CD44 is a cell surface HA‐binding glycoprotein that is overexpressed to some extent by almost all tumors of epithelial origin and plays an important role in tumor initiation and metastasis. CD44 is a compelling marker for cancer stem cells of many solid malignancies. In addition, interaction of HA and CD44 promotes EGFR‐mediated pathways, consequently leading to tumor cell growth, tumor cell migration, and chemotherapy resistance in solid cancers. Accumulating evidence indicates that major HA‐CD44 signaling pathways involve a specific variant of CD44 isoforms; however, the particular variant almost certainly depends on the type of tumor cell and the stage of the cancer progression. Research to date suggests use of monoclonal antibodies against different CD44 variant isoforms and targeted inhibition of HA/CD44‐mediated signaling combined with conventional radio/chemotherapy may be the most favorable therapeutic strategy for future treatments of advanced stage malignancies. Thus, this paper briefly focuses on the association of the major CD44 variant isoforms in cancer progression, the role of HA‐CD44 interaction in oncogenic pathways, and strategies to target CD44‐overexpressed tumor cells.

Purpose: Randomized controlled trials have generally shown a benefit from accelerated radiotherapy in head and neck squamous cell carcinoma (HNSCC). However, the large randomized United Kingdom trial CHART (Continuous Hyperfractionated Accelerated Radiotherapy) failed to show a benefit of strongly accelerated over standard radiotherapy (RT) in 918 patients with HNSCC. In this study, we investigated the impact of tumor hypoxia on the outcome of HNSCC patients in the CHART trial. There are two distinct hypoxia inducible factors (HIFs) that control different gene response pathways and we assessed them both with endogenous markers of hypoxia, hypoxia inducible factor HIF-2 alpha (HIF-2) and carbonic anhydrase CA9, an indicator of HIF-1 alpha (HIF-1) function.Methods: Tissue from pre-RT biopsies performed in 198 of 918 patients recruited was analyzed for the immunohistochemical expression of HIF-2 and CA9.Results: A significant association of high HIF2 and of high CA9 reactivity with poor locoregional control (P < .0001 and P = .0002, respectively) and poor survival (P = .0004 and 0.002, respectively) was noted. In multivariate analysis, HIF-2 and CA9 maintained their independent prognostic significance. Coexpression of both pathways had an additive effect, supporting their independent role. The uni-directional hypothesis, that a benefit from randomization to CHART should be seen in the nonhypoxic tumors, was supported by the data (one-tailed P = .04).Conclusion: Expression of endogenous markers of hypoxia for the HIF-1 and HIF-2 pathway is strongly associated with radiotherapy failure. Using immunohistochemical methods it is possible to identify subgroups of HNSCC patients who are highly curable with radiotherapy, or who are excellent candidates for clinical trials on hypoxia-targeting drugs in two distinct pathways.

Purpose Accelerated repopulation is a main reason for locoregional failure after fractionated radiotherapy for head and neck squamous cell carcinoma (HNSCC). Epidermal growth factor receptor (EGFR) is a key controller of cellular proliferation in HNSCC, which stimulated the current study to look for a direct link between EGFR status and a possible clinical advantage of accelerated radiotherapy. Patients and Methods Immunohistochemical staining for EGFR was performed in 304 patients with available pretreatment tumor biopsy material among 918 patients randomized to receive continuous hyperfractionated accelerated radiotherapy versus conventionally fractionated radiotherapy. The EGFR index was estimated as the proportion of tumor cells with EGFR membrane staining. Results Significant benefit in locoregional tumor control from continuous hyperfractionated accelerated radiotherapy was seen in patients with HNSCC with high EGFR expression (2P = .010) but not in those with low EGFR expression (2P = .85). EGFR status had no significant effect on survival or rate of distant metastases. The EGFR index was significantly associated with histologic grade and microvessel density. There was moderate support for an association between EGFR status and subsite within the head and neck region but no significant association with Ki-67 index, Ki-67 pattern, p53 index, p53 intensity, bcl-2 expression, or cyclin D1 index. Conclusion This study indicates a key role for the EGFR receptor in determining the proliferative cellular response to fractionated radiotherapy in HNSCC. It also shows that we can select the dose-fractionation regime that has the greatest chance of benefiting the patient. These results also encourage further development of EGFR targeting combined with fractionated radiotherapy in HNSCC.

Glucose transporter-1 protein (GLUT1) and carbonic anhydrase IX (CAIX) are regulated by hypoxia inducible factor-1 (HIF-1) and have been studied as putative intrinsic cellular markers for hypoxia. This study directly compares CAIX and GLUT1 with pimonidazole binding in a prospective series of bladder cancer patients and also studies the prognostic significance of the markers, in combination with vascularity and proliferation, in a retrospective series of bladder cancer patients treated in a phase II trial of radical radiotherapy with carbogen and nicotinamide (ARCON). A total of 21 patients with a diagnosis of transitional cell carcinoma of the bladder received 0.5 g m(-2) pimonidazole. Serial tumour sections were stained for pimonidazole, GLUT1 and CAIX and compared. Tissue sections obtained from a series of 64 patients previously treated for invasive bladder cancer using ARCON were stained for GLUT1 and CAIX together with Ki-67 and CD31/34. There was a good geographical colocalisation of both intrinsic markers with pimonidazole and a highly significant agreement in individual patients; correlation coefficients were 0.82 (P=0.0001) for GLUT1 and 0.74 (P<0.0001) for CAIX. In both series of patients, the intrinsic hypoxia markers were highly correlated with each other and a correlation with proliferation was also evident in the retrospective study. In univariate and multivariate analyses, GLUT1 and CAIX were independent predictors for overall and cause specific survival. The hypoxia markers did not predict for local control or metastases-free survival although higher Ki-67 indices showed a trend towards local failure. The data suggest that both hypoxia modification and accelerated treatment may be valid treatment options in bladder cancer.

In vivo bromodeoxyuridine (BrdUrd) labelling of the human large bowel was performed and a detailed histochemical localisation of label in sections of crypts was undertaken using a monoclonal antibody to BrdUrd containing DNA. Flow cytometric studies on extracted nuclei were also performed (data presented elsewhere). The average crypt in the human large bowel (excluding the rectum) was 82 cells in height and 41 cells in circumference, with a total of about 2000 cells (assuming a topographical correction factor of 0.6). Ten per cent of the cells were replicating their DNA--that is, were in the S phase of the cell cycle--and 0.4% were in mitosis. The median position for the labelling index versus cell position frequency plot is at the 20th cell position--at a quarter of the crypt height. The lower and upper limits of the cell proliferation are given by the 5th and 95th percentiles at cell positions 4 and 43 respectively. The peak labelling index is about 30% and it occurs at cell position 15. The labelling index at the crypt base, the probable stem cell zone, is about 14%, suggesting that these cells have a longer cell cycle. Taking a value of 8.6 hours for the duration of the S phase (deduced from the flow cytometric data) and assuming a growth fraction of 1.0 for the mid-crypt, these data provide an estimate of about 30 hours for the cell cycle time. The rectal crypts are about the same size but contain about 30% fewer S phase cells. The data also yielded a per cent BrdUrd labelled mitosis curve.