Relationship of histologic grade and histologic subtype with oncotype Dx recurrence score; retrospective review of 863 breast cancer oncotype Dx results.

PURPOSE: Oncotype Dx (ODx) is a multigene assay that is prognostic and predictive in estrogen receptor (ER) positive early breast cancer. ODx recurrence score (RS) is reported to be histologic grade dependent. Relationship of RS with breast cancer histologic subtypes is unknown. This study was designed to assess the relationship of histologic subtype with RS. Histologic grade dependence of RS was also investigated. METHODS: Results of consecutive ODx tests (1/2007-7/2016) from two institutions were reviewed. Histologic subtypes (in: Lakhani et al., WHO classification, IARC Press, Lyon, 2012), combined Nottingham histologic grade, age and tumor size were recorded from pathology reports. Univariate and multivariate analysis was performed to investigate the relationship between RS and ODx risk categories and histologic subtypes, grade, age and tumor size. RESULTS: RS was grade dependent. RS of grade 1 and grade 2 tumors were significantly lower than grade 3 tumors. There was no high-risk grade 1 tumor. In favorable histologic subtypes there was no high-risk tumor. Mean RS of grade 1 lobular tumors was significantly higher than grade 1 ductal tumors. Using newer ODx cut-offs, 5 grade 1 tumors were reclassified as high risk (RS > 25) and grade 3 lobular tumors showed significantly higher rate of reclassification as high-risk than grade 3 ductal tumors. In a multivariate analysis, only grade showed a significant positive correlation with RS. Adding dichotomous histologic subtyping (favorable vs. non-favorable) to grade further improved correlation with RS. CONCLUSIONS: The Oncotype Dx result is impacted by histologic grade and histologic subtype. Tumors with favorable histologic subtypes and histologic grade 1 tumors do not have high-risk RS. High RS in a grade 1 tumor or in a tumor with favorable histology is unusual that warrants further investigation. Invasive lobular carcinomas rarely show high-risk RS. Histologic grade and histologic subtype should be considered while ordering ODx testing.

Evaluating agreement, histological features, and relevance of separating pleomorphic and florid lobular carcinoma in situ subtypes.

Morphological variants of lobular carcinoma in situ (LCIS) include classical (CLCIS), pleomorphic (PLCIS) and florid type (FLCIS). Treatment guidelines suggest managing PLCIS and FLCIS like ductal carcinoma in situ (DCIS); therefore accurate identification of LCIS subtypes is critical. However, the significance of separating PLCIS from FLCIS is not clear. Also, interobserver agreement in identifying LCIS subtypes, using contemporary criteria, is not known. We aimed to evaluate interobserver agreement amongst breast pathologists in diagnosing LCIS subtypes and use the agreement data to justify LCIS classification for management purposes. Six breast pathologists independently reviewed 50 hematoxylin and eosin-stained slides comprised of a mix of LCIS subtypes. After reviewing published criteria, participants diagnosed PLCIS, CLCIS and apocrine change in a marked region of interest and FLCIS based on entire section. PLCIS was identified in 8 to 37 slides with overall moderate agreement (Fleiss' κ = 0.565) and pairwise κ (Cohen's) ranging from -.008 to 0.492. FLCIS was diagnosed in 15-26 slides with overall substantial agreement (Fleiss' κ = 0.687) and pairwise κ ranging from -.068 to 0.706. Both FLCIS and PLCIS coexisted in 45% of slides with consensus on non-classical LCIS. Comedo-type necrosis (odds ratio = 5.5) and apoptosis (odds ratio = 1.8) predicted FLCIS. We found moderate and substantial agreement in diagnosing PLCIS and FLCIS respectively. Objective histological features linked with aggressive behavior were more frequent with FLCIS. PLCIS and FLCIS patterns frequently coexist, contain similar molecular aberrations, and are managed similarly (like DCIS); therefore, combining FLCIS and PLCIS into one category (non-classical LCIS) should be considered.

Comparison of estrogen receptor, progesterone receptor and HER2 results in concurrent ipsilateral samples with invasive breast carcinoma: a retrospective study of 246 biopsies from 119 patients.

There is no consensus regarding biomarker testing on the ipsilateral breast carcinomas present in separate biopsies, irrespective of whether the biopsies are performed concurrently or consecutively. We aimed to investigate estrogen receptor (ER), progesterone receptor (PR) and HER2 concordance in ipsilateral concurrent biopsies with invasive breast tumors. Consecutive ipsilateral concurrent biopsies with invasive breast tumors were identified retrospectively. Biomarker results, histologic grade and histologic subtype among the tumors in concurrent samples were compared. ER, PR, and HER2 expression was different in 3 (2.5%), 11 (9.2%) and 7 (5.9%) cases, respectively. All ER-discordant cases were sets of ER-negative (ER-) and weak-low ER-positive (ER+), ductal subtype and histologic grade 2 or 3 tumors. All PR-discordant cases were ER+, and comprised of histologic grades 1 to 3 ductal as well as lobular tumors. All HER2 discordant cases were histologic grade 2 to 3 ductal tumors. Biomarker discordance was independent of grade and subtype discordance. We found very low biomarker discordance among tumors in concurrent samples from ipsilateral breast. Our results suggest that ER and HER2 discordance in concurrent samples is predictable. ER discordance is present only in a setting of low ER+ tumors. Low-grade ductal and/or lobular tumors are ER and HER2 concordant. HER2 discordance is noted in grade 2 to 3 ductal tumors only. Histologic subtype and grade may guide extent of biomarker testing in concurrent ipsilateral breast biopsies.

HLA Alleles Associated with Delayed Progression to AIDS Contribute Strongly to the Initial CD8(+) T Cell Response against HIV-1.

BACKGROUND: Very little is known about the immunodominance patterns of HIV-1-specific T cell responses during primary HIV-1 infection and the reasons for human lymphocyte antigen (HLA) modulation of disease progression. METHODS AND FINDINGS: In a cohort of 104 individuals with primary HIV-1 infection, we demonstrate that a subset of CD8(+) T cell epitopes within HIV-1 are consistently targeted early after infection, while other epitopes subsequently targeted through the same HLA class I alleles are rarely recognized. Certain HLA alleles consistently contributed more than others to the total virus-specific CD8(+) T cell response during primary infection, and also reduced the absolute magnitude of responses restricted by other alleles if coexpressed in the same individual, consistent with immunodomination. Furthermore, individual HLA class I alleles that have been associated with slower HIV-1 disease progression contributed strongly to the total HIV-1-specific CD8(+) T cell response during primary infection. CONCLUSIONS: These data demonstrate consistent immunodominance patterns of HIV-1-specific CD8(+) T cell responses during primary infection and provide a mechanistic explanation for the protective effect of specific HLA class I alleles on HIV-1 disease progression.

Limited durability of viral control following treated acute HIV infection.

BACKGROUND: Early treatment of acute HIV infection with highly active antiretroviral therapy, followed by supervised treatment interruption (STI), has been associated with at least transient control of viremia. However, the durability of such control remains unclear. Here we present longitudinal follow-up of a single-arm, open-label study assessing the impact of STI in the setting of acute HIV-1 infection. METHODS AND FINDINGS: Fourteen patients were treated during acute HIV-1 infection and subsequently subjected to an STI protocol that required retreatment if viral load exceeded 50,000 RNA copies/ml plasma or remained above 5,000 copies/ml for more than three consecutive weeks. Eleven of 14 (79%) patients were able to achieve viral loads of less than 5,000 RNA copies/ml for at least 90 d following one, two, or three interruptions of treatment. However, a gradual increase in viremia and decline in CD4+ T cell counts was observed in most individuals. By an intention-to-treat analysis, eight (57%), six (43%), and three (21%) of 14 patients achieved a maximal period of control of 180, 360, and 720 d, respectively, despite augmentation of HIV-specific CD4+ and CD8+ T cell responses. The magnitude of HIV-1-specific cellular immune responses before treatment interruption did not predict duration of viremia control. The small sample size and lack of concurrent untreated controls preclude assessment of possible clinical benefit despite failure to control viremia by study criteria. CONCLUSIONS: These data indicate that despite initial control of viremia, durable viral control to less than 5,000 RNA copies/ml plasma in patients following treated acute HIV-1 infection occurs infrequently. Determination of whether early treatment leads to overall clinical benefit will require a larger and randomized clinical trial. These data may be relevant to current efforts to develop an HIV-1 vaccine designed to retard disease progression rather than prevent infection since they indicate that durable maintenance of low-level viremia may be difficult to achieve.

De novo generation of escape variant-specific CD8+ T-cell responses following cytotoxic T-lymphocyte escape in chronic human immunodeficiency virus type 1 infection.

Human immunodeficiency virus type 1 (HIV-1) evades CD8(+) T-cell responses through mutations within targeted epitopes, but little is known regarding its ability to generate de novo CD8(+) T-cell responses to such mutants. Here we examined gamma interferon-positive, HIV-1-specific CD8(+) T-cell responses and autologous viral sequences in an HIV-1-infected individual for more than 6 years following acute infection. Fourteen optimal HIV-1 T-cell epitopes were targeted by CD8(+) T cells, four of which underwent mutation associated with dramatic loss of the original CD8(+) response. However, following the G(357)S escape in the HLA-A11-restricted Gag(349-359) epitope and the decline of wild-type-specific CD8(+) T-cell responses, a novel CD8(+) T-cell response equal in magnitude to the original response was generated against the variant epitope. CD8(+) T cells targeting the variant epitope did not exhibit cross-reactivity against the wild-type epitope but rather utilized a distinct T-cell receptor Vbeta repertoire. Additional studies of chronically HIV-1-infected individuals expressing HLA-A11 demonstrated that the majority of the subjects targeted the G(357)S escape variant of the Gag(349-359) epitope, while the wild-type consensus sequence was significantly less frequently recognized. These data demonstrate that de novo responses against escape variants of CD8(+) T-cell epitopes can be generated in chronic HIV-1 infection and provide the rationale for developing vaccines to induce CD8(+) T-cell responses directed against both the wild-type and variant forms of CD8 epitopes to prevent the emergence of cytotoxic T-lymphocyte escape variants.

The majority of currently circulating human immunodeficiency virus type 1 clade B viruses fail to prime cytotoxic T-lymphocyte responses against an otherwise immunodominant HLA-A2-restricted epitope: implications for vaccine design.

Human immunodeficiency virus type 1 (HIV-1) mutates to escape immune selection pressure, but there is little evidence of selection mediated through HLA-A2, the dominant class I allele in persons infected with clade B virus. Moreover, HLA-A2-restricted responses are largely absent in the acute phase of infection as the viral load is being reduced, suggesting that circulating viruses may lack immunodominant epitopes targeted through HLA-A2. Here we demonstrate an A2-restricted epitope within Vpr (Vpr59-67) that is targeted by acute-phase HIV-1-specific CD8+ T cells, but only in a subset of persons expressing HLA-A2. Individuals in the acute stage of infection with viruses containing the most common current sequence within this epitope (consensus sequence) were unable to mount epitope-specific T-cell responses, whereas subjects infected with the less frequent I60L variant all developed these responses. The I60L variant epitope was a stronger binder to HLA-A2 and was recognized by epitope-specific T cells at lower peptide concentrations than the consensus sequence epitope. These data demonstrate that HLA-A2 is capable of contributing to the acute-phase cytotoxic T-lymphocyte response in infected subjects, but that most currently circulating viruses lack a dominant immunogenic epitope presented by this allele, and suggest that immunodominant epitopes restricted by common HLA alleles may be lost as the epidemic matures.

Selective escape from CD8+ T-cell responses represents a major driving force of human immunodeficiency virus type 1 (HIV-1) sequence diversity and reveals constraints on HIV-1 evolution.

The sequence diversity of human immunodeficiency virus type 1 (HIV-1) represents a major obstacle to the development of an effective vaccine, yet the forces impacting the evolution of this pathogen remain unclear. To address this issue we assessed the relationship between genome-wide viral evolution and adaptive CD8+ T-cell responses in four clade B virus-infected patients studied longitudinally for as long as 5 years after acute infection. Of the 98 amino acid mutations identified in nonenvelope antigens, 53% were associated with detectable CD8+ T-cell responses, indicative of positive selective immune pressures. An additional 18% of amino acid mutations represented substitutions toward common clade B consensus sequence residues, nine of which were strongly associated with HLA class I alleles not expressed by the subjects and thus indicative of reversions of transmitted CD8 escape mutations. Thus, nearly two-thirds of all mutations were attributable to CD8+ T-cell selective pressures. A closer examination of CD8 escape mutations in additional persons with chronic disease indicated that not only did immune pressures frequently result in selection of identical amino acid substitutions in mutating epitopes, but mutating residues also correlated with highly polymorphic sites in both clade B and C viruses. These data indicate a dominant role for cellular immune selective pressures in driving both individual and global HIV-1 evolution. The stereotypic nature of acquired mutations provides support for biochemical constraints limiting HIV-1 evolution and for the impact of CD8 escape mutations on viral fitness.

Selection, transmission, and reversion of an antigen-processing cytotoxic T-lymphocyte escape mutation in human immunodeficiency virus type 1 infection.

Numerous studies now support that human immunodeficiency virus type 1 (HIV-1) evolution is influenced by immune selection pressure, with population studies showing an association between specific HLA alleles and mutations within defined cytotoxic T-lymphocyte epitopes. Here we combine sequence data and functional studies of CD8 T-cell responses to demonstrate that allele-specific immune pressures also select for mutations flanking CD8 epitopes that impair antigen processing. In persons expressing HLA-A3, we demonstrate consistent selection for a mutation in a C-terminal flanking residue of the normally immunodominant Gag KK9 epitope that prevents its processing and presentation, resulting in a rapid decline in the CD8 T-cell response. This single amino acid substitution also lies within a second HLA-A3-restricted epitope, with the mutation directly impairing recognition by CD8 T cells. Transmission of the mutation to subjects expressing HLA-A3 was shown to prevent the induction of normally immunodominant acute-phase responses to both epitopes. However, subsequent in vivo reversion of the mutation was coincident with delayed induction of new CD8 T-cell responses to both epitopes. These data demonstrate that mutations within the flanking region of an HIV-1 epitope can impair recognition by an established CD8 T-cell response and that transmission of these mutations alters the acute-phase CD8(+) T-cell response. Moreover, reversion of these mutations in the absence of the original immune pressure reveals the potential plasticity of immunologically selected evolutionary changes.