Engineering of Fc Multimers as a Protein Therapy for Autoimmune Disease.

The success of Intravenous Immunoglobulin in treating autoimmune and inflammatory processes such as immune thrombocytopenia purpura and Kawasaki disease has led to renewed interest in developing recombinant molecules capable of recapitulating these therapeutic effects. The anti-inflammatory properties of IVIG are, in part, due to the Fc region of the IgG molecule, which interacts with activating or inhibitory Fcγ receptors (FcγRs), the neonatal Fc Receptor, non-canonical FcRs expressed by immune cells and complement proteins. In most cases, Fc interactions with these cognate receptors are dependent upon avidity-avidity which naturally occurs when polyclonal antibodies recognize unique antigens on a given target. The functional consequences of these avid interactions include antibody dependent cell-mediated cytotoxicity, antibody dependent cell phagocytosis, degranulation, direct killing, and/or complement activation-all of which are associated with long-term immunomodulatory effects. Many of these immunologic effects can be recapitulated using recombinant or non-recombinant approaches to induce Fc multimerization, affording the potential to develop a new class of therapeutics. In this review, we discuss the history of tolerance induction by immune complexes that has led to the therapeutic development of artificial Fc bearing immune aggregates and recombinant Fc multimers. The contribution of structure, aggregation and N-glycosylation to human IgG: FcγR interactions and the functional effect(s) of these interactions are reviewed. Understanding the mechanisms by which Fc multimers induce tolerance and attempts to engineer Fc multimers to target specific FcγRs and/or specific effector functions in autoimmune disorders is explored in detail.

An analysis of facial nerve function in irradiated and unirradiated facial nerve grafts.

PURPOSE: The effect of high-dose radiation therapy on facial nerve grafts is controversial. Some authors believe radiotherapy is so detrimental to the outcome of facial nerve graft function that dynamic or static slings should be performed instead of facial nerve grafts in all patients who are to receive postoperative radiation therapy. Unfortunately, the facial function achieved with dynamic and static slings is almost always inferior to that after facial nerve grafts. In this retrospective study, we compared facial nerve function in irradiated and unirradiated nerve grafts. METHODS AND MATERIALS The medical records of 818 patients with neoplasms involving the parotid gland who received treatment between 1974 and 1997 were reviewed, of whom 66 underwent facial nerve grafting. Fourteen patients who died or had a recurrence less than a year after their facial nerve graft were excluded. The median follow-up for the remaining 52 patients was 10.6 years. Cable nerve grafts were performed in 50 patients and direct anastomoses of the facial nerve in two. Facial nerve function was scored by means of the House-Brackmann (H-B) facial grading system. Twenty-eight of the 52 patients received postoperative radiotherapy. The median time from nerve grafting to start of radiotherapy was 5.1 weeks. The median and mean doses of radiation were 6000 and 6033 cGy, respectively, for the irradiated grafts. One patient received preoperative radiotherapy to a total dose of 5000 cGy in 25 fractions and underwent surgery 1 month after the completion of radiotherapy. This patient was placed, by convention, in the irradiated facial nerve graft cohort. RESULTS: Potential prognostic factors for facial nerve function such as age, gender, extent of surgery at the time of nerve grafting, preoperative facial nerve palsy, duration of preoperative palsy if present, or number of previous operations in the parotid bed were relatively well balanced between irradiated and unirradiated patients. However, the irradiated graft group had a greater proportion of patients with pathologic evidence of nerve invasion (p = 0.007) and unfavorable type of nerve graft (p = 0.04). Although the irradiated graft cohort had more potentially negative prognostic factors, there was no difference in functional outcome (H-B Grade III or IV) between irradiated and unirradiated graft patients. H-B Grades III, IV, V, and VI were the best postoperative facial nerve functions achieved in 35%, 39%, 13%, and 13% of patients, respectively. The patient with preoperative radiotherapy never recovered any facial nerve function (H-B Grade VI). Median time to best facial nerve function after surgery was longer in the irradiated patients (13.1 vs. 10.8 months), but this was not statistically significant (p = 0.10). Presence of preoperative facial nerve palsy (p = 0.005), duration of preoperative palsy (p = 0.003), and age greater than 60 years at the time of grafting (p = 0. 04) were all negative prognostic factors for achieving a functional facial nerve on univariate analysis. Analysis of age as a continuous variable (p = 0.12) and pathologic evidence of nerve invasion (p = 0. 1) revealed a trend toward negative prognostic factors. Gender, number of previous operations in the parotid bed, extent of surgery at the time of nerve grafting, and type of grafting procedure were not significant prognostic factors. Whether radiotherapy was delivered less than 6 weeks after nerve grafting or more than 6 weeks had no impact on achievement of a functional facial nerve. CONCLUSION: Negative prognostic factors for achieving a functional facial nerve in our series include the presence of preoperative facial nerve palsy, duration of preoperative palsy, and age greater than 60 years. Radiotherapy was not a negative prognostic factor. Comparing irradiated and unirradiated grafts revealed no difference in best facial nerve function achieved, despite the presence of a greater proportion of negative prognostic factors in

Epstein-Barr virus DNA is not increased in tonsillar carcinoma.

Epstein-Barr virus (EBV) is a known oncogenic virus associated with a wide variety of cancers, including nasopharyngeal carcinoma. Waldeyer's ring, a collection of lymphoid tissues, includes the nasopharynx, pharyngeal, and lingual tonsils. To determine if EBV plays a causative role in carcinomas arising from other tissues in Waldeyer's ring, we examined pharyngeal tonsillar carcinomas for evidence of EBV infection. As previously reported, DNA was extracted from 53 consecutive tonsil cancers, as well as from age- and gender-matched non-cancerous tonsillectomy specimens. Three different sets of primers for discrete exons of EBV were then used to determine if active or latent EBV infection was expressed in the extracted DNA using the polymerase chain reaction (PCR). All positive bands were then sequenced to confirm the presence of amplified EBV fragments. None of the samples showed evidence for active EBV infection. In primers demonstrating latent infection, 1 of 53 (1.9%) of tumors were positive, versus 6 of 53 (11.3%) of the controls. These results indicate that EBV expression is not increased in DNA from tonsil cancers and that EBV infection does not have a causal relationship with tonsil cancer.

Immunobiologic effects of cytokine gene transfer of the B16-BL6 melanoma.

OBJECTIVE: The genetic modification of tumors offers an approach to modulate the host immune response to relatively weak native tumor antigens. We examined the immunobiologic effects of various cytokine genes transferred into the poorly immunogenic B16-BL6 murine melanoma. DESIGN: Retroviral expression vectors containing cDNAs for interleukin 2, interleukin 4, interferon gamma, or a neomycin-resistant control were electroporated into a B16-BL6 tumor clone. Selected transfected clones were examined for in vitro cytokine secretion and in vivo tumorigenicity. RESULTS: When cells from individual clones were injected intradermally into syngeneic mice, the interleukin 4-secreting clone grew significantly slower than did the neomycin-resistant transfected control, while the growth of the interleukin 2- and interferon gamma-expressing clones was not affected. Despite minimal cytokine secretion by interferon gamma-transfected cells, these cells expressed upregulated major histocompatibility class I antigen and were more susceptible to lysis by allosensitized cytotoxic T lymphocytes compared with parental or neomycin-resistant transfected tumor targets. CONCLUSIONS: We observed diverse immunobiologic effects associated with cytokine gene transfer into the B16-BL6 melanoma. Interleukin 4 transfection of tumor resulted in decreased in vivo tumorigenicity that may be related to a host immune response. Further studies to evaluate the host T-cell response to these gene-modified tumors are being investigated.

Squamous cell carcinoma of the buccal mucosa.

The purpose of this study was to establish treatment criteria for patients with early-stage squamous cell carcinoma of the buccal mucosa. Thirty-one patients were analyzed in a retrospective fashion. Distribution of patients according to tumor stage was relatively even. Within 5 years recurrent disease developed in nearly 80% of evaluable patients. There was a 100% overall incidence of local disease recurrence for patients with stage I and II tumors treated with wide local excision alone and followed up for more than 2 years. On the basis of these data, we conclude that wide local excision for early-stage buccal carcinoma is associated with a high local failure rate. Possible causes for failure and alternative treatment approaches are discussed.

Neonatal choristoma of the tongue containing glial tissue: diagnosis and surgical considerations.

There are only six case reports documenting the presence of glial tissue in the tongue. Because of the small number of cases, the presentation and biologic behavior of these lesions is poorly characterized. We present the case of a 10-day-old male infant who arrived at the University of Michigan Medical Center with a history of positional dyspnea, with resultant cyanosis and bradycardia, dysphagia, and a mass at the base of the tongue. Histopathologically, this lesion was initially labeled as a hamartoma, but was ultimately defined as a choristoma based on the exclusive presentation of glial tissue in the specimen. This paper will discuss the presentation, diagnostic evaluation, and therapeutic management of this case. In addition, the role of intraoperative electrodiagnostic monitoring to preserve neuromuscular function will be addressed.

Subglottic carcinoma: review of a series and characterization of its patterns of spread.

The rarity of primary subglottic malignancies, along with the varied definitions of the anatomic confines of this region, have limited our understanding of the patterns of tumor spread within the subglottis. We conducted a retrospective chart review to analyze clinical and pathologic data in patients with subglottic carcinoma. A pattern of disease progression was identified, which is defined by the cartilaginous laryngeal framework, with the fibroelastic barriers susceptible to tumor invasion. We conclude that although cartilaginous laryngeal structures are preserved until late in the disease course, the ability of tumors to invade the fibroelastic membranes provides them with an insidious means of escape. Specifically, tumor progression occurs primarily within the paraglottic space and extralaryngeal compartments; the potential for mucosal spread is limited. The lack of mucosal disease in patients whose cartilaginous laryngeal structures are intact may present a facade of normality in patients with advanced disease, and perhaps delay the early diagnosis of subglottic malignancies by physical and radiologic examination.

Sphenoid sinus mucocele: a rare complication of transsphenoidal hypophysectomy.

Only seven cases of a sphenoid mucocele occurring after transsphenoidal hypophysectomy have been previously reported in the world literature. In this article, we report a new case, which occurred in a 67-year-old man. The sphenoid sinus mucocele developed 12 years following transsphenoidal hypophysectomy and adjunctive radiotherapy. The patient was successfully managed with incision and drainage. Although transsphenoidal hypophysectomy is a common operation, this particular complication appears to be rare or at least under-reported. Sphenoid sinus mucocele deserves consideration in the differential diagnosis of a sphenoidal parasellar mass in a patient who has undergone an earlier transsphenoidal hypophysectomy.

Vaccine-based approaches to squamous cell carcinoma of the head and neck.

Vaccine-based approaches for the treatment of advanced squamous cell carcinoma of the head and neck have achieved very limited success. Improvement in vaccine efficacy for both diseases control and survival is predicated on a careful analysis of the root causes for successes and failures to date. In this review, we analyse the utility and limitations of select protective and therapeutic vaccine strategies for tumour prevention and therapy. Based on this characterisation, we define potential directions which are meritorious of future study.

Down syndrome: an otolaryngologic perspective.

The otolaryngologic manifestations of Down syndrome often masquerade as common and seemingly trivial conditions. Behind this facade lies a broad spectrum of disease which can detract from an inherently diminished level of function and jeopardize survival. This paper examines the pertinent associations between Down syndrome and various pathological entities related to the head and neck. Evaluation of current therapeutic modalities reinforces the need for early diagnosis and treatment so that the potential of this patient group is fulfilled.

Enhancement of immune reactivity in the lymph nodes draining a murine melanoma engineered to elaborate interleukin-4.

The adoptive transfer of immune T cells has been demonstrated to mediate regression of established tumors in animals, with encouraging results in human clinical trials. In animal studies, lymph nodes (LN) draining a progressively growing immunogenic tumor contain tumor-sensitized but functionally deficient T cells. These "preeffector" cells can be activated in vitro by sequential stimulation with anti-CD3 and interleukin (IL)-2 to differentiate into mature effector cells capable of mediating the regression of disseminated tumor. However, the preeffector cell response is weak during the growth of poorly immunogenic tumors such as the B16-BL6 melanoma. In this study, a clone of B16-BL6, A9, was transfected with the cDNA encoding for murine IL-4, in an attempt to enhance tumor immunogenicity. IL-4 secreting clones grew significantly slower than controls after intradermal (i.d.) inoculation, but all animals eventually succumbed to the progressive tumor. The ability of IL-4-secreting tumor cells to stimulate a preeffector cell response was then investigated. LN draining the IL-4-secreting tumors for 10 days were activated by the anti-CD3/IL-2 method. The resulting lymphocytes were adoptively transferred into animals bearing 3-day established parental pulmonary metastases. The transfer of cells derived from sensitization with the IL-4-secreting tumors was capable of significantly reducing the numbers of pulmonary metastases more effectively than cells sensitized to the parental tumor. Thus, genetic modification of tumor cells to secrete IL-4 can stimulate an increase in the preeffector cell response in the tumor-draining LN, suggesting an enhancement in T-cell-mediated immune function against the parental tumor.

Secretion of both IL-2 and IL-4 by tumor cells results in rejection and immunity.

The generation of a therapeutic immune response to malignancy is critically dependent on the inherent immunogenicity of the tumor. Our study demonstrates that secretion of both interleukin-2 (IL-2) and IL-4 by a seemingly nonimmunogenic tumor abrogates tumorigenicity, and mice that have rejected the genetically modified tumor are immune to challenges with the parental tumor. The induction of immunity by the IL-2/IL-4-secreting tumor was significantly better than that achieved with the admixture of tumor cells and the classic adjuvant, Corynebacterium parvum. To elicit a primary immune response, the majority of cells needed to secrete both cytokines. Ad-mixture of IL-2-secreting cells with IL-4-secreting cells did not result in tumor cell rejection. The IL-2/IL-4-secreting tumor cells were efficiently rejected in animals immunosuppressed by total body irradiation. Depletion of CD4+ or CD8+ T cells did not abrogate rejection of the tumor cells, but the animals depleted of CD4 cells failed to generate protective immunity. Our study demonstrates that secretion of the combination of IL-2 and IL-4 significantly enhances tumor immunogenicity. The requirement of cells secreting both cytokines suggests an intricate mechanism different from the mere presence of both cytokines at the tumor-inoculation site.

Anterior ischemic optic neuropathy following neck dissection.

BACKGROUND: Ischemic optic neuropathy (ION) is a rare but devastating complication of surgery. It has traditionally been associated with intraoperative hypotension in patients with underlying arteriosclerosis. METHODS: We present a case of ION following bilateral neck dissections in which there was minimal intraoperative hypotension and preservation of both internal jugular veins. The potential etiology of this disease is discussed along with a review of the literature. RESULTS: Five cases of ION following neck dissection have been documented. This complication is associated with the combination of intraoperative hypotension and anemia in the setting of prolonged bilateral neck dissection. Treatment is supportive, and final visual prognosis is variable. CONCLUSIONS: Ischemic optic neuropathy following neck dissection is best avoided by intraoperative blood pressure and anemia management. When it does occur, supportive therapy must be given. Final visual outcome is variable.

Diverse manifestations of tumorigenicity and immunogenicity displayed by the poorly immunogenic B16-BL6 melanoma transduced with cytokine genes.

We evaluated the in vivo response to the poorly immunogenic B16-BL6 (BL6) murine melanoma genetically altered to secrete interleukin-2 (IL-2), IL-4, interferon gamma (IFN gamma) and granulocyte/macrophage-colony-stimulating factor (GM-CSF). Three parameters were evaluated: (1) tumorigenicity, (2) vaccination of naive animals, and (3) assessment of antitumor reactivity of T cells derived from tumor-draining lymph nodes (TDLN). Secretion of IL-2 abrogated the tumorigenicity of BL6, while IFN gamma and IL-4 partially reduced tumorigenicity, and GM-CSF had no effect. Protective immunity to wild-type tumor challenge could not be achieved by vaccination with irradiated cytokine-secreting tumors, although IL-2 and IL-4 secretion appeared to retard the growth of the challenge inoculum significantly. An alternative method to evaluate the immunogenicity of the cytokine-secreting tumors was to measure the ability of T cells obtained from TDLN to mediate regression of wild-type tumor in adoptive immunotherapy. Neither IL-2 nor IFN gamma secretion resulted in the induction of immune T cells. By contrast, GM-CSF and IL-4 secretion were found to induce immune T cells in the TDLN with GM-CSF being superior to IL-4. The combined secretion of GM-CSF and IL-4 did not lead to enhanced induction of immune T cells. GM-CSF secretion was found to upregulate B7-1 expression in TDLN, consistent with an increase in the population of antigen-presenting cells. These studies demonstrated that reduced tumorigenicity by cytokine secretion did not correlate with increased immunogenicity. With the cytokines examined, there was limited capability of developing protective immunity against the BL6 tumor. Nevertheless, GM-CSF and IL-4 secretion significantly enhanced T cell immune reactivity to the poorly immunogenic BL6 tumor.

Endoscopic transnasal pituitary surgery: report on 180 cases.

The endoscopic transnasal approach is an evolving technique for treating lesions in the sella turcica. Since this method was introduced at our institution 4 years ago, the majority of transsphenoidal procedures are performed with it. The records of all patients having endoscopic transnasal hypophysectomy at the Mayo Clinic during the last 4 years were reviewed retrospectively. The criteria analyzed were safety, functional and cosmetic outcome, and complications. During the 4-year period, the operative procedure was modified to improve operative exposure and safety. The results of our review showed a significant decrease in length of hospital stay, reduced operative time, reduced need for nasal packing, and elimination of a sublabial incision. The complication rate was equivalent to that reported for the traditional transseptal transsphenoidal approach. As the neurosurgeons at our institution gained experience with this approach, an increasing number of pituitary microadenomas were resected safely and successfully. In addition, because of the limited septal dissection, this approach is particularly helpful for revision operations. This approach also can be used for the full range of pituitary lesions and in conjunction with adjunctive techniques, including frontal craniotomy and gamma-knife irradiation. Currently, the endoscopic transsphenoidal approach is the method preferred for surgically treating pituitary lesions in adults at our institution.

Enhanced therapeutic potential of adoptive immunotherapy by in vitro CD28/4-1BB costimulation of tumor-reactive T cells against a poorly immunogenic, major histocompatibility complex class I-negative A9P melanoma.

Costimulation plays a critical role in T-cell activation and amplification of anti-tumor immunity. Although CD28 engagement triggers an early activation signal, activation-induced 4-1BB molecule on T cells transmits a crucial signal for further expansion and maturation of effector cells. In this report, the authors show that costimulation through CD28 and 4-1BB pathways synergistically enhances the therapeutic efficacy of T cells from tumor-draining lymph nodes. Intravenous adoptive transfer of costimulated T cells into mice bearing disseminated micrometastasis of a poorly immunogenic, major histocompatibility complex class I-negative A9P melanoma results in a 60% cure rate. Autopsy of mice that died after unsuccessful treatment revealed tumor growth in the liver, spleen, and skin with minimal or no evidence of pulmonary disease. In contrast, mice that received no treatment or noncostimulated T cells had massive pulmonary tumors, suggesting that adoptively transferred T cells are less effective against growth of extrapulmonary tumors. These results show that costimulation of tumor-draining lymph node T cells through CD28 and 4-1BB increases their potential for cancer immunotherapy and suggests that improper trafficking of tumor-reactive T cells to extrapulmonary sites must be improved to enhance clinical efficacy.

Generation of therapeutic T-lymphocytes after in vivo tumor transfection with an allogeneic class I major histocompatibility complex gene.

In an effort to enhance the generation of tumor-reactive T-lymphocytes for adoptive immunotherapy, we examined the effects of in vivo transfection of an allogeneic major histocompatibility complex (MHC) class I gene (H-2Ks) of the poorly immunogenic B16BL6 (BL6) melanoma of H-2b origin. Cells from lymph nodes (LNs) draining these tumors after transfection were assessed in adoptive immunotherapy experiments for tumor reactivity after sequential activation with anti-CD3 monoclonal antibody (mAb) followed by culture in interleukin (IL)-2. H-2Ks lipofection of progressively growing BL6 subcutaneous tumors did not reduce tumorigenicity. However, in vivo lipofection of BL6 by intratumor inoculation or admixture of H-2Ks cDNA/liposome complexes and tumor cells prior to inoculation resulted in enhanced development of sensitized T-lymphocytes in the draining LN, which mediated the reduction of the numbers of established 3-day parental lung metastases in six of six experiments. In subsequent studies, in vivo transfection of BL6 with naked H-2Ks cDNA was found to be more effective than lipofection in eliciting sensitized T-cells in the draining LN. Admixture of liposomes alone or control plasmid DNA did not have an adjuvant effect similar to H-2Ks cDNA. Relative tumor transfection efficiency was assessed by an indirect assay with the chloramphenicol acetyltransferase (CAT) reporter gene. BL6 tumors were more efficiently transfected by intratumor inoculation with naked cDNA compared with lipofection. In summary, in vivo allogenization of the poorly immunogenic BL6 tumor resulted in enhanced generation of therapeutic T-cells effective in the treatment of parental tumor.

Open trial of methotrexate as treatment for autoimmune hearing loss.

OBJECTIVE: To assess the efficacy of low-dose methotrexate (MTX) administered for the treatment of autoimmune hearing loss. METHODS: This was a prospective, 12-month, open-label study of 17 patients with refractory autoimmune hearing loss. All patients had ongoing episodic worsening of hearing in one or both ears prior to enrollment despite traditional medical therapy. The MTX dose was 7.5-25 mg/week. Hearing loss and vertigo were evaluated at baseline and at completion of the study. Hearing improvement was defined as an improvement in pure tone threshold (PT) average of >10 dB or an increase in speech discrimination (SD) of >15%; worsening was defined as a decrease of >10 dB in PT or a decrease of >15% in SD in at least one ear. RESULTS: MTX was well tolerated. Among patients with Meniere's disease, 5 of 9 had improvement or resolution of vertigo. Equilibrium improved in all 3 patients with Cogan's syndrome and improved in 2 out of 3 patients with idiopathic hearing loss and this symptom. According to the parameters defined above, hearing improved in 11 patients (65%), was unchanged in 4 patients (23%), and worsened in 2 patients (12%). CONCLUSION: Long-term low-dose MTX therapy may be a useful therapy for at least some patients who have hearing loss with a presumptively autoimmune-mediated component that is refractory to traditional therapies.

Characterization of a spontaneously arising murine squamous cell carcinoma (SCC VII) as a prerequisite for head and neck cancer immunotherapy.

BACKGROUND: To develop novel therapeutic approaches for patients with head and neck malignancies, poorly immunogenic murine models of squamous cell carcinoma (SCC) need to be defined. METHODS: The phenotype, growth characteristics, and responsiveness to tumor-specific T-cell transfer of a spontaneously arising murine SCC (SCC VII) were characterized. RESULTS: SCC VII expresses major histocompatibility complex (MHC) class I molecules yet is resistant to tumor-specific T-cell killing and relatively insensitive to killing mediated by lymphokine-activated killer (LAK) cells. Intradermal tumors are reproducibly established after vaccination of 5 x 10(4) cells, and systemic micrometastases are apparent after intravenous administration of 2.5 x 10(4) cells. Immunotherapy of 3-day lung metastases using tumor-specific T cells and systemic interleukin-2 (IL-2) was ineffective in reducing the number of metastases in vivo. CONCLUSIONS: SCC VII is a poorly immunogenic murine squamous cell cancer, which represents an ideal model for preclinical testing of immunotherapeutic approaches for patients with SCC of the upper aerodigestive tract.