Increased frequency of CD45 negative T cells (T zone cells) in older Golden retriever dogs.

T zone lymphoma (TZL) is characterized by the clonal expansion of T cells lacking expression of the pan-leukocyte antigen CD45 (TZ cells). A strong breed predisposition is observed in Golden retrievers. This study aimed to confirm aberrant CD45 mRNA expression and determine if Golden retrievers without clinical lymphoma have an increased frequency of circulating TZ cells. Gene expression analysis on confirmed TZL cases showed a significant decrease in CD45 expression compared to normal dogs. Peripheral blood samples from senior dogs, 242 Golden retrievers and 42 non-Golden retrievers, without evidence of lymphoproliferative disease were assessed for the presence of TZ cells by flow cytometry. Thirty-one percent of Golden retrievers had TZ cells compared to 14% of non-Golden retrievers. Thirty-four percent of Golden retrievers with TZ cells had a clonal T cell receptor gamma (TRG) gene rearrangement. Interestingly, 20% of Golden retrievers without TZ cells also had a clonal TRG rearrangement. Golden retrievers may have an increased risk of TZL due to an increased frequency of TZ cells.

Clinicopathologic features of lingual canine T-zone lymphoma.

Canine T-zone lymphoma (TZL) is a subtype of T-cell lymphoma characterized by unique histologic pattern and cytomorphology, immunophenotypic loss of CD45 expression, and an indolent clinical behaviour. Dogs with TZL typically present with 1 or more enlarged lymph nodes and/or lymphocytosis. We describe a novel extranodal presentation of TZL involving the tongue. Twelve dogs with tongue masses were diagnosed with lingual TZL based on a variable combination of immunophenotyping via flow cytometry, cytology, histopathology, immunohistochemistry and/or PCR for antigen receptor rearrangement (PARR) assay. Eleven dogs exhibited concurrent lymphocytosis and/or lymph node enlargement. Three cases were initially diagnosed as plasma cell tumours based on histology alone, thereby revealing a potential diagnostic challenge. Seven dogs achieved clinical remission and 4 achieved stable disease following variable treatment, consistent with the indolent nature of typical TZL involving the lymph nodes and peripheral blood. In 1 case the TZL resulted in progressive disease and failure to respond to treatment. In this case, the TZL exhibited histologic features of a higher grade neoplasm. This case series highlights a unique presentation of TZL and identifies a new differential diagnosis for lingual neoplasia. In this study, we characterize the clinical presentation, diagnostic features and patient outcomes of 12 dogs with lingual TZL.

c-Kit Mutation and Localization Status as Response Predictors in Mast Cell Tumors in Dogs Treated with Prednisone and Toceranib or Vinblastine.

BACKGROUND: KIT inhibitors, such as toceranib (TOC), and vinblastine (VBL) have not been prospectively compared in the treatment of macroscopic mast cell tumors (MCTs). Also, it is unknown whether VBL or TOC is superior for treating MCT without c-kit mutations. HYPOTHESIS/OBJECTIVES: To determine the value of KIT genotyping and localization in treatment decisions for dogs with macroscopic MCT. We hypothesized that c-kit mutated MCT would have a better response to TOC than VBL. ANIMALS: Eighty-eight client-owned dogs with macroscopic MCT. METHODS: Prospective, randomized trial. Dogs were randomized to TOC (2.75 mg/kg EOD) or VBL (2.5 mg/m2 weekly × 4 then EOW) by KIT localization and c-kit mutation status using an adaptive randomization scheme. RESULTS: Sixty dogs were allocated to TOC and 28 to VBL. Of the dogs receiving TOC, 20% had c-kit mutations, compared to 30% receiving VBL (P = 0.74). Overall response rates were 46% (TOC) and 30% (VBL) (odds ratio = 1.56 [0.62-3.92]; P = 0.28). Median progression-free survival (PFS) for dogs receiving VBL was 78 days (7-1,521) and for TOC 95.5 (14-990); hazard ratio (HR) = 1.34 [0.72-2.50]; P = 0.36. Median overall survival (OS) was 241.5 days (10-1,521) for the VBL group and 159 (20-990) for the TOC group; HR = 0.80 ([0.45-1.41]; P = 0.44). CONCLUSIONS AND CLINICAL IMPORTANCE: Neither PFS nor OS was significantly different between treatment groups. As the proportion of dogs with c-kit mutations was not different between treatment groups in this population of dogs, c-kit mutation status did not predict treatment response.

Expression of Phosphorylated KIT in Canine Mast Cell Tumor.

Canine cutaneous mast cell tumor (MCT) is the most common canine skin tumor and exhibits variable biologic behavior. Signaling through the KIT receptor tyrosine kinase promotes cellular proliferation and survival and has been shown to play a role in MCT progression. Despite investigations into numerous biomarkers and the proposal of several grading schemas, no single marker or grading system can accurately predict outcome in canine MCT. The first aim of this study was to develop an immunohistochemical assay to measure phosphorylated KIT (pKIT) to investigate its association with 2 commonly used grading systems and other established prognostic markers for canine MCT. Thirty-four archived MCTs were evaluated for expression of pKIT and Ki-67, KIT localization, mitotic count, mutations in exons 8 and 11 in c-kit, and grading by the Patnaik and 2-tier systems. Expression of pKIT was significantly ( P < .05) correlated with the 2-tier grading scheme and c-kit mutation. Correlation approached significance ( P = .06) with Mitotic Index (MI) and Ki-67. An additional aim was to determine whether pKIT labeling provides a pharmacodynamic marker for predicting response to the receptor tyrosine kinase inhibitor toceranib (TOC). MCTs from 4 of 7 patients demonstrated a partial response to TOC. pKIT expression was assessed by immunohistochemistry in biopsies obtained before and 6 hours after the patients were treated with TOC. Reduced pKIT expression after TOC treatment was demonstrated in 3 of the 4 patients with a partial response compared to 1 of the 3 nonresponders. Collectively, these results demonstrate that immunohistochemical detection of pKIT may be a clinically relevant assay to evaluate the activation status of the major oncogenic pathway in canine MCT.

Differences in the geographic distribution of lymphoma subtypes in Golden retrievers in the USA.

The purpose of this cross-sectional study was to examine differences in the geographic distribution of two distinct subtypes of canine lymphoma (CL), B-cell lymphoma (BCL) and T-zone lymphoma (TZL), in the USA while accounting for heritable risks associated with the outcome of disease through inclusion of only one breed of dog. This study included 454 Golden retrievers and associations between geographic areas of the USA and the phenotypic variant of lymphoma were examined using multivariable logistic regression. There was a detectable difference in the geographic distribution of BCL and TZL with dogs in the Northeast [odds ratio (OR) = 3.4, 95% confidence interval (CI) = 1.6-7.0] and East North Central regions (OR = 12.1, 95%CI = 3.6-40.5) being more likely to be diagnosed with TZL as compared to dogs in the Mountain region of the USA. The finding of non-random geographic distribution of lymphoma subtypes suggest that environmental risk factors may contribute to the development of different types of CL.

Breed Distribution and Clinical Characteristics of B Cell Chronic Lymphocytic Leukemia in Dogs.

BACKGROUND: B-cell chronic lymphocytic leukemia (B-CLL) is the most common hematopoietic malignancy in humans in the developed world and the primary risk factor is genetic. Dogs also develop B-CLL, but there is no systematic description of the disease in dogs. Understanding the epidemiology of B-CLL in dogs may help practitioners recognize the disease and position the dog as a model for future genetic studies. OBJECTIVES: To describe B-CLL presentation in dogs, its clinicopathologic findings, and breed predisposition. ANIMALS: Four hundred and ninety-one dogs with B-CLL and 5,673 control dogs with suspicion of a lymphoproliferative disorder (LPD). METHODS: Retrospective cross-sectional study of dogs for which samples were submitted to the Colorado State University Clinical Immunology Laboratory for immunophenotyping between 2010 and 2014. To assess breed predilection, dogs with B-CLL were compared to those with suspicion of other LPDs using logistic regression. RESULTS: The median age was 11 years with no sex predilection. Half of the dogs presented with peripheral lymphadenopathy or splenomegaly and 26% had anemia. Eleven small-breed dogs had significantly increased odds of B-CLL. In addition, English Bulldogs had an increased risk and a unique presentation: these dogs were diagnosed at a median of 6 years and expressed lower class II MHC and CD25. CONCLUSIONS: B-cell chronic lymphocytic leukemia is overrepresented in small-breed dogs. Future genetic studies of these breeds may identify genetic risk factors. The unique presentation of English Bulldogs provides evidence of multiple forms of this disease. Additional studies are necessary to determine whether presenting signs are associated with survival.

Pulse-Administered Toceranib Phosphate Plus Lomustine for Treatment of Unresectable Mast Cell Tumors in Dogs.

BACKGROUND: Nonresectable mast cell tumors (MCT) in dogs remain a therapeutic challenge, and investigation of novel combination therapies is warranted. Intermittent administration of tyrosine kinase inhibitors (TKI) combined with cytotoxic chemotherapy may effectively chemosensitize canine MCT while decreasing cost and adverse effects associated with either agent administered as monotherapy. HYPOTHESIS/OBJECTIVES: The primary study objectives were to (1) identify the maximally tolerated dose (MTD), (2) determine the objective response rate (ORR) and (3) describe the adverse event profile of pulse-administered toceranib phosphate (TOC) combined with lomustine. ANIMALS: Forty-seven client-owned dogs with measurable MCT. METHODS: Toceranib phosphate was given PO on days 1, 3 and 5 of a 21-day cycle at a target dosage of 2.75 mg/kg. Lomustine was given PO on day 3 of each cycle at a starting dosage of 50 mg/m(2) . All dogs were concurrently treated with diphenhydramine, omeprazole, and prednisone. RESULTS: The MTD of lomustine was established at 50 mg/m(2) when combined with pulse-administered TOC; the dose-limiting toxicity was neutropenia. Forty-one dogs treated at the MTD were evaluable for outcome assessment. The ORR was 46% (4 complete response, 15 partial response) and the overall median progression-free survival (PFS) was 53 days (1 to >752 days). On multivariate analysis, variables significantly associated with improved PFS included response to treatment, absence of metastasis, and no previous chemotherapy. CONCLUSIONS AND CLINICAL IMPORTANCE: Combined treatment with pulse-administered TOC and lomustine generally is well tolerated and may be a reasonable treatment option for dogs with unresectable or metastatic MCT.

Canine T-zone lymphoma: unique immunophenotypic features, outcome, and population characteristics.

BACKGROUND: Canine T-cell lymphoma (TCL) is clinically and histologically heterogeneous with some forms, such as T-zone lymphoma (TZL), having an indolent course. Immunophenotyping is an important tool in the classification of TCL in people, and can be equally useful in dogs. HYPOTHESIS/OBJECTIVES: We hypothesized that loss of expression of the CD45 antigen is a specific diagnostic feature of TZL. ANIMALS: Twenty dogs with concurrent histology and immunophenotyping by flow cytometry were studied in depth. An additional 494 dogs diagnosed by immunophenotyping were used to characterize the population of dogs with this disease. METHODS: Lymph node biopsies from 35 dogs with TCL were classified by 2 pathologists using WHO criteria. Twenty lymph nodes were from dogs with CD45- TCL and 15 were from CD45+ TCL. The pathologists were blinded to the flow cytometry findings. Outcome information was sought for the 20 dogs with CD45- lymphoma, and population characteristics of the additional 494 dogs were described. RESULTS: All 20 CD45- cases were classified as TZL. The 15 CD45+ cases were classified as aggressive TCL and are described in an accompanying paper. TZL cases had a median survival of 637 days. Examination of 494 additional dogs diagnosed with TZL by immunophenotyping demonstrated that 40% of cases are in Golden Retrievers, are diagnosed at a median age of 10 years, and the majority have lymphadenopathy and lymphocytosis. CONCLUSIONS: TZL has unique immunophenotypic features that can be used for diagnosis.

Flow cytometric characterization and clinical outcome of CD4+ T-cell lymphoma in dogs: 67 cases.

BACKGROUND: Canine T-cell lymphoma (TCL) is conventionally considered an aggressive disease, but some forms are histologically and clinically indolent. CD4 TCL is reported to be the most common subtype of TCL. We assessed flow cytometric characteristics, histologic features when available, and clinical outcomes of CD4+ TCL to determine if flow cytometry can be used to subclassify this group of lymphomas. OBJECTIVE: To test the hypothesis that canine CD4+ T-cell lymphoma (TCL) is a homogeneous group of lymphomas with an aggressive clinical course. ANIMALS: Sixty-seven dogs diagnosed with CD4+ TCL by flow cytometry and treated at 1 of 3 oncology referral clinics. METHODS: Retrospective multivariable analysis of outcome in canine CD4+ TCL including patient characteristics, treatment, and flow cytometric features. RESULTS: The majority of CD4+ TCL were CD45+, expressed low class II MHC, and exhibited an aggressive clinical course independent of treatment regimen (median survival, 159 days). Histologically, CD4+ TCL were classified as lymphoblastic or peripheral T cell. Size of the neoplastic lymphocytes had a modest effect on both PFI and survival in this group. A small number of CD4+ TCL were CD45- and class II MHC high, and exhibited an apparently more indolent clinical course (median survival not yet reached). CONCLUSIONS AND CLINICAL IMPORTANCE: Although the majority of CD4+ TCL in dogs had uniform clinical and flow cytometric features and an aggressive clinical course, a subset had a unique immunophenotype that predicts significantly longer survival. This finding strengthens the utility of flow cytometry to aid in the stratification of canine lymphoma.

Multicenter prospective trial of hypofractionated radiation treatment, toceranib, and prednisone for measurable canine mast cell tumors.

BACKGROUND: Mast cell tumors (MCT) are common cutaneous tumors in dogs and when not amenable to surgical excision can present a therapeutic challenge. New treatment protocols for unresectable MCT are needed. HYPOTHESIS: The combination of toceranib, prednisone, and hypofractionated radiation treatment (RT) will be well tolerated and efficacious. ANIMALS: Seventeen client-owned dogs with measurable MCT amenable to RT. METHODS: Prospective clinical trial. All dogs received prednisone, omeprazole, diphenhydramine, and toceranib. Toceranib was administered for 1 week before initiating RT, consisting of 24 Gy delivered in 3 or 4 fractions. RESULTS: On an intent-to-treat basis, the overall response rate was 76.4%, with 58.8% of dogs achieving a complete response and 17.6% a partial response. The median time to best response was 32 days, and the median progression-free interval was 316 days. The overall median survival time was not reached with a median follow-up of 374 days. The most common toxicoses were gastrointestinal and hepatic. CONCLUSIONS AND CLINICAL IMPORTANCE: The combination of hypofractionated RT, toceranib, and prednisone was tolerated and efficacious in the majority of dogs. Response rates and durations were higher than those reported for toceranib as a single-agent treatment for MCT. This combination is a viable treatment option for unresectable MCT.

A tumor-related lymphoid progenitor population supports hierarchical tumor organization in canine B-cell lymphoma.

BACKGROUND: Tumors have heterogeneous properties, which could be explained by the existence of hierarchically and biologically distinct tumor cells such as tumor-initiating cells (TICs). This model is clinically important, as TICs are promising targets for cancer therapies. However, TICs in spontaneous B-cell lymphoma have not been conclusively identified. HYPOTHESIS/OBJECTIVES: Tumor cells with a progenitor phenotype exist in B-cell lymphoma, reflecting a hierarchical organization. ANIMALS: Twenty-eight client-owned dogs with previously untreated B-cell lymphoma and 6 healthy dogs. METHODS: This was a prospective study. Flow cytometry was used to identify lymphoid progenitor cells (LPCs) that coexpressed hematopoietic progenitor antigens CD34, CD117, and CD133, with lymphoid differentiation markers CD21 and/or CD22 in B-cell lymphoma. The polymerase chain reaction for antigen receptor rearrangements was used to analyze clonality and relatedness of tumor populations. A xenograft model with NOD/SCID/IL-2Rγ(-/-) mice was adapted to expand and serially transplant primary canine B-cell lymphoma. RESULTS: LPCs were expanded in lymph nodes from 28 dogs with B-cell lymphoma compared with 6 healthy dogs (P= .0022). LPCs contained a clonal antigen receptor gene rearrangement identical to that of the bulk of tumor cells. Canine B-cell lymphoma xenografts in recipient mice that maintained LPCs in the tumors were recurrently observed. CONCLUSIONS AND CLINICAL IMPORTANCE: These results suggest the presence of a hierarchy of tumor cells in B-cell lymphoma as has been demonstrated in other cancers. These findings have the potential to impact not only the understanding of lymphoma pathogenesis but also the development of lymphoma therapies by providing novel targets for therapy.

Class II major histocompatibility complex expression and cell size independently predict survival in canine B-cell lymphoma.

BACKGROUND: Class II major histocompatibility complex (MHC) is an independent predictor of outcome in human B-cell lymphoma. We assessed class II expression together with other markers for their impact on prognosis in canine B-cell lymphoma. HYPOTHESIS: Low class II MHC expression, large cell size, and expression of CD34 will predict a poorer outcome in canine B-cell lymphoma. Expression of CD5 and CD21 on tumor cells also may be associated with outcome. ANIMALS: One hundred and sixty dogs with cytologically confirmed lymphoma. METHODS: Patient signalment, treatment type, and flow cytometry characteristics were analyzed for their influence on outcome. A multivariable predictive model of survival was generated using 2/3 of the patients and validated on the remaining 1/3 of the dataset. RESULTS: Class II MHC expression had a negative association with mortality and relapse. Treatment type also influenced relapse and mortality, whereas cell size and patient age was only associated with mortality. CD34, CD21, and CD5 expression was not associated with disease outcome. The constructed model performed variably in predicting the validation group's outcome at the 6-month time point. CONCLUSIONS AND CLINICAL IMPORTANCE: Low levels of class II MHC expression on B-cell lymphoma predict a poor outcome, as in human B-cell lymphoma. This finding has implications for the use of dogs to model human lymphomas. Class II expression, cell size, treatment, and age can be combined to predict mortality with a high level of specificity.

Increased monocyte chemotactic protein-1 concentration and monocyte count independently associate with a poor prognosis in dogs with lymphoma.

Overexpression of the chemokine monocyte chemotactic protein-1 (MCP-1) has been associated with a poor prognosis in many human cancers. Increased MCP-1 concentrations may promote tumour progression by increasing mobilization of myeloid derived suppressor cells such as immature monocytes and neutrophils. We hypothesized that increased numbers of peripheral neutrophils or monocytes and increased MCP-1 concentrations would predict a worse outcome in dogs with multicentric lymphoma. In this retrospective study involving 26 client-owned dogs diagnosed with lymphoma, we show that peripheral neutrophil and monocyte counts as well as serum MCP-1 concentrations were significantly elevated relative to healthy control animals, and that such increases were associated with a decreased disease-free interval in dogs treated with chemotherapy based on cyclophosphamide, vincristine, doxorubicin and prednisone (CHOP). To our knowledge, this is the first study showing that pretreatment evaluation of monocyte and neutrophil counts can provide important prognostic information in dogs with lymphoma. The mechanisms underlying these observations remain to be determined.

Paraneoplastic T cell lymphocytosis associated with a thymoma in a dog.

A four-year-old male neutered Australian shepherd dog was diagnosed with a thymoma and concurrent mature T cell lymphocytosis. The lymphocytosis consisted of a mixed population of T cells expressing either CD4 or CD8 or neither marker, and the result of polymerase chain reaction for antigen receptor rearrangement was negative. The peripheral lymphocytosis resolved within 24 hours following thoracotomy and thymectomy. Similar cases have been reported in man, but the aetiology of the increased circulating lymphocytes remains unclear. Although peripheral lymphocytosis is an uncommon paraneoplastic syndrome associated with thymomas, thymoma should be considered as a differential when the increased lymphocytes consist of a mixed population of T cells.

Anemia is associated with decreased survival time in dogs with lymphoma.

BACKGROUND: Anemia is a common complication in human patients with neoplasia and has been associated with decreased survival time and a poorer quality of life. HYPOTHESIS: The presence of anemia at diagnosis is negatively associated with survival and remission times in dogs with lymphoma, but not in dogs with osteosarcoma. ANIMALS: Eighty-four dogs with lymphoma and 91 dogs with osteosarcoma that presented for treatment at the Animal Cancer Center, Colorado State University. METHODS: Retrospective, case-control study. Medical records were reviewed to determine the presence or absence of anemia (PCV < 40) at initial presentation. Median survival and remission times were identified by the Kaplan-Meier product limit method and the association between anemia and survival was determined by a multivariable Cox proportional hazard regression analysis. RESULTS: Cancer-related anemia is more frequent in dogs with lymphoma than in control dogs or dogs with osteosarcoma. Dogs with lymphoma and anemia had a significantly decreased survival time compared with dogs without anemia. There was no effect of anemia on remission time in dogs with lymphoma. Anemic dogs with osteosarcoma did not have decreased survival or remission time compared with nonanemic dogs with osteosarcoma. CONCLUSIONS AND CLINICAL IMPORTANCE: Shortened survival time in dogs with lymphoma and anemia at initial presentation has important prognostic significance. Understanding cancer-related anemia in dogs might offer new opportunities to improve quality of life and survival times in these patients.

Canine lymphoproliferative disease characterized by lymphocytosis: immunophenotypic markers of prognosis.

BACKGROUND: Canine lymphoproliferative disease often presents with lymphocytosis and is immunophenotypically diverse. HYPOTHESIS: Immunophenotype predicts prognosis in canine lymphoproliferative disorders involving circulating lymphocytosis. ANIMALS: Dogs that had peripheral blood evaluation performed by flow cytometry by the Clinical Immunology Service at Colorado State University between 2003 and 2005. METHODS: Outcome data regarding treatment and survival were sought on patients with lymphocytosis comprising a single lymphocyte subset. Ninety-six patients that met the inclusion criteria had sufficient follow-up information to be included in the study. RESULTS: Four main phenotypic classifications were found: CD8+ T-cell, CD21+ B-cell, CD4-8-5+ (aberrant T-cell phenotype), and CD34+ (undifferentiated progenitor). Expression of CD34 predicted poor outcome with median survival of 16 days (P < .0001) compared with other phenotypes. Within the CD8+ phenotype, dogs presenting with a lymphocytosis >30,000 lymphocytes/muL had significantly shorter median survival (131 days) than those presenting with <30,000 lymphocytes/muL (1098 days, P < .0008). Within the T-cell leukemias, there was no difference in outcome between dogs with CD4-8-5+ leukemia and dogs with the CD8+ T-cell phenotype nor was the loss of expression of the pan-leukocyte marker CD45 associated with decreased survival time. A CD21+ lymphocytosis composed of large cells was associated with shorter survival time (129 days) than those with smaller circulating cells (median survival not reached, P < .01). CONCLUSIONS AND CLINICAL IMPORTANCE: Immunophenotyping provides an objective method for determining prognosis in lymphoproliferative disorders characterized by lymphocytosis.

Use of FoxP3 expression to identify regulatory T cells in healthy dogs and dogs with cancer.

Regulatory T cells (Treg) are a distinct group of T lymphocytes with immunosuppressive properties that serve normally to prevent harmful autoimmune responses. However, Tregs can also interfere with beneficial immune responses such as anti-tumor and anti-viral immunity in humans and rodents. Given the overall importance of Tregs, it is likely that they play an important role in diseases of dogs as well. However, at present reagents required for identification of Tregs in dogs are not available. Therefore, we investigated whether expression of FoxP3, a transcription factor that is highly expressed in Tregs in humans and rodents could also be used to identify Tregs in dogs. We found that a cross-reactive FoxP3 antibody identified a subset of CD4(+) T cells in blood and lymph nodes of dogs. By flow cytometry the mean percentage of FoxP3(+)CD4(+) T cells in normal dogs was 4.3% in blood and 9.8% in the lymph nodes. In dogs with cancer, there was a significant increase in numbers of Treg in blood (7.5%) and tumor-draining lymph nodes (17.1%) compared to age-matched healthy control dogs. We also found that FoxP3(+)CD4(+) T cells in dogs could be significantly expanded in vitro by TCR activation together with addition of TGF-beta and IL-2. Treated cells also significantly increased expression of TGF-beta and IL-10mRNA. We conclude from these studies that a cross-reactive FoxP3 antibody can be used to identify Tregs in dogs and that this reagent may serve as a useful tool for investigating the role of Treg in a variety of diseases of dogs.

Diagnosis of canine lymphoid neoplasia using clonal rearrangements of antigen receptor genes.

Although the diagnosis of canine leukemia and lymphoma in advanced stages is usually uncomplicated, some presentations of the disease can be a diagnostic challenge. In certain situations, lymphoma and leukemia can be difficult to distinguish from a benign reactive proliferation of lymphocytes. Because clonality is the hallmark of malignancy, we have developed an assay that uses the polymerase chain reaction to amplify the variable regions of immunoglobulin genes and T-cell receptor genes to detect the presence of a clonal lymphocyte population. The assay detected clonally rearranged antigen receptor genes in 91% of the 77 dogs with lymphoid malignancy. Of the 24 dogs tested, that were either healthy or had clearly defined conditions not related to lymphoid malignancy, a clonally rearranged antigen receptor gene was found in one (a dog with Ehrlichia canis infection). Gene rearrangement was appropriate for the immunophenotype (immunoglobulin gene rearrangement in B-cell leukemias and T-cell receptor gene rearrangement in T-cell leukemias). Dilution analysis showed that the clonal rearrangement could be detected when 0.1-10% of the DNA was derived from neoplastic cells, depending on the source tissue. Potential applications of this assay include the diagnosis of lymphoma or leukemia in biopsy samples, cavity fluids, fine needle aspirates, bone marrow and peripheral blood; the determination of lineage (B or T cell); staging of lymphoma; and detection of residual disease after chemotherapy.

Plasmodium yoelii: resistance to disease is linked to the mtv-7 locus in BALB/c mice.

We have identified congenic mouse strains that differ dramatically in resistance to infection with the murine malaria parasite Plasmodium yoelii 17X. After infection, BALB/c mice develop severe anemia and a high degree of parasitemia which sometimes results in death. The mtv-7 congenic strain BALB.D2.mlsa, however, develops only a mild degree of anemia and parasitemia. In this paper we describe the course of the disease and discuss the potential role of mtv-7 and linked loci in control of this infection. These mice differ in their response to anemia, which may contribute to their differential susceptibility to disease; however, no influence of mtv-7 reactive T cells was documented.

Resistance to herpes stromal keratitis conferred by an IgG2a-derived peptide.

Not all peripheral tissue antigens enter the thymus and it is unclear how the immune system remains tolerant to this class of self antigen. As tolerance to self peptides can generate gaps in the T-cell repertoire for cross-reactive foreign antigens, we investigated whether this mechanism might also diminish autoimmune reactions to similar peptides expressed by peripheral tissues. Herpes stromal keratitis (HSK) is a virally induced autoimmune reaction against corneal tissues mediated by T cells, and is a leading cause of human blindness. Resistance to HSK in mice is associated with allotypic variation in immunoglobulin genes, possibly because circulating immunoglobin-derived peptides can cross-tolerize T cells specific for corneal tissue autoantigens. Here we show that HSK is mediated by T-cell clones specific for corneal self antigens which also recognize an allotype-bearing peptide derived from IgG2a, and that exposure of HSK-susceptible mice to a soluble form of this peptide confers resistance to HSK. Shared expression of peptide subsequences between sequestered tissue proteins and circulating proteins may be important for maintenance of self-tolerance and prevention of autoimmunity.