Prognostic factors in tongue cancer relative importance of demographic, clinical and histopathological factors

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1 British Journl of Cncer (2000) 83(5), doi: / bjoc , vilble online t on Prognostic fctors in tongue cncer reltive importnce of demogrphic, clinicl nd histopthologicl fctors S Kntol 1, M Prikk 2, K Jokinen 3, K Hyrynkngs 3, Y Soini 4, O-P Alho 3 nd T Slo 2,4 Deprtments of 1 Orl & Mxillofcil Surgery nd 2 Dignostics nd Orl Medicine, Institute of Dentistry, Oulu University Hospitl, University of Oulu, Apistie 3, Oulu, Finlnd; 3 Deprtments of Otorhinolryngology nd 4 Pthology, Oulu University Hospitl, P.O. Box 22, Oulu, Finlnd Summry The incidence of nd mortlity from squmous cell crcinom (SCC) of the tongue hve incresed during the recent decdes in the Western world. Much effort hs been mde to predict tumour behviour, but we still lck specific prognostic indictors. The im of our study ws to evlute the reltive importnce of the known demogrphic, clinicl nd histologicl fctors in homogeneous popultion-bsed group of ptients with SCC of the mobile tongue. The demogrphic nd clinicl fctors were reviewed retrospectively from primry nd tertiry cre ptient files. Histologicl prognostic fctors were determined from pre-tretment biopsies. The TNM stge ws found to be the most importnt prognostic fctor. In prticulr, locl spred outside the tongue rther thn spred to regionl lymph nodes ws relted to poor prognosis. Severl demogrphic nd histopthologicl fctors were closely relted to TNM stge. When the cses were divided into stge I II crcinoms nd stge III IV crcinoms, it ppered tht the ptient s older ge (> 65 yers), high mlignncy score nd n bsence of overexpressed p53 protein were ssocited with poorer prognosis in stge I II crcinoms. Such cses my require more ggressive tretment. Among ptients with stge III IV crcinoms, hevy use of lcohol ws significntly ssocited with poor disese-specific survivl time. Keywords: squmous cell crcinom; survivl; immunohistochemistry The incidence of orl squmous cell crcinom (SCC) hs incresed over the pst decdes in Europe nd in the United Sttes. At the sme time, mortlity from the disese hs incresed s well (Depue, 1986; Dvis nd Severson, 1987; Mcfrlne et l, 1996). The mngement of orl SCC is notbly dependent on the TNM stging system, which is bsed on the clinicl evlution. The stge, however, is not lwys sufficient for prognostiction. For exmple, some smll T1 tumours behve in n ggressive mnner nd hve n unexpectedly poor prognosis. Thus, it would be of gret benefit to be ble to identify the more ggressive tumours t the time of dignosis. There re numerous previous publictions deling with the identifiction of demogrphic, clinicl nd histologicl prognostic fctors (Jnot et l, 1996; Gluckmn et l, 1997). However, prt from TNM stge, there is controversy over the reltive importnce of different prognostic fctors. Since the TNM stge hs been found to be the most importnt prognostic fctor for orl SCC, there re studies of specificlly TNM stge I tumours (Högmo et l, 1999) or T1 2 tumours (Leedy et l, 1994), for exmple. Nevertheless, to our knowledge, there re no previous studies in which the prognostic fctors would hve been compred in ll stges. The im of the present pper ws to ssess the reltive importnce of the known prognostic demogrphic, clinicl nd histologicl fctors in popultionbsed smple of 105 ptients with SCC of the mobile tongue. Furthermore, we investigted whether distinctive prognostic Received 13 October 1999 Revised 8 My 2000 Accepted 8 My 2000 Correspondence to: S Kntol fctors could be found for stge I II nd for stge III IV crcinoms. PATIENTS AND METHODS Ptients The study re comprised the two northernmost provinces of Finlnd with popultion of pproximtely The Oulu University Hospitl is the only tertiry centre in the re, nd ll the ptients with tongue cncer re treted there. All the ptients who lived in the re nd were dignosed with cncer of the tongue (Interntionl Clssifiction of Diseses, version 9, code 141) between Jnury 1st 1974 nd December 31st 1994 were mnully identified from the surgicl, rdiotherpy nd dischrge registers of the Oulu University Hospitl. Crcinoms extending over vrious ntomicl structures were included, whenever the tongue ws cliniclly nd without doubt detected to be the site of origin. Tumours of the bse of the tongue (Interntionl Clssifiction of Diseses, version 9, code 1410) were excluded from the study. Altogether 108 new cses of tongue cncer were detected during the period Three cses were excluded becuse of insufficient clinicl dt. Demogrphic fctors Dt concerning the following demogrphic fctors were gthered from the clinicl records: sex, ge t the time of dignosis (44 yers or under, yers nd over 65 yers), socio-economic Lst two uthors hve eqully contributed to the work. 614

2 Prognostic fctors in tongue cncer 615 sttus (United Ntions, 1978), plce of residence (urbn vs. rurl) (Sttistics Finlnd, 1993), smoking (current or former smoker vs. non-smoker) nd lcohol consumption (none, light, moderte vs. hevy). Alcohol consumption ws defined s hevy if the ptient drnk dily or he/she hd been dignosed s n lcoholic. Clinicl fctors The following clinicl fctors were gthered from the ptient s primry cre or hospitl files: clinicl ppernce of the crcinom (exophytic vs. indurted, ulcertive, crter), crcinom spred outside the tongue (yes vs. no), neck nodes t the time of dignosis (yes vs. no), nd the TNM stge clssifiction of the tumour (Interntionl Union Aginst Cncer, 1987). Histopthologicl fctors All the histologicl fctors were determined from prffin sections of pre-tretment biopsies. Eight cses were excluded becuse of poor specimen qulity. During the study, unfortuntely, severl prffin blocks were exhusted, nd only 55 smples were vilble for nlysis of poptosis. The histologicl sections (with the exception of WHO grding) were nlysed by two of the uthors suitbly trined in scoring. The determintion ws done in blinded mnner without knowledge of the ptient outcome. The scores were bsed on consensus between the two uthors. The following fctors were determined from the sections. Tumour grde ccording to the WHO clssifiction (well differentited, modertely differentited or poorly differentited, World Helth Orgniztion, 1997), s given by the pthologist, ws retrospectively reviewed from the ptients hospitl files. The mlignncy scores of the crcinoms were determined using the method introduced by Bryne et l (1992). In this method, the score is determined from the deep invsive tumour mrgins. We determined the score from the most lterl invsive mrgin of the tumour present on the smple, s the deep invsive mrgin of the tumour ws only present in 26 (29%) of the biopsy smples. A score from 1 to 4 is given for five morphologicl fetures: degree of kertiniztion, nucler polymorphism, number of mitoses, pttern of invsion nd lymphoplsmcytic infiltrtion. The subscores re then summed up into totl mlignncy score (5 20). In the nlysis, the cses were divided into three groups bsed on the mlignncy score: 5 10 points, points nd points. Tumour ngiogenesis ws determined from sections immunohistochemiclly stined ginst fctor VIII (Dko, A/S, Glostrup, Denmrk), the method used being similr to tht of Leedy et l (1994). The microvessels, excluding single cells, were counted in ten high-power fields within the tumour (objective 40; dimeter of the field 400 µm). The men vlue of the vessels in the ten fields ws counted, nd the results were divided into three groups bsed on the qurtiles of the mens for the nlysis of tumour ngiogenesis. Inflmmtory cells (polymorphonucler, neutrophilic nd eosinophilic leucocytes, lymphocytes nd plsm cells) were counted on the tumour mrgin (objective 40, dimeter of the field 400 µm). The totl extent of the inflmmtory infiltrte ws ctegorised s light (+), moderte (++) or hevy (+++). Nerve invsion ws determined by using S-100-stined sections (Dko, A/S, Glostrup, Denmrk). Nerve invsion ws considered positive if there were clerly recognisble SCC cells in ny of the nerves present in the biopsy section. To determine tumour poptosis, 3 -end lbelling of poptotic cell DNA ws performed using the ApopTg in situ poptosis detection kit (Oncor, Githersburg, MD) nd following the mnufcturer s instructions with few modifictions s described by Soini nd Pääkkö (1996). Apoptotic cells were counted using similr method s pplied to microvessels. For the nlysis of poptosis, the results were divided into three groups bsed on the qurtiles of the mens. The presence of humn ppillom virus DNA (HPV 6, 11, 16, 18, 31 nd 33 subtypes) in the sections ws scertined by in situ hybridiztion s described by Soini et l (1996). Humn ppillom virus ws lso detected by PCR, using the HPV consensus primers MY 09 nd MY 11 (Soini et l, 1996), β-ctin by 5 sense nd 3 ntisense primers ccording to Ponte et l (1984). Overexpression of p53 protein ws determined from sections using similr method s described by Soini nd Pääkkö (1996). Immunohistochemicl stining ws done using the polyclonl rbbit nti-humn p53 ntibody CM1 diluted 1:1000 nd the secondry nti-rbbit ntibody s well s the vidin-biotin peroxidse complex (Vectstin ABC kit, Vector lbortories, Burlingme, CA). The expression of p53 ws recorded s negtive ( ), wekly positive (+) or strongly positive (++). Tretment of ptients Smll T1N0M0 crcinoms were treted with locl rdicl surgery without neck dissection. In the cse of T2N0M0 crcinoms, modified upper neck dissection ws included in the tretment. Lrger nd more widely spred crcinoms were treted with combintion of hemiglossectomy nd rdicl neck dissection nd djcent postopertive rdiotherpy. The most dvnced, inoperble cses were treted with rdiotherpy only. Sixty-six (63%) of the ptients hd surgery, 20 (19%) rdiotherpy nd 11 (10%) both surgery nd djuvnt rdiotherpy. Five (5%) of the ptients refused surgery nd were thus treted with rdiotherpy lone, nd three (3%) ptients did not receive ny tretment. Sttisticl methods The inter-reltionships between the vrious prognostic fctors were ssessed with the chi-squre method. The overll nd disese-specific men survivl nd recurrence-free times nd their 95% confidence intervls (95% CI) were clculted ccording to the Kpln-Meier method. Furthermore, disese-specific 75th percentile survivl times nd their rnges were clculted nd compred with Breslow s test. The disese-specific survivl time ws determined from the dte the ptient first received tretment for the cncer. The tumour sttus of ech ptient ws recorded bsed on the dt of the ltest control visit t the referrl centre. The prognostic effects of demogrphic nd histopthologicl fctors were studied seprtely in stge I II nd stge III IV crcinoms. In the nlyses, P vlues 0.05 were considered sttisticlly significnt. RESULTS Overll prognosis The men overll survivl time ws 99 months (95% CI ) nd the disese-specific survivl time 145 months (95% British Journl of Cncer (2000) 83(5),

3 616 S Kntol et l Tble 1 The disese-specific 75th percentile survivl times (months) nd rnge for ptient-relted prognostic fctors in series of 105 ptients with cncer of the tongue Fctors (n) 75th percentile Rnge P survivl time Gender 0.94 Femle (60) 19 (2 43) Mle (45) 17 (3 91) Age group yrs nd under (12) >60 b yrs (48) 22 (3 91) over 65 yrs (45) 13 (2 43) Socio-economic sttus 0.31 Low (71) 19 (2 91) High (21) >60 b Domicile 0.66 Urbn (68) 19 (3 91) Rurl (37) 17 (2 32) Smoking* 0.49 No (38) 91 (6 91) Yes (49) 32 (5 32) Alcohol consumption 0.05 None, seldom, moderte (54) >60 b Hevy (15) 12 (3 17) Clculted for those with dt vilble. b The disese-specific survivl rte ws over 75% Tble 2 The disese-specific 75th percentile survivl times (months) nd rnge for clinicl prognostic fctors in series of 105 ptients with cncer of the tongue Fctors (n) 75th percentile Rnge P survivl time Clinicl stge I (14) 36 (14 91) II (30) >60 b III (41) 19 (8 43) IV (16) 5 (3 32) Crcinom spred outside 0.04 the tongue No (82) 31 (2 91) Yes (23) 12 (3 32) Size of the crcinom mm or under (20) 91 (8 91) mm (62) 17 (2 32) over 40 mm (20) 12 (4 22) Neck nodes t the time of dignosis 0.25 No (61) 31 (2 91) Yes (43) 14 (3 32) Appernce of the crcinom 0.93 Exophytic (32) >60 b Ulcertive/crter (34) 22 (9 32) Clculted for those with dt vilble. b The disese-specific survivl rte ws over 75% CI ). The 75th percentile disese-specific survivl time ws 19 months (rnge 2, 91). A totl of 42 (40%) ptients developed recurrence during the follow-up. The men recurrence-free rte ws 126 months (95% CI ). The recurrences were locl in 47% of the cses, in the neck in 35% nd both locl nd in the neck in 18%. Distributions nd prognostic effects of demogrphic, clinicl nd histopthologicl fctors in univrite nlyses Sixty (55%) of the ptients were femle (Tble 1). The medin ge of the ptients ws 64 yers (rnge 26 90). Seventy-seven percent of the ptients hd low socio-economic sttus nd 65% lived in n urbn domicile. Over hlf of the ptients were smokers nd 22% were hevy lcohol users. The ptient s older ge nd hevy use of lcohol decresed significntly the disese-specific survivl time (P vlues 0.02 nd 0.05, respectively). None of the other demogrphic fctors hd significnt effect on survivl (Tble 1). There were 44 (44%) stge I II crcinoms in the study, nd in forty-three (41%) cses the crcinom hd spred to the neck nodes by the time of dignosis (Tble 2). The medin size of the crcinoms ws 30 m. The TNM stge hd the most significnt effect on the ptient s men disese-specific survivl time (P = ) (Tble 2). Prticulrly locl spred outside the tongue hd significnt effect on prognosis (P = 0.04). Tumour size, the presence of positive neck nodes t the time of dignosis or the clinicl ppernce of the crcinom did not hve sttisticlly significnt effect on disese-specific survivl (Tble 2). Three (3%) of the ptients hd poorly differentited neoplsm (Tble 3). Only five (6%) of the cses hd high (16 20 points) mlignncy score. None of the histopthologicl fctors hd sttisticlly significnt effect on disese-specific survivl (Tble 3). Nevertheless, low mlignncy score nd hevy tumour ngiogenesis (men of 8.5 vessels or over) tended to hve Tble 3 The disese-specific 75th percentile survivl times (months) nd rnge for histopthologicl prognostic fctors in series of 105 ptients with cncer of the tongue Fctors (n) 75th percentile Rnge P survivl time Grde 0.59 Well differentited (44) 29 (2 36) Modertely differentited (40) >60 Poorly differentited (3) 16 (16 17) Mlignncy score (38) 43 (6 43) (47) 22 (3 32) (5) 14 (14 16) Tumour ngiogenesis vessels nd under (17) 17 (8 22) vessels (31) 31 (6 91) 8.5 vessels nd over (18) >60 Inflmmtory response 0.61 Light (12) 12 (8 14) Moderte (47) 22 (4 91) Hevy (38) >60 Nerve invsion 0.35 No (49) 29 (5 43) Yes (7) >60 Apoptosis nd under (14) 16 (8 16) (25) 91 (4 91) 1.4 nd over (16) 22 (3 22) Overexpression of p53 protein 0.90 (37) 19 (6 32) + (26) 22 (4 91) ++ (22) 36 (3 36) The disese-specific survivl rte ws over 75%. positive influence on survivl time. Also, when there ws hevy inflmmtory response t the tumour edge, the disesespecific survivl time tended to be longer. We were unble to detect humn ppillom virus in ny of the biopsies nlysed. British Journl of Cncer (2000) 83(5),

4 Prognostic fctors in tongue cncer 617 DEMOGRAPHIC FACTORS Mle sex Older ge Smoking Excessive lcohol use CLINICAL FACTORS Crcinom spred outside the tongue Lrge crcinom size Crcinom spred to neck nodes HISTOPATHOLOGICAL FACTORS High mlignncy score Low tumour ngiogenesis Moderte or hevy inflmmtory response Incresed poptosis Absence of overexpressed p53 protein Figure 1 Sttisticlly significnt (P 0.05, chi-squre nlysis) interctions between demogrphic, clinicl nd histopthologicl fctors mrked with rrows in series of 105 ptients with cncer of the tongue Interction between demogrphic, clinicl nd histopthologicl fctors As shown in Figure 1, severl demogrphic nd histopthologicl fctors were significntly relted to the clinicl stge t the time of dignosis. We found tht mle sex, older ge, smoking nd hevy use of lcohol correlted significntly with the stge. Furthermore, it ppered tht low microvessel density, high mlignncy score of the crcinom, hevy inflmmtory response t the edge of the crcinom, incresed poptosis nd n bsence of overexpressed p53 protein correlted significntly with the clinicl distribution of the crcinom. In contrst, there were no sttisticlly significnt correltions between ny of the demogrphic nd histopthologic fctors studied. Prognostic fctors in stge I II crcinoms Ptient s ge over 65 yers, high mlignncy score (16 20 points) nd n bsence of the overexpressed p53 protein significntly shortened the men disese-specific survivl in stge I II crcinoms (Tble 4). The findings were similr, lthough not sttisticlly significnt, when the prognostic fctors for recurrence free times were determined (dt not shown). Tble 4 The disese-specific 75th percentile survivl times (ST) in months nd rnge ccording to ptient-relted nd histopthologic prognostic fctors in stge I II crcinoms nd in stge III IV crcinoms in 101 ptients with cncer of the tongue (stge not vilble for four ptients) Stge I II crcinoms Stge III IV crcinoms (n) ST (rnge) P (n) ST (rnge) P All ptients (44) 91 (6 91) (57) 14 (3 43) Gender Femle (27) >60 b (30) 19 (5 43) Mle (17) 31 (6 91) (27) 12 (3 17) Age group yrs nd under (24) >60 b (32) 14 (3 22) over 65 yrs (20) 29 (6 36) (25) 13 (5 43) Smoking No (21) 36 (6 91) (17) >60 b Yes (20) >60 b (28) 16 (5 32) Alcohol consumption None moderte (30) 91 (6 91) (23) >60 b Hevy (5) >60 b (10) 12 (3 17) Grde Well-differentited (24) 36 (6 36) (19) 19 (5 22) Modertely diff. (16) >60 b (21) >60 b Poorly differentited (3) 16 (16 17) Mlignncy score (20) >60 b (18) 43 (8 43) (17) >60 b (27) 17 (3 32) (3) 14 (14 16) (2) >60 b Angiogenesis Under 8.5 vessels (24) 31 (6 91) (22) 17 (9 22) 8.5 vessels nd over (7) >60 b (10) >60 b Inflmmtory response Light (3) >60 b (7) 9 (8 14) Moderte or hevy (40) 91 (6 91) (43) 22 (3 43) Apoptosis Under 1.4 (16) 91 (16 91) (21) 17 (5 17) 1.4 nd over (6) 17 (6 17) (9) 22 (3 22) Overexpression of p53 protein (16) 17 (6 29) (21) 19 (9 32) + ++ (21) >60 b (25) 17 (3 43) P for the difference between stge I II nd stge III IV crcinoms ws b The disese-specific survivl rte ws over 75% British Journl of Cncer (2000) 83(5),

5 618 S Kntol et l Prognostic fctors in stge III IV crcinoms Hevy use of lcohol ws significntly relted to poorer prognosis in stge III IV crcinoms (Tble 4). Furthermore, smoking, low or moderte tumour ngiogenesis nd slight inflmmtory response t the edge of the crcinom tended to correlte with poor prognosis, even though the correltion ws not significnt. Agin, similr findings were chieved when the prognostic fctors for recurrence-free times were determined (dt not shown). DISCUSSION We nlysed the reltive importnce of demogrphic, clinicl nd histopthologicl fctors in popultion-bsed study of 105 tongue SCC ptients. SCC of the tongue is the most common introrl mlignncy nd is usully detected t reltively erly stge. In the present ptient popultion, however, there were 57 (56%) ptients with stge III IV crcinoms. The prognosis of tongue cncer is better thn the prognosis of hed nd neck SCCs in generl. The prognostic importnce of the TNM stge ws once gin confirmed by the present findings. It ws clerly shown tht the ptients with TNM Stge IV crcinoms hd significntly poorer prognosis thn the ptients with smller nd more loclized tumours. One must remember, however, tht in the present ptient mteril the tretment ws plnned ccording to the TNM stge. The result confirms the findings of severl previous studies (Ghouri et l, 1993; Bundgrd et l, 1996; Jnot et l, 1996). Prticulrly locl spred outside the tongue rther thn spred to regionl lymph nodes ws significntly relted to poor prognosis. It ws noted tht severl demogrphic nd histopthologicl fctors were significntly relted to the clinicl distribution of the crcinom. From clinicl point of view, it is of gret interest to find out the most importnt prognostic fctors in stge I II cses. Ptient s older ge, high Bryne s mlignncy score nd n bsence of overexpressed p53 protein predicted poorer prognosis in stge I II crcinoms. In the cse of stge III IV crcinoms, hevy use of lcohol ws significntly ssocited with short survivl time. Since the overll survivl of the disese ws good, 63%, medin survivl times could not be clculted. Insted, we used 75th percentile survivl times nd rnges, which were more pproprite here. The ptient s older ge significntly shortened the disesespecific survivl time in the present study. The result is in ccordnce with previous study, where hed nd neck SCC ptients ged over 60 yers hd significntly poorer prognosis thn younger ptients (Jnot et l, 1996). However, contrdictory dt lso exist. Friedlnder et l (1997) noted tht the ptient s younger ge ws ssocited with ggressive tumour behviour in tongue cncers. The prognostic vlue of Bryne s mlignncy score in orl cncers hs been shown erlier by other uthors (Piffkò et l, 1997). The present study is the first to our knowledge to estblish the prognostic significnce of the mlignncy score, specificlly in stge I II tongue cncers. There were, however, only three stge I II crcinoms in the group with the highest mlignncy scores. This might reflect the less ggressive nture of tongue cncer compred to the mjority of hed nd neck cncers. In study by Högmo et l (1999) of stge I tongue cncer ptients no correltion between the mlignncy score nd regionl recurrence of the disese ws found. In tht prticulr study, however, Bryne s mlignncy score ws nlysed fctor by fctor nd not s totl score. It is thus impossible to conclude whether the score itself would hve hd prognostic vlue. According to the present study it seems tht n bsence of the overexpressed p53 leds to poorer prognosis in stge I II crcinoms. The finding is quite the opposite compred to recent Swedish study, where the presence of p53 predicted regionl recurrence in erly stge tongue crcinoms (Högmo et l, 1999). According to Leedy et l (1994), however, p53 did not hve ny prognostic vlue in T1 nd T2 tongue crcinoms. Similrly, the study by Gluckmn et l (1997) reveled no ssocition between the presence of p53 nd tumour ggressiveness. In the present study the overexpression of p53 protein ws determined with immunohistochemicl (IHC) stining. Flse negtive results my occur when the PCR method is used, s it tests only certin exons (5 8) of the protein, leving the muttions possibly occurring in other exons undetected (Soussi et l, 1994). Sunders et l (1999) performed full-length gene sequencing nd IHC stining to determine p53 in lryngel crcinoms. Nevertheless, they did not find correltion between the presence of p53 nd the recurrence rte of crcinoms. There ws trend to suggest tht hevy ngiogenesis is relted to good prognosis in stge III IV crcinoms, contrdicting the erlier findings (Gsprini et l, 1993; Shpitzer et l, 1996), but greeing with the study by Jnot et l (1996), where hed nd neck SCC ptients with higher microvessel counts hd better two-yer overll survivl rtes nd lower rtes of locoregionl filures nd metstsis. The number of microvessels ws smll in the present study compred to the previous studies (Leedy et l, 1994; Gluckmn et l, 1997). This ws most probbly due to the fct tht the microvessels were counted within the crcinom nd not in the hot spot re, nor were single stined cells considered microvessels. None of the stining methods presently in use (fctor VIIIAg, CD-31 nd CD-34) re ble to distinguish between neovsculriztion nd the vessels tht existed before tumour development. Moreover, it hs been suggested tht due to the bundnt vsculrity of the tongue nd the orl cvity, crcinoms of this region might be less dependent on neovsculriztion for growth nd metstsis (Gleich et l, 1997). Humn ppillomvirus (HPV) ws not found in single cse in this ptient popultion. This ws quite surprising in view of how much published dt re vilble concerning the correltion between the expression of HPV nd orl SCC. However, there re lso some previous studies with similr results (Zeuss et l, 1991; Mtzow et l, 1998). In recent study by Pintos et l (1999) on upper erodigestive trct neoplsms, HPV ws detected in three out of 26 orl SCCs. The uthors noted tht the HPV detection rte ws higher for phryngel crcinoms thn for tumours of the mouth or the lrynx. We used two sensitive methods to detect HPV, PCR nd in situ hybridiztion, which mkes our results relible. In the nlysis PCR by β-ctin primers gve positive result in every smple. This indictes tht the mplifiction of genomic DNA ws successful nd flse negtive results were excluded. Furthermore, the detection of HPV ws done with exctly similr techniques s in the study by Soini et l (1996), where HPV DNA ws found in 30% of the lung crcinom smples studied. As conclusion, ccording to the present study, the TNM stge nd prticulrly the locl spred of the crcinom t the time of dignosis significntly correlted with the disese-specific British Journl of Cncer (2000) 83(5),

6 Prognostic fctors in tongue cncer 619 survivl time in ptients with SCC of the tongue. In stge I II crcinoms the ptient s older ge (> 65 yers), high mlignncy score nd n bsence of overexpressed p53 protein hd profound effect on disese-specific survivl. According to this study, smll, loclized crcinoms in such cses my require more ggressive tretment. REFERENCES Bryne M, Koppng HS, Lilleng R nd Kjærheim Å (1992) Mlignncy grding of the deep invsive mrgins of orl squmous cell crcinoms hs high prognostic vlue. J Pthol 166: Bundgrd T, Benzen SM, Wildt J, Sørensen FB, Søgrd H nd Nielsen JE (1996) Histopthologic, stereologic, epidemiologic, nd clinicl prmeters in the prognostic evlution of squmous cell crcinom of the orl cvity. Hed Neck 18: Dvis S nd Severson RK (1987) Incresing incidence of cncer of the tongue in the United Sttes mong young dults. Lncet 2(8564): Depue RH (1986) Rising mortlity from cncer of the tongue in young white mles. N Engl J Med 315: 647 Friedlnder PL, Schntz SP, Shh AR, Yu G nd Shh JP (1997) Squmous cell crcinom of the tongue in young ptients: mtched-pired nlysis. Hed Neck 20: Gsprini G, Weidner N, Mlut S, Pozz F, Borcchi P, Mezzetti M, Testolin A nd Bevilcqu P (1993) Intrtumorl microvessel density nd p53 protein: correltion with metstsis in hed nd neck squmous-cell crcinom. Int J Cncer 55: Ghouri AF, Zmor RL, Hrvey JE, Spitzngel Jr EL, Sessions DG (1993) Epidermoid crcinom of the orl cvity nd orophrynx: vlidity of the current AJCC stging system nd new sttisticl tools for the prediction of subclinicl neck disese. Otolryngol Hed Neck Surg 108(3): Gleich LL, Biddinger PW, Duperier FD nd Gluckmn JL (1997) Tumor ngiogenesis s prognostic indictor in T2 T4 orl cvity squmous cell crcinom: clinicopthologic correltion. Hed Neck 19: Gluckmn JL, Pvelic ZP, Welkoborsky H-J, Mnn W, Stmbrook P, Gleich L, Wilson K, Righi P, Portugl LG, McDonld J, Biddinger P, Stewrd D nd Grtside P (1997) Prognostic indictors for squmous cell crcinom of the orl cvity: A clinicopthologic correltion. Lryngoscope 107: Högmo A, Kuylentiern R, Lindholm J nd Munck-Wiklnd E (1999) Predictive vlue of mlignncy grding systems, DNA content, p53, nd ngiogenesis for stge I tongue crcinoms. J Clin Pthol 52: Interntionl Union Aginst Cncer (1987) TNM clssifiction of mlignnt tumours 4th edn. Springer-Verlg: Berlin Jnot F, Klijnienko J, Russo A, Mmet J-P, de Brud F, EI-Nggr AK, Pignon J-P, Luboinski B nd Cvikovic E (1996) Prognostic vlue of clinicopthologicl prmeters in hed nd neck squmous cell crcinom: prospective nlysis. Br J Cncer 73: Leedy DA, Trune DR, Kronz JD, Weidner N nd Cohen JI (1994) Tumor ngiogenesis, the p53 ntigen, nd cervicl metstsis in squmous cell crcinom of the tongue. Otolryngol Hed Neck Surg 111: Mcfrlne GJ, Shrp L, Porter S nd Frnceschi S (1996) Trends in survivl from cncers of the orl cvity nd phrynx in Scotlnd: clue s to why the disese is becoming more common? Br J Cncer 73: Mtzow T, Boysen M, Klntri M, Johnsson B nd Hgmr B (1998) Low detection rte of HPV in orl nd lryngel crcinoms. Act Oncol 37: Piffkò J, Bànkflvi À, Öfiner D, Bryne M, Rsch D, Joos U, Böcker W nd Schmid KW (1997) Prognostic vlue of histobiologicl fctors (mlignncy grding nd AgNOR content) ssessed t the invsive tumour front of orl squmous cell crcinoms. Br J Cncer 75: Pintos J, Frnco EL, Blck MJ, Bergeron J nd Arell M (1999) Humn ppillomvirus nd prognoses of ptients with cncers of the upper erodigestive trct. Cncer 85: Ponte P, Ng SY, Engel J, Gunning P nd Kedes L (1984) Evolutionry conservtion in the untrnslted regions of ctin mrnas: DNA sequence of humn betctin cdna. Nucleic Acids Res 12: Sunders ME, McKenzie R, Shipmn R, Frnsen E, Gilbert R nd Jordn CK (1999) Ptterns of p53 gene muttions in hed nd neck cncer: full-length gene sequencing nd results of primry rdiotherpy. Clin Cncer Res 5: Shpitzer T, Chimoff M, Gl R, Stern Y, Feinmesser R nd Segl K (1996) Tumor ngiogenesis s prognostic fctor in erly orl tongue cncer. Arch Otolryngol Hed Neck Surg 122: Soini Y nd Pääkkö P (1996) Extent of poptosis in reltion to p53 nd bcl 2 expression in germ cell tumours. Hum Pthol 27: Soini Y, Nuorv K, Kmel D, Pöllänen R, Vähäkngs K, Lehto V-P, Pääkkö P (1996) Presence of humn ppillom virus DNA nd bnorml p53 protein ccumultion in lung crcinom. Thorx 51: Soussi T, Legros Y, Lubkin R, Ory K nd Schlichtolz B (1994) Multifctoril nlysis of p53 ltertion in humn cncer: review. Int J Cncer 57: 1 9 Sttistics Finlnd (1993) Communities Helsinki United Ntions (1978) Recommendtions for the 1980 censuses of popultion nd housing: the ECE region. In: Sttisticl stndrds nd studies. No. 31: New York World Helth Orgniztion (1997) Histologicl typing of cncer nd precncer of the orl mucos (Second Edition). Pindborg J, Reichrt P, Smith C nd vn der Wl I (eds), Springer-Verlg, Berlin Heidelberg Zeuss MS, Miller CS nd White DK (1991) In situ hybridiztion nlysis of humn ppillomvirus DNA in orl mucosl lesions. Orl Surg Orl Med Orl Pthol 71: British Journl of Cncer (2000) 83(5),

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