Stress Keratin 17 Is a Predictive Biomarker Inversely Associated with Response to Immune Check-Point Blockade in Head and Neck Squamous Cell Carcinomas and Beyond.

Low response rates in immune check-point blockade (ICB)-treated head and neck squamous cell carcinoma (HNSCC) drive a critical need for robust, clinically validated predictive biomarkers. Our group previously showed that stress keratin 17 (CK17) suppresses macrophage-mediated CXCL9/CXCL10 chemokine signaling involved in attracting activated CD8+ T cells into tumors, correlating with decreased response rate to pembrolizumab-based therapy in a pilot cohort of ICB-treated HNSCC (n = 26). Here, we performed an expanded analysis of the predictive value of CK17 in ICB-treated HNSCC according to the REMARK criteria and investigated the gene expression profiles associated with high CK17 expression. Pretreatment samples from pembrolizumab-treated HNSCC patients were stained via immunohistochemistry using a CK17 monoclonal antibody (n = 48) and subjected to spatial transcriptomic profiling (n = 8). Our findings were validated in an independent retrospective cohort (n = 22). CK17 RNA expression in pembrolizumab-treated patients with various cancer types was investigated for predictive significance. Of the 48 patients (60% male, median age of 61.5 years), 21 (44%) were CK17 high, and 27 (56%) were CK17 low. A total of 17 patients (35%, 77% CK17 low) had disease control, while 31 patients (65%, 45% CK17 low) had progressive disease. High CK17 expression was associated with a lack of disease control (p = 0.037), shorter time to treatment failure (p = 0.025), and progression-free survival (PFS, p = 0.004), but not overall survival (OS, p = 0.06). A high CK17 expression was associated with lack of disease control in an independent validation cohort (p = 0.011). PD-L1 expression did not correlate with CK17 expression or clinical outcome. CK17 RNA expression was predictive of PFS and OS in 552 pembrolizumab-treated cancer patients. Our findings indicate that high CK17 expression may predict resistance to ICB in HNSCC patients and beyond.

Betapapillomaviruses in p16-Negative Vulvar Intraepithelial Lesions Associated with Squamous Cell Carcinoma.

Approximately 40% of vulvar squamous cell carcinoma (vSCC) cases are etiologically associated with high-risk human papillomaviruses (HPVs) of the alpha genera (α-HPV) that cause other anogenital cancers; however, the etiology of α-HPV-negative vSCC is poorly understood. HPVs of the beta genera (ß-HPV) are risk factors for cutaneous squamous cell carcinoma (cSCC) and may be related to carcinomas originating in other cutaneous sites such as the vulva. In this study, we investigate the presence of ß-HPVs, with an emphasis on p16-negative squamous lesions adjacent to vSCC. We subjected 28 vulvar squamous intraepithelial lesions adjacent to vSCC for comprehensive HPV genotyping, p16 and p53 immunohistochemistry, and consensus morphology review. Selected cases were subjected to qPCR and RNA in situ hybridization. Clinical data were obtained from medical records. ß-HPV DNA was detected in eight of ten p16-negative lesions and three of fourteen p16-positive high-grade squamous intraepithelial lesions. The HPV DNA loads in vulvar squamous intraepithelial lesions ranged between less than 1 HPV DNA copy per cell to more than 100 HPV DNA copies per cell. This is, to the best of our knowledge, the first report of the association of p16-negative vulvar intraepithelial squamous lesions with detection of ß-HPVs. These findings expand possible etiologic mechanisms that may contribute to p16-negative lesions of the vulva.

Stress Keratin 17 Expression in Head and Neck Cancer Contributes to Immune Evasion and Resistance to Immune-Checkpoint Blockade.

PURPOSE: We investigated whether in human head and neck squamous cell carcinoma (HNSCC) high levels of expression of stress keratin 17 (K17) are associated with poor survival and resistance to immunotherapy. EXPERIMENTAL DESIGN: We investigated the role of K17 in regulating both the tumor microenvironment and immune responsiveness of HNSCC using a syngeneic mouse HNSCC model, MOC2. MOC2 gives rise to immunologically cold tumors that are resistant to immune-checkpoint blockade (ICB). We engineered multiple, independent K17 knockout (KO) MOC2 cell lines and monitored their growth and response to ICB. We also measured K17 expression in human HNSCC of patients undergoing ICB. RESULTS: MOC2 tumors were found to express K17 at high levels. When knocked out for K17 (K17KO MOC2), these cells formed tumors that grew slowly or spontaneously regressed and had a high CD8+ T-cell infiltrate in immunocompetent syngeneic C57BL/6 mice compared with parental MOC2 tumors. This phenotype was reversed when we depleted mice for T cells. Whereas parental MOC2 tumors were resistant to ICB treatment, K17KO MOC2 tumors that did not spontaneously regress were eliminated upon ICB treatment. In a cohort of patients with HNSCC receiving pembrolizumab, high K17 expression correlated with poor response. Single-cell RNA-sequencing analysis revealed broad differences in the immune landscape of K17KO MOC2 tumors compared with parental MOC2 tumors, including differences in multiple lymphoid and myeloid cell types. CONCLUSIONS: We demonstrate that K17 expression in HNSCC contributes to immune evasion and resistance to ICB treatment by broadly altering immune landscapes of tumors.

The immunoregulatory landscape of human tuberculosis granulomas.

Tuberculosis (TB) in humans is characterized by formation of immune-rich granulomas in infected tissues, the architecture and composition of which are thought to affect disease outcome. However, our understanding of the spatial relationships that control human granulomas is limited. Here, we used multiplexed ion beam imaging by time of flight (MIBI-TOF) to image 37 proteins in tissues from patients with active TB. We constructed a comprehensive atlas that maps 19 cell subsets across 8 spatial microenvironments. This atlas shows an IFN-γ-depleted microenvironment enriched for TGF-ß, regulatory T cells and IDO1+ PD-L1+ myeloid cells. In a further transcriptomic meta-analysis of peripheral blood from patients with TB, immunoregulatory trends mirror those identified by granuloma imaging. Notably, PD-L1 expression is associated with progression to active TB and treatment response. These data indicate that in TB granulomas, there are local spatially coordinated immunoregulatory programs with systemic manifestations that define active TB.

Intravascular adenomyomatosis: a morphologic variant of intravenous leiomyomatosis associated with endometriosis and potential for misdiagnosis.

Intravenous leiomyomatosis (IVL) is a quasi-malignant smooth muscle tumor involving lymphatic and venous spaces of the myometrium. Rare cases of IVL with admixed endometrial glands and stroma have been described, termed intravascular adenomyomatosis. We report four additional cases of intravascular adenomyomatosis and expand the clinicopathologic features of these rare tumors. Patients were 39-45 years old and presented with symptoms of dysmenorrhea, postmenopausal bleeding, or pelvic mass. All cases were associated with endometriosis. Three cases comprised intravascular bland smooth muscle tumors with plexiform features, and in some foci, the intravascular tumor contained endometrial type glands and stroma. In one case, there was extensive (>10) foci of intravascular adenomyomatosis without evidence of associated smooth muscle neoplasm but did have an endometrial polyp with adenomyomatous features. None of the cases had nuclear atypia, increased mitotic activity, or tumor cell necrosis. The endometrial stromal components were positive for CD10 and negative or weakly positive for desmin by immunohistochemistry. Two cases underwent molecular testing for JAZF1 and PHF1 rearrangements with negative results. Three patients had no evidence of disease at the time of the last follow-up, and one had persistent but stable disease 7 years after incomplete surgical removal and megestrol acetate treatment. Intravascular adenomyomatosis is a variant morphology rarely seen in IVL that lacks characteristic morphologic and molecular features of endometrial stromal sarcoma. Similar to IVL, prognosis is likely linked to completeness of surgical resection. In this study, we found that intravascular adenomyomatosis is frequently associated with endometriosis, a novel finding to add to the literature on this rare IVL variant.

Pregnancy-associated Inflammatory Myofibroblastic Tumors of the Uterus Are Clinically Distinct and Highly Enriched for TIMP3-ALK and THBS1-ALK Fusions.

As inflammatory myofibroblastic tumors (IMTs) have become more widely recognized in the female genital tract, an intriguing subset of uterine tumors associated with pregnancy has emerged. Whether uterine IMTs occurring in the setting of pregnancy are clinically or biologically distinct from other uterine IMTs is unknown. Furthermore, little is known about the perinatal factors that may influence the development of these tumors. Here, we report the largest case series of 8 pregnancy-associated IMTs. All pregnancy-associated IMTs in this series occurred in association with pregnancy complications, including abnormal implantation (n=1), gestational diabetes (n=2), preeclampsia and/or HELLP syndrome (n=2), antiphospholipid syndrome (n=1), premature rupture of membranes (n=1), and hepatitis B (n=1). Notably, all IMTs were expelled at the time of delivery or immediately postpartum and were either adherent to the placenta or presented as separate, detached tissue. Tumors ranged from 2.0 to 6.0 cm (median, 3.9 cm), were well-circumscribed and showed classic histologic features of IMTs, including myxoid stroma and a lymphoplasmacytic infiltrate. Seven of 8 cases were positive by ALK immunohistochemistry and confirmed to have an ALK gene rearrangement by fluorescent in situ hybridization and RNA sequencing. The ALK-rearranged IMTs were found to be particularly enriched for TIMP3-ALK (n=5) and THBS1-ALK (n=2) fusions. The single case that was negative for an ALK rearrangement exhibited the classic morphology of an IMT. None of the 4 cases with available clinical follow-up recurred. The clinicopathologic features of pregnancy-associated IMTs in this series in conjunction with those reported in the literature suggests that these may be transient tumors that develop during pregnancy and shed at parturition; they appear to have a relatively indolent clinical course and favorable outcome, although studies with a longer duration of follow-up are still required.

Whole-Genome Analysis of Cervical Human Papillomavirus Type 35 from rural Zimbabwean Women.

Human papillomavirus (HPV) types differ by geographic location and the ethnicity of the human host, which may have implications for carcinogenicity. HPV35 is one of the least frequently identified high-risk types in North America and Europe but was the most common high-risk HPV (hrHPV) infection in a cohort in rural Zimbabwe. Whole genome analysis is limited for HPV35; no such studies have been performed in Zimbabwe. Of 648 women in the initial cohort in Zimbabwe, 19 (19/648, 2.9%) tested positive for HPV35, and eight samples were successfully sequenced for HPV35. The maximum number of sequence variants for the whole genome was 58 nucleotides (0.7%) compared to the prototype (58/7879). The maximum number of sequence variants in E6 and E7 was 3 (3/450, 0.7%) 2 (2/300, 0.7%), respectively. These are the first HPV35 whole genome sequences from Zimbabwe, and these data further lend support to the carcinogenicity of HPV35 despite limited sequence heterogeneity. Further studies to determine carcinogenic effects and impact of HPV vaccinations are warranted, especially in sub-Saharan Africa.

Knowledge and awareness of cervical cancer in Southwestern Ethiopia is lacking: A descriptive analysis.

PURPOSE: Cervical cancer remains the second most common cancer and cancer-related death among women in Ethiopia. This is the first study, to our knowledge, describing the demographic, and clinicopathologic characteristics of cervical cancer cases in a mainly rural, Southwestern Ethiopian population with a low literacy rate to provide data on the cervical cancer burden and help guide future prevention and intervention efforts. METHODS: A descriptive analysis of 154 cervical cancer cases at the Jimma University Teaching Hospital in Southwestern Ethiopia from January 2008 -December 2010 was performed. Demographic and clinical characteristics were obtained from patient questionnaires and cervical punch biopsies were histologically examined. RESULTS: Of the 154 participants with a histopathologic diagnosis of cervical cancer, 95.36% had not heard of cervical cancer and 89.6% were locally advanced at the time of diagnosis. Moreover, 86.4% of participants were illiterate, and 62% lived in a rural area. CONCLUSION: A majority of the 154 women with cervical cancer studied at the Jimma University Teaching Hospital in Southwestern Ethiopia were illiterate, had not heard of cervical cancer and had advanced disease at the time of diagnosis. Given the low rates of literacy and knowledge regarding cervical cancer in this population which has been shown to correlate with a decreased odds of undergoing screening, future interventions to address the cervical cancer burden here must include an effective educational component.

Community-based self-collected human papillomavirus screening in rural Zimbabwe.

BACKGROUND: In low- and middle-income countries (LMIC), women have limited access to and uptake of cervical cancer screening. Delayed diagnosis leads to poorer outcomes and early mortality, and continues to impede cancer control disproportionately in LMIC. Integrating self-collected, community-based screening for High Risk-Human Papilloma Virus (HR-HPV) into existent HIV programs is a potential screening method to identify women at high risk for developing high-risk cervical lesions. METHODS: We implemented community-based cross-sectional study on self-collection HR-HPV screening in conjunction with existing community outreach models for the distribution of antiretroviral therapy (ART) and the World Health Organization Expanded Program on Immunization (EPI) outreach in villages in rural Zimbabwe from January 2017 through May 2017. RESULTS: Overall, there was an 82% response rate: 70% of respondents participated in self-collection and 12% were ineligible for the study (inclusion criteria: age 30-65, not pregnant, with an intact uterus). Women recruited in the first 2-3 months of the study had more opportunities to participate and therefore significantly higher participation: 81% participation (additional 11% ineligible), while those with fewer opportunities also had lower participation: 63% (additional 13% ineligible) (p < 0.001). Some village outreach centers (N = 5/12) had greater than 89% participation. CONCLUSIONS: Integration of HR-HPV screening into existing community outreach models for HIV and immunizations could facilitate population-based screening to scale cancer control and prevention programs in sub-Saharan Africa. Community/village health workers (CHW/VHW) and village outreach programs offer a potential option for cervical cancer screening programs to move towards improving access of sexual and reproductive health resources for women at highest risk.

hrHPV prevalence and type distribution in rural Zimbabwe: A community-based self-collection study using near-point-of-care GeneXpert HPV testing.

OBJECTIVES: High-risk human papilloma viruses (hrHPV) are the causative agents of cervical cancer, the leading cause of cancer deaths among Zimbabwean women. The objective of this study was to describe the hrHPV types found in Zimbabwe for consideration in cervical cancer screening and vaccination efforts. DESIGN AND METHODS: To determine hrHPV prevalence and type distribution in Zimbabwe we implemented a community-based cross-sectional study of self-collected cervicovaginal samples with hrHPV screening using near-point-of-care Cepheid GeneXpert HPV. RESULTS: The hrHPV prevalence was 17% (112/643); 33% (41/123) vs. 14% (71/520) among HIV-1-positive and -negative participants, respectively (p=2.3E-07). Typing via Xpert HPV showed very good overall agreement (77.2%, kappa=0.698) with the Seegene Anyplex II HPV HR Detection kit. The most common types were HPV16, HPV18, HPV35, HPV52, HPV58, HPV68, HPV18, and HPV51, each of which appeared in 14-20% of infections. 37% (28/76) of women with positive cytology results (ASCUS+) had a type not included in the basic vaccine and 25% (19/76) had a type not currently in the nine-valent vaccine. CONCLUSIONS: hrHPV type distribution includes less common high-risk types in rural Zimbabwe. The distribution and carcinogenicity of hrHPV type distribution should be considered during screening assay design, program development, as well as vaccine distribution and design.