tscc patients : patients ( n=7 ) with histopathologically confirmed inoperable tongue ( primary site anterior 2/3 of tongue ) squamous cell carcinoma ( tnm stage iv ) ( table i ) were included in this preliminary study . all these patients attended the out patient department of chittaranjan national cancer institute ( cnci ) , kolkata , during 2006 - 2008 . these patients were treated with cisplatin ( 70 mg / m i.v . on day 1 ) + 5-fu ( 500 mg / mi.v . on day 1 - 3 ) in two cycles in 21 days interval and ifn2b ( 3 miu twice wkly for 3 wk ) was administered in between two cycles of chemotherapy . haematological and immune parameters were checked before initiation of the treatment , after first cycle of chemotherapy and after ifn2b therapy . study protocol was approved by institutional review board ( irb ) of cnci , kolkata . characteristics of inoperable stage iv tscc patients response assessment and follow up : clinical features of patients were assessed regularly by monitoring tumour size , ankyloglossia , node involvement and general health . the objective responses , evaluation of time to progression , duration of response , time to treatment failure and overall survival on this treatment were recorded . leucocyte , platelet and haemoglobin status : blood ( 6 ml ) was collected from tscc patients and used for white blood cell ( wbc ) , platelet count and determination of haemoglobin percentage . wbc differential count was performed by microscopic examination on leishman 's stained blood smear on slides . reagents and antibodies : recombinant human ifn2b was gifted by shanferon , santha - biotech ; india . cd4 , cd8 , cd56 monoclonal antibodies , opteia ifn , interleukin ( il)-12 , il-10 , il-4 estimation kits and tmb substrate solution were procured from bd - pharmingen ( san diego , ca , usa ) . tumour cell lines : hep-2 ( epidermoid carcinoma of larynx ) cells were maintained in dulbecco 's modified eagle medium ( dmem ) ( life technologies , ny , usa ) , supplemented with 10 per cent ( v / v ) heat inactivated foetal bovine serum ( fbs ) , 2 mm l - glutamine and gentamycin ( 0.052 mg / ml ) at 37c without the supply of 5 per cent co2 . the nk sensitive k562 ( erythro - leukemic ) cell line was maintained in roswell park memorial institute 1640 ( rpmi 1640 ) medium ( life technologies , ny , usa ) , supplemented with fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . pbmc culture : peripheral blood mononuclear cells ( pbmc ) from tscc patients were isolated from heparinized venous blood by density gradient centrifugation over ficoll hypaque . isolated pbmc were cultured in rpmi 1640 , supplemented with 10 per cent fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . cells and supernatants ( stored at -80c ) were used in different assays mentioned below . extracellular secretion of cytokines : type 1 ( ifn and il-12 ) and type 2 ( il-4 and il-10 ) cytokines were measured in pbmc culture supernatant by elisa using commercially available kits ( bd pharmingen , san dieago , usa ) . in brief , 96 well microtitre plates were coated with capture antibody ( anti - ifn/anti - il-12/anti - il-4/anti - il-10 ) , incubated overnight at 4c and blocked with 5 per cent bsa for 1 h. after washing , 100 l of cell free supernatant was added into each well to incubate for 2 h. bound cytokines were detected by using biotinylated mouse anti - human ifn/il-12/il-4/il-10 and avidin - horse radish peroxidase subsequently . reaction was stopped with 2n h2so4 and absorbance was measured at 450 nm using microplate reader ( tecan spectra , grodig , austria ) . flow cytometric analysis of immune cellular markers : blood ( 100 l ) was labelled with 20 l of different anti - human fluorescence labelled antibodies ( cd4-fitc , cd8-pe and cd56-pe ) for 30 min as per manufacturer 's recommendation ( bd pharmingen , san diego , usa ) . after labelling , rbc was lysed by facs lysing solution ( bd pharmingen , usa ) , washed , fixed in 1 per cent paraformaldehyde in pbs and cytometry was performed by using cell quest software on a facscan flow cytometer ( becton dickinson , mountainview , usa ) . cytotoxicity assay in vitro : cytotoxicity of pbmc , after removal of adherent fraction by plastic adherence technique , against different cancer cells , was tested by lactate dehydrogenase ( ldh ) release assay using commercially available cytotoxicity detection kit ( roche diagnostics , mannheim , germany ) . pbmc ( 1x10 ) were added as effector in three effector : target ( e : t ) ratios ( 10:1 , 50:1 , 100:1 ) in each well and co - cultured for 4 h. cell free supernatant was used to measure the level of released ldh . tscc patients : patients ( n=7 ) with histopathologically confirmed inoperable tongue ( primary site anterior 2/3 of tongue ) squamous cell carcinoma ( tnm stage iv ) ( table i ) were included in this preliminary study . all these patients attended the out patient department of chittaranjan national cancer institute ( cnci ) , kolkata , during 2006 - 2008 . these patients were treated with cisplatin ( 70 mg / m i.v . on day 1 ) + 5-fu ( 500 mg / mi.v . on day 1 - 3 ) in two cycles in 21 days interval and ifn2b ( 3 miu twice wkly for 3 wk ) was administered in between two cycles of chemotherapy . haematological and immune parameters were checked before initiation of the treatment , after first cycle of chemotherapy and after ifn2b therapy . study protocol was approved by institutional review board ( irb ) of cnci , kolkata . characteristics of inoperable stage iv tscc patients response assessment and follow up : clinical features of patients were assessed regularly by monitoring tumour size , ankyloglossia , node involvement and general health . the objective responses , evaluation of time to progression , duration of response , time to treatment failure and overall survival on this treatment were recorded . leucocyte , platelet and haemoglobin status : blood ( 6 ml ) was collected from tscc patients and used for white blood cell ( wbc ) , platelet count and determination of haemoglobin percentage . wbc differential count was performed by microscopic examination on leishman 's stained blood smear on slides . reagents and antibodies : recombinant human ifn2b was gifted by shanferon , santha - biotech ; india . cd4 , cd8 , cd56 monoclonal antibodies , opteia ifn , interleukin ( il)-12 , il-10 , il-4 estimation kits and tmb substrate solution were procured from bd - pharmingen ( san diego , ca , usa ) . tumour cell lines : hep-2 ( epidermoid carcinoma of larynx ) cells were maintained in dulbecco 's modified eagle medium ( dmem ) ( life technologies , ny , usa ) , supplemented with 10 per cent ( v / v ) heat inactivated foetal bovine serum ( fbs ) , 2 mm l - glutamine and gentamycin ( 0.052 mg / ml ) at 37c without the supply of 5 per cent co2 . the nk sensitive k562 ( erythro - leukemic ) cell line was maintained in roswell park memorial institute 1640 ( rpmi 1640 ) medium ( life technologies , ny , usa ) , supplemented with fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . pbmc culture : peripheral blood mononuclear cells ( pbmc ) from tscc patients were isolated from heparinized venous blood by density gradient centrifugation over ficoll hypaque . isolated pbmc were cultured in rpmi 1640 , supplemented with 10 per cent fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . cells and supernatants ( stored at -80c ) were used in different assays mentioned below . extracellular secretion of cytokines : type 1 ( ifn and il-12 ) and type 2 ( il-4 and il-10 ) cytokines were measured in pbmc culture supernatant by elisa using commercially available kits ( bd pharmingen , san dieago , usa ) . in brief , 96 well microtitre plates were coated with capture antibody ( anti - ifn/anti - il-12/anti - il-4/anti - il-10 ) , incubated overnight at 4c and blocked with 5 per cent bsa for 1 h. after washing , 100 l of cell free supernatant was added into each well to incubate for 2 h. bound cytokines were detected by using biotinylated mouse anti - human ifn/il-12/il-4/il-10 and avidin - horse radish peroxidase subsequently . reaction was stopped with 2n h2so4 and absorbance was measured at 450 nm using microplate reader ( tecan spectra , grodig , austria ) . flow cytometric analysis of immune cellular markers : blood ( 100 l ) was labelled with 20 l of different anti - human fluorescence labelled antibodies ( cd4-fitc , cd8-pe and cd56-pe ) for 30 min as per manufacturer 's recommendation ( bd pharmingen , san diego , usa ) . after labelling , rbc was lysed by facs lysing solution ( bd pharmingen , usa ) , washed , fixed in 1 per cent paraformaldehyde in pbs and cytometry was performed by using cell quest software on a facscan flow cytometer ( becton dickinson , mountainview , usa ) . cytotoxicity assay in vitro : cytotoxicity of pbmc , after removal of adherent fraction by plastic adherence technique , against different cancer cells , was tested by lactate dehydrogenase ( ldh ) release assay using commercially available cytotoxicity detection kit ( roche diagnostics , mannheim , germany ) . pbmc ( 1x10 ) were added as effector in three effector : target ( e : t ) ratios ( 10:1 , 50:1 , 100:1 ) in each well and co - cultured for 4 h. cell free supernatant was used to measure the level of released ldh . tscc patients : patients ( n=7 ) with histopathologically confirmed inoperable tongue ( primary site anterior 2/3 of tongue ) squamous cell carcinoma ( tnm stage iv ) ( table i ) were included in this preliminary study . all these patients attended the out patient department of chittaranjan national cancer institute ( cnci ) , kolkata , during 2006 - 2008 . these patients were treated with cisplatin ( 70 mg / m i.v . on day 1 ) + 5-fu ( 500 mg / mi.v . on day 1 - 3 ) in two cycles in 21 days interval and ifn2b ( 3 miu twice wkly for 3 wk ) was administered in between two cycles of chemotherapy . haematological and immune parameters were checked before initiation of the treatment , after first cycle of chemotherapy and after ifn2b therapy . study protocol was approved by institutional review board ( irb ) of cnci , kolkata . characteristics of inoperable stage iv tscc patients response assessment and follow up : clinical features of patients were assessed regularly by monitoring tumour size , ankyloglossia , node involvement and general health . the objective responses , evaluation of time to progression , duration of response , time to treatment failure and overall survival on this treatment were recorded . leucocyte , platelet and haemoglobin status : blood ( 6 ml ) was collected from tscc patients and used for white blood cell ( wbc ) , platelet count and determination of haemoglobin percentage . wbc differential count was performed by microscopic examination on leishman 's stained blood smear on slides . reagents and antibodies : recombinant human ifn2b was gifted by shanferon , santha - biotech ; india . cd4 , cd8 , cd56 monoclonal antibodies , opteia ifn , interleukin ( il)-12 , il-10 , il-4 estimation kits and tmb substrate solution were procured from bd - pharmingen ( san diego , ca , usa ) . tumour cell lines : hep-2 ( epidermoid carcinoma of larynx ) cells were maintained in dulbecco 's modified eagle medium ( dmem ) ( life technologies , ny , usa ) , supplemented with 10 per cent ( v / v ) heat inactivated foetal bovine serum ( fbs ) , 2 mm l - glutamine and gentamycin ( 0.052 mg / ml ) at 37c without the supply of 5 per cent co2 . the nk sensitive k562 ( erythro - leukemic ) cell line was maintained in roswell park memorial institute 1640 ( rpmi 1640 ) medium ( life technologies , ny , usa ) , supplemented with fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . pbmc culture : peripheral blood mononuclear cells ( pbmc ) from tscc patients were isolated from heparinized venous blood by density gradient centrifugation over ficoll hypaque . isolated pbmc were cultured in rpmi 1640 , supplemented with 10 per cent fbs , penicillin ( 50 units / ml ) , streptomycin ( 50 g / ml ) and gentamycin ( 0.052 mg / ml ) at 37c in a humidified atmosphere with 5 per cent co2 . cells and supernatants ( stored at -80c ) were used in different assays mentioned below . extracellular secretion of cytokines : type 1 ( ifn and il-12 ) and type 2 ( il-4 and il-10 ) cytokines were measured in pbmc culture supernatant by elisa using commercially available kits ( bd pharmingen , san dieago , usa ) . in brief , 96 well microtitre plates were coated with capture antibody ( anti - ifn/anti - il-12/anti - il-4/anti - il-10 ) , incubated overnight at 4c and blocked with 5 per cent bsa for 1 h. after washing , 100 l of cell free supernatant was added into each well to incubate for 2 h. bound cytokines were detected by using biotinylated mouse anti - human ifn/il-12/il-4/il-10 and avidin - horse radish peroxidase subsequently . reaction was stopped with 2n h2so4 and absorbance was measured at 450 nm using microplate reader ( tecan spectra , grodig , austria ) . flow cytometric analysis of immune cellular markers : blood ( 100 l ) was labelled with 20 l of different anti - human fluorescence labelled antibodies ( cd4-fitc , cd8-pe and cd56-pe ) for 30 min as per manufacturer 's recommendation ( bd pharmingen , san diego , usa ) . after labelling , rbc was lysed by facs lysing solution ( bd pharmingen , usa ) , washed , fixed in 1 per cent paraformaldehyde in pbs and cytometry was performed by using cell quest software on a facscan flow cytometer ( becton dickinson , mountainview , usa ) . cytotoxicity assay in vitro : cytotoxicity of pbmc , after removal of adherent fraction by plastic adherence technique , against different cancer cells , was tested by lactate dehydrogenase ( ldh ) release assay using commercially available cytotoxicity detection kit ( roche diagnostics , mannheim , germany ) . hep-2 and k562 cells ( 1x10 of each ) pbmc ( 1x10 ) were added as effector in three effector : target ( e : t ) ratios ( 10:1 , 50:1 , 100:1 ) in each well and co - cultured for 4 h. cell free supernatant was used to measure the level of released ldh . patients included in this study presented with ankyloglossia and n1/n2 regional nodes . among seven patients , 3 & 6 ) responded completely [ complete response ( cr)-no growth in tongue on palpation , no ankyloglossia , no node palpable , no growth seen in ct scan of face and neck ] , four patients ( nos . 1 , 2 , 5 & 7 ) responded partially [ partial response ( pr)-downgrading of ankyloglossia grade iii to grade i , < 50% response to tumour growth ] . 4 . no such changes were noticed in the tongue of patient without response ( table i ) . these patients were followed for 6 to 12 months with median follow up of 9 months . total leucocyte count was decreased after cisplatin + 5-fu therapy in all seven patients studied ( table ii ) higher grade of leukepenia was controlled by gcsf . the extent of recovery was much higher in two patients with complete response . in case of platelet count chemotherapy induced decrease was observed in four patients among five patients examined for platelet . ifn2b treatment helped to recover this count in three cases within four . in patient 3 , where complete response was detected , significant recovery of platelet count from 72 , immunological and haematological status of tscc patients under chemo - immunotherapy cisplatin + 5-fu therapy along with ifn2b treatment increased the number of cd8 t cells in five patients among seven patients studied . two patients have not shown much change before and after treatment . in case of cd4 t cells , increase in the number no change was reflected in cd4:cd8 ratio , however , this ratio was slightly decreased after chemotherapy and it was increased after ifn2b treatment . in addition to t cells , another important cytotoxic cells , cd56 nk cells , also demonstrated similar profile of changes as observed in case of cd8 t cells . number of cd56 cells was increased in five of the seven patients . among patients with higher expression of cd8 and cd56 markers , complete response was noticed in patient nos . 3 and 6 , with longest survival . cytotoxic ability of pbmc obtained from ifn2b exposed tscc patients against tumour cells was studied ( table ii ) . larynx cancer hep-2 cells were used to get reflection of t cell cytotoxicity and nk sensitive erythroleukemic cells k562 were used to assess nk cell cytotoxicity . four among six patients studied have demonstrated increase in the cytotoxicity towards hep-2 cells , in comparison to their pretreatment values . it was noticed that per cent of cytotoxicity reduced after cis+5-fu treatment was recovered in all patients after ifn2b treatment . the patient ( no . 3 ) with complete response has showed increase in hep-2 cell cytotoxicity in maximum extent ( post - chemotherapy , 7.2% to post - ifn2b , 21.3% ) . cytotoxicity of nk sensitive k562 cells demonstrated similar pattern , as observed in case of hep-2 cells . type 1 cytokine status of tscc patients , undergoing chemo - immunotherapy was studied by monitoring ifng and il-12 , secreted from pbmc at different points of the treatment . 3 with complete response maintained high secretary level of ifn till day 20 after radiotherapy . in case of il-12 type 2 cytokine status of tscc patients was studied by monitoring il-10 and il-4 , secreted from pbmc at different points of the treatment ( table ii ) . ifn2b therapy reduced secretary il-10 level in four of seven patients ; including those two patients with complete response . other three patients showed no change in comparison to their pre - treatment values . in case of il-4 downregulation patients included in this study presented with ankyloglossia and n1/n2 regional nodes . among seven patients , 3 & 6 ) responded completely [ complete response ( cr)-no growth in tongue on palpation , no ankyloglossia , no node palpable , no growth seen in ct scan of face and neck ] , four patients ( nos . 1 , 2 , 5 & 7 ) responded partially [ partial response ( pr)-downgrading of ankyloglossia grade iii to grade i , < 50% response to tumour growth ] . 4 . no such changes were noticed in the tongue of patient without response ( table i ) . these patients were followed for 6 to 12 months with median follow up of 9 months . total leucocyte count was decreased after cisplatin + 5-fu therapy in all seven patients studied ( table ii ) higher grade of leukepenia was controlled by gcsf . the extent of recovery was much higher in two patients with complete response . in case of platelet count chemotherapy induced decrease was observed in four patients among five patients examined for platelet . ifn2b treatment helped to recover this count in three cases within four . in patient 3 , where complete response was detected , significant recovery of platelet count from 72 , immunological and haematological status of tscc patients under chemo - immunotherapy cisplatin + 5-fu therapy along with ifn2b treatment increased the number of cd8 t cells in five patients among seven patients studied . two patients have not shown much change before and after treatment . in case of cd4 t cells , increase in the number no change was reflected in cd4:cd8 ratio , however , this ratio was slightly decreased after chemotherapy and it was increased after ifn2b treatment . in addition to t cells , another important cytotoxic cells , cd56 nk cells , also demonstrated similar profile of changes as observed in case of cd8 t cells . number of cd56 cells was increased in five of the seven patients . among patients with higher expression of cd8 and cd56 markers , complete response was noticed in patient nos . 3 and 6 , with longest survival . cytotoxic ability of pbmc obtained from ifn2b exposed tscc patients against tumour cells was studied ( table ii ) . larynx cancer hep-2 cells were used to get reflection of t cell cytotoxicity and nk sensitive erythroleukemic cells k562 were used to assess nk cell cytotoxicity . four among six patients studied have demonstrated increase in the cytotoxicity towards hep-2 cells , in comparison to their pretreatment values . it was noticed that per cent of cytotoxicity reduced after cis+5-fu treatment was recovered in all patients after ifn2b treatment . the patient ( no . 3 ) with complete response has showed increase in hep-2 cell cytotoxicity in maximum extent ( post - chemotherapy , 7.2% to post - ifn2b , 21.3% ) . cytotoxicity of nk sensitive k562 cells demonstrated similar pattern , as observed in case of hep-2 cells . type 1 cytokine status of tscc patients , undergoing chemo - immunotherapy was studied by monitoring ifng and il-12 , secreted from pbmc at different points of the treatment . 3 with complete response maintained high secretary level of ifn till day 20 after radiotherapy . in case of il-12 type 2 cytokine status of tscc patients was studied by monitoring il-10 and il-4 , secreted from pbmc at different points of the treatment ( table ii ) . ifn2b therapy reduced secretary il-10 level in four of seven patients ; including those two patients with complete response . other three patients showed no change in comparison to their pre - treatment values . in case of il-4 downregulation patients included in this study presented with ankyloglossia and n1/n2 regional nodes . among seven patients , 3 & 6 ) responded completely [ complete response ( cr)-no growth in tongue on palpation , no ankyloglossia , no node palpable , no growth seen in ct scan of face and neck ] , four patients ( nos . 1 , 2 , 5 & 7 ) responded partially [ partial response ( pr)-downgrading of ankyloglossia grade iii to grade i , < 50% response to tumour growth ] . 4 . no such changes were noticed in the tongue of patient without response ( table i ) . these patients were followed for 6 to 12 months with median follow up of 9 months . total leucocyte count was decreased after cisplatin + 5-fu therapy in all seven patients studied ( table ii ) higher grade of leukepenia was controlled by gcsf . the extent of recovery was much higher in two patients with complete response . in case of platelet count chemotherapy induced decrease was observed in four patients among five patients examined for platelet . ifn2b treatment helped to recover this count in three cases within four . in patient 3 , where complete response was detected , significant recovery of platelet count from 72 , cisplatin + 5-fu therapy along with ifn2b treatment increased the number of cd8 t cells in five patients among seven patients studied . two patients have not shown much change before and after treatment . in case of cd4 t cells , increase in the number no change was reflected in cd4:cd8 ratio , however , this ratio was slightly decreased after chemotherapy and it was increased after ifn2b treatment . in addition to t cells , another important cytotoxic cells , cd56 nk cells , also demonstrated similar profile of changes as observed in case of cd8 t cells . number of cd56 cells was increased in five of the seven patients . among patients with higher expression of cd8 and cd56 markers , complete response was noticed in patient nos . 3 and 6 , with longest survival . cytotoxic ability of pbmc obtained from ifn2b exposed tscc patients against tumour cells was studied ( table ii ) . larynx cancer hep-2 cells were used to get reflection of t cell cytotoxicity and nk sensitive erythroleukemic cells k562 were used to assess nk cell cytotoxicity . four among six patients studied have demonstrated increase in the cytotoxicity towards hep-2 cells , in comparison to their pretreatment values . it was noticed that per cent of cytotoxicity reduced after cis+5-fu treatment was recovered in all patients after ifn2b treatment . the patient ( no . 3 ) with complete response has showed increase in hep-2 cell cytotoxicity in maximum extent ( post - chemotherapy , 7.2% to post - ifn2b , 21.3% ) . cytotoxicity of nk sensitive k562 cells demonstrated similar pattern , as observed in case of hep-2 cells . type 1 cytokine status of tscc patients , undergoing chemo - immunotherapy was studied by monitoring ifng and il-12 , secreted from pbmc at different points of the treatment . 3 with complete response maintained high secretary level of ifn till day 20 after radiotherapy . in case of il-12 , type 2 cytokine status of tscc patients was studied by monitoring il-10 and il-4 , secreted from pbmc at different points of the treatment ( table ii ) . ifn2b therapy reduced secretary il-10 level in four of seven patients ; including those two patients with complete response . other three patients showed no change in comparison to their pre - treatment values . in case of il-4 downregulation we have reported earlier that pbmc isolated from immunosuppressed hnscc patients appear immunoefficient after in vitro stimulation of these cells with ifn2b4 . the detailed haematological and immune functions of patients with tscc were assessed and all patients exhibited poor immune functions in terms of the poor cytotoxic function of pbmc , less number of cytotoxic t and nk cells and type1/type2 cytokine imbalance . these immunocompromised patients appeared more immunologically unstable , when immune parameters were examined after cisplatin+5-fu therapy . patients at this point were immunocompromised with leukepenia and low platelet count . in spite of the efficacy of cisplatin+5-fu treatment in lowering tumour burden of hnscc patients , it is often associated with immune paralysis19 . following cisplatin+5-fu treatment , ifn2b therapy was initiated and patients were examined clinically and immunologically upon completion of the treatment . two patients responded completely and their survival was recorded 28 months after initiation of the treatment . robust immunostimulation by ifn2b was demonstrated in these two patients , particularly in patient no . particular patients reported in clinic as tumor free and clinically fit 50 months after treatment initiation . four patients with partial clinical response also demonstrated immune response following ifn2b therapy and no difference was noted between patients with cr and pr . improved prognosis and prolonged survival of head and neck cancer patients using different chemotherapeutic regimens , along with ifn2b was reported from an initial multicentre open trial15 , however , no effort was done to know the immune status of these patients . volck et al16 treated 14 patients of recurrent hnscc and reported clinical benefit , including complete response in one . they checked the nk cell activity of these patients and observed a superior correlation of nk cell function with clinical outcome . we also found enhancement of the nk cell activity and nk derived cytokine milieu after ifn2b therapy . it was noted that proportion of cd8 cells decreased after chemotherapy , but was increased after completion of chemo - immunotherapy . in most of the cases shah et al20 reported that in cancer cervix patients , survival rate was significantly higher in patients with a high cd4/cd8 ratio as compared to patients who had a low cd4/cd8 ratio . unitt et al21 observed that a high cd4/cd8 ratio was associated with a reduced risk of tumour recurrance after liver transplantation in hepatocellular carcinoma . ifn2b activated t cells can kill cancer cells , possibly by the induction of cytotoxic t lymphocyte ( ctl ) response and antibody dependent cellular cytotoxicity ( adcc ) . ifn2b mediated upregulation of the perforin , granzymeb synthesis and expression was associated with either antibody or ctl mediated tumour cell killing17 . patients with cr were able to maintain the immune system in activated state , till the completion of radiotherapy , 20 days after the completion of the ifn2b therapy . neoadjuvant chemotherapy with platinum and taxens or concomitant chemo - radiotherapy in advanced inoperable tscc patients is a usual clinical practice . results obtained from this preliminary study with cisplatin+5-fu followed by ifn2b suggest that inclusion of ifn2b in the therapeutic protocol enhances the immune response of immunosuppressed patients , that may ultimately enhance the clinical outcome . no mucositis was experienced in any patient after ifn2b therapy possibly due to the use of ifn2b for a short duration ( 3 wk ) . leukepenia and neutropenia were monitored on a regular basis and controlled by the use of gcsf as and when required . similar studies with large number of tscc patients are required to be conducted , where repeated ifn2b therapy can be given to maintain long term immune activation and concomitant radiotherapy can be tested .
background & objectives : interferon alpha 2b ( ifn2b ) has been reported to regulate several immune functions efficiently to enhance the cytotoxic activity of nk and t cells towards various forms of tumours . 
 the objective of the present study was to evaluate the efficacy of ifn2b in overcoming disease induced and/or treatment associated imunosuppression of tongue squamous cell carcinoma ( tscc ) patients undergoing chemotherapy for better clinical outcome.methods:seven tscc patients under cisplatin + 5-fluorouracil chemotherapy in combination with ifn2b were assessed for various immunohaematological parameters before treatment , after chemotherapy and after ifn2b therapy.results:deterioration of the haematological and immune responses was detected in immunosuppressed tscc patients after chemotherapy . 
 ifn2b treatment led to a recovery in these parameters in most of the patients . 
 greater number of t / nk cells and enhanced secretion of type 1 cytokines were also noted . 
 haematological complications were reduced after completion of the therapy . 
 immune- and haematostimulation were also observed in patients with partial response . 
 no positive clinical response was detected in one patient.interpretation & conclusions : ifn2b appears to be an effective immunostimulator having clinical impact to combat the immunosuppression in tscc patients . 
 successful immunostimulation by ifn2b may help tscc patients in clinical improvement . 
 the findings of this preliminary study need to be confirmed on a large number of patients with tscc .