Clonal expansion of T lymphocytes in human melanoma metastases after treatment with a hapten-modified autologous tumor vaccine.

Sensi, Marialuisa; Farina, Cinthia; Maccalli, Cristina; Lupetti, Raffaella; Nicolini, Gabriella; Anichini, Andrea; Parmiani, Giorgio; Berd, David

doi:10.1172/jci119215

Cited by 50 publications

(34 citation statements)

References 33 publications

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“…9 Moreover, they represent expansion of T-cell clones with novel T-cell receptor structures. 10 Previously we have reported that administration of DNP vaccine to melanoma patients with bulky, resectable nodal metastases produced relapse-free and overall survivals that appeared to be as good as or better than those reported with the standard postsurgical therapy, high-dose interferon-␣. 11 In that study, the induction of DTH to autologous, unmodified melanoma cells was a significant predictor of clinical outcome.…”

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confidence: 89%

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Treatment of metastatic melanoma with autologous, hapten‐modified melanoma vaccine: Regression of pulmonary metastases

et al. 2001

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Key words: melanoma; vaccine; immunotherapy; autologous; cyclophosphamideStudies performed and published over the past 30 years have made it clear that immunizing a tumor-bearing patient to his or her own tumor cells is exceedingly difficult. An appealing explanation for this difficulty is that tumor antigens are weakly immunogenic and often induce tolerance, i.e., paralysis of the immune response, rather than effective immunity. 1 Fortunately, advances in basic immunology have disclosed a number of ways of breaking immunologic tolerance. One way is the judicious use of cytotoxic drugs. In 1967, Maguire and Ettore 2 made the surprising discovery that administration of cyclophosphamide to animals prior to an immunization caused the cell-mediated immune response to be augmented rather than suppressed. We have shown that this phenomenon applies to cancer-bearing humans as well. Thus, the administration of low-dose (300 mg/m 2 ) cyclophosphamide prior to an immunization with autologous tumor cells greatly increased the development of cell-mediated immunity to those cells. 3 A second approach to circumventing tolerance is the use of haptens-small molecules that are incapable of inducing an immune response unless they are coupled to a larger molecule. Coupling of a hapten to a protein, i.e., hapten modification, often results in the induction of an immune response to the protein that would otherwise not be possible. For example, Weigle, 4 in a series of now classic experiments, immunized mice with hapten-modified thyroglobulin. This resulted in the induction of an immune response not only to the hapten-modified thyroglobulin, but also to unmodified, i.e., native, thyroglobulin and produced autoimmune thyroiditis. A similar phenomenon was observed more recently by Neurath et al. 5 They found that application of the hapten dinitrophenyl to the colonic mucosa of mice resulted in the development of an autoimmune colitis, similar to human ulcerative colitis. Initially, these mice developed cellular immunity to the haptenmodified mucosa. However, as the chemically modified mucosa cells were shed, a potent immune response to the normal, unmodified mucosa developed, which led to the autoimmune pathology.The immunologic basis of these interesting phenomena is now clear: there are T lymphocytes generated by immunization with hapten-modified cells that are reactive to both modified and unmodified cells. For example, class I MHC-restricted T-cell clones generated from mice immunized with TNP-modified syngeneic lymphocytes respond to MHC-associated, TNP-modified "self" peptides. Furthermore, some TNP-reactive clones respond to certain MHC-binding, unmodified peptides as well. 6 These observations in animal models provide a strong rationale for a human cancer vaccine consisting of autologous tumor cells modified with a hapten, DNP, preceded by the administration of low-dose cyclophosphamide. We have observed that administration of DNP vaccine to patients with metastatic melanoma induces a unique reaction-the development of inflamma...

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confidence: 89%

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confidence: 97%

Treatment of metastatic melanoma with autologous, hapten‐modified melanoma vaccine: Regression of pulmonary metastases

et al. 2001

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“…After an ethanol precipitation, the cDNA was resuspended in 20 L of water. Semiquantitative reverse transcription ( RT ) PCR for BV repertoire analysis was performed as reported, 21 with the exception that amplifications were conducted according to the stepdown PCR parameters. 35,36 Briefly, reaction mixtures consisted of 20 L containing 1 L of diluted cDNA, 200 M of each dNTP, 0.5 M of each primer, and 0.6 U of Ampli -Taq Gold ( Applied Biosystems, Norwalk, CT ).…”

Section: Bv Repertoire Analysismentioning

confidence: 99%

“…In order to normalize TCR cDNA among samples, cDNA, starting from a concentration corresponding to 100 ng of total RNA, was serially diluted and quantitated by amplification with oligonucleotide specific for the TCR -chain constant region (BC ). 21 After an initial preincubation at 948C for 3 minutes, cycling was divided into five groups consisting of an initial three -cycles group followed by four six -cycles groups with a denaturation step of 20 seconds at 948C and an elongation step of 1 minute at 728C. Annealing lasted for 40 seconds, started at temperatures of 648C in the first three -cycles group, and decreased by 38C in each of the later groups.…”

Section: Bv Repertoire Analysismentioning

confidence: 99%

“…Individual bands were digitised and integrated with the use of the Imagequant software package provided by the manufacturer. The same amount of BC cDNA template for specimens of each patient was used for BV repertoire analysis performed with a panel of described oligonucleotide primers; 21,37 amplification was performed as above with the exception that the last group of cycles consisted of nine, instead of six, cycles. The level of specific amplification was measured by densitometry following hybridization with a BC -specific probe as described above and each BV band was expressed as a percentage of the sum of all BV signals detected in the repertoire analysis.…”

Section: Bv Repertoire Analysismentioning

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Comparative assessment of TCRBV diversity in T lymphocytes present in blood, metastatic lesions, and DTH sites of two melanoma patients vaccinated with an IL-7 gene-modified autologous tumor cell vaccine

et al. 2002

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A phase I clinical trial using autologous, IL -7 gene -modified tumor cells in patients with disseminated melanoma has been recently completed. Although no major clinical responses were observed, increased antitumor cytotoxicity was measured in postvaccine peripheral blood lymphocytes in a subset of treated patients. To analyze the in situ immune response, the T cell receptor -chain variable region ( BV ) repertoire of T cells infiltrating postvaccine lesions was studied in two patients, and compared with that of T cells present in prevaccine ones, in peripheral blood lymphocytes, and, in one patient, in delayed type hypersensitivity ( DTH ) sites of autologous melanoma inoculum. A relative expansion of T cells expressing few BVs was observed in all postvaccine metastases, and their intratumoral presence was confirmed by immunohistochemistry. Length pattern analysis of the complementarity determining region 3 ( CDR3 ) indicated that the repertoire of T cells expressing some of these BVs was heterogeneous. At difference, CDR3, -chain joining region usage, and sequence analysis enabled us to demonstrate, within a T -cell subpopulation commonly expanded at DTH sites and at the postvaccine lesion of patient 1, that both DTH sites contained identical dominant T -cell clonotypes. One of them was also expressed at increased relative frequency in the postvaccine lesion compared to prevaccine specimens. These results provide evidence for immunological changes, including in situ clonally expanded T cells, in metastases of patients vaccinated with IL -7 gene -transduced cells.

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Clonal expansions of 6‐thioguanine resistant T lymphocytes in the blood and tumor of melanoma patients

Albertini

Macklin

Zuleger

et al. 2008

Environ and Mol Mutagen

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The identification of specific lymphocyte populations that mediate tumor immune responses is required for elucidating the mechanisms underlying these responses and facilitating therapeutic interventions in humans with cancer. To this end, mutant hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficient (HPRT-) T-cells were employed as probes to detect T-cell clonal amplifications and trafficking in vivo in patients with advanced melanoma. Mutant T-cells from peripheral blood were obtained as clonal isolates or in mass cultures in the presence of 6-thioguanine (TG) selection, and from tumor-bearing lymph nodes or metastatic melanoma tissues by TG-selected mass cultures. Non-mutant (wild-type) cells were obtained from all sites by analogous means, but without TG selection. cDNA sequences of the T-cell receptor (TCR) beta chains (TCR-β), determined directly (clonal isolates) or following insertion into plasmids (mass cultures), were used as unambiguous biomarkers of in vivo clonality of mature T-cell clones. Clonal amplifications, identified as repetitive TCR-β V-region, complementarity determining region 3 (CDR3), and J-region gene sequences, were demonstrated at all sites studied, i.e., peripheral blood, lymph nodes, and metastatic tumors. Amplifications were significantly enriched among the mutant compared with the wild-type T-cell fractions. Importantly, T-cell trafficking was manifest by identical TCR-β cDNA sequences, including the hyper-variable CDR3 motifs, being found in both blood and tissues in individual patients. The findings described herein indicate that the mutant T-cell fractions from melanoma patients are enriched for proliferating T-cells that infiltrate the tumor, making them candidates for investigations of potentially protective immunological responses.

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Clonal expansion of T lymphocytes in human melanoma metastases after treatment with a hapten-modified autologous tumor vaccine.

Cited by 50 publications

References 33 publications

Treatment of metastatic melanoma with autologous, hapten‐modified melanoma vaccine: Regression of pulmonary metastases

Treatment of metastatic melanoma with autologous, hapten‐modified melanoma vaccine: Regression of pulmonary metastases

Comparative assessment of TCRBV diversity in T lymphocytes present in blood, metastatic lesions, and DTH sites of two melanoma patients vaccinated with an IL-7 gene-modified autologous tumor cell vaccine

Clonal expansions of 6‐thioguanine resistant T lymphocytes in the blood and tumor of melanoma patients

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