Sample average treatment effect on the treated (SATT) analysis using counterfactual explanation identifies BMT and SARS-CoV-2 vaccination as protective risk factors associated with COVID-19 severity and survival in patients with multiple myeloma.

Patients with multiple myeloma (MM), an age-dependent neoplasm of antibody-producing plasma cells, have compromised immune systems and might be at increased risk for severe COVID-19 outcomes. This study characterizes risk factors associated with clinical indicators of COVID-19 severity and all-cause mortality in myeloma patients utilizing NCATS' National COVID Cohort Collaborative (N3C) database. The N3C consortium is a large, centralized data resource representing the largest multi-center cohort of COVID-19 cases and controls nationwide (>16 million total patients, and >6 million confirmed COVID-19+ cases to date). Our cohort included myeloma patients (both inpatients and outpatients) within the N3C consortium who have been diagnosed with COVID-19 based on positive PCR or antigen tests or ICD-10-CM diagnosis code. The outcomes of interest include all-cause mortality (including discharge to hospice) during the index encounter and clinical indicators of severity (i.e., hospitalization/emergency department/ED visit, use of mechanical ventilation, or extracorporeal membrane oxygenation (ECMO)). Finally, causal inference analysis was performed using the Coarsened Exact Matching (CEM) and Propensity Score Matching (PSM) methods. As of 05/16/2022, the N3C consortium included 1,061,748 cancer patients, out of which 26,064 were MM patients (8,588 were COVID-19 positive). The mean age at COVID-19 diagnosis was 65.89 years, 46.8% were females, and 20.2% were of black race. 4.47% of patients died within 30 days of COVID-19 hospitalization. Overall, the survival probability was 90.7% across the course of the study. Multivariate logistic regression analysis showed histories of pulmonary and renal disease, dexamethasone, proteasome inhibitor/PI, immunomodulatory/IMiD therapies, and severe Charlson Comorbidity Index/CCI were significantly associated with higher risks of severe COVID-19 outcomes. Protective associations were observed with blood-or-marrow transplant/BMT and COVID-19 vaccination. Further, multivariate Cox proportional hazard analysis showed that high and moderate CCI levels, International Staging System (ISS) moderate or severe stage, and PI therapy were associated with worse survival, while BMT and COVID-19 vaccination were associated with lower risk of death. Finally, matched sample average treatment effect on the treated (SATT) confirmed the causal effect of BMT and vaccination status as top protective factors associated with COVID-19 risk among US patients suffering from multiple myeloma. To the best of our knowledge, this is the largest nationwide study on myeloma patients with COVID-19.

Single-Cell Proteomics and Tumor RNAseq Identify Novel Pathways Associated With Clofazimine Sensitivity in PI- and IMiD- Resistant Myeloma, and Putative Stem-Like Cells.

Multiple myeloma (MM) is an incurable plasma cell malignancy with dose-limiting toxicities and inter-individual variation in response/resistance to the standard-of-care/primary drugs, proteasome inhibitors (PIs), and immunomodulatory derivatives (IMiDs). Although newer therapeutic options are potentially highly efficacious, their costs outweigh the effectiveness. Previously, we have established that clofazimine (CLF) activates peroxisome proliferator-activated receptor-γ, synergizes with primary therapies, and targets cancer stem-like cells (CSCs) in drug-resistant chronic myeloid leukemia (CML) patients. In this study, we used a panel of human myeloma cell lines as in vitro model systems representing drug-sensitive, innate/refractory, and clonally-derived acquired/relapsed PI- and cereblon (CRBN)-negative IMiD-resistant myeloma and bone marrow-derived CD138+ primary myeloma cells obtained from patients as ex vivo models to demonstrate that CLF shows significant cytotoxicity against drug-resistant myeloma as single-agent and in combination with PIs and IMiDs. Next, using genome-wide transcriptome analysis (RNA-sequencing), single-cell proteomics (CyTOF; Cytometry by time-of-flight), and ingenuity pathway analysis (IPA), we identified novel pathways associated with CLF efficacy, including induction of ER stress, autophagy, mitochondrial dysfunction, oxidative phosphorylation, enhancement of downstream cascade of p65-NFkB-IRF4-Myc downregulation, and ROS-dependent apoptotic cell death in myeloma. Further, we also showed that CLF is effective in killing rare refractory subclones like side populations that have been referred to as myeloma stem-like cells. Since CLF is an FDA-approved drug and also on WHO's list of safe and effective essential medicines, it has strong potential to be rapidly re-purposed as a safe and cost-effective anti-myeloma drug.

Risk and Outcome of Breakthrough COVID-19 Infections in Vaccinated Patients With Cancer: Real-World Evidence From the National COVID Cohort Collaborative.

PURPOSE: To provide real-world evidence on risks and outcomes of breakthrough COVID-19 infections in vaccinated patients with cancer using the largest national cohort of COVID-19 cases and controls. METHODS: We used the National COVID Cohort Collaborative (N3C) to identify breakthrough infections between December 1, 2020, and May 31, 2021. We included patients partially or fully vaccinated with mRNA COVID-19 vaccines with no prior SARS-CoV-2 infection record. Risks for breakthrough infection and severe outcomes were analyzed using logistic regression. RESULTS: A total of 6,860 breakthrough cases were identified within the N3C-vaccinated population, among whom 1,460 (21.3%) were patients with cancer. Solid tumors and hematologic malignancies had significantly higher risks for breakthrough infection (odds ratios [ORs] = 1.12, 95% CI, 1.01 to 1.23 and 4.64, 95% CI, 3.98 to 5.38) and severe outcomes (ORs = 1.33, 95% CI, 1.09 to 1.62 and 1.45, 95% CI, 1.08 to 1.95) compared with noncancer patients, adjusting for age, sex, race/ethnicity, smoking status, vaccine type, and vaccination date. Compared with solid tumors, hematologic malignancies were at increased risk for breakthrough infections (adjusted OR ranged from 2.07 for lymphoma to 7.25 for lymphoid leukemia). Breakthrough risk was reduced after the second vaccine dose for all cancers (OR = 0.04; 95% CI, 0.04 to 0.05), and for Moderna's mRNA-1273 compared with Pfizer's BNT162b2 vaccine (OR = 0.66; 95% CI, 0.62 to 0.70), particularly in patients with multiple myeloma (OR = 0.35; 95% CI, 0.15 to 0.72). Medications with major immunosuppressive effects and bone marrow transplantation were strongly associated with breakthrough risk among the vaccinated population. CONCLUSION: Real-world evidence shows that patients with cancer, especially hematologic malignancies, are at higher risk for developing breakthrough infections and severe outcomes. Patients with vaccination were at markedly decreased risk for breakthrough infections. Further work is needed to assess boosters and new SARS-CoV-2 variants.

secDrug: a pipeline to discover novel drug combinations to kill drug-resistant multiple myeloma cells using a greedy set cover algorithm and single-cell multi-omics.

Multiple myeloma, the second-most common hematopoietic malignancy in the United States, still remains an incurable disease with dose-limiting toxicities and resistance to primary drugs like proteasome inhibitors (PIs) and Immunomodulatory drugs (IMiDs).We have created a computational pipeline that uses pharmacogenomics data-driven optimization-regularization/greedy algorithm to predict novel drugs ("secDrugs") against drug-resistant myeloma. Next, we used single-cell RNA sequencing (scRNAseq) as a screening tool to predict top combination candidates based on the enrichment of target genes. For in vitro validation of secDrugs, we used a panel of human myeloma cell lines representing drug-sensitive, innate/refractory, and acquired/relapsed PI- and IMiD resistance. Next, we performed single-cell proteomics (CyTOF or Cytometry time of flight) in patient-derived bone marrow cells (ex vivo), genome-wide transcriptome analysis (bulk RNA sequencing), and functional assays like CRISPR-based gene editing to explore molecular pathways underlying secDrug efficacy and drug synergy. Finally, we developed a universally applicable R-software package for predicting novel secondary therapies in chemotherapy-resistant cancers that outputs a list of the top drug combination candidates with rank and confidence scores.Thus, using 17AAG (HSP90 inhibitor) + FK866 (NAMPT inhibitor) as proof of principle secDrugs, we established a novel pipeline to introduce several new therapeutic options for the management of PI and IMiD-resistant myeloma.

Irg1/itaconate metabolic pathway is a crucial determinant of dendritic cells immune-priming function and contributes to resolute allergen-induced airway inflammation.

Itaconate is produced from the mitochondrial TCA cycle enzyme aconitase decarboxylase (encoded by immune responsive gene1; Irg1) that exerts immunomodulatory function in myeloid cells. However, the role of the Irg1/itaconate pathway in dendritic cells (DC)-mediated airway inflammation and adaptive immunity to inhaled allergens, which are the primary antigen-presenting cells in allergic asthma, remains largely unknown. House dust mite (HDM)-challenged Irg1-/- mice displayed increases in eosinophilic airway inflammation, mucous cell metaplasia, and Th2 cytokine production with a mechanism involving impaired mite antigen presentations by DC. Adoptive transfer of HDM-pulsed DC from Irg1-deficient mice into naïve WT mice induced a similar phenotype of elevated type 2 airway inflammation and allergic sensitization. Untargeted metabolite analysis of HDM-pulsed DC revealed itaconate as one of the most abundant polar metabolites that potentially suppress mitochondrial oxidative damage. Furthermore, the immunomodulatory effect of itaconate was translated in vivo, where intranasal administration of 4-octyl itaconate 4-OI following antigen priming attenuated the manifestations of HDM-induced airway disease and Th2 immune response. Taken together, these data demonstrated for the first time a direct regulatory role of the Irg1/itaconate pathway in DC for the development of type 2 airway inflammation and suggest a possible therapeutic target in modulating allergic asthma.

Short palate, lung, and nasal epithelial clone 1 (SPLUNC1) level determines steroid-resistant airway inflammation in aging.

Asthma and its heterogeneity change with age. Increased airspace neutrophil numbers contribute to severe steroid-resistant asthma exacerbation in the elderly, which correlates with the changes seen in adults with asthma. However, whether that resembles the same disease mechanism and pathophysiology in aged and adults is poorly understood. Here, we sought to address the underlying molecular mechanism of steroid-resistant airway inflammation development and response to corticosteroid (Dex) therapy in aged mice. To study the changes in inflammatory mechanism, we used a clinically relevant treatment model of house-dust mite (HDM)-induced allergic asthma and investigated lung adaptive immune response in adult (20-22 wk old) and aged (80-82 wk old) mice. Our result indicates an age-dependent increase in airway hyperresponsiveness (AHR), mixed granulomatous airway inflammation comprising eosinophils and neutrophils, and Th1/Th17 immune response with progressive decrease in frequencies and numbers of HDM-bearing dendritic cells (DC) accumulation in the draining lymph node (DLn) of aged mice as compared with adult mice. RNA-Seq experiments of the aged lung revealed short palate, lung, and nasal epithelial clone 1 (SPLUNC1) as one of the steroid-responsive genes, which progressively declined with age and further by HDM-induced inflammation. Moreover, we found increased glycolytic reprogramming, maturation/activation of DCs, the proliferation of OT-II cells, and Th2 cytokine secretion with recombinant SPLUNC1 (rSPLUNC1) treatment. Our results indicate a novel immunomodulatory role of SPLUNC1 regulating metabolic adaptation/maturation of DC. An age-dependent decline in the SPLUNC1 level may be involved in developing steroid-resistant airway inflammation and asthma heterogeneity.

In vitro and ex vivo gene expression profiling reveals differential kinetic response of HSPs and UPR genes is associated with PI resistance in multiple myeloma.

Extensive inter-individual variation in response to chemotherapy (sensitive vs resistant tumors) is a serious cause of concern in the treatment of multiple myeloma (MM). In this study, we used human myeloma cell lines (HMCLs), and patient-derived CD138+ cells to compare kinetic changes in gene expression patterns between innate proteasome inhibitor (PI)-sensitive and PI-resistant HMCLs following test dosing with the second-generation PI Ixazomib. We found 1553 genes that changed significantly post treatment in PI-sensitive HMCLs compared with only seven in PI-resistant HMCLs (p < 0.05). Genes that were uniquely regulated in PI-resistant lines were RICTOR (activated), HNF4A, miR-16-5p (activated), MYCN (inhibited), and MYC (inhibited). Ingenuity pathway analysis (IPA) using top kinetic response genes identified the proteasome ubiquitination pathway (PUP), and nuclear factor erythroid 2-related factor 2 (NRF2)-mediated oxidative stress response as top canonical pathways in Ix-sensitive cell lines and patient-derived cells, whereas EIF2 signaling and mTOR signaling pathways were unique to PI resistance. Further, 10 genes were common between our in vitro and ex vivo post-treatment kinetic PI response profiles and Shaughnessy's GEP80-postBz gene expression signature, including the high-risk PUP gene PSMD4. Notably, we found that heat shock proteins and PUP pathway genes showed significant higher upregulation in Ix-sensitive lines compared with the fold-change in Ix-resistant myelomas.

Molecular signatures of multiple myeloma progression through single cell RNA-Seq.

We used single cell RNA-Seq to examine molecular heterogeneity in multiple myeloma (MM) in 597 CD138 positive cells from bone marrow aspirates of 15 patients at different stages of disease progression. 790 genes were selected by coefficient of variation (CV) method and organized cells into four groups (L1-L4) using unsupervised clustering. Plasma cells from each patient clustered into at least two groups based on gene expression signature. The L1 group contained cells from all MGUS patients having the lowest expression of genes involved in the oxidative phosphorylation, Myc targets, and mTORC1 signaling pathways (p < 1.2 × 10-14). In contrast, the expression level of these pathway genes increased progressively and were the highest in L4 group containing only cells from MM patients with t(4;14) translocations. A 44 genes signature of consistently overexpressed genes among the four groups was associated with poorer overall survival in MM patients (APEX trial, p < 0.0001; HR, 1.83; 95% CI, 1.33-2.52), particularly those treated with bortezomib (p < 0.0001; HR, 2.00; 95% CI, 1.39-2.89). Our study, using single cell RNA-Seq, identified the most significantly affected molecular pathways during MM progression and provided a novel signature predictive of patient prognosis and treatment stratification.

A de novo splice site mutation in EHMT1 resulting in Kleefstra syndrome with pharmacogenomics screening and behavior therapy for regressive behaviors.

BACKGROUND: Kleefstra syndrome (KS) is a rare autosomal dominant developmental disability, caused by microdeletions or intragenic mutations within the epigenetic regulator gene EHMT1 (euchromatic histone lysine N-methyltransferase 1). In addition to common features of autism, young adult regressive behaviors have been reported. However, the genetic downstream effects of the reported deletions or mutations on KS phenotype have not yet been completely explored. While genetic backgrounds affecting drug metabolism can have a profound effect on therapeutic interventions, pharmacogenomic variations are seldom considered in directing psychotropic therapies. METHODS: In this report, we used next-generation sequencing (exome sequencing and high-throughput RNA sequencing) in a patient and his parents to identify causative genetic variants followed by pharmacogenomics-guided clinical decision-making for making positive changes toward his treatment strategies. The patient had an early autism diagnosis and showed significant regressive behaviors and physical aberrations at age 23. RESULTS: Exome sequencing identified a novel, de novo splice site variant NM_024757.4: c.2750-1G>T in EHMT1, a candidate gene for Kleefstra syndrome, in the patient that results in exon skipping and downstream frameshift and termination. Gene expression results from the patient showed, when compared to his parents, there was a significant decreased expression of several reported gene variants associated with autism risk. Further, using a pharmacogenomics genotyping panel, we discovered that the patient had the CYP2D6 nonfunctioning variant genotype *4/*4 that results in very low metabolic activity on a number of psychotropic drugs, including fluvoxamine which he was prescribed. As reported here, a change in psychotropic drugs and intense behavior therapies resulted in a significant reversal of the regressive behaviors and physical aberrations. CONCLUSION: These results demonstrate an individualized approach that integrated genetic information and behavior therapies, resulting in a dramatic improvement in regressive behaviors associated with KS.

Single cell sequencing reveals heterogeneity within ovarian cancer epithelium and cancer associated stromal cells.

OBJECTIVES: The purpose of this study was to determine the level of heterogeneity in high grade serous ovarian cancer (HGSOC) by analyzing RNA expression in single epithelial and cancer associated stromal cells. In addition, we explored the possibility of identifying subgroups based on pathway activation and pre-defined signatures from cancer stem cells and chemo-resistant cells. METHODS: A fresh, HGSOC tumor specimen derived from ovary was enzymatically digested and depleted of immune infiltrating cells. RNA sequencing was performed on 92 single cells and 66 of these single cell datasets passed quality control checks. Sequences were analyzed using multiple bioinformatics tools, including clustering, principle components analysis, and geneset enrichment analysis to identify subgroups and activated pathways. Immunohistochemistry for ovarian cancer, stem cell and stromal markers was performed on adjacent tumor sections. RESULTS: Analysis of the gene expression patterns identified two major subsets of cells characterized by epithelial and stromal gene expression patterns. The epithelial group was characterized by proliferative genes including genes associated with oxidative phosphorylation and MYC activity, while the stromal group was characterized by increased expression of extracellular matrix (ECM) genes and genes associated with epithelial-to-mesenchymal transition (EMT). Neither group expressed a signature correlating with published chemo-resistant gene signatures, but many cells, predominantly in the stromal subgroup, expressed markers associated with cancer stem cells. CONCLUSIONS: Single cell sequencing provides a means of identifying subpopulations of cancer cells within a single patient. Single cell sequence analysis may prove to be critical for understanding the etiology, progression and drug resistance in ovarian cancer.

LIG1 polymorphisms: the Indian scenario.

Elucidation of the genetic diversity and relatedness of the subpopulations of India may provide a unique resource for future analysis of genetic association of several critical community-specific complex diseases.We performed a comprehensive exploration of single nucleotide polymorphisms (SNPs) within the gene DNA ligase 1 (LIG1) among a multiethnic panel of Indian subpopulations representative of the ethnic, linguistic and geographical diversity of India using a two-stage design involving DNA resequencing-based SNP discovery followed by SNP validation using sequenom-based genotyping. Thirty SNPs were identified in LIG1 gene using DNA resequencing including three promoter SNPs and one coding SNP. Following SNP validation, the SNPs rs20580/C19008A and rs3730862/C8804T were found to have the most widespread prevalence with noticeable variations in minor allele frequencies both between the Indian subpopulation groups and also from those reported on other major world populations. Subsequently, SNPs found in Indian subpopulations were analysed using bioinformatics-based approaches and compared with SNP data available on major world populations. Further, we also performed genotype-phenotype association analysis of LIG1 SNPs with publicly available data on LIG1 mRNA expression in HapMap samples. Results showed polymorphisms in LIG1 affect its expression and may therefore change its function. Our results stress upon the uniqueness of the Indian population with respect to the worldwide scenario and suggest that any epidemiological study undertaken on the global population should take this distinctiveness in consideration and avoid making generalized conclusions.

Protective association exhibited by the single nucleotide polymorphism (SNP) rs1052133 in the gene human 8-oxoguanine DNA glycosylase (hOGG1) with the risk of squamous cell carcinomas of the head & neck (SCCHN) among north Indians.

BACKGROUND & OBJECTIVES: Imbalances in compactly regulated DNA repair pathways in the form of single nucleotide polymorphisms (SNPs) within vital DNA repair genes may result in insufficient DNA repair and increase in DNA breaks thus rendering the human system vulnerable to the debilitatory effects of grave diseases like cancers. The present study involves investigation of association of the non-synonymous SNP rs1052133 (C8069G/Ser326Cys) located in the exonic region of the gene human 8-oxoguanine DNA glycosylase (hOGG1) with the risk of squamous cell carcinomas of the head and neck (SCCHN). METHODS: Case-control based genetic association study was performed among 575 (250 SCCHN cases and 325 normal healthy controls) sub-population cluster-matched (Indo-Europeans linguistic subgroup + Caucasoid morphological subtype) samples from the north Indian States of Uttar Pradesh and Uttarakhand using polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP) and DNA sequencing analysis. RESULTS: Our results demonstrated statistically significant protective association for the heterozygous CG [Odds Ratio (OR) 0.6587, 95% Confidence Interval (CI) 0.4615 to 0.9402, P=0.0238], homozygous mutant GG (OR 0.2570, 95% CI 0.1070 to 0.6175, P=0.0013) and combined mutant CG + GG (OR 0.6057, 95% CI 0.4272 to 0.8586, P=0.0059) genotypes. INTERPRETATION & CONCLUSIONS: The results indicate that the polymorphism rs1052133 is strongly associated with SCCHN susceptibility and the mutant (G) allele might be a protective factor for SCCHN among north Indian subpopulations.

Statistically significant association of the single nucleotide polymorphism (SNP) rs13181 (ERCC2) with predisposition to Squamous Cell Carcinomas of the Head and Neck (SCCHN) and Breast cancer in the north Indian population.

BACKGROUND: Non-synonymous single nucleotide polymorphisms (SNPs) within vital DNA repair genes may cause reduction of activity leaving the genome unrepaired resulting in genomic instability and cancer. MATERIALS AND METHODS: The present endeavour involved study on the association of the SNP rs13181 (Lys751Gln/A18911C) in the Nucleotide Excision Repair (NER) pathway gene ERCC2 (excision repair cross-complementing rodent repair deficiency, complementation group 2) with the risks of Squamous Cell Carcinomas of the Head and Neck (SCCHN) and Breast cancer using a case-control based association study among 685 (400 controls and 285 SCCHN-affected cases) and 395 (227 normal healthy female controls and 168 breast cancer cases) ethnically-matched samples, respectively from north India using Polymerase Chain Reaction followed by Restriction Fragment Length Polymorphism (PCR-RFLP) analysis. RESULTS: Results showed significant association of rs13181 homozygous mutant (CC) [Odds Ratio (OR) 4.412, 95% Confidence Interval (CI) 2.413 to 8.068], heterozygous (AC) (OR 2.086, 95% CI 1.246 to 3.492) and combined mutant (AC + CC) (OR 2.672, 95% CI 1.647 to 4.334) genotypes with predisposition to Breast cancer. Statistically significant increase in SCCHN risk was also associated with the mutant genotypes of rs13181 (ERCC2), viz. homozygous mutant (CC) (OR 1.680, 95% CI 1.014 to 2.784), heterozygous (AC) (OR 1.531, 95% CI 1.092 to 2.149) and combined mutant (AC + CC) (OR 1.560, 95% CI 1.128 to 2.158) genotypes. CONCLUSION: The results of this case-control study indicate that the polymorphism rs13181 might be a risk factor for predisposition towards SCCHN and breast cancer among north Indian subpopulations.

Association of polymorphisms in base excision repair genes with the risk of breast cancer: a case-control study in North Indian women.

Inheritance of common genetic variants at one or more base excision repair (BER) genes may result in a reduced DNA repair capacity and in an increased risk of cancers like breast cancer. The present case-control study with 390 north Indian women (155 breast cancer cases and 235 controls) was aimed to investigate the association of seven nonsynonymous BER gene polymorphisms viz. rs1130409/T1865G (APEX1), rs1799782/T22142C (XRCC1), rs25487/G23990A (XRCC1), rs4989588/T3337A (FEN1), rs4989586/ G3259A (FEN1), rs4989587/C3315T (FEN1), and rs1050525/G6941T (PCNA) with breast cancer susceptibility. Statistically significant association with breast cancer risk was observed for rs1130409 homozygous mutant GG [odds ratio (OR) 3.35, 95% confidence interval (CI) 1.36-8.26), heterozygous GT (OR 2.42, 95% CI 1.56-3.76), and combined mutant (GT + GG) (OR 2.52, 95% CI 1.65-3.86] genotypes and rs25487 homozygous mutant AA (OR 2.91, 95% CI 1.66-5.10) and combined mutant (AA + AG) (OR 1.41, 95% CI 0.903-2.19) genotypes, whereas protective association was exhibited by rs1799782 homozygous mutant CC (OR 0.413, 95% CI 0.082-2.08), heterozygous TC (OR 0.351, 95% CI 0.189-0.650), and combined mutant (TC + CC) (OR 0.357, 95% CI 0.199-0.641) genotypes. Association study using reconstructed haplotypes of XRCC1 gene showed positive association for the TA haplotype (OR 2.014, 95% CI 1.462-2.775) and a protective association for the CG haplotype (OR 0.173, 95% CI 0.052-0.576) pertaining to breast cancer risk. The results indicate that the polymorphisms rs1130409 (APEX1) and rs25487 (XRCC1) might be involved in contributing towards breast cancer susceptibility, while rs1799782 (XRCC1) might have protective influence.

Association of CYP1A1 polymorphisms with breast cancer in North Indian women.

Cytochrome P-450 (CYP) 1A1 is a candidate gene for low penetrance breast cancer (BC) susceptibility. Evidences demonstrate that ethnic differences in BC incidence may be partly due to genetic factors, including polymorphisms in the genes. In the present case control study four CYP1A1 gene polymorphisms, m1 (T6235C), m2 (A4889G), m3 (T5639C), and m4 (C4887A) were studied for their association with BC conjointly with the known risk factors such as age, menopausal status, diet, and life style. Polymorphisms of CYP1A1 gene were detected by PCR-RFLP method. The homozygous mutant (G/G) of m2 polymorphism was significantly associated with BC. Consequently, association of both m2 heterozygous mutant genotype (A/G) and combined group [homozygous (G/G) plus heterozygous (A/G) mutant genotype] showed association with postmenopausal women. Incidences of BC were also found to be independent of clinicopathological factors except heterozygous mutant genotype (A/G) m2 showed association with dietary factors and high grade tumors while homozygous mutant (G/G) m2 showed association with ER/PR-positive BC cases. Wild-type m3 was observed in all the subjects in cases as well as in controls. No significant association was observed between m1 and m3 polymorphisms and BC risk in all the subjects as well as when stratified into pre- and postmenopausal subjects. This indicates that out of ml and m2 polymorphisms that have been reported in Asians, only m2 is associated with North Indians.