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Kenneth L Rock

Publications and source records attributed to Kenneth L Rock.

At least 19 recordsLinked to original sources
PubMed2005

Priming of T cells by exogenous antigen cross-presented on MHC class I molecules.

Cross-presentation is the process whereby bone-marrow-derived antigen-presenting cells acquire, process and present exogenous antigen as peptides bound to MHC class I molecules to CD8(+) T cells. Professional antigen-presenting cells acquire cell-associated antigen predominantly in the form of protein, then process and present antigenic peptides on their surface MHC class I molecules via several mechanisms and efficiently cross-prime naïve CD8(+) T cells in vivo. This pathway is of considerable interest because it has an important role in the immune surveillance of tissues for pathogens and cancers.

Animals↗
PubMed2005

Leucine aminopeptidase is not essential for trimming peptides in the cytosol or generating epitopes for MHC class I antigen presentation.

To detect viral infections and tumors, CD8+ T lymphocytes monitor cells for the presence of antigenic peptides bound to MHC class I molecules. The majority of MHC class I-presented peptides are generated from the cleavage of cellular and viral proteins by the ubiquitin-proteasome pathway. Many of the oligopeptides produced by this process are too long to stably bind to MHC class I molecules and require further trimming for presentation. Leucine aminopeptidase (LAP) is an IFN-inducible cytosolic aminopeptidase that can trim precursor peptides to mature epitopes and has been thought to play an important role in Ag presentation. To examine the role of LAP in generating MHC class I peptides in vivo, we generated LAP-deficient mice and LAP-deficient cell lines. These mutant mice and cells are viable and grow normally. The trimming of peptides in LAP-deficient cells is not reduced under basal conditions or after stimulation with IFN. Similarly, there is no reduction in presentation of peptides from precursor or full-length Ag constructs or in the overall supply of peptides from cellular proteins to MHC class I molecules even after stimulation with IFN. After viral infection, LAP-deficient mice generate normal CTL responses to seven epitopes from three different viruses. These data demonstrate that LAP is not an essential enzyme for generating most MHC class I-presented peptides and reveal redundancy in the function of cellular aminopeptidases.

Amino Acid Sequence↗
PubMed2005

Both dendritic cells and macrophages can stimulate naive CD8 T cells in vivo to proliferate, develop effector function, and differentiate into memory cells.

The generation of T cell immunity requires the acquisition and presentation of Ag on bone marrow-derived APCs. Dendritic cells (DC) are believed to be the most potent bone marrow-derived APCs, and the only ones that can stimulate naive T cells to productively respond to Ags. Because macrophages (Mphi) are bone marrow-derived APCs that are also found in tissues and lymphoid organs, can acquire and present Ag, and can express costimulatory molecules, we have investigated their potential to stimulate primary T cell responses in vivo. We find that both injected Mphi and DCs can migrate from peripheral tissues or blood into lymphoid organs. Moreover, injection of peptide-pulsed Mphi or DCs into mice stimulates CD8 T cells to proliferate, express effector functions including cytokine production and cytolysis, and differentiate into long-lived memory cells. Mphi and DCs stimulate T cells directly without requiring cross-presentation of Ag on host APCs. Therefore, more than one type of bone marrow-derived APC has the potential to prime T cell immunity. In contrast, another bone marrow-derived cell, the T lymphocyte, although capable of presenting Ag and homing to the T cell areas of lymphoid organs, is unable to stimulate primary responses. Because Mphi can be very abundant cells, especially at sites of infection and inflammation, they have the potential to play an important role in immune surveillance and the initiation of T cell immunity.

Adoptive Transfer↗
PubMed2005

Characterization of a proapoptotic antiganglioside GM2 monoclonal antibody and evaluation of its therapeutic effect on melanoma and small cell lung carcinoma xenografts.

Monoclonal antibodies have begun to show great clinical promise for the treatment of cancer. Antibodies that can directly affect a tumor cell's growth and/or survival are of particular interest for immunotherapy. Previously, we described monoclonal antibody DMF10.62.3 that had antiproliferative and proapoptotic effects when it bound an antigen of unknown identity on tumor cells in vitro. In this report, we determined that DMF10.62.3 and a clonally related antibody DMF10.167.4 recognize the ganglioside GM2. These antibodies react with a GM2 epitope that is expressed on a large number of tumor cell lines, including human melanoma and small cell lung carcinoma, but not on normal primary lines or most normal tissues. Interestingly, this pattern of cellular reactivity is distinct from that reported for other previously described GM2 antibodies, a difference that is presumably due to DMF10.167.4's binding to a unique GM2-associated epitope. Additional characterization of DMF10.167.4 revealed that this antibody was able to induce apoptosis and/or block cellular proliferation when cultured in vitro with the human Jurkat T lymphoma, CHL-1 melanoma, and SBC-3 small cell lung carcinoma lines. In vivo, DMF10.167.4 antibody was well tolerated in mice and did not detectably bind to or damage normal tissues. However, this antibody was able to prevent murine E710.2.3 lymphoma, human CHL-1 melanoma, and SBC-3 small cell lung carcinoma lines from establishing tumors in vivo and blocked progression of established CHL-1 and SBC-3 tumors in vivo. Therefore, monoclonal antibody DMF10.167.4 has immunotherapeutic potential.

Animals↗
PubMed2005

A mutant cell with a novel defect in MHC class I quality control.

COS7 (African Green Monkey kidney) cells stably transfected with the mouse MHC class I allele H-2K(b) were mutagenized, selected for low surface expression of endogenous MHC class I products, and subcloned. A mutant cell line, 4S8.12, expressing very low surface MHC class I (approximately 5% of parental levels) was identified. This cell line synthesized normal levels of the MHC class I H chain and beta(2)-microglobulin, as well as normal levels of TAP, tapasin, GRP78, calnexin, calreticulin, ERp57, and protein disulfide isomerase. Full-length OVA was processed to generate presented H-2K(b)-SIINFEKL complexes with equal efficiency in wild-type and mutant cells, demonstrating that proteasomes, as well as TAP and tapasin, functioned normally. Therefore, all the known components of the MHC class I Ag presentation pathway were intact. Nevertheless, primate (human and monkey) MHC class I H chain and beta(2)-microglobulin failed to associate to form the normal peptide-receptive complex. In contrast, mouse H chains associated with beta(2)-microglobulin normally and bound peptide at least as well as in wild-type cells. The 4S8.12 cells provide strong genetic evidence for a novel component in the MHC class I pathway. This as-yet unidentified gene is important in early assembly of primate, but not mouse, MHC class I complexes.

Animals↗
PubMed2005

KOC (K homology domain containing protein overexpressed in cancer): a novel molecular marker that distinguishes between benign and malignant lesions of the pancreas.

KOC (K homology domain containing protein overexpressed in cancer) is a novel oncofetal RNA-binding protein highly expressed in pancreatic carcinomas. Recently, Corixa Corporation developed a monoclonal antibody specific for KOC that can be used with standard immunohistochemical techniques. The purposes of this study were 1) to assess KOC mRNA expression in pancreatic carcinoma, 2) to determine the pattern of KOC immunoexpression among benign, borderline, and malignant pancreatic epithelial lesions, and 3) to evaluate the utility of the KOC antibody in distinguishing between these entities. mRNA was isolated from fresh pancreatic tissues (19 carcinomas, 2 normal pancreas, 1 chronic pancreatitis) and amplified using standard RT-PCR techniques. Fifteen of 19 (79%) carcinomas overexpressed KOC mRNA relative to non-neoplastic tissue samples and expression increased progressively with tumor stage: the mean copy number of KOC mRNA transcripts was 1.5, 11.1, 31, and 28 for stage I, II, III, and IV carcinomas, respectively, compared with 0.9 and 1 for normal pancreatic tissue and chronic pancreatitis, respectively. Immunostains using the KOC antibody were performed on 50 surgical resection specimens (38 invasive adenocarcinomas, 3 intraductal papillary-mucinous neoplasms, 2 mucinous cystic neoplasms, 7 chronic pancreatitis). KOC staining was present in 37 of 38 (97%) carcinomas: the staining reaction was moderate or strong in 36 of 38 (94%) and present in >50% of the tumor cells in 35 of 38 (92%) cases. Severe dysplasia of the ductal epithelium, present in 19 foci of intraductal papillary mucinous carcinoma, mucinous cystadenocarcinoma, and grade 3 pancreatic intraepithelial neoplasia (PanIN3) showed strong or moderate staining in 15 (79%) cases, whereas foci of mild and moderate dysplasia (intraductal papillary-mucinous neoplasms and mucinous cystic neoplasms with adenoma and/or moderate dysplasia, PanIN1, and PanIN2) were uniformly negative for this marker in 25 and 22 cases, respectively. In the normal pancreas, weak background staining of acini was present in 12 of 50 (24%) cases but was easily distinguishable from the type of staining identified in neoplastic epithelium, and benign ducts and ductules were negative in all cases. Four of 38 (11%) foci of chronic pancreatitis, present in the 7 resections performed for chronic pancreatitis as well as 31 foci of peritumoral chronic pancreatitis, showed weak staining in <10% of the ductules. We conclude that KOC is a sensitive and specific marker for carcinomas and high-grade dysplastic lesions of the pancreatic ductal epithelium. Therefore, immunostains directed against KOC may be of diagnostic utility in the evaluation of pancreatic lesions, particularly when biopsy material is limited.

Animals↗
PubMed2005

Cross-presentation: underlying mechanisms and role in immune surveillance.

It was originally thought that a cell's major histocompatibility complex (MHC) class I molecules presented peptides derived exclusively from proteins synthesized by the cell itself. However, in some circumstances, antigens from the extracellular environment can be presented on MHC class I molecules and stimulate CD8(+) T-cell immunity, a process termed cross-presentation. Cross-presentation was originally discovered as an obscure phenomenon in transplantation immunity. However, it is now clear that it is a major mechanism by which the immune system monitors tissues and phagocytes for the presence of foreign antigen. Cross-presentation is the only pathway by which the immune system can detect and respond to viral infections or mutations that exclusively occur in parenchymal cells rather than in bone marrow-derived antigen-presenting cells (APCs). Professional APCs, such as dendritic cells, are the principal cells endowed with the capacity to cross-present antigens. In this process, the APCs acquire proteins from other tissue cells through endocytic mechanisms, especially phagocytosis or macropinocytosis. The internalized antigen can then be processed through at least two different mechanisms. In one pathway, the antigen is transferred from the phagosome into the cytosol, where it is hydrolyzed by proteasomes into oligopeptides that are then transported by the transporter associated with antigen processing to MHC class I molecules in the endoplasmic reticulum or phagosomes. In a second pathway, the antigen is cleaved into peptides by endosomal proteases, particularly cathepsin S, and bound by class I molecules probably in the endocytic compartment itself. Depending on the nature of the antigen, one or both of these pathways can contribute to cross-presentation in vivo. The outcome of cross-presentation can be either tolerance or immunity. Which of these outcomes occurs is thought to depend on whether antigens are acquired by themselves alone, leading to tolerance, or with immunostimulatory signals, leading to immunity. One source of such signals is from dying cells that release immunostimulatory 'danger' signals that promote the generation of immunity to their cellular antigens. In addition to the critical role of cross-presentation in normal immune physiology, this pathway has considerable potential for being exploited for developing subunit vaccines that elicit both CD4(+) and CD8(+) T-cell immunity.

Animals↗
PubMed2004

Mammaglobin and CRxA-01 in pleural effusion cytology: potential utility of distinguishing metastatic breast carcinomas from other cytokeratin 7-positive/cytokeratin 20-negative carcinomas.

BACKGROUND: The most common causes of malignant pleural effusions in women are metastatic lung carcinomas and breast carcinomas. It is often very difficult to distinguish between breast carcinomas and other metastatic carcinomas when they share a similar morphology and a similar cytokeratin profile (CK7-positive/CK20-negative [CK7+/CK20-]). To better differentiate between metastatic mammary carcinomas and other metastatic carcinomas in pleural effusion cytology, the authors studied the potential use of a novel antibody, CRxA-01, which was identified by a cDNA subtraction library, together with a well characterized antibody against mammaglobin. METHODS: A computer search for patients with malignant pleural effusion specimens between January 1992 and November 2002 generated 228 patients, 71 of whom had cell block material and a known clinical history. Primary malignancies among these patients included 20 breast carcinomas, 32 lung carcinomas, 4 endometrial carcinomas, 9 ovarian carcinomas, 4 gastrointestinal carcinomas, and 2 genitourinary carcinomas. All specimens were immunostained with anti-CK7, CK20, CRxA-01, and mammaglobin antibodies. Only CK7-positive/CK20-negative (CK7+/CK20-) specimens were included in the current study, and only definitive membranous staining for CRxA-01 and cytoplasmic staining for mammaglobin were considered to be positive. RESULTS: For patients with metastatic breast carcinomas, mammaglobin was positive in 11 of 20 (55%) tissue specimens and CRxA-01 was positive in 12 of 20 (60%) tissue specimens. When CRxA-01 and mammaglobin were used together, 16 of 20 (80%) tissue specimens were positive for mammaglobin or/and CRxA-01 antibodies. This staining pattern was not seen for tissue specimens from patients with other metastatic carcinomas. Two of 4 (50%) uterine carcinoma specimens and 6 of 9 (67%) ovarian carcinoma specimens were positive for CRxA-01 only. CONCLUSIONS: CRxA-01 and mammaglobin were expressed in most metastatic breast carcinoma specimens. Other CK7+/CK20- carcinoma specimens did not express mammaglobin and showed weak or negative staining for CRxA-01. When used together, CRxA-01 and mammaglobin greatly improved the sensitivity and specificity for the detection of metastatic breast carcinoma in pleural effusion specimens.

Antibodies, Neoplasm↗
PubMed2004

Natural endogenous adjuvants.

It has long been known that immunization with a protein by itself is often not sufficient to stimulate immunity, and may instead induce tolerance. To elicit productive immune responses exogenous adjuvants need to be co-injected with an antigen. One important class of adjuvants are the unique (non-mammalian) components of microbes. It is now believed that an adjuvant is required for immunity because the immune system evolved to respond to dangerous situations such as infections, and the presence of an adjuvant is the mechanism used to identify these situations. However, there are some circumstances where immune responses are generated in the apparent absence of any microbial or other exogenous adjuvant. Such situations include immune responses to transplants, tumors, autoimmunity and possibly certain viral infections. It has been postulated that in these situations the danger signals come from endogenous adjuvants that are released from dying cells. There is abundant evidence that dead cells are immunogenic, and recently it has been shown that cells contain endogenous adjuvant activities that are released after death. Some actual and putative endogenous adjuvants, such as monosodium urate and heat shock proteins, have been identified and there are others whose identities are not yet known. The potential biological roles of this class of adjuvants are discussed.

Adjuvants, Immunologic↗
PubMed2004

Cellular protein is the source of cross-priming antigen in vivo.

Cross-priming is essential for generating cytotoxic T lymphocytes to viral, tumor, and tissue antigens that are expressed exclusively in parenchymal cells. In this process, the antigen-bearing parenchymal cells must somehow transfer their antigens to bone marrow-derived professional antigen-presenting cells. Although intact proteins, small peptides, or peptide-heat shock protein complexes can all be acquired and presented by antigen-presenting cells, the physiologically relevant form of antigen that is actually transferred from parenchymal cells and cross-presented in vivo is unknown and controversial. To address this issue we have investigated the ability of fibroblasts stably expressing chicken ovalbumin constructs targeted to different subcellular compartments to cross-prime cytotoxic T lymphocytes. Although these transfectants generated similar amounts of the immunogenic ovalbumin peptide, their cross-priming activity differed markedly. Instead, the cells cross-priming ability correlated with their steady-state levels of ovalbumin protein and/or the physical form/location of the protein. Moreover, in subcellular fractionation experiments, the cross-priming activity colocalized with antigenic protein. In addition, depletion of intact protein antigen from these cell fractions eliminated their cross-priming activity. In contrast, the major heat shock protein candidates for cross-presentation were separable from the cell's main sources of cross-priming antigen. Therefore, cellular proteins, rather than peptides or heat shock protein/peptide complexes, are the major source of antigen that is transferred from antigen-bearing cells and cross-presented in vivo.

Animals↗
PubMed2004

Important role of cathepsin S in generating peptides for TAP-independent MHC class I crosspresentation in vivo.

The immune system detects viral infections and mutations in parenchymal cells when antigens from these cells are crosspresented on MHC class I molecules of professional antigen-presenting cells (APC). Exogenous antigens are crosspresented through TAP-dependent (cytosolic) or poorly understood TAP-independent (vacuolar) pathways. The TAP-independent pathway is blocked by the cysteine protease inhibitor, leupeptin, but not by proteasome inhibitors, which is opposite to the effects of these agents on the TAP-dependent pathway. Dendritic cells lacking the cysteine protease cathepsin S lack the TAP-independent pathway. Mice whose APC lack cathepsin S have reduced crosspriming to particulate and cell-associated antigens, as well as to influenza virus. Cathepsin S-deficient phagosomes generate a class I-presented peptide poorly. In contrast, cathepsin S-sufficient phagosomes and recombinant cathepsin S produce the mature epitope. Therefore, cathepsin S plays a major role in generating presented peptides for the vacuolar pathway of crosspresentation, and this mechanism is active in vivo.

Animals↗
PubMed2004

Post-proteasomal antigen processing for major histocompatibility complex class I presentation.

Peptides presented by major histocompatibility complex class I molecules are derived mainly from cytosolic oligopeptides generated by proteasomes during the degradation of intracellular proteins. Proteasomal cleavages generate the final C terminus of these epitopes. Although proteasomes may produce mature epitopes that are eight to ten residues in length, they more often generate N-extended precursors that are too long to bind to major histocompatibility complex class I molecules. Such precursors are trimmed in the cytosol or in the endoplasmic reticulum by aminopeptidases that generate the N terminus of the presented epitope. Peptidases can also destroy epitopes by trimming peptides to below the size needed for presentation. In the cytosol, endopeptidases, especially thimet oligopeptidase, and aminopeptidases degrade many proteasomal products, thereby limiting the supply of many antigenic peptides. Thus, the extent of antigen presentation depends on the balance between several proteolytic processes that may generate or destroy epitopes.

Animals↗
PubMed2003

Molecular identification of a danger signal that alerts the immune system to dying cells.

In infections, microbial components provide signals that alert the immune system to danger and promote the generation of immunity. In the absence of such signals, there is often no immune response or tolerance may develop. This has led to the concept that the immune system responds only to antigens perceived to be associated with a dangerous situation such as infection. Danger signals are thought to act by stimulating dendritic cells to mature so that they can present foreign antigens and stimulate T lymphocytes. Dying mammalian cells have also been found to release danger signals of unknown identity. Here we show that uric acid is a principal endogenous danger signal released from injured cells. Uric acid stimulates dendritic cell maturation and, when co-injected with antigen in vivo, significantly enhances the generation of responses from CD8+ T cells. Eliminating uric acid in vivo inhibits the immune response to antigens associated with injured cells, but not to antigens presented by activated dendritic cells. Our findings provide a molecular link between cell injury and immunity and have important implications for vaccines, autoimmunity and inflammation.

3T3 Cells↗
PubMed2003

A dietary enzyme: alpha-methylacyl-CoA racemase/P504S is overexpressed in colon carcinoma.

Epidemiological studies have shown that consumption of red meat increases the risk of developing colon cancer. An enzyme, alpha-methylacyl CoA racemase (AMACR), also known as P504S, plays an important role in peroxisomal beta-oxidation of branched-chain fatty acids from red meat and dairy products. High expression of AMACR was recently found in prostate cancer. In this study, we investigated expression of AMACR in 242 cases of colonic tumors including 176 colorectal carcinomas, 38 colon adenomas and 28 hyperplastic (non-neoplastic) polyps by immunohistochemical analysis. The mRNA levels of AMACR expression in normal and colon cancer tissues were assessed by real-time PCR. Significant up-regulation of AMACR mRNA was found in colon carcinomas compared to normal tissue. There was very low or no expression of AMACR protein in normal colon, but AMACR was highly expressed in 76 and 75% of well and moderately differentiated colon carcinomas, respectively, and in 79% of adenomas. In contrast, only 4% of hyperplastic polyps expressed AMACR. Since this enzyme is involved in the metabolism of branched-chain fatty acids from beef, milk and dairy products, our results provide important molecular information regarding a possible link between diet and development of colon cancer. AMACR may also serve as a molecular marker for colon cancers and its precursor lesions.

Adenocarcinoma↗
PubMed2003

The cytosolic endopeptidase, thimet oligopeptidase, destroys antigenic peptides and limits the extent of MHC class I antigen presentation.

Most antigenic peptides presented on MHC class I molecules are generated by proteasomes during protein breakdown. It is unknown whether these peptides are protected from destruction by cytosolic peptidases. In cytosolic extracts, most antigenic peptides are degraded by the metalloendopeptidase, thimet oligopeptidase (TOP). We therefore examined whether TOP destroys antigenic peptides in vivo. When TOP was overexpressed in cells, class I presentation of antigenic peptides was reduced. In contrast, TOP overexpression didn't reduce presentation of peptides generated in the endoplasmic reticulum or endosomes. Conversely, preventing TOP expression with siRNA enhanced presentation of antigenic peptides. TOP therefore plays an important role in vivo in degrading peptides released by proteasomes and is a significant factor limiting the extent of antigen presentation.

Animals↗
PubMed2002

An IFN-gamma-induced aminopeptidase in the ER, ERAP1, trims precursors to MHC class I-presented peptides.

Precursors to major histocompatibility complex (MHC) class I-presented peptides with extra NH2-terminal residues can be efficiently trimmed to mature epitopes in the endoplasmic reticulum (ER). Here, we purified from liver microsomes a lumenal, soluble aminopeptidase that removes NH2-terminal residues from many antigenic precursors. It was identified as a metallopeptidase named "adipocyte-derived leucine" or "puromycin-insensitive leucine-specific" aminopeptidase. However, because we localized it to the ER, we propose it be renamed ER-aminopeptidase 1 (ERAP1). ERAP1 is inhibited by agents that block precursor trimming in ER vesicles and although it trimmed NH2-extended precursors, it spared presented peptides of 8 amino acid and less. Like other proteins involved in antigen presentation, ERAP1 is induced by interferon-gamma. When overexpressed in vivo, we found that ERAP1 stimulates the processing and presentation of an antigenic precursor in the ER.

Animals↗
PubMed2002

The ER aminopeptidase ERAP1 enhances or limits antigen presentation by trimming epitopes to 8-9 residues.

Endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) appears to be specialized to produce peptides presented on class I major histocompatibility complex molecules. We found that purified ERAP1 trimmed peptides that were ten residues or longer, but spared eight-residue peptides. In vivo, ERAP1 enhanced production of an eight-residue ovalbumin epitope from precursors extended on the NH2 terminus that were generated either in the ER or cytosol. Purified ERAP1 also trimmed nearly half the nine-residue peptides tested. By destroying such nine-residue peptides in normal human cells, ERAP1 reduced the overall supply of antigenic peptides. However, after interferon-gamma treatment, which causes proteasomes to produce more NH2-extended antigenic precursors, ERAP1 increased the supply of peptides for MHC class I antigen presentation.

Animals↗