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PubMed2001

Low-copy-number human transgene is recognized as an X inactivation center in mouse ES cells, but fails to induce cis-inactivation in chimeric mice.

X chromosome inactivation is initiated from a segment of the mammalian X chromosome called the X inactivation center. Transgenes from this region of the murine X chromosome are providing the means to identify the DNA needed for cis inactivation in mice. We recently showed that chimeric mice carrying transgenes from the human X inactivation center (XIC) region also provide a functional assay for human XIC activity; approximately 6 copies of a 480-kb human transgene (ES-10) were sufficient to initiate random X inactivation in cells of male chimeric mice (Migeon et al., 1999, Genomics, 59, 113-121). Now, we report studies of another human transgene (ES-5), which contains less than 300 kb of the human XIC region on Xq13.2 including an intact XIST locus and which has inserted in one or two copies into mouse chromosome 6. The ES-5 transgene is recognized as an X inactivation center in mouse embryonic stem cells, but is not sufficient to induce random X inactivation in somatic cells of highly chimeric mice. Human transgenes in chimeric mice provide a means to uncouple the key steps in this complex pathway and facilitate the search for essential components of the human XIC region.

Animals↗
PubMed1998

Identification of the nuclease active site in the multifunctional RecBCD enzyme by creation of a chimeric enzyme.

The recombinational hot spot chi modulates the nuclease and helicase activities of the RecBCD enzyme, leading to generation of an early DNA intermediate for homologous recombination. Here we identify the subunit location of the nuclease active site in RecBCD. The isolated RecB protein cleaves circular single-stranded M13 phage DNA, but RecB1-929, comprising only the 100 kDa N-terminal domain of RecB, does not. We reported previously that the reconstituted RecB1-929CD enzyme also is not a nuclease, suggesting that the C-terminal 30 kDa domain of RecB is a non-specific ssDNA endonuclease. However, we were unable to detect nuclease activity with the subtilisin-generated C-terminal 30 kDa fragment of RecB. Since the subtilisin-generated fragment did not bind to a ssDNA-agarose column, we designed a chimeric enzyme by attaching the C-terminal 30 kDa domain of RecB to the gene 32 protein of T4 phage, a ssDNA binding protein that does not have strand scission ability. In addition, Asp427 in the chimeric enzyme (Asp1080 in RecB), a residue that is conserved among several RecB homologs, was substituted to alanine (the D427A mutant). The wild-type chimeric enzyme cleaves the M13 DNA and the D427A mutation abolishes the endonuclease activity of the chimeric enzyme but does not affect its DNA binding ability. This finding indicates an unusual bipartite nature in the structural organization of RecB, in which the DNA-binding function is located in the N-terminal 100 kDa domain and the nuclease catalytic domain is located in the C-terminal 30 kDa domain. The purified RecBD1080ACD mutant is a processive helicase but not a nuclease, demonstrating that RecBCD has a single nuclease active site in the C-terminal 30 kDa domain of RecB.

Amino Acid Sequence↗
PubMed1994

Vpu-mediated proteolysis of gp160/CD4 chimeric envelope glycoproteins in the endoplasmic reticulum: requirement of both the anchor and cytoplasmic domains of CD4.

The Vpu protein of HIV-1 induces degradation of CD4 in the endoplasmic reticulum. Previous studies have elucidated the role of the CD4 cytoplasmic domain in the Vpu-induced degradation process, and the minimal Vpu responsive element mapped to a small region in the CD4 tail. In the present study, we have carried out both biochemical and biological experiments to analyze the role of the CD4 anchor domain in the Vpu-induced degradation process. We generated chimeric proteins that possessed the ecto-anchor domains of gp160 and the cytoplasmic domain of CD4. The chimeric envelope glycoproteins were functionally active in the fusion of HeLa CD4+ cells, with the exception of those having the arginine to isoleucine (R to I) substitution in the gp160 anchor domain. Coexpression studies revealed that these chimeric glycoproteins were stable and functionally active in the presence of Vpu, as opposed to those having the anchor-cytoplasmic domains of CD4. The half-life of Vpu-sensitive chimeric glycoproteins was calculated to be approximately 60-90 min, whereas Vpu-resistant envelope glycoproteins exhibited relatively longer half-lives in the presence of Vpu. Taken together, these studies strongly suggest that the CD4 anchor domain appears to provide critical sequence or structural elements through which the Vpu protein could access CD4 or glycoproteins bearing the Vpu responsive element for degradation in the endoplasmic reticulum.

Amino Acid Sequence↗
PubMed2001

Use of chimeric nectin-1(HveC)-related receptors to demonstrate that ability to bind alphaherpesvirus gD is not necessarily sufficient for viral entry.

Human nectin-1 (HveC, Prr1), a member of the immunoglobulin superfamily and a receptor for the entry of herpes simplex viruses 1 and 2 (HSV-1, HSV-2), pseudorabies virus (PRV), and bovine herpesvirus 1 (BHV-1), binds to viral gD. For HSV-1, HSV-2, and PRV, the gD-binding region of nectin-1 has been localized to the N-terminal V-like domain. To determine whether the two C-like domains of nectin-1 influenced gD binding and entry activity, genes encoding chimeric proteins were constructed. Portions of nectin-1 were replaced with homologous regions from nectin-2 (HveB, Prr2), a related protein with ability to mediate the entry of PRV, HSV-2, and Rid mutants of HSV-1, but not HSV-1 or BHV-1. Also, one or more domains of nectin-1 were fused to the two membrane-proximal Ig domains of CD4, a protein with no herpesvirus entry or gD-binding activity. The chimeric proteins were expressed in Chinese hamster ovary cells, which normally lack alphaherpesvirus entry receptors, and detected on the cell surface by one or more anti-nectin-1 monoclonal antibodies. One chimeric protein (nectin-1 amino acids 1-124 fused to CD4) failed to bind to soluble forms of HSV-1, HSV-2, PRV, and BHV-1 gD and, as expected, also failed to mediate entry of the viruses from which these gDs were derived. The other chimeric receptors bound all forms of gD. Some mediated the entry of all the viruses tested but others mediated entry of some but not all the viruses. We conclude that binding of gD to the nectin-1 V domain is not sufficient for entry activity, that there are structural requirements for entry activity independent of gD binding, and that these requirements are different for the several alphaherpesviruses that can use nectin-1 as a receptor.

Amino Acid Sequence↗
PubMed2001

The 3' stem loop of the West Nile virus genomic RNA can suppress translation of chimeric mRNAs.

Cis-acting elements that regulate translation have been identified in the 3' noncoding regions (NCRs) of cellular and viral mRNAs. As one means of analyzing the effect on translation of the conserved 3' terminal RNA structure of the West Nile virus (WNV) genome, the translation efficiencies of chimeric mRNAs composed of a CAT reporter gene flanked by viral or nonviral 5' and 3' terminal sequences were compared. In vitro, the WNV 3'(+) stem loop (SL) RNA reduced the translation efficiencies of chimeric mRNAs with either viral or nonviral 5' NCRs, suggesting that a specific 3'-5' RNA-RNA interaction was not involved. In contrast, the 3' terminal sequence of a togavirus, rubella virus, enhanced translation efficiency. The WNV 3'(+)SL reduced translation efficiency both in cis and in trans and of both capped and uncapped chimeric mRNAs. We have previously reported that three cellular proteins bind specifically to the WNV 3'(+)SL RNA. Competition between the WNV 3'(+)SL and the 5' terminus of the chimeric mRNAs for proteins involved in translation initiation could explain the translation inhibition observed.

Animals↗
PubMed2002

Characterization of complex B cell epitopes on woodchuck hepatitis virus surface antigens by using plasmids encoding chimeric proteins and DNA immunization.

The conformational nature of the B cell epitopes on the hepadnavirus surface antigens makes its characterization difficult. Here, a new approach by DNA vaccination with plasmids expressing chimeric hepadnavirus surface antigens was explored to determine B cell epitopes on the surface antigens of woodchuck hepatitis virus (WHsAg). A series of chimeric genes consisting of complementary fragments of WHsAg and hepatitis B virus surface antigens (HBsAg) was constructed. These plasmids expressed the following: (i) middle chimeric surface antigens (MCSAgs), including pre-S2 region and small surface antigens; (ii) small chimeric surface antigens (CSAgs); (iii) a mutated WHsAg with two amino acid substitutions, the Leu 136 to Thr and Ala 140 to Asp, within the central immunogenic region. The mutated region from amino acid 135 to 143 within WHsAg mimics the second loop of the HBsAg a-determinant. MCSAgs and CSAgs were expressed in transiently transfected mammalian cells and were reactive to anti-HBsAg and anti-WHsAg, as shown by indirect immunofluorescence staining and ELISA. Vaccination with plasmids encoding MCSAgs induced strong antibody responses to the pre-S2 region. Anti-pre-S2 antibodies were directed to a linear, immunodominant region within the amino-terminal region of the pre-S2 region and were able to precipitate serum WHsAg. Vaccinations with the plasmids expressing the CSAgs led to the conclusion that an extended region aa 116-169 of WHsAg, analogous to the HBsAg a-determinant, was sufficient for the induction of anti-WHsAg antibodies. The mutated WHsAg with the second loop of the HBsAg a-determinant efficiently induced anti-WHsAg antibodies, but also a low titer of anti-HBsAg. Thus, multiple B cell epitopes of a linear and conformational nature are present on WHsAg. We presented an efficient and broadly applicable strategy for analysis of complex immunogenic determinants of natural or mutated viral antigens.

Amino Acid Sequence↗
PubMed2002

Chimeric ecotropic MLV envelope proteins that carry EGF receptor-specific ligands and the Pseudomonas exotoxin A translocation domain to target gene transfer to human cancer cells.

Redirecting retroviral vector transduction simply by insertion of a ligand into the envelope (Env) protein has met with several obstacles. For example, virions targeted to epidermal growth factor receptor (EGFR), after receptor binding, rapidly traffic to the lysosomes, where they are degraded. Exotoxin A of Pseudomonas aeruginosa has the ability to translocate from endosomes to the cytoplasm by means of a translocation domain (TLD). We generated a series of chimeric Env proteins of Moloney murine leukemia virus containing EGFR ligands, where TLD was inserted into different regions. These chimeric proteins were successfully produced, if the translocation domain was not located at the immediate N-terminus of Env. The ability to transduce murine cells via the ecotropic receptor varied but correlated with the amount of Env proteins incorporated into the virions. Chimeric vector particles could bind to EGFR, demonstrating the functional exposure of the peptide ligand. However, transduction of human cells expressing EGFR but not the ecotropic receptor by virions carrying the chimeric protein was not observed.

ADP Ribose Transferases↗
PubMed2002

Chimeric human papilloma virus-simian/human immunodeficiency virus virus-like-particle vaccines: immunogenicity and protective efficacy in macaques.

Vaccines to efficiently block or limit sexual transmission of both HIV and human papilloma virus (HPV) are urgently needed. Chimeric virus-like-particle (VLP) vaccines consisting of both multimerized HPV L1 proteins and fragments of SIV gag p27, HIV-1 tat, and HIV-1 rev proteins (HPV-SHIV VLPs) were constructed and administered to macaques both systemically and mucosally. An additional group of macaques first received a priming vaccination with DNA vaccines expressing the same SIV and HIV-1 antigens prior to chimeric HPV-SHIV VLP boosting vaccinations. Although HPV L1 antibodies were induced in all immunized macaques, weak antibody or T cell responses to the chimeric SHIV antigens were detected only in animals receiving the DNA prime/HPV-SHIV VLP boost vaccine regimen. Significant but partial protection from a virulent mucosal SHIV challenge was also detected only in the prime/boosted macaques and not in animals receiving the HPV-SHIV VLP vaccines alone, with three of five prime/boosted animals retaining some CD4+ T cells following challenge. Thus, although some immunogenicity and partial protection was observed in non-human primates receiving both DNA and chimeric HPV-SHIV VLP vaccines, significant improvements in vaccine design are required before we can confidently proceed with this approach to clinical trials.

Animals↗
PubMed2002

Chimeric arteriviruses generated by swapping of the M protein ectodomain rule out a role of this domain in viral targeting.

Arteriviruses are enveloped, positive-strand RNA viruses for which the two major envelope proteins GP(5) and M occur as disulfide-linked heterodimers. These were assumed to serve the viral targeting functions, but recent ectodomain swapping studies with equine arteritis virus (EAV) indicate that the GP(5) protein does not determine arteriviral tropism. Here, we focused on the short, 13- to 18-residue ectodomain of the M protein. Using an infectious cDNA clone of the Lelystad virus isolate of porcine reproductive and respiratory syndrome virus (PRRSV), we substituted the genomic sequence encoding the M ectodomain by that of murine lactate dehydrogenase-elevating virus, EAV, and the US PRRSV-isolate, VR2332. Viable viruses with a chimeric M protein were obtained in all three cases, but for the latter two only after removal of the genomic overlap between the M and GP(5) genes. Characterization of the chimeric viruses revealed that they could be distinguished immunologically from wild-type virus, that they were genetically stable in vitro, but that they were impaired in their growth, reaching lower titers than the parental virus. The latter appeared to be due to an increased particle-to-infectivity ratio of the chimeric virus particles. Interestingly, the chimeric viruses had retained their ability to infect porcine cells and had not acquired tropism for cells susceptible to the viruses from which the foreign ectodomains were derived. We conclude that the surface structures composed by the arterivirus M and GP(5) ectodomains do not determine viral tropism.

Amino Acid Sequence↗
PubMed1994

Control of gene expression in plant cells using a 434:VP16 chimeric protein.

The herpes simplex virus type 1 VP16 polypeptide is a potent trans-activator of viral gene expression. We have tested the ability of the VP16 activation domain to activate gene expression in plant cells. A plasmid encoding a translational fusion between the full-length 434 repressor and the C-terminal 80 amino acids of VP16, was constructed. When expressed in Escherichia coli, the chimeric protein binds efficiently to 434-binding motifs (operators). For expression in plant cells, the chimeric activator gene was placed between the cauliflower mosaic virus (CaMV) 35S promoter and nos terminator sequences in a pUC-based plasmid. The 434 operators were placed upstream of a minimal CaMV 35S promoter linked to the E. coli gus reporter gene. This reporter-expression cassette was then incorporated into the same plasmid as the 434 cI/VP16 activator-expression cassette. Two control plasmids were also constructed, one encoding the 434 protein with no activator domain and the second a chimeric activator with no DNA-binding domain. The chimeric activator was tested for its ability to activate gene expression in a tobacco protoplast transient assay system. Results are presented to show that we can obtain in plant cells significant activation of gene expression that is dependent on both DNA-binding and the presence of the activator domain.

Base Sequence↗
PubMed1992

Comparative efficiencies of bispecific F(ab'gamma)2 and chimeric mouse/human IgG antibodies in recruiting cellular effectors for cytotoxicity via Fc gamma receptors.

The three forms of Fc gamma receptor carried by monocytes (Fc gamma RI, II) and natural killer (NK) cells (Fc gamma RIII) are all capable of mediating cell lysis. Here we compare the use of F(ab'gamma)2 bispecific antibodies, specifically targetting individual Fc gamma R, and chimeric IgG mouse/human antibodies which are capable of targetting all Fc gamma R, for their ability to mediate target cell destruction. The derivatives are prepared by linking hinge sulphydryl residues via tandem thioether bonds, using a bismaleimide crosslinker: Fab' from an anti-Fc gamma R mAb linked to Fab' from a common anti-target mAb (BsAb), or Fab' from the common anti-target mouse antibody linked to human Fc gamma (FabFc or bisFabFc). All the derivatives targetting chick red blood cells gave efficient lysis, although different effector cell donors yielded differences in both the lytic levels achieved and the comparative efficiencies of derivatives. In contrast, significant lysis of the guinea pig lymphoblastic leukaemia, L2C, regularly resulted only via the anti-Fc gamma RIII BsAb and the chimeric derivatives. These results suggest that the chimeric, Fc-containing derivatives mediate tumour cell lysis principally through Fc gamma RIII on NK cells. This is in contrast to the situation with the chick red blood cells where the chimeric derivatives appear capable of lysing erythrocytes by utilizing either monocytes or NK cells, because significant (approximately 50%) lysis occurred with effector cell populations magnetically depleted through either Fc gamma RII or Fc gamma RIII. A major difference between these two types of antibody derivative was their ability to function in the presence of high concentrations of normal human Fc gamma. The lysis mediated by BsAb reactive with Fc gamma RI or II was unaffected by the presence of human Fc gamma at 2.5 mg/ml (a concentration comparable with that yielded by IgG in plasma) whereas the BsAb recognizing Fc gamma RIII and all the Fc-containing derivatives were completely inhibited.

Animals↗
PubMed1991

Chimeric antibody: potential applications for drug delivery and immunotherapy.

Antibodies, because of their inherent specificity, seem ideal agents for recognizing and destroying malignant cells. When monoclonal antibodies became available, they appeared ideal candidates for use as anti-cancer drugs. However, monoclonal antibodies as currently constituted still have certain inherent limitations. Transfectomas provide an approach to overcoming some of these limitations. Genetically engineered antibodies can be expressed following gene transfection into lymphoid cells. One of the major advantages of expressing genetically engineered antibodies, is that one is not limited to using antibodies as they occur in nature. In particular, non-immunoglobulin sequences can be joined to antibody sequences creating multi-functional chimeric antibodies. Creation of a family of multi-functional chimeric antibodies with a growth factor joined to a combining specificity may be useful in targeting therapy to malignant cells and delivering drugs into specific locales in the human body. Presence of the growth factor may facilitate transcytosis of chimeric antibody across the blood-brain barrier using growth factor receptors. These novel chimeric antibodies constitute a new family of immunotherapeutic molecules for cancer therapy.

Animals↗
PubMed1996

Combined immunophenotyping and FISH with sex chromosome-specific DNA probes for the detection of chimerism in epidermal Langerhans cells after sex-mismatched bone marrow transplantation.

Langerhans cells (LC) of the skin represent bone marrow-derived dendritic antigen-presenting cells and are therefore important in pathophysiological processes such as rejection, graft-versus-host disease, and graft-versus-leukemia-reaction after bone marrow transplantation (BMT). For understanding of these diseases, the evaluation of the chimeric status of LC following BMT is of great interest. To analyze the sex chromosome constitution of LC in the skin, we established a modified and refined technique of combined immunophenotyping and fluorescence in situ hybridization (FISH) and investigated frozen sections of skin biopsies from nine patients after allogeneic sex-mismatched BMT and of two healthy donors for control. LC were specifically labeled using a fluorescent CD1 a antibody and hybridized simultaneously with X and Y chromosome-specific DNA probes. The results of this practical application on nine leukemia patients show the appearance of donor-type LC and the persistence of host-type LC at various times (36 up to 1395 days) after sex-mismatched BMT. Complete chimerism of LC could not be detected in any case. The frequency of recipient-specific LC ranged from 7% to 92% and showed no correlation with time postgrafting. We conclude from our results of 1461 analyzed LC that combined immunophenotyping and interphase cytogenetic analysis by FISH is the method of choice for the assessment of chimerism in a particular cell type after sex-mismatched BMT. Its practical application on other tissues affected by BMT-related pathophysiological processes reveals further knowledge of the time-dependent course of chimeric patterns after BMT.

Acute Disease↗
PubMed1999

Linker-based GnRH-PE chimeric proteins inhibit cancer growth in nude mice.

Since the number of cancer-related deaths has not decreased in recent years, major efforts are being made to find new drugs for cancer treatment. In this report we introduce the gonadotropin releasing hormone-Pseudomonas exotoxin (GnRH-PE) based chimeric proteins L-GnRH-PE66 and L-GnRH-PE40. These proteins are composed of a GnRH moiety attached to modified forms of Pseudomonas exotoxin via a polylinker (gly4ser)2. The chimeric proteins L-GnRH-PE66 and L-GnRH-PE40 have the ability to target and kill adenocarcinoma cell lines in vitro, whereas non-adenocarcinoma cell lines are not affected. We demonstrate that L-GnRH-PE66 and L-GnRH-PE40 efficiently inhibit cancer growth. Nude mice were injected subcutaneously with the SW-48 adenocarcinoma cell line to produce xenograft tumours. When the tumours were established and visible, the animals were injected with chimeric proteins for 10 days. At the end of this period, a reduction of up to 3-fold in tumor size was obtained in the treated mice, as compared with the control group, which received equivalent amounts of GnRH; the difference was even greater 13 days after termination of treatment. Thus, the chimeric proteins L-GnRH-PE66 and L-GnRH-PE40 are promising candidates for treatment of a variety of adenocarcinomas and their use in humans should be considered.

ADP Ribose Transferases↗
PubMed2004

Construction of chimeric cytosolic fructose-1,6-bisphosphatases by insertion of a chloroplastic redox regulatory cluster.

In order to transform cytosolic fructose-1,6-bisphosphatases (FBPase)(EC 3.1.3.11) into potential reductively-modulated chloroplast-type enzymes, we have constructed four chimeric FBPases, which display structural viability as deduced by previous modelling. In the X1-type BV1 and HL1 chimera the N-half of cytosolic sugar beet (Beta vulgaris L.) and human FBPases was fused with the C-half of the pea (Pisum sativum L.) chloroplast enzyme, which carries the cysteine-rich light regulatory sequence. In the X2-type BV2 and HL2 chimera this regulatory fragment was inserted in the corresponding site of the sugar beet cytosolic and human enzymes. Like the plant cytosolic FBPases, the chimeric enzymes show a low rise of activity by dithiothreitol. Both BV1 and BV2, but not HL1 and HL2, display a negligible activation by Trx f, but neither of them by Trx m. Antibodies raised against the pea chloroplast enzyme showed a positive reaction against the four chimeric FBPases and the human enzyme, but not against the sugar beet one. The four chimera display typical kinetics of cytosolic FBPases, with Km values in the 40-140 microM range. We conclude the existence of a structural capacity of cytosolic FBPases for incorporating the redox regulatory cluster of the chloroplast enzyme. However, the ability of these chimeric FBPases for an in vitro redox regulation seems to be scarce, limiting their use from a biotechnology standpoint in in vivo regulation of sugar metabolism.

Amino Acid Sequence↗
PubMed2000

Evaluation of chimerism in DNA samples by PCR amplification of D1S80 with detection by capillary electrophoresis.

BACKGROUND: Analysis of the relative amounts of donor and recipient DNA in bone marrow after bone marrow transplantation is frequently used to determine the status of the transplant. We studied the performance of an assay to quantify chimerism based on amplification of the D1S80 variable number tandem repeat marker by PCR with detection of PCR products by capillary electrophoresis (CE). METHODS AND RESULTS: Samples from potential bone marrow donors and recipients were analyzed separately and in mixtures to simulate various degrees of chimerism from 10% to 90% and subjected to PCR/CE analysis. There was excellent agreement between the measured and known relative proportions of DNA components in chimeric samples. The lower limit of sensitivity for detection of chimerism was 1%; between-runs coefficients of variation were <5%. CONCLUSIONS: Amplification of the D1S80 minisatellite by PCR with CE detection is a reliable method for determination of the relative contribution of different DNAs in mixed samples. This method is fast, quantitative, and extremely reproducible.

Alleles↗
PubMed2002

Minimal conditioning required in a murine model of T cell depletion, thymic irradiation and high-dose bone marrow transplantation for the induction of mixed chimerism and tolerance.

In a recently developed murine model for the induction of mixed chimerism and tolerance, hosts are treated with T cell depleting monoclonal antibodies (TCD mAbs; days -5, -1 and +7), thymic irradiation (TI) (7 Gy), and a high dose of fully allogeneic bone marrow cells (BMC, 200 x 10(6)). To find the minimum amount of each treatment required for success with this approach, we treated groups with (1) a lower dose of TI (3.5 Gy), (2) fewer BMC (100 x 10(6)), (3) no TI, (4) no TI plus additional TCD mAbs on day +14, or (5) fewer injections of TCD mAbs (day -5 only). Chimerism was followed by flow cytometry (FCM), and tolerance was assessed by skin grafting. Without TI, no long-term chimerism or tolerance could be induced, even when an additional dose of TCD mAbs was administered on day +14. A reduction in the dose of either BMC or TI led to substantially reduced effectiveness, as demonstrated by lower levels of chimerism and poorer donor skin graft survival. However, the dose of TCD mAbs and hence the duration of recipient T cell depletion could be safely reduced and thus the potential toxicity of the conditioning regimen lowered.

Animals↗
PubMed2004

Targeting of tumor cells by lymphocytes engineered to express chimeric receptor genes.

Adoptive cellular immunotherapy of cancer has been limited to date mostly due to the poor immunogenicity of tumor cells, the immunocompromised status of cancer patients in advanced stages of their disease, and difficulties in raising sufficient numbers of autologous tumor-specific T lymphocytes. On the other hand, the slow tumor penetration and short half-life of exogenously administered tumor-specific monoclonal antibodies have provided major obstacles for an effective destruction of tumor cells by the humoral effector arm of the immune system. Attempts to improve the efficacy of adoptive cellular cancer immunotherapy have led to the development of novel strategies that combine advantages of T cell-based (i.e., efficient tumor penetration, cytokine release and cytotoxicity) and antibody-based (high specificity for tumor-associated antigens) immunotherapy by grafting cytotoxic T lymphocytes (CTLs) with chimeric receptors composed of antibody fragments (which recognize tumor-cell antigens) and a cellular activation motif. Antigen recognition is therefore not restricted by major histocompatibility genes, as the physiological T-cell receptor, but rather is directed to native cell surface structures. Since the requirements of major histocompatibility complex (MHC) restriction in the interaction of effector cells with target cells are bypassed, the tumor cell-binding of CTLs grafted with chimeric receptors is not affected by down-regulation of HLA class I antigens and by defects in the antigen-processing machinery. Ligand binding by the chimeric receptor triggers phosphorylation of immunoglobulin tyrosine activation motifs (ITAMs) in the cytoplasmic region of the molecule and this activates a signaling cascade that is required for the induction of cytotoxicity, cytokine secretion and proliferation. Here, the authors discuss the potential of lymphocytes grafted with chimeric antigen receptors in the immunotherapy of malignant disease.

Amino Acid Motifs↗