Faculty Bibliography

Abstract Background: Lymphedema (LE) is a frequent complication following breast cancer treatment. While progress is being made in the identification of phenotypic risk factors for the development of LE, little information is available on the molecular characterization of LE. The purpose of this study was to determine if variations in pro- and anti-inflammatory cytokine genes were associated with LE following breast cancer treatment. Methods and Results: Breast cancer patients completed a number of self-report questionnaires. LE was evaluated using bioimpedance spectroscopy. Genotyping was done using a custom genotyping array. No differences were found between patients with (n=155) and without LE (n=387) for the majority of the demographic and clinical characteristics. Patients with LE had a significantly higher body mass index, more advanced disease, and a higher number of lymph nodes removed. Genetic associations were identified for three genes (i.e., interleukin (IL4) 4 (rs2227284), IL 10 (rs1518111), and nuclear kappa factor beta 2 (NFKB2 (rs1056890)) associated with inflammatory responses. Conclusions: These genetic associations suggest a role for a number of pro- and anti-inflammatory genes in the development of LE following breast cancer treatment.

Abstract Preoperative breast pain in women with breast cancer may result from a number of causes. Previous work from our team found that breast pain occurred in 28.2% of women (n = 398) who were about to undergo breast cancer surgery. The occurrence of preoperative breast pain was associated with a number of demographic and clinical characteristics, as well as variation in two cytokine genes. Given that ion channels regulate excitability of sensory neurons, we hypothesized that variations in potassium channel genes would be associated with preoperative breast pain in these patients. Therefore, in this study, we evaluated for associations between single-nucleotide polymorphisms and inferred haplotypes among 10 potassium channel genes and the occurrence of preoperative breast pain in patients scheduled to undergo breast cancer surgery. Multivariable logistic regression analyses were used to identify those genetic variations that were associated with the occurrence of preoperative breast pain while controlling for age and genomic estimates of and self-reported race/ethnicity. Variations in four potassium channel genes: (1) potassium voltage-gated channel, delayed rectifier, subfamily S, member 1 (KCNS1); (2) potassium inwardly rectifying channel, subfamily J, member 3 (KCNJ3); (3) KCNJ6; and (4) potassium channel, subfamily K, member 9 (KCNK9) were associated with the occurrence of breast pain. Findings from this study warrant replication in an independent sample of women who report breast pain following one or more breast biopsies.

BACKGROUND: Cancer pain severely limits function and significantly reduces quality of life. Subtypes of sensory neurons involved in cancer pain and proliferation are not clear. METHODS: We produced a cancer model by inoculating human oral squamous cell carcinoma (SCC) cells into the hind paw of athymic mice. We quantified mechanical and thermal nociception using the paw withdrawal assays. Neurotoxins isolectin B4-saporin (IB4-SAP), or capsaicin was injected intrathecally to selectively ablate IB4(+) neurons or TRPV1(+) neurons, respectively. JNJ-17203212, a TRPV1 antagonist, was also injected intrathecally. TRPV1 protein expression in the spinal cord was quantified with western blot. Paw volume was measured by a plethysmometer and was used as an index for tumor size. Ki-67 immunostaining in mouse paw sections was performed to evaluate cancer proliferation in situ. RESULTS: We showed that mice with SCC exhibited both mechanical and thermal hypersensitivity. Selective ablation of IB4(+) neurons by IB4-SAP decreased mechanical allodynia in mice with SCC. Selective ablation of TRPV1(+) neurons by intrathecal capsaicin injection, or TRPV1 antagonism by JNJ-17203212 in the IB4-SAP treated mice completely reversed SCC-induced thermal hyperalgesia, without affecting mechanical allodynia. Furthermore, TRPV1 protein expression was increased in the spinal cord of SCC mice compared to normal mice. Neither removal of IB4(+) or TRPV1(+) neurons affected SCC proliferation. CONCLUSIONS: We show in a mouse model that IB4(+) neurons play an important role in cancer-induced mechanical allodynia, while TRPV1 mediates cancer-induced thermal hyperalgesia. Characterization of the sensory fiber subtypes responsible for cancer pain could lead to the development of targeted therapeutics.

Persistent pain following breast cancer surgery is a significant clinical problem. Although immune mechanisms may play a role in the development and maintenance of persistent pain, few studies have evaluated for associations between persistent breast pain following breast cancer surgery and variations in cytokine genes. In this study, associations between previously identified extreme persistent breast pain phenotypes (ie, no pain vs severe pain) and single nucleotide polymorphisms (SNPs) spanning 15 cytokine genes were evaluated. In unadjusted analyses, the frequency of 13 SNPs and 3 haplotypes in 7 genes differed significantly between the no pain and severe pain classes. After adjustment for preoperative breast pain and the severity of average postoperative pain, 1 SNP (ie, interleukin [IL] 1 receptor 2 rs11674595) and 1 haplotype (ie, IL10 haplotype A8) were associated with pain group membership. These findings suggest a role for cytokine gene polymorphisms in the development of persistent breast pain following breast cancer surgery. PERSPECTIVE: This study evaluated for associations between cytokine gene variations and the severity of persistent breast pain in women following breast cancer surgery. Variations in 2 cytokine genes were associated with severe breast pain. The results suggest that cytokines play a role in the development of persistent postsurgical pain.

Individual bacteria and shifts in the composition of the microbiome have been associated with human diseases including cancer. To investigate changes in the microbiome associated with oral cancers, we profiled cancers and anatomically matched contralateral normal tissue from the same patient by sequencing 16S rDNA hypervariable region amplicons. In cancer samples from both a discovery and a subsequent confirmation cohort, abundance of Firmicutes (especially Streptococcus) and Actinobacteria (especially Rothia) was significantly decreased relative to contralateral normal samples from the same patient. Significant decreases in abundance of these phyla were observed for pre-cancers, but not when comparing samples from contralateral sites (tongue and floor of mouth) from healthy individuals. Weighted UniFrac principal coordinates analysis based on 12 taxa separated most cancers from other samples with greatest separation of node positive cases. These studies begin to develop a framework for exploiting the oral microbiome for monitoring oral cancer development, progression and recurrence.

Cisplatin resistance in head and neck squamous cell carcinoma (HNSCC) reduces survival. In this study we hypothesized that methylation of key genes mediates cisplatin resistance. We determined whether a demethylating drug, decitabine, could augment the anti-proliferative and apoptotic effects of cisplatin on SCC-25/CP, a cisplatin-resistant tongue SCC cell line. We showed that decitabine treatment restored cisplatin sensitivity in SCC-25/CP and significantly reduced the cisplatin dose required to induce apoptosis. We then created a xenograft model with SCC-25/CP and determined that decitabine and cisplatin combination treatment resulted in significantly reduced tumor growth and mechanical allodynia compared to control. To establish a gene classifier we quantified methylation in cancer tissue of cisplatin-sensitive and cisplatin-resistant HNSCC patients. Cisplatin-sensitive and cisplatin-resistant patient tumors had distinct methylation profiles. When we quantified methylation and expression of genes in the classifier in HNSCC cells in vitro, we showed that decitabine treatment of cisplatin-resistant HNSCC cells reversed methylation and gene expression toward a cisplatin-sensitive profile. The study provides direct evidence that decitabine restores cisplatin sensitivity in in vitro and in vivo models of HNSCC. Combination treatment of cisplatin and decitabine significantly reduces HNSCC growth and HNSCC pain. Furthermore, gene methylation could be used as a biomarker of cisplatin-resistance.

Cisplatin is the primary chemotherapy for head and neck squamous cell carcinoma (HNSCC). No equally effective chemotherapeutics are available when cisplatin resistance occurs. We hypothesize that DNA methylation of key genes mediates cisplatin resistance; moreover, pretreatment with decitabine, a demethylating agent, restores cisplatin sensitivity by mediating expression of genes that are instrumental to cisplatin resistance. Objectives: 1) Determine whether decitabine treatment of a cisplatin-resistant HNSCC cell line restores the anti-proliferative and apoptotic effects of cisplatin; 2) Evaluate the anti-proliferative effect of decitabine and cisplatin (i.e. combination treatment) on a preclinical HNSCC model; 3) Determine whether combination treatment reduces cancer pain; and 4) Create a "gene expression profile of cisplatin resistance" by analyzing cisplatinsensitive and cisplatin-resistant HNSCC in patients. Methods: SCC-25, a cisplatin-sensitive HNSCC cell line, and SCC-25/CP, a cisplatin-resistant cell line, were pre-treated with 5mM decitabine and then treated with cisplatin (3-300 mM) for 48 hours. Proliferation was quantified with an MTS assay. Apoptosis was quantified with a caspase 3/7 assay. A preclinical model was created by inoculating SCC-25/CP cells into the hind-paw of BALB/ c mice. Twenty-four mice were placed into one of four treatment groups: control sham, decitabine-only, cisplatin- only, or combination treatment. Decitabine (6 mg/kg) was administered on post-inoculation days (PID) seven and nine, and cisplatin (6 mg/kg) was administered on PID 12, 15, 18, and 21. Tumor growth was quantified. Mechanical allodynia (i.e. pain) was quantified with a paw withdrawal assay. Formalin-fixed, paraffin- embedded biopsies were obtained from HNSCC patients who underwent chemotherapy with cisplatin. Tumors were classified as either cisplatin-sensitive (RECIST 3 or 4) or cisplatin-resistant (RECIST 1 or 2). Gene expression was quantified in these two sets of samples. Results: In the in !

PURPOSE: Surgical resection of midface neoplasms and subsequent reconstruction have been shown to have significant negative effects on quality of life (QOL). The purpose of this pilot study was to assess individuals' health-related QOL after maxillectomy and reconstruction with a prosthetic obturator. MATERIALS AND METHODS: The QOL of 25 of 43 patients who underwent maxillectomy and prosthetic obturator reconstruction at the University of California-San Francisco was assessed using 3 questionnaires: University of Washington Quality of Life version 4 (UWQOL), Obturator Functioning Scale (OFS), and Mental Health Inventory (MHI). RESULTS: The response rate to the QOL questionnaires was 92% (23 of 25 patients). Time elapsed from maxillectomy and prosthetic obturator reconstruction to the QOL survey response ranged from 0.3 to 6.6 years (mean, 2.7 years; standard deviation [SD], 1.9 years). The post-treatment mean QOL scores were 77.3 (SD, 13.6) for UWQOL, 72.0 (SD, 12.6) for OFS, and 4.5 (SD, 0.9) for Mental Health Inventory. Individuals who received adjuvant radiation scored lower for speech and appearance (OFS, P = .05, P = .03, respectively) as well as for saliva and overall QOL (UWQOL, P = .02, P = .08, respectively). There was a strong correlation between QOL scores in OFS and UWQOL questionnaires (r = 0.78, P < .001). CONCLUSION: The results of this pilot study suggest that postoperative radiation therapy was the strongest variable affecting QOL in patients with maxillectomy and prosthetic obturator reconstruction. There is further need for a multicenter trial with a larger sample to identify how factors affecting QOL of patients after maxillectomy might influence the choice of reconstruction.