drugs for the treatment of haematological malignancies.

We analyzed the effect of FK 506 on the production of nitric oxide by macrophages. Isolated rat peritoneal macrophages were cultured for 24 h with or without lipopolysaccharide (LPS) (5 microg/ml) and in the absence or presence of FK 506 (0.1 and 1 microg/ml). The concentration of NO2- in culture supernatants was taken as a measure of nitric oxide production. FK 506 (0.1 and 1 microg/ml) reduced the LPS-induced increase of NO2- levels by 68% and 81%, respectively. The impact of cyclosporin A (CsA) was studied in order to compare their effects. CsA (0.1 and 1 microg/ml) decreased the levels of nitrites by 39% and 69%, respectively. The results obtained suggest that both immunosuppressive drugs exhibit a dose-dependent inhibitory effect on nitric oxide production and that FK 506 is a more potent agent than CsA in this respect.

Antitumor effect of N-9-[2-(phosphonomethoxy) ethyl]-2,6-diaminopurine (PMEDAP) was studied in an in vivo model of s.c. transplanted Sprague-Dawley (SD/cub) rat T-cell lymphomas. Three individual SD/cub neoplasias (SD10/96, SD14/97, SD1/90) of different phenotypes were used. During the treatment, survival of the rats, increase of lymphoma mass, and DNA fragmentation detected by APO/BRDU kit, as well as Bcl2 and p53 protein expression, were followed. The study gives evidence of the positive therapeutic effect of PMEDAP in two of the three tested lymphomas, SD10/96 and SD14/97. Slowly growing SD1/90 lymphoma differs from the others in a uniform karyotype with trisomy of chromosome 11, CD4- immunophenotype, heterogeneous cellular morphology and constitutive expression of p53 protein found in some neoplastic cells. Thus, the diverse anticancer efficacy of PMEDAP treatment among SD/cub lymphomas could be associated with the different phenotypes of individual neoplasias.

Anticancer effect of heat shock, either alone or in combination with the drug PMEDAP, and cold water immersion stress were studied in an in vivo model of s.c. transplanted rat T-cell lymphomas in an inbred Sprague-Dawley rat line (SD/cub). Significant anticancer effect was induced by repeated sessions of heat shock; decrease of s.c. lymphoma weight and prolongation of survival time of treated rats was found to be dependent on the number of HS sessions. Much stronger therapeutic effect was observed after repeated heat shock in combination with PMEDAP administration. Light and electron microscopy studies were performed to characterize the alterations within the lymphomas. Morphologically, cellular alterations corresponding with apoptosis were observed in lymphoma cells after repeated heat shock. Indirect immunoperoxidase technique was used to detect HSP 72/73 protein(s), p53 and Bcl2 proteins in lymphomas heated directly or indirectly. The induction of HSP 72/73 protein(s) was found in the lymphoma tissues from autopsied animals exposed to heat shock; the intensity of its expression was dependent on the experimental design. The expression of p53 and BcL2 proteins was not changed in lymphoma cells of HS treated animals as compared to that of untreated lymphoma bearing controls; the Bcl2 protein was present in both treated and untreated lymphomas, and the p53 protein remained undetectable in all samples. Contrary to the heat shock, the cold stress did not suppress growth of lymphomas and, furthermore, accelerated the infiltration of parenchymatous organs with lymphoma cells.

Changes in the karyotype in three spontaneous Sprague-Dawley rat lymphomas (SD7/95, SD8/96, SD9/96) have been studied in the course of in vivo passages. In individual lymphomas karyological findings of primary disease from lymph nodes were compared with changes found in the 1st and 10th passages of the lymphoma and bone marrow samples. Chromosome studies were performed on direct preparations using the G-banding technique. Chromosome counts of all specimens studied were near diploidy, the majority of metaphase cells being pseudodiploid. In later passages of two lymphomas, the tendency in selection to hyperdiploid karyotype, particularly in bone marrow was observed. The examination revealed an increased percentage of breaks in lymph node cells of primary disease and the existence of nonrandom change, derivative chromosome 11, which occurred in structural variability in all three lymphomas studied. The aberration involving chromosome 11 was evaluated as the addition of unknown material at chromosome band 11q11 or as a duplication or triplication of segment 11q12-q23. If this structural aberration was not found, the excessive derivative chromosome 11 or translocation t(11;13) was proved to be present. Further, rearrangements of chromosomes 13 and 7 were nonrandom chromosome abnormalities revealed in later passages of the lymphomas. The results are in accordance with our previous observations in 14 cases of SD lymphomas that showed nonrandom occurrence of rearrangements concerning chromosome 11 and also relatively frequent translocation involving chromosome 13.

The diphosphoryl derivative of the acyclic nucleotide phosphonate analog 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA), found previously to weakly inhibit DNA pol delta/proliferating cell nuclear antigen, was studied as a substrate for pol alpha, delta, epsilon, and epsilon*. A comparison of the Vmax and Km for this derivative (PMEApp) and dATP demonstrated that the relative efficiency of the incorporation of this analog into the DNA chain is decreasing in the following order: pol delta approximately equal to pol epsilon approximately equal to pol epsilon*pol alpha. Under the reaction conditions, this incorporation amounted to 4.4 to 0.7% of dAMP molecules. Similar Km values for PMEApp and dATP in pol epsilon and pol epsilon* catalyzed reactions revealed that proteolysis of the enzyme probably does not affect the dNTP binding site. The DNA polymerases tested were inhibited by the reaction product (PMEA terminated DNA chain) with similar Ki/Km ratios (pol alpha 0.2; pol delta, 0.1; pol epsilon 0.05; and pol epsilon*, 0.06). The associated 3'-5'-exonuclease activity of pol delta, epsilon, and epsilon* was able to excise PMEA from the 3'-OH end of DNA with a rate one order of magnitude lower than that of the dAMP residue.

PMEDAP and/or PMEA treatment of SD rat lymphomas significantly prolonged the mean survival time of tumor-bearing animals. Dose-dependent genotoxicity of both PMEDAP and PMEA was not observed in in vitro tests on stabilized diploid MRC-5 cell line. The mitotic activity of MRC-5 cells was completely inhibited after 48 hours exposure in culture medium containing PMEDAP (10 micrograms/ml), or PMEA (25 micrograms/ml), respectively. Significant concentration dependent inhibition of cell proliferation caused by PMEDAP and/or PMEA was also observed in murine splenocytes. The analogs specifically inhibit proliferation of mitogen-activated T-lymphocytes. Modulation of subpopulations of peripheral blood cells under in vivo conditions was found in inbred SD animals. Intraperitoneal administration of PMEDAP to young healthy SD animals induced the decrease of the CD4+/CD8+ value from 1.3-1.6 to 0.72 while i.p. application of PMEA caused a decrease of the same ratio to 0.62.

To verify the penetration of liposomes through skin, we have used liposomes with encapsulated protein G-gold conjugate in a gel vehicle. Skin samples were examined 2, 4, 24, 48, and 72 h after liposome application. Our findings show that the penetration of liposomes through skin depends mainly on their size. Liposomes up to 600 nm in diameter penetrate through skin rather easily, whereas liposomes 1000 nm and more in diameter remain interiorized in the stratum corneum. The main penetration of liposomes proceeds along the hair sheaths as indicated by larger amounts of free liposomes in the corium of guinea pigs.

Spontaneous rat CD4+CD8-T-cell leukaemia transplanted in syngeneic recipients served as an experimental model system for IL-2 therapy. As a source of IL-2, supernatants from in vitro cultured plasmacytoma cell line X63-m-IL2 secreting constitutively recombinant murine IL-2 were utilized. Administration of IL-2, s.c. to the vicinity of the tumour inoculum, suppressed tumour growth. The tumour-inhibitory IL-2 effects were time- and dose-dependent. When the treatment has started 10 days after the challenge with 10(4) leukaemia cells, IL-2 inhibitory effects on the lymphoma growth in situ were demonstrated by lower tumour weight combined with necrotic changes. No histological signs of lymphoma generalization were found in parenchymatous organs of IL-2-treated rats in contrast to the untreated controls. No histological or functional injuries to the kidneys due to IL-2 administration were found. The results of effector cell phenotyping demonstrated the kinetics of the CD4+/CD8+ ratio characterized by CD4+ T-cell depletion and resulting increase in the percentage of CD8+ PBL.

We have investigated the interaction of liposomes with the continuous cell lines P388D1 and L-132 and mouse peritoneal macrophages. To distinguish the liposomes from other vesicular structures, we have used liposomes with incorporated protein G and gold. A heterogeneous mixture of multilamellar liposomes 30 nm up to 1000 nm in size has been employed. Samples were examined at different time intervals. We found differences in the uptake of liposomes with regard to size and rate. Cells of a P388D1 monolayer took up liposomes by endocytosis very early after addition of liposomes and the number of lysosomes in their cytoplasm increased. In L-132 cells, first a fusion occurred between liposomes and the cell cytoplasmic protrusions, in the cytoplasm of which the mitochondria had multiplied. Peritoneal macrophages phagocytosed mainly large multilamellar liposomes and the membranous system of Golgi apparatus was the most prominent structure in the cytoplasm. Phagocytosis in P388D1 and L-132 cells was noted sporadically as late as 24 h after addition of liposomes to the cells.

The genotoxic and embryotoxic effects of phosphonomethoxyalkylpurines, a new group of antiviral agents, decrease in the following order: PMEGPMEthioGPMEDAPPMEA(R)-PMPDAP = (R)-PMPA. Results of the present study are fully consistent with the previously found efficacy of their diphosphates to inhibit the replicative DNA polymerases. The marked genotoxicity of PMEG and PMEthioG is comparable to that of mitomycin C, whereas the moderate genotoxicity of PMEA is comparable to that of AZT. (R)-PMPDAP and (R)-PMPA did not induce any structural aberrations of chromosomes under the experimental conditions.

The antiphlogistic Ibuprofen incorporated in liposomes caused a decrease of the inflammatory edema induced by Carrageenan in the distal part of the rat's hind leg after both the intramuscular and percutaneous administration. The antiphlogistic effect of free Ibuprofen in the cream was weaker. Intramuscular administration of empty liposomes slowed down in the initial stages the development of inflammation and slightly diminished the size of edema.

Based on the powerful virostatic potency and cytostatic activity of adenine, 2,6-diaminopurine, and guanine derivatives of acyclic phosphonate nucleotide analog (S)-1-(3-hydroxy-2-phosphonomethoxypropyl) and 9-(2-phosphonomethoxyethyl) series, we examined the inhibitory potencies of their diphosphates [(S)-9-(3-hydroxy-2-phosphonomethoxypropyl)adenine diphosphate (HPMPApp), 9-(2-phosphonomethoxyethyl)adenine diphosphate, 9-(2-phosphonomethoxyethyl)-2,6-diaminopurine diphosphate (PMEDAPpp), and 9-(2-phosphonomethoxyethyl)guanine diphosphate, analogs of nucleoside 5'-triphosphates] toward cellular DNA polymerases alpha, delta, and epsilon (isolated from tumors of T cell spontaneous acute lymphoblastic leukemia in Sprague-Dawley inbred rats). Kinetic measurements (K(m), K(i), and V(max)) of synthetic homopolymeric template primers have shown that HPMPApp is a selective and potent inhibitor of polymerase epsilon, whereas PMEDAPpp strongly inhibits polymerase delta. These two compounds may be useful for elucidating the roles of polymerases delta and epsilon. Of the nucleotide analogs tested, 9-(2-phosphonomethoxyethyl) guanine diphosphate is the most efficient inhibitor of polymerases alpha and epsilon, whereas the diphosphate of 9-(2-phosphonomethoxyethyl) adenine, the therapeutically important agent adefovir, inhibits polymerases alpha and epsilon relatively poorly and exerts only moderate inhibition of polymerase delta. These data are quite consistent with previously reported cytostatic activity of these nucleotide analogs. All of the enzymes studied catalyze the incorporation of 9-(2-phosphonomethoxyethyl)adenine, 9-(2-phosphonomethoxyethyl)-2, 6-diaminopurine, and (S)-9-(3-hydroxy-2-phosphonomethoxypropyl)adenine into DNA chain. 9-(2-Phosphonomethoxyethyl)adenine diphosphate and PMEDAPpp were confirmed to be DNA chain terminators. On the other hand, HPMPApp formed poly(dT)/oligo(dA(18)-[(S)-9-(3-hydroxy-2-phosphonomethoxypropyl)a denine]2-4 structures.

Five cases of spontaneous acute lymphoblastic leukaemias (ALL) of Sprague-Dawley (SD) inbred rats have been studied cytogenetically. Chromosome studies were performed on direct preparations of the lymphomas using common cytogenetic methods. For detailed karyology, G-banded preparations were used. The chromosome numbers of metaphase cells in all leukaemias were near diploidy, the majority of metaphases being pseudodiploid with 1-2 stable marker chromosomes. The most frequent change involved chromosome 11, where the translocation form of trisomy 11 or 11q+ aberration was observed. Chromosome 11 abnormalities were found in all five leukaemias studied, suggesting to be a nonrandom change. 11 q11-q12 is supposed to be the critical region involved in the genesis or progression of SD ALL. These results are in agreement with our previous observations in nine cases of SD ALL (Sladká et al., 1988).

Retrovirus-like particles were detected by the negative staining method in supernatants of lymph node and spleen cell suspensions prepared from Sprague-Dawley rats with spontaneous acute lymphoblastic leukaemia (ALL). Similar particles were found in supernatants of cell suspensions from a lymphoma that developed after inoculation of lymph node and spleen cell suspension prepared from animals with spontaneous ALL into the subcutis of Sprague-Dawley recipients. On ultrathin sections, budding forms of the virus particles were seen as a dot at the periphery of neoplastically transformed cells.

Severe graft-versus-host disease (GVHD) developed following intravenous administration of parental BN.lx lymphoid cells into (SHR x BN.lx)F1 animals. Clinical symptoms, including foot-pad hyperemia, slobbering, melaena, progressive weight loss leading to death within 3 to 4 weeks, can be completely abrogated by 8 injections of PMEA starting 1 h after cell administration. Practically normal histological findings in PMEA-treated animals contrasted with nearly complete damage of bone marrow, lymphoid infiltration of salivary glands and even ulceration with hemorrhage of the epidermoid part of the stomach in untreated control GVHD animals.

The acyclic nucleotide analogues, HPMPC and PMEA, differ in their in vitro effect on the genetic material of eukaryotic cells. While HPMPC exerts a cytostatic effect on eukaryotic cells in vitro, PMEA has a genotoxic activity. These results correspond with the mode of embryotoxic action of these compounds: HPMPC exhibits a general embryolethal effect, whereas PMEA is apparently teratogenic and interacts with the mutant allele producing preaxial polydactyly of the hind limbs.

Two cases of spontaneous acute lymphoblastic leukemia in the inbred strain of Sprague-Dawley (Prague) rats have been observed. Since its reliable transplantability in syngeneic recipients was established, leukemic animals were used to test the cytostatic effect of PMEA. The treatment resulted in a significant prolongation of survival time of the treated animals. At the same time histological examination of PMEA-treated and untreated animals indicated that the drug effectively slows down the growth of lymphoma at the site of inoculation and inhibits the subsequent progression of tumor cells in the lung, liver, spleen and lymph nodes.

PMEA was administered i.p. daily on 10 consecutive days to inbred LEW rats inoculated with leukemic lymphoblastic cells KPH-Lw-I. The treatment was started 24 h after the inoculation. The observed cytostatic effects consisted in a significant prolongation of survival time associated with a suppression of the number of bone marrow leukemia cells with characteristic chromosomal marker of KPH-Lw-I leukemia as well as with a depression of the number of lymphoblasts in the blood.

The cytostatic potency of PMEA was evaluated on five rat and mouse experimental tumors. The compound significantly inhibits three spontaneously arisen (LW13K2, A870N and LLC) tumors. Two chemically induced tumors (FBN, HEP-LEW) were not influenced by PMEA.

1-(S)-(3-Hydroxy-2-phosphonylmethoxypropyl)cytosine (HPMPC) inhibits efficiently human cytomegalovirus (HCMV) replication. The drug exhibited marked antiviral effect in vitro not only at single application after virus adsorption, but also if given 48 hr post-infection. The inhibition of HCMV replication by HPMPC occurred after mere pretreatment of the host cells with the drug followed by its removal from the medium before virus infection. No free or host cell bound infectious virus was detected in the drug pretreated (1.5 micrograms/ml) virus infected cells. This effect was drug concentration dependent.

The antiviral efficacy of phosphonoformic acid (PFA) was examined on tissue cultures of the human embryonal lungs against the viruses herpes simplex type 1 and 2, herpes zoster, and the human cytomegalovirus using the method of the inhibition of the cytopathic effect and against herpes simplex type 1 and 2 on the tissue cultures of Vero cells using the methods of the inhibition of plaque formation. PFA was demonstrated to inhibit the reproduction of the viruses under study in both tests on tissue cultures. In persistence studies, the cytomegalovirus was not isolated back from the cells of the human embryonal lungs, which gave evidence of its greater sensitivity to PFA. In in vivo experiments, PFA in the form of a 3% ointment suppressed herpetic dermatitis on the guinea-pig skin, when treatment started 6, 24 and 48 hours after infection. In a preliminary experiment on rabbit corneas, an ointment with 3% PFA inhibited herpetic keratoconjunctivitis in time intervals of 1 and 3 hours after infection.

Rapid detection of human cytomegalovirus early nuclear protein in human diploid cells by immunofluorescence (IF) using commercial (Du Pont) monoclonal antibody was compared with the classical virus isolation in these cells. From 30 specimens tested early nuclear protein was detected in 16 cases but the virus was isolated out of 4 samples only.

10 diabetics with kidney and pancreas transplantation (6) and with isolated kidney transplantation (4) underwent examination with the aim to produce evidence of cytomegalovirus excretion (CMV) by the urine. To prove the presence of CMV antigen, the authors used monoclonal antibody produced by DuPont. CMV was found to be present in the urine of 4 patients. The examination lasts max. 48 hrs. It yielded no evidence of interdependence between viruria and the serum levels of the CMV antibodies. Tests for CMV in biological material by means of monoclonal antibody is a modern and quick method which, unlike conventional virological procedures (serology, virus cultivation), plays an important part in clinical medicine.

Raft-versus-host disease (GVHD) is known to be an extremely complex alloantigenic reaction. 1 In our work, we intended to analyze the contribution of RT1 and non-RT1 genetic factors to the course and features of the disease. We used separate lymph node (LN) cells and splenocyte suspensions to induce systemic GVHD in a fully (RT1 and multiple non-RT1) semiallogeneic P→F 1 model with BN. Lx and SHR strains as donors. Recently, a similar model with mixed lymphoid cell inoculum has been exploited for the testing of immunomodulating effects of a new acyclic adenine nucleotide analog. 2,3 These strain combinations are particularly useful when considering the possibility of demonstrating the role of genes of non-RT1 genetic background by inducing GVHD in F 1 (SHR × BN. Lx ) hybrids with lymphoid cells originating from individual recombinant inbred (RI) strains of BXH and HXB sets derived from (BN. Lx × SHR)F 2 and (SHR × BN. Lx )F 2 crosses. 4,5

MRC OX-3, MRC OX-6, and MRC OX-17 antibodies directed against rat MHC class II antigens were tested on a panel of rat inbred and congenic strains with different haplotypes using indirect immunoperoxidase and immunofluorescence techniques. In accordance with reports of others the MRC OX-17 was found to define RT1 class II monomorphic and the MRC OX-3 polymorphic determinants. MRC OX-6, giving no reaction with RT1d and RT1dv1 haplotypes, was shown to be directed against a rat MHC class II polymorphic determinant.

In rats, two alloantigenic loci, RT4 and RT6, were separated by recombination in a backcross population of (LEW X LEW.C-4A) X LEW hybrids using cytotoxicity tests with alloantisera and immunofluorescence with monoclonal antibody P4/16 in combination with histogenetic (skin graft) testing. Two new recombinant congenic strains were established, LEW.C-4A/I of the genotype CCRT4aaRT6aa and LEW.6B of the genotype ccRT4bbRT6bb. In the rat, transplantation results localize the histocompatibility locus RT4 closer to the locus albino, the T-cell RT6 locus without any proven histocompatibility effect is closely linked to the haemoglobin locus Hbb, in contrast to the mouse in which the haemoglobin (Hbb) and histocompatibility H-1 loci are closely linked.

From a panel of seven mouse monoclonal antibodies against human class II molecules only one, HL-37, directed against the beta chain of human DQ1 and DQ3 antigens cross-reacted in membrane immunofluorescence with the RT1b haplotype of the rat major histocompatibility complex. From the results obtained using an indirect immunoprecipitation method with 35S- and 125I-labelled rat samples from RT1b-carrying lymph node cells it can be supposed that the rat homologue of the human DQ beta chain was detected.

LEW rat lymphatic leukemia/lymphoma was antigenically phenotyped by means of W3/13, OX7, P4/16 and F 17-23-2 MoAbs. T-cell lineage related markers were proven to be expressed by leukemia cells. AAS prepared in congenic rat strains have shown the following pattern: alpha RT1 (MHC) AAS directed against RT1 antigenic specificities both "public" and "private" gave positive reactions with 100% of leukemia cells, all cross-reacting AAS directed against "public" specificities only, reacted positively too with 17-100% of leukemia cells and no alien specificities have been detected when LEW antisera were tested. The expression of RT5 differentiation antigen was proved on leukemia cells by means of alpha RT5 congenic AAS. T-cell differentiation antigen RT6 was also detected by means of alpha RT6 AAS with closely similar specificity as MoAb P4/16 which also positively reacted with KPH-Lw-I cells. Leukemia T-cell origin is also supported by the absence of class II antigens (F 17-23-2 MoAb) and SIg receptors. A presence of leukemia/lymphoma associated antigen was indicated by AAS absorption analysis.

The antigenic phenotyping of four individual spontaneous ALLs of SD strain revealed practically complete concordance of RT1 class I expression with PBL of SD strain. RT5 antigen as well as class II and SIg markers were not proven. T cell markers defined by MoAbs OX7 (Thy 1.1) and W3/13 (T cell common) were detected on all four ALL tested.