MicroRNAs for Modulating Herpes Virus Gene Expression

A complex comprising an mRNA hybridization target to which is hybridized a miRNA or siRNA derivative thereof, wherein the hybridization of the miRNA or siRNA derivative thereof to the mRNA hybridization target is predicted by a method comprising the steps of:

• a) providing one or more databases comprising selected miRNA sequences and sequences representing 3′ untranslated regions (3′UTRs) of the population of mRNA molecules;
• b) determining one or more seed oligomers for each of the selected miRNA molecules;
• c) computing the probability (p) of finding an oligomer complementary to a seed oligomer at any position of a random background sequence generated using a kth order Markov model based on the sequence composition of the 3′ UTRs;
• d) counting the number (c) of occurrences of an oligomer in each 3′UTR that is complementary to a seed oligomer, thereby creating a collection of miRNA-3′UTR pairs;
• e) providing a score for each miRNA-3′UTR pair, wherein the score is determined by a single hypothesis p-value PVSH of a binomial distribution, computed by
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• f) ranking the miRNA-3′UTR pairs according to their score PVSH, wherein the highest rank corresponds to the smallest PVSH;
• g) evaluating the statistical significance of the t highest-ranking microRNA-target pairs, wherein t is an integer number between 1 and the total number of pairs tested, by generating N random genomes analogous to the selected genome, wherein each random genome comprises the same number of 3′UTRs as the selected genome, and each corresponding 3′UTR is of the same length and is based on the same kth Markov model as the corresponding 3′UTR in the selected genome;
• h) repeating steps c) through f) for each of the N random genomes;
• i) evaluating the statistical significance of the t highest-ranking miRNA-3′UTR pairs from step f) for the selected genome by (1) counting the number Nt of the randomly generated genomes in which the tth pair exhibits PVSH smaller than the tth pair in the selected genome and (2) computing the p-value PVMH(t) corrected for Multiple Hypothesis Testing from the formula
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• wherein PVMH(t) is the probability of finding higher scores for the t highest-ranking miRNA-3′UTR pairs in the random genome as compared with the selected genome;
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• j) identifying the miRNA hybridization targets by assessing each PVMH(t), wherein a smaller PVMH(t), correlates with a higher probability that the predicted targets are miRNA hybridization targets.

A siRNA or a chemically modified analog of a miRNA, which hybridizes with one or more mRNA targets selected from:

• a) HSV 3′UTRs RL1 (ICP 34.5), RL2 (ICP0), UL1, UL2, UL5, UL9, UL11, UL13, UL14, UL16, UL20, UL24, UL34, UL35, UL37, UL39, UL42, UL47, UL49A, UL51, UL52, US1 (US1.5, ICP22), US8, US8A, US9, US11, or US12 (ICP47);
• b) EBV 3′UTRs BALF2, BALF3, BALF5, BARF0, BaRF1, BARF1, BBLF4, BDLF 3.5, BDLF4, BFRF2, BGLF1, BGLF2, BGLF3, BGLF 3.5, BHLF1, BHRF1, BLLF3, BMRF1, BNRF1, BOLF1, BRLF1, BSLF2/BMLF1, BVLF1, BXLF1, BXRF1, BZLF1, BZLF2, LF3, LMP-1, LMP-2A, or LMP-2B;
• c) HCMV 3′UTRs IE1 (UL123), IE2 (UL122), RL1, RL10, UL3, UL16, UL17, UL20, UL26, UL29, UL31, UL32, UL33, UL34, UL37, UL38, UL40, UL43, UL44, UL45, UL50, UL51, UL52, UL54, UL57, UL60, UL61, UL67, UL69, UL78, UL79, UL80, UL86, UL87, UL91, UL92, UL95, UL97, UL98, UL10, UL103, UL105, UL107, UL112-113, UL117, UL120, UL137, UL141a, UL151, UL151a, UL153, US7, US10, US12, US14, US24, US26, US27, US28, New ORF1, or New ORF3;
• d) KSHV 3′UTRs ORF6, ORF7, ORF8, ORF9, ORF16, ORF18, ORF21, ORF25, ORF26, ORF28, ORF32, ORF40, ORF47, ORF49, ORF 50 (Rta), ORF56, ORF57, ORF58, ORF59, ORF63, ORF72, ORF73 (LANA), ORF74, ORF75, ORFK4, ORFK8 (Zta), ORFK13, and ORFK14;
• or
• e) VZV 3′UTRs ORF16, ORF47, ORF52, ORF55, ORF59, ORF61, or ORF62.

A vector comprising a polynucleotide which, when expressed in a mammalian cell, produces a transcript that is processed within the cell to form a miRNA or a siRNA derivative thereof, which is capable of binding to a viral 3′UTR selected from:

• a) HSV 3′UTRs RL1 (ICP 34.5), RL2 (ICP0), UL1, UL2, UL5, UL9, UL11, UL13, UL14, UL16, UL20, UL24, UL34, UL35, UL37, UL39, UL42, UL47, UL49A, UL51, UL52, US1 (US1.5, ICP22), US8, US8A, US9, US11, or US12 (ICP47);
• b) EBV 3′UTRs BALF2, BALF3, BALF5, BARF0, BaRF1, BARF1, BBLF4, BDLF 3.5, BDLF4, BFRF2, BGLF1, BGLF2, BGLF3, BGLF 3.5, BHLF1, BHRF1, BLLF3, BMRF1, BNRF1, BOLF1, BRLF1, BSLF2/BMLF1, BVLF1, BXLF1, BXRF1, BZLF1, BZLF2, LF3, LMP-1, LMP-2A, or LMP-2B;
• c) HCMV 3′UTRs IE1 (UL123), IE2 (UL122), RL1, RL10, UL3, UL16, UL17, UL20, UL26, UL29, UL31, UL32, UL33, UL34, UL37, UL38, UL40, UL43, UL44, UL45, UL50, UL51, UL52, UL54, UL57, UL60, UL61, UL67, UL69, UL78, UL79, UL80, UL86, UL87, UL91, UL92, UL95, UL97, UL98, UL100, UL103, UL105, UL107, UL112-113, UL117, UL120, UL137, UL141a, UL151, UL151a, UL153, US7, US10, US12, US14, US24, US26, US27, US28, New ORF1, or New ORF3;
• d) KSHV 3′UTRs ORF6, ORF7, ORF8, ORF9, ORF16, ORF18, ORF21, ORF25, ORF26, ORF28, ORF32, ORF40, ORF47, ORF49, ORF 50 (Rta), ORF56, ORF57, ORF58, ORF59, ORF63, ORF72, ORF73 (LANA), ORF74, ORF75, ORFK4, ORFK8 (Zta), ORFK13, and ORFK14;
• or
• e) VZV 3′UTRs ORF16, ORF47, ORF52, ORF55, ORF59, ORF61, or ORF62.

A pharmaceutical composition for treatment of herpes virus infection caused by HSV, EBV, HCMV, KSHV or VSV, comprising a pharmaceutical carrier and miRNA comprising one or more of:

• a) HSV miRNAs hsv1-miR-H1, or hsv1-miR-LAT;
• b) EBV miRNAs ebv-miR-BART1-3p, ebv-miR-BART1-5p, ebv-miR-BART2, ebv-miR-BART3-3p, ebv-miR-BART3-5p, ebv-miR-BART4, ebv-miR-BART5, ebv-miR-BART6-3p, ebv-miR-BART6-5p, ebv-miR-BART7, ebv-miR-BART8-3p, ebv-miR-BART8-5p, ebv-miR-BART9, ebv-miR-BART10, ebv-miR-BART11-3p, ebv-miR-BART11-5p, ebv-miR-BART12, ebv-miR-BART13, ebv-miR-BART14-3p, ebv-miR-BART14-5p, ebv-miR-BART15, ebv-miR-BART16, ebv-miR-BART17-3p, ebv-miR-BART17-5p, ebv-miR-BART18, ebv-miR-BART19, ebv-miR-BART20-3p, ebv-miR-BART20-5p, ebv-miR-BHRF1-1, ebv-miR-BHRF1-2*, or ebv-miR-BHRF1-3;
• c) HCMV miRNAs hcmv-miR-UL22-1, hcmv-miR-UL22A-1*, hcmv-miR-UL31-1, hcmv-miR-UL36-1, hcmv-miR-UL36-1-N, hcmv-miR-UL53-1, hcmv-miR-UL54-1, hcmv-miR-UL70-3p, hcmv-miR-UL70-5p, hcmv-miR-UL102-1, hcmv-miR-UL102-2, hcmv-miR-UL111a-1, hcmv-miR-UL112-1, hcmv-miR-UL148D-1, hcmv-miR-US4, hcmv-miR-US5-1, hcmv-miR-US5-2, hcmv-miR-US5-2-N, hcmv-miR-US25-1, hcmv-miR-US25-2-5p, hcmv-miR-US25-2-3p, hcmv-miR-US29-1, or hcmv-miR-US33-1;
• d) KSHV miRNAs kshv-miR-K12-1, kshv-miR-K12-2, kshv-miR-K12-3, kshv-miR-K12-3*, kshv-miR-K12-4-5p, kshv-miR-K12-4-3p, kshv-miR-K12-5, kshv-miR-K12-6-5p, kshv-miR-K12-6-3p, kshv-miR-K12-7, kshv-miR-K12-8, kshv-miR-K12-9*, kshv-miR-K12-9, kshv-miR-K12-10a, kshv-miR-K12-10b, kshv-miR-K12-11, or kshv-miR-K12-12;
• or
• e) human miRNAs
• (i) targeting HSV:
• hsa-miR-138, hsa-miR-205, hsa-miR-326, hsa-miR-381, hsa-miR-425, hsa-miR-492, or hsa-miR-522;
• (ii) targeting EBV:
• hsa-miR-24, hsa-miR-214, hsa-miR-296, hsa-miR-328, hsa-miR-346, or hsa-miR-502;
• (iii) targeting HCMV:
• hsa-miR-15a, hsa-miR-15b, hsa-miR-16, hsa-miR-103, hsa-miR-107, hsa-miR-126, hsa-miR-142-5p, hsa-miR-184, hsa-miR-194, hsa-miR-195, hsa-miR-200b, hsa-miR-200c, hsa-miR-202, hsa-miR-326, hsa-miR-330-5p, hsa-miR-367, hsa-miR-424, hsa-miR-429, hsa-miR-450-b-3p, hsa-miR-497, hsa-miR-503, hsa-miR-548d-3p, hsa-miR-548k, hsa-miR-551a, hsa-miR-551b, hsa-miR-552, hsa-miR-592, hsa-miR-598, hsa-miR-652, hsa-miR-769-3-p, or hsa-miR-1226;
• (iv) targeting KSHV:
• hsa-let-7a, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f, hsa-let-7g, hsa-let-7i, hsa-miR-1, hsa-miR-9, hsa-miR-15a, hsa-miR-15b, hsa-miR-16, hsa-miR-17-5p, hsa-miR-18a, hsa-miR-18b, hsa-miR-20a, hsa-miR-20b, hsa-miR-23a, hsa-miR-23b, hsa-miR-30a-5p, hsa-miR-30a-3p, hsa-miR-30b, hsa-miR-30c, hsa-miR-30e-5p, hsa-miR-30e-3p, hsa-miR-93, hsa-miR-98, hsa-miR-105, hsa-miR-106a, hsa-miR-106b, hsa-miR-125a, hsa-miR-125b, hsa-miR-129, hsa-miR-134, hsa-miR-137, hsa-miR-141, hsa-miR-142-3p, hsa-miR-145, hsa-miR-150, hsa-miR-154, hsa-miR-181a, hsa-miR-181b, hsa-miR-181c, hsa-miR-181d, hsa-miR-182*, hsa-miR-194, hsa-miR-195, hsa-miR-196a, hsa-miR-196b, hsa-miR-199a, hsa-miR-199b, hsa-miR-200a, hsa-miR-205, hsa-miR-206, hsa-miR-210, hsa-miR-213, hsa-miR-299-3p, hsa-miR-302a, hsa-miR-302b, hsa-miR-302c, hsa-miR-302d, hsa-miR-324-3p, hsa-miR-326, hsa-miR-329, hsa-miR-337, hsa-miR-338, hsa-miR-340, hsa-miR-346, hsa-miR-372, hsa-miR-373, hsa-miR-424, hsa-miR-448, hsa-miR-450, hsa-miR-453, hsa-miR-455, hsa-miR-490, hsa-miR-491, hsa-miR-492, hsa-miR-497, hsa-miR-518b, hsa-miR-518c, hsa-miR-518d, hsa-miR-519d, hsa-miR-520a, hsa-miR-520b, hsa-miR-520c, hsa-miR-520d, hsa-miR-520g, hsa-miR-520h, hsa-miR-525, or hsa-miR-526b;
• or
• (v) targeting VZV:
• hsa-miR-99a, hsa-miR-99b, hsa-miR-100, hsa-miR-124a, hsa-miR-132, hsa-miR-141, hsa-miR-150, hsa-miR-197, hsa-miR-200a, hsa-miR-212, hsa-miR-219, hsa-miR-330, hsa-miR-374, hsa-miR-371, hsa-miR-339, hsa-miR-451, hsa-miR-495, and hsa-miR-510.