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Prognostic information is essential for the evaluation, judgement and optimal treatment of patients with squamous cell cancers (SCCs) of the upper aerodigestive tract. Using immunohistochemical and flow cytometric techniques, we have studied the significance of cellular expression of the Ki-67 antigen, epidermal growth factor receptor (EGFR), the transferrin receptor (TFR) and DNA ploidy status in a prospective analysis of patients with SCCs of the head and neck region. All 42 fresh tumour samples (five well differentiated; 28 moderately differentiated; nine poorly differentiated) expressed both EGFR and TFR to varying degrees. Receptor expression was most marked on the peripheral invading margin of cancer cell islands although staining was also demonstrated in a random fashion within cellular islands and consistently along the basal cell layer of overlying stratified squamous epithelium. The percentage of cancer cells that reacted with the Ki-67 monoclonal antibody was assessed as low (less than 10%) in 15 samples (35.8%), intermediate (10-30%) in 19 samples (45.2%) and high (greater than 30%) in eight samples (19.0%). Eleven of 15 samples (73%) with a low percentage reactivity were DNA diploid, whereas seven of eight samples (87.5%) with a high percentage reactivity were DNA aneuploid. Poorly differentiated SCCs were significantly more often aneuploid than were either moderately or well differentiated tumours. Our results suggest that EGFR and TFR are widely distributed on SCCs, especially on proliferating cells at the invading tumour margin. In addition, there is a close spatial correlation between cells expressing EGFR, TFR and those expressing the Ki-67 antigen. Tumours in which the staining intensity for both EGFR and TFR was intense invariably expressed the Ki-67 antigen in a high proportion of cells. Further patient follow-up will be important in determining whether intense EGFR and TFR staining, combined with a high percentage reactivity with Ki-67 antibody and DNA aneuploidy, will ultimately define a subset of head and neck cancer patients with a poor clinical outcome.

Neoplastic transformation has been associated with a variety of structural changes, among which are changes in membrane carbohydrates. Not much is known, though, e.g., how early in the tumourogenic event these changes take place and what effect these changes have on cell growth, invasion, and ability to metastasize. We were able to identify the B-D-Gal(1-3)DGal-NAc as a membrane carbohydrate component present in malignant laryngeal tissue, but not on adjacent normal mucous membrane. This carbohydrate structure was found to be present in metastatic as well as in nonmetastatic tumours. It was also found in well-differentiated as well as poorly differentiated carcinomas. We suggest that changes in carbohydrate components on the cell membrane of the laryngeal cancer cell are an early event in tumour progression and probably are not related to the degree of invasion or the ability to metastasize.

Prostaglandin levels in serum, tumor specimens, and normal larynx mucosa were measured before and after laryngectomy in 43 patients with larynx cancer. Related findings described elsewhere in the literature are discussed.

In previous work a new method of histologic grading of malignancy in squamous cell carcinomas of the larynx was described. It was based on morphologic criteria representing the tumor cell population itself and the tumor host relationship. Nuclear polymorphism was revealed to be a very important factor for determining prognosis. This method of histologic grading of malignancy was in this work further analyzed by cytophotometric DNA determinations on individual cells obtained by scraping the surface of squamous cell carcinomas of the larynx and the tongue. Increased heterogenous cellular DNA distributions were found in cases with high histological malignancy grading. Cells in the carcinoma cases with a normal epithelial morphology had the same DNA content as that of control cells of normal buccal mucosa. Cells with higher DNA values all had malignant or suspicious morphology.

Radioimmunological assay of carcino-embryonic antigen (CEA) was performed in 238 patients with cancer of the upper respiratory and digestive tracts after a minimum follow-up period of 3 years. Secretion of CEA was observed in only 1 out of 3 cases, levels, always being low and only slightly influenced by the localization or size of the tumor, and the condition of the lymph nodes. Initial levels are of no prognostic significance. The appearance of high levels during the course of the disease corresponds to shorter survival, but as it does not precede clinical evidence of recurrence or metastases this assay cannot play a role as a tumor marker.

The stratified squamous epithelium covering vocal cord polyps can vary from normal to hyperplastic and keratinized. We have analyzed the histology and cytokeratins of 11 of these polyps. Cytokeratins were separated on one-dimensional SDS-acrylamide gels to determine molecular weights. There was significant heterogeneity in the cytokeratins present in the different polyps. Comparison of histologic findings and cytokeratin content showed a correlation between extent of keratinization and presence of high molecular weight cytokeratins. We conclude that the epithelium of vocal cord polyps exhibit patterns of cytokeratins characteristic of both hyperproliferation and abnormal states of differentiation, which are mirrored by histologic variations, and that vocal cord polyps are a heterogeneous group of benign lesions.

A chondrosarcoma arising in the posterior cricoid plate is presented. The tumour gave rise to increasing inspiratory stridor: laryngectomy was performed. The tumour consisted of loose cartilaginous tissue with great predominance of highly differentiated cartilage cells and only small areas with nuclear polymorphism. This complicated the differential diagnosis between chondroma and highly differentiated chondrosarcoma. From the fact that the patient died from massive pulmonary metastases 3 1/2 years later, it is evident that the degree of malignancy of cartilaginous tumours should be determined even on small polymorphic foci. The DNA histogram for the foci of the laryngeal tumour with atypia differed distinctly from those for benign chondroma and normal cartilage, but resembled those of the pulmonary and splenic metastases and of a nasal chondrosarcoma. Photometric examination may provide an aid in the difficult differential diagnosis between chondroma and highly differentiated chondrosarcoma.

A new cell line designated 584A2 has been recently established from a patient with squamous cell carcinoma of the larynx. Cytogenetic analysis of the cell line revealed multiple copies of chromosome 7, as well as a homogeneous staining region (HSR) on one chromosome 7. Since overexpression of epidermal growth factor (EGF) cell surface receptors (EGFr) often occurs in other squamous carcinoma cell lines, it was predicted that 584A2 might overexpress EFGr. This was confirmed by: (1) metabolic labeling, with subsequent immunoprecipitation of EGFr and comparing autoradiographs to a cell line without an HSR and fewer copies of chromosome 7, and (2) performing EGF binding assays with Scatchard analysis. Since overexpression of EGFr correlates with an inhibitory effect of EGF on cell culture, the biological effects of EGF on 584A2 were examined in this study. At 5 ng/ml (serum-free medium), EFG stimulated incorporation of [3H] thymidine into trichloroacetic acid-precipitable material compared with controls. Incorporation increased between days 0 to 1 and 1 to 2 days with a 6- to 7-fold maximum. Dose-response studies (0 to 100 ng/ml) indicated maximum incorporation (6- to 7-fold) occurred between 0.1 ng/ml and 1.0 ng/ml. Cell growth was monitored over 7 days and, during this time, 5 ng/ml EGF produced a 10- to 12-fold increase in absolute cell numbers when compared with controls. We concluded that, unlike other squamous carcinoma lines with elevated EGFr, EGF stimulates rather than inhibits 584A2 cell proliferation.

A hydroxylapatite assay was used to assay androgen receptor levels in 23 primary laryngeal carcinomas, nine metastatic laryngeal carcinomas, and three oropharyngeal carcinomas. The median androgen receptor level in laryngeal carcinoma was 0.7 femtomoles of receptor protein per milligram of cytosol protein. Ten patients with advanced or metastatic laryngeal carcinoma and one patient with metastatic floor of mouth carcinoma were treated with the antiandrogen flutamide. There were three short-lived partial responses (1, 2, and 2.5 months) in nine patients whose tumors could be evaluated for response. Toxic reactions consisted of mild breast tenderness, and hypercalcemia in two patients with bone metastases. Androgen receptor levels did not correlate with response to flutamide. The low androgen receptor levels in laryngeal tumors may account for the relative inactivity of the antiandrogen in laryngeal cancer.

Carcinoid neoplasms, although rare, have to be considered in the differential diagnosis of neoplasms of the larynx. The total number of cases reviewed and listed in the present article is 47. The gross appearance of carcinoid tumours is not characteristic and does not differ from that of other malignant neoplasms of the larynx. Under the light microscope various types can be distinguished, which are described and illustrated in this review article. Available material of five previously published cases and of one new case have been re-examined under the electron microscope and by modern histochemical and immunohistochemical methods. The ultrastructural presence of neurosecretory granules and of argyrophilia by the Grimelius technique were uniformly positive, together with monoclonal antibodies for somatostatin and keratin. These special methods offer a reliable basis for the diagnosis of neuroendocrine neoplasms to which carcinoid tumours belong. The treatment of carcinoid neoplasms of the larynx is surgical.

Occupational exposure to asbestos has been epidemiologically associated with carcinoma of the larynx; however, no search for or quantitation of laryngeal asbestos bodies has been performed. This report concerns an autopsy study of five patients with occupational asbestos exposure and proven asbestos-associated pulmonary disease in whom histologic sections of the larynx were examined and a digestion-concentration technique was used to isolate and quantitate the asbestos bodies in the remaining laryngeal tissues. No dysplastic epithelial changes were present in the mucosa. Asbestos bodies were recovered from two of the five larynges but from none of ten larynges obtained from autopsy controls.

Routinely processed paraffin sections from 20 patients with adult laryngeal papillomas were examined for the presence of human papillomavirus type 11 (HPV-11) DNA and its specific mRNA by in situ hybridization methods using 35S-labeled RNA probes. Immunohistochemical techniques were also used to identify papillomavirus genus-specific common antigen (pgs-antigen). HPV-11 DNA signals and/or papillomavirus genus-specific common antigen were detected in all eight samples of multiple laryngeal papilloma. On the other hand, in 12 samples of single laryngeal papilloma, neither papillomavirus genus-specific common antigen nor HPV-11 DNA were detected. Four patients were positive for both HPV-11 DNA and pgs-antigen. In three of these four patients, HPV-11 mRNA signals were also detected. These results provided direct evidence of the association of HPV and adult multiple laryngeal papilloma.

Biopsy samples from 20 patients with a history of laryngeal papillomas were analyzed by Southern blot hybridization for the presence of human papillomavirus DNA. All papillomas examined contained papillomavirus DNA sequences. Four samples from uninvolved sites in two patients with active disease and eight samples from patients in remission also contained viral DNA. No human papillomavirus DNA was detected in a third patient seen during remission. These results explain the clinical pattern of frequent recurrences even after long-term remission in laryngeal papillomatosis.

The histogenetic origin of the spindle-cell component of spindle-cell carcinoma of the head and neck mucosa remains controversial. The spindle cells have been considered a variant growth pattern of squamous-cell carcinoma, a non-neoplastic mesenchymal reaction, and a malignant admixture of epithelial and mesenchymal neoplasm. To evaluate the spindle-cell component, we studied 25 tumors (18 biphasic and seven monophasic) by utilizing the following: an avidin-biotin complex immunoperoxidase technique with a variety of antikeratin antibodies (AE1, AE3, CAM 5.2, 35BH11, and polyclonal Dako) and a monoclonal antivimentin antibody, and an avidin-biotin alkaline phosphatase double-labeling technique to detect coexpression of keratin and vimentin. The immunohistologic staining pattern was compared with electron-microscopic studies. Eight of 18 biphasic neoplasms contained immunoreactive keratin in the spindle-cell component that was distributed focally in a minority of cells in 3 tumors and diffusely throughout five of the neoplasms. Four of seven ulcerated monophasic spindle-cell tumors devoid of histologic squamous-cell carcinoma also were keratin positive, confirming epithelial differentiation. The majority of the spindle cells in all the tumors contained vimentin intermediate filaments. In three immunoperoxidase keratin positive biphasic tumors examined with alkaline phosphatase double labeling, occasional spindle cells were found that coexpressed keratin and vimentin and were interspersed with cells expressing either intermediate filament. Electron microscopy was performed on the spindle-cell component of 13 tumors, nine biphasic and four monophasic. Of the biphasic tumors, four were immunoperoxidase keratin positive; three of these showed epithelial differentiation by electron microscopy. Five biphasic tumors were keratin negative, and three tumors had epithelial differentiation by electron microscopy. Four monophasic spindle-cell tumors were immunoperoxidase keratin positive, and one of these had epithelial features by electron microscopy. Two monophasic tumors were keratin negative and without ultrastructural evidence of epithelial features. By using a combination of immunohistochemical and electron-microscopic observations, we identified evidence for epithelial differentiation in the spindled cells in 11 of 18 biphasic tumors and four of seven monophasic spindle-cell tumors.

We studied 13 neuroendocrine carcinomas of the larynx. They constituted 59% of the 22 nonepidermoid carcinomas of the larynx seen at Memorial Hospital during a 45-year period, and for which adequate material was available for review. Four tumors were histologically identical to small cell carcinomas of the lung and were classified as small cell neuroendocrine carcinomas (SCNC). One case represents one of the original descriptions of the laryngeal SCNC. No SCNC was argyrophil, and of the three studied immunohistochemically, all contained neuron-specific enolase, one carcinoembryonic antigen (CEA) and one serotonin. Nine tumors were large cell carcinomas (LCNC). Eight LCNC were argyrophil, and all nine contained neuron-specific enolase, six calcitonin, four CEA, one HCG, two serotonin, and two somatostatin. The laryngeal neuroendocrine carcinomas commonly presented in chronic cigarette smokers with mean ages of 63 (SCNC) and 60 (LCNC), were not associated with other endocrine tumors, and proved highly fatal in spite of radical surgery and radiation therapy. At last follow-up only one patient was alive (after 13 months). Patients dying with SCNC survived a mean of 11 months, and those with LCNC, 36 months. To determine whether the laryngeal LCNC most closely resembles pulmonary neuroendocrine tumors, head and neck paragangliomas, or thyroid medullary carcinoma (TMC), they were histologically, histochemically, and immunohistochemically compared with control cases of each group. Overall, LCNC most closely resembles TMC, and given the frequency with which each presents as a neck mass, misinterpretation of one for the other is very possible. Evidence is provided suggesting that some LCNC have also been mistaken for the laryngeal paraganglioma.

Cyclic nucleotides (cAMP, cGMP) are suggested to participate in the regulation of cell growth and differentiation. Guanylate cyclase is the enzyme which catalyzes the synthesis of cGMP. The basal guanylate cyclase activity was slightly higher in well-differentiated squamous cell carcinomas than in normal mucosa, but was two-fold higher in papillomas. Poorly differentiated squamous cell carcinoma did not show any increased basal activity. Stimulation with nitroprusside (NP) resulted in a 20% increased activity for normal mucosa and a 30% increase for poorly differentiated carcinomas, whereas enzyme prepared from well-differentiated squamous carcinomas and papillomas showed a two-fold increase.

A case of laryngeal squamous papilloma in the early stages of development showed histological features suggestive of virus infection. Five years later positive evidence of HPV-6 infection was obtained at a time when the lesion had developed into a squamous cell carcinoma. It is concluded that this case represents a complete example of the virus to papilloma to carcinoma sequence, and as far as is known, is the first reported case of its kind in the larynx.

A laryngeal squamous cell carcinoma in situ with an underlying spindle cell nodule (pseudosarcoma) was immunohistochemically labeled with antibodies to tissue-specific intermediate filament proteins, including desmin, vimentin, and cytokeratin. Two distinct populations of cells were found within the lesion: cytokeratin-positive cells, corresponding to the carcinomatous component of the tumor, and vimentin-positive spindle cells in the subepithelial nodule. In view of the strict specificity of antivimentin and anticytokeratin for cells of mesenchymal and epithelial origin, respectively, it is proposed that the two components of the pseudosarcoma in our case are not morphologic variants of the same tumor, and that the subepithelial nodule represents a mesenchymal lesion. These results can, however, not be extrapolated to other cases since in some the spindle cell component may represent metaplastic epithelial cells. In view of the difficulties encountered in reaching a correct diagnosis in these lesions, it is recommended to use intermediate filament typing to elucidate the nature of the spindle cells in this controversial tumor.

Androgen cytoplasmic (ARc) and nuclear (ARn) receptors were analyzed in normal and cancerous laryngeal tissue obtained from male and female patients. In men, neither ARc nor ARn were detectable in most of the normal larynx tissue specimens, while either ARc, ARn, or both were present in eight of 16 laryngeal carcinomas. In women, all three normal and two of four malignant laryngeal tissue specimens possessed ARc without ARn. The presence of androgen receptors in some laryngeal carcinomas shows that these tumors are possibly hormone sensitive, and hormonal therapy should be considered in treating these tumors.

Normal human laryngeal epithelia and laryngeal squamous cell carcinoma were assayed by Feulgen DNA cytofluorometry using free cell nuclei isolated from carnoy-fixed, paraffin-embedded specimens. In all of the 12 normal specimens, the epithelium showed typical diploid cell clones with low proliferative activity. Polyploid cells were seen in only two specimens from subjects aged 61 and 69 years respectively, and the number of polyploid cells seen in these two specimens was only two. Fifteen cancer cases were divided into three groups: an untreated group (5 cases), a chemotherapy group (5 cases) and a group of cases with recurrence after radiation therapy (5 cases). Among these three groups the DNA ploidy patterns were compared. In the untreated group, all cases showed a two-peak diploid pattern and a high proliferative activity, and polyploid cells were present. In the chemotherapy group, a wide one-peak histogram extending from 2C to about 5C was noted in 4 cases, and an aneuploid pattern in one case. Thus, the DNA ploidy pattern in the chemotherapy group differed from that in the untreated group. Of the 5 cases with recurrence after radiation therapy, one had a tetraploid pattern, but the remaining 4, a two-peak diploid pattern similar to that seen in the untreated group. Polyploid cells were observed in all these cancer cases. However, because they were also seen in some normal subjects, the finding of polyploid cells is not considered to be conclusive of cancer diagnosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Estrogen binding sites were identified on plasma membranes from biopsy materials (N = 16) of normal human larynx; such binding was found to be specific. In tissue from 22 cases of laryngeal papilloma, membrane binding of 17-beta-estradiol was also specific. Although such binding occurred for papilloma membranes at a comparable affinity, it was clear that all membranes from the papillomatous tissue possessed a 148% greater estrogen binding capacity; there were at least 17 times more estrogen binding sites for each mg of receptor protein on papilloma membrane than for normal laryngeal epithelium. These findings suggest a possible receptor marker for the detection and/or prediction of recurrence of laryngeal papilloma.

For diagnostic purposes 14 histologically benign, 15 dysplastic, and 16 malignant squamous epithelial lesions were subjected to DNA cytophotometry. Results were computed according to an algorithm for DNA diagnosis and grading of malignancy. In cases of unequivocally malignant or benign lesions all DNA diagnoses agreed with the histologic diagnoses. In one case the allegedly faulty DNA diagnosis of cancer was proven to be correct on clinical follow-up. Within the group of histologically mild to moderate dysplasias the algorithm identified four cases as malignant that were proved to be malignant either in the follow-up or at another site of the same lesion. With the aid of the DNA malignancy grade two groups of patients with squamous epithelial carcinomas of the larynx could be discerned with a highly significant difference according to their survival times.

Formalin- or alcohol-fixed, paraffin-embedded laryngeal specimens were stained immunohistochemically with several kinds of monoclonal anti-Keratin antibody (KL-1, PKK-1, PKK-2, PKK-3), polyclonal anti-Keratin antibody, and epithelial membrane antigen. Immunohistochemical technique was used by Avidin-Biotin Complex method with the following subjects; normal epithelium 6 cases, dysplasia 15 cases, inflammation and polyp 12 cases, squamous cell carcinoma 47 cases. 6 specimens of squamous cell carcinoma were totally resected and alcohol-fixed, and other specimens were all biopsied and formalin-fixed. Comparing these two methods, the alcohol-fixed specimen gave superior result to those obtained with formalin-fixed in its staining. Non-staining area was recognized around basal layer by KL-1 stain in the normal epithelium. Enlargement of non-staining area was recognized in the dysplasia. Also, KL-1 staining showed positive staining with the keratinizing squamous cell carcinoma, and weak staining with the non-keratinizing squamous cell carcinoma. And, PKK-3 showed weak staining in the keratinizing squamous cell carcinoma. The results indicate that high molecular weight keratin (56 kilodalton) hardly localize around the basal layer in the squamous epithelium, and also in the non-keratinizing squamous cell carcinoma. The other way, this also suggests that low molecular weight keratin (45 kilodalton) hardly localize in the keratinizing squamous cell carcinoma. It is supposed that intracellular localization of keratin of dysplastic epithelium and squamous cell carcinoma varies from that in the normal squamous epithelium. Accordingly, monoclonal keratin stain is thought to be an useful method for the diagnosis of dysplasia and squamous cell carcinoma.

Red blood cell, serum and tissue Mg concentrations in 100 subjects with aged 34-78, with squamous cell carcinoma of the larynx were determined by atomic absorption spectrophotometry. The concentration of this cation in cancerous tissues was found to be about twice as high as in the unchanged homologous tissues of the host. This was accompanied by a systemic deficiency manifesting itself by a decreased Mg concentration in red blood cells, serum and muscles which became more pronounced as the disease advanced, and returned to normal after radical treatment.

Single-cell DNA fluorometry was performed on smear preparations from 57 histologically verified lesions from the vocal cords. Special attention was paid to the detection of hypertetraploid cell nuclei, which may be considered neoplastic markers, since they are not present in normal vocal cord epithelium. Hypertetraploid nuclei were not found in hyperplastic or mildly dysplastic epithelia but were found in 4 of 14 specimens of moderate dysplasia and in 5 of 19 specimens of severe dysplasia. The presence of hypertetraploid nuclei was associated with more frequent recurrences and progression of disease. Hypertetraploid cell nuclei were found in 9 of 15 invasive carcinomas and in this group were associated with a worse prognosis. The proliferative activity was 1.7% for hyperplasia and mild dysplasia, 3.1% and 2.8%, respectively, for moderate and severe dysplasia and 7.6% for squamous-cell carcinoma.

Biopsy specimens from 14 patients treated for laryngeal papillomatosis were tested for the presence of human papillomavirus (HPV) genome by the technique of DNA-DNA hybridisation. According to the age of initial presentation, cases were subdivided into juvenile (less than 16 years) and adult onset (older than 16 years) groups. Histological investigation confirmed that it was impossible to distinguish the groups on this basis. Molecular virology using both dot blot and Southern transfer techniques showed that 10 cases carried the HPV type 6 genome, three cases HPV type 11, and in one case no HPV DNA was detected. All six adult onset cases carried HPV 6 sequences while the juvenile onset group comprised four HPV 6 and three HPV 11 cases. In the juvenile onset group more females were affected; in the adult onset group more males were affected. Two of the patients shown to have HPV type 11 sequences in their biopsy material were the most resistant to treatment. One of the adult onset cases subsequently developed a squamous cell carcinoma of the larynx in which HPV 6 DNA was detected. As far as we know this is first time that HPV-DNA has been confirmed in laryngeal papilloma undergoing malignant change.

Report of a 54 year-old man with a carcinoid tumor in the larynx and cervical lymph node metastasis. This left sided and dum-bell shaped tumor was spreading from the vocal cord to the piriform sinus. It was smooth, reddish and rubbery. Microscopic study showed, near by the mucosa, the tumor cells arranged in a trabecular pattern, but more disseminated and infiltrating in the deepness. Grimelius stain was positive, Masson Fontana stain negative. Electron microscopy showed neuro-secretory type granules. Immuno-histo-chemical study revealed irregularly scattered cells which were positive for ACTH, but negative for somatostatin, calcitonin and gastrin. Review of literature for this extremely rare tumor.

Tyrosine-rich crystalloids were identified in three benign mixed tumors of the parotid gland, one terminal duct adenocarcinoma of a minor salivary gland, and the fibrous connective tissue of two laryngectomy specimens. Light and electron microscopic studies showed the crystalloids to be composed of irregular deposits of amorphous electron-dense material. In the salivary gland tumors this material was commonly associated with interstitial collagen and was found in greatest abundance near myoepithelial cells. This proximity suggests that the tyrosine-rich crystalloids result from the precipitation on stromal collagen of products secreted by neoplastic myoepithelial cells.

A well-differentiated mucoepidermoid carcinoma of the subglottis of a 77-year-old man was studied by light and electron microscopy. The tumor consisted of mucous cells, epidermoid cells, and intermediate cells of both differentiation. Mucous cells formed glands and cystic spaces filled with mucin. Abundant tonofibrils in aggregates were observed in tumor cell cytoplasms when they were differentiated into epidermoid cells. Epidermoid cells varied in differentiation.

In twenty-three cases of laryngeal dysplasia frozen mucosal strips were examined with four monoclonal and one polyclonal keratin antibody. The expression of specific keratin polypeptides was studied in different degrees of dysplasia with regard to the subunits expressed in normal and carcinomatous laryngeal epithelium in the same patient. An alteration in the expression of the subunits of cytokeratin in favour of low molecular weight polypeptides takes place in the transformation of normal epithelium to squamous cell carcinoma. This alteration seems to occur at an early stage and is present already in mild dysplasia. The results suggest that with a suitable antibody dysplastic laryngeal epithelium can be distinguished from normal epithelium, and also on some cases, mild dysplasia from more severe degrees of dysplasia. CAM 5.2, which identifies lower molecular weight cytokeratin proteins (50, 43 and 38 kD), is such an antibody, and can be a valuable diagnostic aid in the histological interpretation of laryngeal dysplasia.

3000 IgE serum concentrations were determine d radioimmunologically in a large group of 1616 tumor patients and a group of 308 controls. Because of a wide scattering the individual IgE levels do not give a diagnostic indication. On the other hand we have found a highly significant difference (p less than 0.001) between the IgE levels of male and female tumor patient-groups, which does not exist among the controls. Female patients tend to lower, male patients to higher levels. In patients with carcinoma of the testis three times higher IgE levels were found. Their relation to a coincidence with high HCG levels is discussed. Furthermore IgE follow up-studies are described. They correlate close by with the clinical picture. Consequently IgE follow up-studies are applicable to tumor prognosis and possibly to tumor therapy.

We have investigated the molecular basis for differences that we observed in the biological activities of genetically related but distinguishable human papillomavirus type 6 (HPV-6) subtypes. To analyse tissue-specific differences in replication and transcription, and to identify viral gene products important in the benign transformation of epithelial cells, we have modified procedures utilizing guanidinium isothiocyanate and density gradient centrifugation to facilitate the extraction of relatively undegraded DNA and RNA from 21 biopsy specimens of respiratory tract papillomata. Southern transfer analysis was used to characterize the viral genome, and to demonstrate that relative quantities of viral DNA in lesions varied but that the range was similar in lesions induced by HPV-6c, -6d, -6e and -6f. Dot blot analysis of the amount of viral RNA in comparison with the amount of 28S ribosomal RNA indicated that the relative level of viral RNA in each lesion varied considerably and that on average there was approximately twice as much viral RNA in HPV-6c-induced lesions as in HPV-6d-, -6e- or -6f-induced lesions. In dot blot and Northern analyses, hybridization of RNA from HPV-6c-induced lesions with HPV-6c DNA gave a stronger signal than hybridization of the same RNA with an HPV-6e probe, and vice versa. These differences in hybridization intensities with subtype-specific probes indicate that the most abundant RNA species are transcribed from parts of the genome that show sequence divergence between these two subtypes. Northern analysis demonstrated the predominant viral transcript to be about 1200 nucleotides in length in lesions induced by each of the four subtypes.

The clinical course and pathology of 57 patients with laryngeal papillomatosis were reviewed. Tissues from 26 patients were analyzed for human papillomavirus (HPV) DNA by Southern blot hybridization. Histopathologic evaluation of the papillomas showed no correlation with age of onset or clinical pattern of remission and recurrence. The pathology was characterized by abnormal squamous maturation with parakeratosis, retardation of superficial cell maturation, papillomatosis, and basal hyperplasia. HPV DNA was present in all lesions, with 92% containing either HPV-6 or 11. Latent HPV DNA was detected in clinically uninvolved tissues of 11 of 14 (78.5%) patients studied. There was no correlation between HPV type, histopathology and/or clinical pattern. Despite homogeneity of pathology, the clinical expression of laryngeal HPV infection varied widely. A mechanism for the pathogenesis of laryngeal papillomatosis, based on the concept of maturational arrest, is proposed.

On the basis of histoautoradiography of 79 laryngeal tumours, the importance of the degree of malignancy and of 3H-TdR labelling index was analysed. The "dynamic histology" (degree of structural differentiation and proliferation activity) seems to be correlated with the frequency of neck lymph node metastases. This relation provides possibilities for an early diagnosis of node involvement and an early adequate treatment.

A case of clear cell carcinoma of the larynx with multiple metastases is reported. The histological and immunohistochemical characteristics of this tumor are discussed as well as the main differential diagnoses. An origin from sero-mucous glands of the larynx is proposed. A literature review showed that clear cell carcinoma of the larynx is rare and is characterized by extremely high aggressiveness.

Five cases of monophasic and 7 cases of biphasic spindle-cell carcinomas were analyzed immunohistochemically for the presence of vimentin and keratin type intermediate filaments in the pleomorphic spindle cells. Vimentin reactivity proved to be a consistent feature but keratin reactivity was more variable, this latter filament being lost in two cases initially presenting as pure squamous cell carcinomas showing dedifferentiation towards a pure monophasic spindle-cell tumour when recurring. The converse was also noted: acquisition of keratin in a monophasic spindle-cell tumour that recurred as squamous cell carcinoma. These results were considered to support the concept that spindle-cell tumours of the upper aerodigestive tract are a peculiar type of carcinoma and not a product of a pluripotent stem cell exhibiting bidirectional differentiation. Diagnostic implications are as follows: keratin positivity in a spindle-cell tumour substantiates its carcinomatous nature but its absence does not rule out a diagnosis of spindle-cell carcinoma.

The flow cytometer is capable of analyzing a large number of cells or cell nuclei. The flow cytometer has been used to analyze the DNA content of tumor cells from squamous cell carcinomas of the upper aerodigestive tract using a modification of the Vindelov staining procedure. The main difficulty in analyzing solid tumors using flow cytometry technology is the disaggregation of the cells and the quantification of the diploid point (2N point). A set of internal standards was developed for establishing the 2N point, as well as a technique for disaggregation, which is described. A histogram characteristic of a more aggressive squamous cell carcinoma which tends to recur more quickly and more frequently than the other squamous cell carcinoma histogram types was identified. The flow cytometer proved to be a practical method of analysis allowing for quantitative measurement of DNA content from a large number of cells originating from a squamous cell carcinoma. The trend between recurrence and histogram type will require further study.

Forty laryngeal carcinomas were studied by immunofluorescence with specific antisera against components of the basement membrane (type IV collagen and laminin) as well as antisera against connective tissue antigens (type V collagen and fibronectin). The basement membrane surrounding well-differentiated squamous cell carcinomas showed an appearance similar to that seen beneath normal epithelium. In contrast, marked alterations of the basement membrane were constantly observed around infiltrating and poorly-differentiated carcinomas. The staining of connective tissue components in most cases was as intense in carcinomas as in normal laryngeal mucosa. The use of antibodies to basement membrane components may help to elucidate the mechanism of invasion of connective tissues by malignant cells.

For six patients with partial or total laryngectomies with extensive mucosal hyperplasias, we generated DNA histograms from multiple mucosal sites using Feulgen-stained tissue sections and microspectrophotometric microscopy. Aneuploid DNA histograms were identified in the mucosa of all six specimens, indicating that neoplastic transformation had occurred. The histologic characteristics of neoplastic change included thickened or hyperplastic epithelium, surface maturation or keratinization, often a proliferation of small, immature basallike cells in the depths of the epithelium, and evidence of abnormal epithelial maturation as evidenced by focal areas of cytoplasmic keratinization in the lower portions of the mucosa. We think this histologic expression of intraepithelial neoplasia is more common than the "classic" form of carcinoma in situ with full mucosal replacement by proliferating immature basallike cells. Keratin is a common reaction in laryngeal mucosa, and its presence on the surface or in the depths of the epithelium does not militate against the diagnosis of severe intraepithelial neoplastic transformation.

The existence of hormone receptors on or within neoplastic tissue has potential diagnostic, therapeutic, and prognostic importance. It was the purpose of this study to measure estrogen and progesterone receptors in a large group of patients with head and neck cancer to determine their frequency. Sixty-five patients with head and neck tumors underwent a total of 75 estrogen and 50 progesterone receptor assays. In this group, 87.7% were squamous cell carcinoma. In the estrogen receptor assays, 89.3% (67/75) were negative, 8% (6/75) were borderline, and only 2.7% (2/75) were positive. In the progesterone receptor assays, 78% (39/50) were negative, 22% (11/50) were borderline, and there were no positive results. There were no changes in assays of tissue removed at biopsy v tissue removed during surgery. There was no impact with chemotherapy. In conclusion, head and neck cancers do not appear to possess estrogen or progesterone receptors and can be considered to be hormonally independent.

It's possible papillomas of oral, pharyngeal and laryngeal regions concerning clinical and prognostical aspects to divide under a unit principle. In addition to the criterion of juvenile/adult you must consider solitary or multiple appearance, as well as the macroscopical keratinization. An adequate classification under the aspect of the different malignancy on the basis of morphological and clinical examination of 30 papillomas is proposed. The group of adult, multiple and unkeratinized papillomas of the larynx shows in opposite of opinions of today a tendency of malignancy. Infusion therapy with Bleomycin leads to a volume reduction and to a prolongation of the interval on the papillomas of the larynx, who tend to a malignant degeneration and to a short interval.

We have examined the distribution of human papillomavirus (HPV) DNA in paraffin sections of humans warts by in situ hybridization with biotin-labeled DNA probes. Recombinant plasmid DNAs (HPV-1, -6, -11, -16) were labeled by nick translation with biotinylated deoxyuridine triphosphate. Paraffin sections were hybridized with the probes for 18 h in stringent or non-stringent conditions, and DNA-DNA hybrids were detected by immunocytochemistry. Paraffin sections of warts were also examined for the presence of HPV capsid antigen with the avidin-biotin peroxidase complex method for immunocytochemistry. HPV DNA was detected and localized in paraffin sections from a plantar wart, a laryngeal papilloma, and seven anogenital condylomas. The specific HPV type present in each lesion was determined by hybridization under stringent conditions with the homologous DNA probe. The papillomas were found to contain many more cells with replicating virus DNA, as demonstrated by in situ hybridization, than was apparent from the number of cells containing detectable virus antigen. In situ hybridization with biotin-labeled probes is an effective technique for the identification of HPV infection in routinely collected and processed tissue specimens.